involvement of the caspase enzymes in satellite cells

Volume 5, Issue 3J Tissue Sci Eng 2014

ISSN: 2157-7552, JTSE an open access journalTissue Science-2014

September 24-26, 2014

September 24-26, 2014 Valencia Convention Centre, Spain

3rd International Conference on

Tissue Science & Regenerative MedicineInvolvement of the caspase enzymes in satellite cells apoptosisRoberta Di PietroG. d’Annunzio University of Chieti-Pescara, Italy

Ageing is characterized by an impairment of muscle regenerative potential and a progressive loss of skeletal muscle. This condition, called sarcopenia, has important health care implications for humans, as it contributes to frailty, functional

loss and premature death. The ability of skeletal muscle to regenerate is owed to a population of myogenic stem cells called satellite cells (SCs). In previous papers of our research group we demonstrated an age-related decrease in the antioxidant capacity of human SCs that may negatively affect the ageing SCs ability to repair muscle. Despite the involvement of caspases in muscle wasting, the real role of these enzymes is still controversial. To test the possibility of caspases involvement in SCs death in human ageing muscles, we cultured in vitro primary cells derived from vastus lateralis of young and aged subjects. We analysed both initiator caspases and effector caspases in the presence or absence of specific or broad pharmacologic inhibitors. Furthermore, we evaluated the expression of various genes that play a critical role in oxidative stress and cell death. Our findings highlighted an increased rate of spontaneous apoptosis and an up-regulation of CASP2, 6, 7, 9 and other cell death genes in aged SCs, supporting the hypothesis of an intrinsic ageing of SCs and previous reports demonstrating an increased susceptibility to apoptosis of SCs from old animals. These results suggest that a greater proportion of SCs from old subjects might undergo programmed cell death in vivo in response to stressful stimuli, thereby impairing skeletal muscle regeneration.

BiographyRoberta Di Pietro got the degree in Medicine with Honors in 1985 and the PhD in Sports Medicine with Honors in 1988, University of Chieti, Italy. She worked as a visiting scientist in UK at the Biochemistry Department, AFRC, and Cambridge; in USA, at the Pathology Department, USUHS, Bethesda and at the Institute of Human Virology, University of Maryland at Baltimore. She currently works at the G. d’Annunzio University of Chieti, Italy, as a Full Professor of Histology. Since 2007 she joined the Editorial Board of Current Pharmaceutical Design as an Executive Guest Editor and since 2010 she was recognized as a Registered Referee for Archives of Ophthalmological Reviews and Reproductive Biology and Endocrinology. She is now author of 157 scientific publications plus international e-book and Italian textbook chapters.

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Roberta Di Pietro, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineCross linkers for heart valve tissue engineeringSomers PamelaGhent University, Belgium

Background: Currently, an effective cross linking reagent to treat xenogenic decellularized heart valve matrices is lacking. Matrices still elicit an intense cell-mediated immune response and calcification. The aim of this study was to evaluate the cross linking effect of Quercetin, Catechin, Caffeic Acid and Tannic acid on porcine aortic valve matrices.

Materials and methods: Cytotoxicity of the different cross linkers was evaluated. Mechanical properties of cross linked porcine matrices and control matrices (non-fixed) were examined by tensile strength testing. Cytocompatibility of the fixed matrices was examined. Cross linked and control matrices were implanted subcutaneously in Wistar rats (n=9). After 2 weeks the explants were examined by light microscopy. Calcium content was determined using inductively coupled plasma-mass spectrometry. Antibody reaction against porcine tissue in the rat serum was determined.

Results: Cytotoxicity studies demonstrated that cross linkers, even at high concentrations, did not inhibit cell viability. All cross linkers, except Tannic acid, improve mechanical strength of acellular porcine matrices. Moreover, tensile strength of Quercetin fixed matrices was comparable with glutaraldehyde-fixed leaflets. Light microscopic evaluation showed that cross linked matrices showed only a mild lymphocytic inflammatory reaction. Furthermore, Quercetin fixed leaflets exhibited a well preserved matrix without infiltration of CD3+ cells. Calcium levels after 2 weeks were for controls (non-fixed): 206.33µg/mg; Quercetin: 151.33µg/mg; Catechin: 181µg/mg and Caffeid acid fixed matrices: 163.66µg/mg.

Conclusion: Quercetin is the most suitable candidate for heart valve cross linking and could be used as alternative for glutaraldehyde. Whether, Quercetin allows for autologous cell repopulation in order to create a viable tissue engineered heart valve still needs to be investigated.

BiographyPamela Somers has completed her PhD in Medical Sciences at the age of 29 years from Ghent University. At the moment she performs postdoctoral research in cardiac tissue engineering at Ghent University. She has published more than 29 papers in reputed journals.

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Somers Pamela, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014

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Tissue Science & Regenerative MedicineNew chitosan-based scaffolds for peripheral nerve regeneration – Pre clinical studiesKirsten Haastert-TaliniHannover Medical School, Germany

There is a need for new conduits that could provide a replacement strategy to autologous nerve grafting in the reconstruction of peripheral nerve injuries with substance loss. The natural biopolymer chitosan has gained increasing interest in

biomedical and tissue engineering applications because of its biocompatibility, biodegradability and low toxicity. Initial cytotoxicity tests were performed in vitro and followed by bridging of 10 mm sciatic nerve gaps in adult rats with chitosan tubes. Out of medical grade chitosan the tubes were manufactured by a proprietary extrusion process followed by adjustment of different degrees of acetylation (DA): DAI (~2%), DAII (~5%), and DAIII (~ 20%). Multidisciplinary and comprehensive in vivo evaluations revealed that while growth factor regulation is not altered by chitosan tube implantation, DAI and DAIII tubes display reduced support of early regeneration events. Functional and structural regeneration was most similar to autologous nerve graft reconstruction when DAII chitosan tubes were used. Furthermore, DAIII tubes displayed much less stability and a too fast degradation (Haastert-Talini et al., Biomaterials 2013, Dec; 34(38):9886-904). Fine-tuned chitosan tubes with a DA of ~5% are currently processed for the development of more complex artificial nerve devices. The optimal luminal enrichment for the tubes is currently investigated again in vitro and in vivo. With luminal enrichment also the reconstruction of longer nerve defects will be addressable. Therefore, we investigate combinations of regeneration promoting hydrogels with primary or genetically modified Schwann cells or bone marrow derived mesenchymal stromal cells or growth-factor-conjugated-nanoparticles. The new results will be presented with this paper.

BiographyKirsten Haastert-Talini has completed her doctoral degree in 2002 from University of Veterinary Medicine Hannover, Germany, and postdoctoral studies from Hannover Medical School, Germany. She is Associate Professor at the Institute of Neuroanatomy, Hannover Medical School. Her research focuses on tissue engineering strategies for peripheral nerve repair. She has published more than 35 original and review articles in reputed journals. The work presented has received funding from the European Community’s Seventh Frame work Programme (FP7-HEALTH-2011) under grant agreement n° 278612 (BIOHYBRID, coordinator C. Grothe).

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Kirsten Haastert-Talini, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineApplication of bioactive hydrogels to the regeneration of tissues: A feasible approach to connective tissueJulio San RomanInstitute of Polymer Science & Technology, Spain

Acute wounds in normal, healthy individuals heal through an orderly sequence of physiological events that include hemostasis, inflammation, epithelialization, fibroplasia, and maturation. When this process is altered or stalled, a chronic

wound may develop and is more likely to occur in patients with underlying disorders such as peripheral artery disease, diabetes, venous insufficiency, nutritional deficiencies, and other disease states. Vascular diseases in combination with concomitant pathologies such as diabetes and/or cardiovascular or cerebrovascular disorders are frequent problems commonly treated in clinic. Vascular pathologies lead to cutaneous lesions in the lower limbs which are often complicated by ischemia. Actual treatments for venous and diabetic ulcers include the application of hydrogels in combination with cells or biological skin substitutes.

The application of biodegradable polymeric hydrogels offers the opportunity to contribute in one formulation to the activation of the healing processes in compromise healing processes. The present contribution deals with the design and application of new approaches to the healing with biodegradable hydrogels based on bilayered or multilayered membranes loaded with complementary and coordinate bioactive agents. The application bioactive compounds that promote the vascularization such as proteins like proadenomedulin PAMP, in connection with the activity produced by activators of growth factors such as heparin, have been loaded in a two layered polymeric dressing for application in complicated wounds. The respond of the connective or dermal tissue to the application of hydrophilic membranes is very positive, and for controlled degradation processes has been obtained an excellent response in cell cultures and in vivo experiments.

The experiments carried out in a normalized model using the ear of rabbits proved the excellent biocompatibility of the polymeric membrane, as well as the effect of the loaded PAM and heparin in the healing process. Results of the evolution of the healing process with a quantification of the contraction and epitelization as well as the control of inflammation in the animal model, demonstrate that any scaffold or support for cell proliferation and tissue regeneration has to offer not only the adequate medium for the regeneration of the natural extracellular matrix, but also a controlled system for the release in a targeting way of the element necessary to activate the regenerative processes. Data of experiments carried out with normal and ischemic tissues of rabbits will be presented and discussed considering the effect of the bioactive compounds loaded in the hydrogels in the regenerative processes.

BiographyJulio San Roman has completed his Doctor in Chemistry (Polymers Science) in 1975 and currently Heads the Group of Biomaterials, Instituto de Ciencia y Tecnología de Polímeros, Consejo Superior de Investigaciones Científicas, CSIC at Madrid, Spain. He is a Full Professor in the Consejo Superior de Investigaciones Científicas and a Member of the European Society for Biomaterials ESB, since 1987 and of the Council of the Society since 2006. He is the Fellow of the Biomaterials Societies since 2004. He held the President position of Iberian Society for Biomechanics and Biomaterials, Polymer Section of the Spanish Royal Society of Chemistry and European Polymer Federation. He has been a member of the Scientific Committee of ESB meetings in the last 10 years and the chairman of the European Polymer Congress EPF 2011 organised in Granada, Spain, and chairman of the 2013 congress of the European Society for Biomaterials organised in Madrid, Spain. The Scientific activities are centred in the study and development of polymer systems for biomedical applications, and specifically in Tissue Engineering, Polymer Drugs and Drug Delivery Systems. He has published more than 400 refereed articles in specialised journals of Polymer Science, Biomaterials, and the Biomedical field. He has contributed with more than 40 Chapters in specialised books and is co-editor of two books on biodegradable polymers for biomedical applications and Biomaterials.

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Julio San Roman, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThe use of autologous stem cells in the treatment of a soccer player with ankle AVNDennis M. LoxFlorida Spine Center, USA

The treatment of avascular necrosis (AVN) can be a difficult dilemma for the clinician. When avascular necrosis (AVN) occurs in a competitive athlete the situation can easily become the end of a sporting career. This case will reflect the

difficulties encountered when the early diagnosis of AVN is critical and various treatment options are suggested by different health care providers. The ramifications of these recommendations also impact upon the future goals of the athlete. A teenage soccer player injured her ankle while playing soccer. This was initially felt to be a simple ankle sprain that failed to resolve with conservative measures including, rest, ice, elevation and anti-inflammatories. Subsequent x-rays failed to reveal bony abnormalities. Running and playing soccer were unable to be continued secondary to pain and swelling. An MRI was obtained which revealed AVN of the navicular bone without joint or bone degeneration. Two orthopedic surgeons were consulted and both recommended discontinuing soccer and having an ankle fusion. This option was unacceptable to the patient and her family. Alternative strategies were explored and the option of utilizing autologous stem cells was considered. The patient and family consulted Dr. Dennis Lox and it was determined she was a reasonable candidate for autologous stem cell injection as an alternative treatment to having her ankle fused. This was done and she responded favorable. Her pain and swelling subsided. Repeat MRI’s were consistent with some resolution of her area of AVN. At 6 months she returned to competitive soccer play. The following year she was the leading scorer on her team and being scouted by Division I colleges. This case reflects the ability of Regenerative medicine to alter the consequences of irreversible surgical interventions. In this case, the surgical option of ankle fusion would have ended the patients sports career, and as a teenager girl, the psychosocial implications of an ankle fusion may have lasting effects.

BiographyDennis Lox practices Sports and Regenerative Medicine in the Tampa Bay, Florida and the Washington D.C. metro areas in the United States. Dr. Lox has treated numerous patients with AVN. As well as patients from all levels of sports competition including the professional ranks. Lox frequently presents his findings at national and International Conferences on Regenerative Medicine and Tissue Engineering.

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Dennis M. Lox, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineEpigentic factors involved in the “mineralizing/demineralizing” capability of cell phenotypes: Histone deacetylases (HDACS), transcription factors (TFs), microRNAs, and vitamin K2 (MK-7)Jan O. GordeladzeUniversity of Oslo, Norway

The Epigenator-Initiator-Maintainer axis determines the ultimate fate of the phenotype characteristics of the cellular functions within different organs of the body. The initiator signals (pertained within the realm of histone modifications,

transcriptional control through transcription factors, and microRNAs) represent slowly “dissipating” forces tilting the cell phenotype towards a more or less stable profile.

However, phenotypic characteristics may be altered, i.e. changed or reinforced, as a result of developing diseases or gene therapy. Here, we present some results of manipulations of HDACs, transcription factors (TFs), microRNAs, as well as vitamin K2 status on mineralizing cells (osteoblasts), and non-mineralizing cells (epithelial cells) expose to both normal growth conditions and inflammation mediators (Th-cells, macrophages or interleukins), showing that it is possible to engineer cells with a phenotype where: 1) wanted mineralization is reinforced, and 2) unwanted mineralization is blocked.

By using various algorithms, one may pinpoint regulatory loops, including TFs (e.g. SP1, SP3, and ETS1) and microRNAs (e.g. miRNAs 23, 27, 29, 133, 149, 204, 211, and 328). The subject TFs and MicroRNAs are also part of an intricate hierarchical regulation by several HDAC classes, including the Sirtuins, which respond to cellular rations of NADH/NAD+ (cellular energy status). Furthermore, it will be demonstrated how vitamin K2 (MK-7, via binding to SXR) interferes with several signaling pathways down-stream of several growth factor signaling mechanisms, thus either reinforcing or suppressing the mineralizing characteristics of different cell phenotypes.

BiographyJan O. Gordeladze, PhD (born 25th of April, 1950), holds a triple professor competence (medical biochemistry, physiology, and pharmacology), and is presently working as a Professor at the Department of Biochemistry, Institute of Basic Medical Science, University of Oslo, Norway. He has previously been employed as the medical director of MSD, Norway, serving two years as a Fulbright scholar at the NIH, Bethesda, Maryland, USA, and from 2006-2009 being employed as Associate Professor at the University of Montpellier, France. He has published more than 100 scientific articles, reviews/book chapters and presented more than 250 abstracts/posters/talks at conferences worldwide.

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Jan O. Gordeladze, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineUse of mesenchymal stem cells and scaffolds for improving tendon and skin regenerationMarco PatrunoUniversity of Padova, Italy

Tissue engineering techniques and cell-based therapies are commonly accepted modus operandi to treat frequently occurring conditions such as traumatic tendon rupture or skin damages since available remedies appear to be ineffective in restoring

the original structure and function of the injured tissue. Moreover, the mechanisms behind effectiveness of novel approaches in treating tendon and skin injuries remain poorly characterised. Our laboratory is interested i) in the characterization of adult mesenchymal stem cells (MSC) from different sources, ii) in studying the tenogenic differentiate pathway and iii) in the application of MSC on bio-scaffolds, taking into account that one of the major aim of regenerative medicine is to realize experimental techniques that take maximal advantage of natural reparative processes. For instance, the regenerative capabilities are formidable in marine invertebrates, especially thanks to their mutable connective tissue that can be isolated and used to prepare film and 3D scaffolds. Therefore, one of our research goals is to mix adult stem cells with an innovative and low-cost source of native intact collagen fibril since these matrices have the potential to be used particularly where high mechanical performances are required. It has also been studied the delivery and viability of cells injected in different kind of bio-scaffolds; in particular, two main types of cells were injected: undifferentiated cells (mesenchymal stem cells) or cells differentiated towards a specific fate with the use of TAT sequence. Here, I summarize and discuss the most significant results of our research obtained in vitro and in vivo, regarding the production of bio-scaffolds together with the use of adult stem cells for treating tendon and skin lesions.

BiographyMarco Patruno is an Associate Professor of the Department of Comparative Biomedicine & Food Science, University of Padova, Italy. Patruno obtained the degree of Animal Science from the University of Milano, Italy in 1995 and his PhD in Developmental Biology from Royal Holloway, University of London, UK, in 2001. He published several international papers and chapters in scientific books. Patruno teaches Veterinary Histology, Embryology and Anatomy and his interests are related to regenerative processes, muscle development and the potential of adult stem cells isolated from different.

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Marco Patruno, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineHealing parenchyma with mesenchyma: A new therapeutic paradigmRamon LlullSTEM Center, Spain

Nature heals damaged specialized tissues by mending torn tissues, feeding hungry tissues, containing lesive processes and replacing damaged units. These self healing functions are carried out by a developmentally defined group of cells

-the stromal cells- comprised by a hetereogeneous population of fibroblasts, endothelial cells, tissue resident mononuclear and progenitor cells. Stromal cells share common essential functions and processes, they also share a common mesenchymal ancestry and early developmental stages, as they remain located within the three-dimensional perivascular structure of different tissues. Indeed, the fact that the above stromal cell functions adapt to resident tissue specificicity unique properties holds true, as it is true the fact that such specificities have been constitutively determined by the surrounding parenchyma and they are not present during earlier differentiation stages. Personal observations and literature review provide ample evidence that stromal mesenchymal cells loose their tissue-specific functions following tissue dissociation when cultured in-vitro under non-specific media conditions; they regain their tissue-specific functions in response to their own specific culture media or when returned to their initial tissue source (orthotopic implantation); and express different tissue-specific functions when exposed to other tissue-specific media conditions or after heterotopic implantation. If so, then, a therapeutically useful question is: If such reparative functions were retained by stromal cells after tissue dissociation, cell selection and concentration:

Would these stromal cells express their essential reparative properties when exchanged among different muscle-skeletal tissues? For instance, would the abundant, expendable adipose derived stromal cells exert a quantifiable therapeutic effect when implanted in let's say muscle, or bone, or cartilage? Would they display reparative functions immediately upon engraftment or rather undergo in-vivo reprogramming (dedifferentiation to an earlier stage and redifferentiation) into tissue-specific supporting cells? If instead, these stromal cells did not express their essential reparative functions when delivered heterotopically: Would it be a common, practical, method to then obtain tissue specific stromal cells which could be reliably engrafted and exert a therapeutic effect following orthotopic implantation? Certainly, stromal cells occupy analogous hystological locations across different muscle-skeletal tissues, and remain integrated into tissues by common anchors, matrix structures, and intercellular bounds. This presentation attempts at unveiling answers to the above questions, thus supporting a new therapeutic paradigm: The Biospace of Cell Surgery evolves as a regulatory friendly therapeutic strategy enabled by surgical procurement of autologous cells, both terminally differentiated and progenitor, minimally processed, and immediately delivered to the same patient to exert the mesenchymal cell's therapeutic capacity for muscle-skeletal tissue disorders.

BiographyRamon Llull obtained his medical degree with honors at University of Navarra, Spain in 1988. His research on immunological aspects of musculo-skeletal tissue transplantation was awarded the Henry Christian Memorial Award for Excellence in Research from the Association of American Physicians, and sponsored by the Plastic Surgery Educational Foundation and the American Association for Hand Surgery. He subsequently entered a plastic surgery clinical training at the University of Pittsburgh residency program. During his training was awarded the 1994 young investigator award of the International Transplantation Society for his continuing studies on musculo-skeletal tissue immunology. He is founder of Stemsource Inc., and holds with Dr. Katz while at the University of Pittsburgh, several US and international patents related to tissue engineering, device biotechnology and stem cells. He is Joint founder, First President and Past-Chairman of the Board of Directors of the International Federation of Adipose Technologies and Science (IFATS). He is member of the Educational Committee of the Tissue Engineering Society, International. In 2009, he enabled the first cell vaccine for brain tumors in Europe as a consultant of Northwest Bio therapeutics (NWBT), a US based Biotechnology Company focused on cancer immunotherapy. Currently, he is founder of The GID Group, Ltd, and holds several patents describing cell surgery methods, devices and sources.

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Ramon Llull, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineRecellularization of vascular and tubular compartments of porcine kidney scaffolds with embryonic stem cells Marina FigliuzziUniversity of Bergamo, Italy

Chronic kidney disease (CKD) is a progressive condition marked by deteriorating kidney function over time. The therapeutic strategies for CKD are currently directed at limiting renal functional decline or at renal replacement therapies with either

dialysis or transplantation. Novel alternative tools help to address this shortcoming by regeneration of damaged kidney using natural occurring scaffolds seeded with stem cells. The aims of the present study were to produce porcine whole-kidney scaffolds by a decellularization process and to test the efficacy of scaffold recellularization using physiological perfusion condition. The efficacy of cellular removal and biocompatibility of the preserved porcine matrices, as well as scaffold reproducibility, are critical to the success of this approach. To this aim, we designed and custom built a chamber to allocate large organs for decellularization and subsequent cell seeding. Complete decellularization of porcine kidneys can be achieved after 48 hr perfusion with sodium dodecyl sulfate 0.5%, as documented by histologic and immunofluorescence findings confirming complete cell removal without loss of structural components. The whole-kidney scaffolds preserved the 3D architecture of vessels, glomeruli and tubuli as shown by SEM analysis. To regenerate functional tissue, we repopulated acellular porcine kidneys with mouse embryonic stem (mES) cells through the renal artery and through the ureter. To obtain cell seeding within the tubular compartment a negative pressure of -70 mmHg was applied to the perfusion chamber. H&E staining and immunofluorescence demonstrated that mES cells infused through the renal artery were uniformly distributed in the vasculature and in glomerular capillaries while cells infused through the collecting system reached tubular compartments. Our findings indicate that porcine kidney can be successfully decellularized to produce intact renal extracellular matrix scaffolds. We also demonstrate the ability of the perfusion protocol to obtain cell engraftment in vascular and tubular structures. Because porcine scaffolds have similar morphology and function as their human counterparts, they certainly represented a platform for clinically applicable renal bioengineering investigations.

BiographyMarina Figliuzzi graduated in Biological Sciences at the University of Milano (Milano, Italy) in 1991 and got specialization in Pharmacological Research at the Mario Negri Institute, Bergamo, Italy in 1994. From 1994-2000 she was a research investigator at the Mario Negri Institute for Pharmacological Research, Bergamo, Italy. From the 2000-2014 she is the Head of the Unit of Tissue Engineering, Department of Biomedical Engineering. From the 2014 she is the Head of the Laboratory of Tissue Engineering for Regenerative Medicine, Department of Biomedical Engineering. Main areas of interest are: Pancreatic cell transplantation in experimental models of insulin dependent diabetes; Immunoisolation devices for pancreatic islets; Differentiation of progenitor pancreatic cells in insulin containing cells; Immunhistochemistry; Cell culture; Organ decellularization.

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Marina Figliuzzi, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThe contribution of fibronectin to cell-induced tissue morphogenesisFoolen JasperETH Zurich, Switzerland

When engineering fibrous tissues, attaining an appropriate distribution of cells and matrix to ascertain in vivo functionality and durability remains a challenge. Since increased cell density towards tissue surfaces was recently correlated with the

distribution of fibronectin in 3D microtissues, we now asked how fibronectin impacts cell-induced tissue morphogenesis and heterogeneity.

To address this question, microtissues were engineered in a high-throughput model system containing rectangular arrays of 12 posts. Posts constrained fibroblast-populated collagen gels that self-assembled into trampoline-shaped tissues with dimensions of ~750x750x40µm. Fibronectin contribution was assessed using fibronectin-knockout mouse embryonic fibroblasts (MEFs-Fn-/-) and floxed equivalents (MEFs-Fnf/f), in fibronectin-depleted growth medium with/without exogenously added plasma or cellular fibronectin.

In the absence of fibronectin, MEF-Fn-/- were homogenously distributed throughout the collagen matrix. Contrary, in the presence of fibronectin, both cell types produced shell-like tissues with a cell-free collagen core and cell/fibronectin-rich surfaces. By incorporating a stretch-sensitive FRET-sensor, (plasma) fibronectin-stretch was observed to increase from the tissue center towards surfaces. We therefore suggest cells are attracted by fibronectinin a stretch-magnitude dependent manner. In addition, excessive assembly of (plasma) fibronectin by MEF-Fn-/- was observed at the tissue surface, speculated to originate from the absence of (cellular) fibronectin, which was indeed abolished after addition of cellular fibronectin.

Results suggest that tissue morphogenesis/heterogeneity is governed by fibronectin, providing stretch-dependent migration cues while cellular fibronectin provides biological cuesreducing excessive ECM assembly leading to tissue homeostasis. Implications can be found in accounting for the role of fibronectin in tissue engineering that may explain developing tissue heterogeneity.

BiographyFoolen Jasper has completed his PhD from the department of biomedical engineering at Eindhoven University of Technology, followed by a postdoctoral study of which part was conducted in the lab of Prof. Chris Chen at the University of Pennsylvania. He is now a Post-doctoral fellow in the lab of Viola Vogel at ETH Zurich and was recently awardeda Marie Curie fellowship.

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Foolen Jasper, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineMultifactorial optimization of cell-cocooned collagen-alginate microspheres for cardiac tissue engineeringMehrdad RafatLinköping University, Sweden

Summary: Tissue-specific delivery of stem cells holds the potential to regenerate damaged heart tissue and to restore its functions after Myocardial Infarction. Here we describe a novel cell encapsulation technique and an optimization method toward delivery of stem cells to damaged heart tissue. Induced pluripotent stem cells (iPS) were encapsulated in hybrid spheres of collagen and alginate and characterized for cell viability and spheres physical properties.

Background/Issue(s)/Objectives: Heart attack and failure is the number one cause of death in developed countries. Stem cell transplantation has been used as a promising therapy for heart disease. However, extensive cell attrition, and loss at the site of transplantation present a limit to therapeutic efficacy. We have hypothesized that cells viability, functionality, and target delivery will be enhanced by encapsulating the cells in hybrid collagen-alginate microspheres. Our main objectives are to develop cocooning techniques for effective delivery of viable stem cells to the damaged heart tissue and a mean to optimize microspheres properties with respect to size, shape, and encapsulated cells viability.

Design/Method: Hybrid spheres of collagen and alginate were produced using a coaxial air jet flow technique. The method involves the gelation of a collagen-alginate-cell solution in a calcium chloride bath. Microspheres loaded with iPS were washed and transferred to a Petri dish containing culture medium and incubated at 37°C. Microspheres formation and morphology (shape and size) and viability of the cells were monitored and evaluated using light microscopy techniques.

A 23 full multifactorial experimental design (MED) was employed to fabricate and test the spheres to determine optimum size, shape, and material composition. The effects of air flow rate, air gap distance and collagen to alginate ratio of the microspheres were studied.

Results: The MED results demonstrated that the air flow rate had a large effect on size, e.g. the higher the flow rate the smaller the spheres. Collagen to alginate ratio and air gap did not have a significant impact on size. The shape was, however, influenced by all factors. These findings allow us to fine-tune spheres to meet the size and shape criteria for different cell types. Spontaneous degradation occurred faster in spheres with higher collagen content compared to samples with less collagen. iPS cells were successfully encapsulated within the spheres and found viable over a 5-day period in culture.

Conclusions/Next steps: Preliminary results reveal that viable iPS cells could be encapsulated in hybrid collagen-alginate spheres. The impact of air flow rate and spheres composition on sphere’s size and shape were determined. The next steps will include extensive biological and physical characterizations of microspheres, i.e. collagenase biodegradation, release mechanism of the cells, and injection in animal models.

BiographyRafat received his Master’s and PhD degrees in Chemical and Biological Engineering from the University of Ottawa (Canada) with specialization in Biomaterials and Tissue Engineering in 2002 and 2007, respectively. He is the co-inventor of four patents and the senior author of several refereed publications. Rafat co-invented the first clinically-tested bioengineered cornea at the University of Ottawa Eye Institute. He has further developed the bioengineered cornea technology in his group at Linköping University and taken it to the next level closer to commercialization. His research group’s interests are mainly focused on Biomaterials Engineering and development of cell-interactive tissue-engineered materials as implantable scaffolds for ocular and cardiovascular applications such as bioengineered corneas and cardiac patches and nano and microencapsulation systems for controlled delivery of stem cells, drugs, and proteins to target tissues. His career spans a broad range of professional experiences-from working as a researcher and lecturer in academia, a medical devices regulator at the Canadian Government, and a consultant to biotech industries. He recently received two prestigious awards/grants from the European Research Agency including FP7 Marie Curie Incoming Fellowship Award (IIF) and FP7 Career Integration Grant. He is also the Co-Founder and Director of R&D, LinkoCare LifeSciences AB (Ltd.), a spinoff of Linköping University focused on commercialization of bioengineered corneas and other bioengineered tissues.

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Mehrdad Rafat, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineGuanidine substituted polyamidoamine hydrogels as scaffolds for cell culturing and tissue regenerationPaolo FerrutiUniversità degli Studi di Milano, Italy

Polyamidoamines (PAAs) are synthetic tert-amine polymers prepared by polyaddition of amines to bisacrylamides. They can be planned to be biocompatible and biodegradable. Guanidine-substituted amines, such as 4-aminobutylguanidine

(agmatine), lead to PAAs carrying guanidine side substituents. Agmatine combined with the carboxylated bisacrylamide 2,2-bisacrylamidoacetic acid gives a PAA called AGMA1 whose repeating unit mimics the RGA peptide:

Crosslinked AGMA1 in aqueous media gives hydrogels that exhibit excellent properties as scaffolds for many cell types culturing in vitro, including neural cells. AGMA1 tubular conduits were also used in vivo as scaffolds for nerve regeneration in a rat sciatic nerve cut model. Good surgical outcomes were achieved with no sign of inflammation or neuroma. Moreover, nerve regeneration was morphologically sound and the quality of functional recovery was satisfactory. However, AGMA1 hydrogels did not possess the mechanical properties needed to promote the osteoblastic differentiation of pre-osteoblastic cells, hence they were unsuitable as scaffolds for hard tissue regeneration. However, AGMA1 hydrogels responded favourably to MMT reinforcement, giving rise in aqueous media to nanocomposite hydrogels with shear storage modulus, G’, in the swollen state up to 20 times higher than those of the corresponding virgin hydrogels. The AGMA1-MMT nanocomposites definitely proved to warrant potential for the osteoblastic differentiation of mouse calvaria-derived pre-osteoblastic cells MC3T3-E1.

BiographyPaolo Ferruti took his degrees at the historical College “Collegio Borromeo” of the University of Pavia, and then was summoned by GiulioNatta at the Polytechnic of Milan. In 1968, he worked with Melvin Calvin in Berkeley at the Lawrence Radiation Laboratory of the University of California. In 1976, he became Full Professor at the University of Naples and then commuted to Bologna, Brescia and finally Milan. He authored more than 400 papers and 50 patents. Functional polymers for tecnica and biotechnological applications are his chief interest. His main scientific achievement has been the discovery of poly (amidoamine)s.

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Paolo Ferruti, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineBeta-catenin signaling regulates the chemotactic responses of mesenchymal stem cellsHuanxiang ZhangMedical College of Soochow University, China

Precise migration of stem cells, either endogenous or transplanted, is crucially important for embryonic development, homeostasis in adults, and tissue repair after injury. However, the detailed mechanisms of the directed migration of these

cells are not clear. During the past few years, our study showed that the differentiating mesenchymal stem cells (MSCs) possess different migratory capacity and the chemotactic responses of these cells correlates closely with their differentiation states. Accordingly, the formation and the asymmetrical distribution of focal adhesions (FAs) between the leading lamella and the cell rear, the phosphorylation of focal adhesion kinase (FAK) and paxillin, as well as the turnover of FAs varied greatly in differentiating MSCs, leading to the most effective chemotactic responses of MSCs in certain differentiation states. Further, we demonstrated that signaling through PI3K/Akt and MAPKs are involved in regulating the directed migration of MSCs. More importantly, we found that beta-catenin signaling is prerequisite for the chemotactic migration of MSCs. In this talk, I will summarize our data regarding the regulatory effects of beta-catenin signaling on MSCs that undergo chemotaxis.

BiographyHuanxiang Zhang has completed his PhD at the age of 28 years from Beijing Normal University, China and postdoctoral studies from Geneva University School of Medicine, Switzerland. He is now working in the Department of Cell Biology, Medical College of Soochow University, China. His research focuses on the control of the directed migration and differentiation of stem cells, including neural stem cells, mesenchymal stem cells and embryonic stem cells, and tissue engineering, especially the interaction between stem cells and the silk fibroin scaffolds with a variety of physical and chemical properties.

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Huanxiang Zhang, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineGuided tissue engineering for articular cartilage regenerationChristoph BrochhausenUniversity Medical Centre Mainz, Germany

The purposes of tissue engineering strategies to treat cartilage defects are the expansion of cells, their maturation, their integration into the defect and their preservation. The rationale for the use of growth factors in tissue engineering

applications is given by their ability to stimulate the controlled proliferation and differentiation of seeded cells in the scaffolds. They should also enhance the migration, proliferation, and differentiation of cells from the edges of the defect. Since regeneration recapitulates in parts embryological development, we used the growth plate of the long bones, which consists of chondrocytes in different proliferation and maturation stages, as a model to target growth factors and signalling molecules for cartilage tissue engineering. Growth factors act in a dose-dependent manner and via receptors on the target cells. Therefore, the kinetics of growth factor release from delivering devices should be adapted to the situation of the microenvironment. The amount of growth factor must reach an optimum for its biological action. Based on our analyses on the growth plate we identify prostaglandin E2 (PGE2) as a promising candidate molecule and integrated it in a release system which guarantied prolonged release of functional PGE2. In a next step the release system was integrated in a three dimensional collagen-based scaffold. We clearly demonstrated that within that scaffold under the presence of PGE2 cultured chondrocytes showed a rapid re-differentiation and relevant synthesis of collage II, the typical collagen for articular cartilage. This combined scaffold-release construct opens innovative perspectives for the future of guided tissue engineering strategies.

BiographyChristoph Brochhausen is Chief Consultant Pathologist and Head of Electron Microscopy at the Institute of Pathology at the University Medical Centre Mainz as well as Group Leader of the Cartilage Tissue Engineering Group at the REPAIR-lap. His research-work was awarded by national and international awards. He has published more than 80 papers in reputed journals and serving as an editorial board member of some reputed journals.

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Christoph Brochhausen, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineBionic materials for neuromuscular restoration and maintenanceRob M.I. Kapsa1,2, Quigley AF1,2, Moulton S1, Cornock R1, Zhang B2, Choong PFM2 and Wallace GG1

1University of Wollongong, Australia2St. Vincent’s Hospital, Australia

Effective engineering of skeletal muscle requires platforms that facilitate proliferation and maintenance of primary muscle stem/precursor cells (eg myoblasts) and muscle fibre maturation in a manner that reflects native muscle structure. In

addition, the supporting scaffold needs to accommodate the correct innervation of the re-engineered muscle tissue by promoting axonal connection and neuromuscular junction formation. We have been investigating the use of micro and nano-structured conducting polymer surfaces and soft gel systems for ex vivo muscle and nerve growth, differentiation and trophic electrical stimulation. Micro-structured platforms were created by localization of wet-spun PLA:PLGA fibers onto polypyrrole substrate, whilst nano-structured platforms were created by orientation of carbon nanotube fibres on a conducting gold mylar surface, over which a layers of polypyrrole were deposited. Human and murine myoblasts, rat dorsal root ganglion explants (sensory nerve) and PC12 neuronal cell lines were grown and/or differentiated on these platforms. A significant effect on myotube orientation was seen on both micro and nano-structured surfaces whilst surface topography similarly influenced the direction in which elements of the neuronal components grew. Growth of muscle cells as well as neuronal components (Scwhann cells and axons) on both nano and micro-structured polypyrrole was increased by electrical stimulation, providing a novel model system by which the effective innervation of regenerating muscle can be explored.

BiographyRobert Kapsa completed his PhD in 1996 at the University of Melbourne Dept. Medicine, St. Vincent’s Hospital (Melbourne). He is Program Director for the Bionics Platform of the ARC Centre for Electromaterials Science (ACES) and concurrent Principal Scientist and Head of Research Neurosciences at St Vincent’s Hospital in Melbourne. He has published 85 peer-reviewed manuscripts including 2 book chapters and one book in the areas of muscle biochemistry, genetics, gene therapy and muscle and nerve engineering. His work is focused on the development of autologous cell-based regenerative therapies for hereditary nerve and muscle disease.

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Rob M.I. Kapsa et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineCollagen-based biomaterials for tissue engineering and regeneration of the corneaNeil LagaliLinköping University, Sweden

There is a pressing global medical need for the development of tissue-engineered alternatives to supplement the scarce supply of human donor corneal tissue for transplantation and vision restorative therapy. There are currently over 10 million

people blind due to corneal scarring or disease, with 1-2 million new cases of corneal blindness arising annually. To meet this enormous global demand, our research has focused on developing transparent, tissue-engineered collagen-based hydrogels for implantation in the cornea. In preclinical studies, designs have evolved with respect to collagen composition and use of various synthetic cross-linking agents. Utilizing advanced in vivo imaging techniques such as optical coherence tomography and in vivo confocal microscopy, we have developed methods to visualize and longitudinally quantify bio-integration, wound healing, and cellular and neural compatibility of the biomaterial implants in rabbit and pig models, in real time. The implantable biomaterials and imaging techniques have also been extended into the clinic, where 10 patients received the first tissue-engineered corneal substitutes, and were followed and documented extensively for over six years postoperatively. In more recent work, our clinical experiences have resulted in refined designs of biomaterials and surgical implantation techniques, with a focus on robustness of materials in the operating room and optimization of the degradation profile of biomaterials in vivo, to facilitate host cell invasion and regeneration of tissue. Considerations of cost, raw material availability, and safety for human use are also becoming increasingly important as future efforts are aiming towards large-scale production of tissue-engineered corneas to meet an increasing global demand.

BiographyNeil Lagali is Associate Professor at the Faculty of Health Sciences, Linköping University, Sweden. He completed his PhD in Canada, and has held postdoctoral fellowships in The Netherlands, Canada, and Sweden. His current research interests include in vivo imaging in ophthalmology, corneal transplantation, and angiogenesis. He has published over 40 papers in international journals, authored several book chapters and one book, holds several patents and serves as editorial board member for BMC Ophthalmology.

[email protected]

Neil Lagali, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThe new challenge, considering adult mesenchymal stem cells (MSC) for articular cartilage repairMiguel G. GarberClinica Quirurgica Quantum, Spain

Articular cartilage plays pivotal roles in securing smooth joint kinetics and act as a shock absorber, however, is a highly organized tissue lacking self-regeneration capacity upon lesion. Chondral injury could lead to the development of

osteoarthritis (OA). Post-lesions defects are a major clinical problem, and there is at present no satisfactory clinical technique to regenerate cartilage. The initial treatment of meniscus tear of the knee is managed conservatively with no steroidal antiinflammatory drugs and physical therapy. When such conservative treatment fails, an arthroscopic resection of the meniscus is necessary, with this procedure, cartilage defects are repaired with fibrocartilage, which is known to be biochemically and biomechanically different from normal hyaline cartilage and degeneration occurs in the reparative tissue. However, the major drawback of the meniscectomy is an early onset of osteoarthritis. Mesenchymal stem cells (MSCs) are defined as pluripotent cells found in numerous human tissues, including bone marrow and adipose tissue, MSCs are known to have a potential for articular cartilage regeneration. Therefore, an effective and noninvasive treatment for patients with continuous knee pain due to damaged meniscus has been sought. Here, we present a review, highlighting the possible regenerative mechanisms of damaged meniscus with MSCs (especially adipose tissue-derived stem cells (ASCs)), along with a case of successful repair of torn meniscus with significant reduction of knee pain by percutaneous injection of autologous ASCs into an adult human knee. MSCs represent a promising new therapy in regenerative medicine.

BiographyMiguel Garber has over 27 years experience in Internal medicine and cardiology, with expertise in regenerative medicine, training and education, research, productdevelopment and senior management. He has more than 10 years working with Stem Cell, including building and managing the stem cell evaluation, explore anddeveloping stem cell therapies for cardiomyopathies, osteoarthritis and regenerative medicine at Stem cell Therapeutics Department of American Medical Information Group and Clinica Quirurgica Quantum. He is currently serving as Medical Director of Clinica Quantum and Clinical Director of regenerative Medicine department at Clinica Quantum, ongoing of several investigative research involved Stem Cells application (ASC) and Drug stimulating stem cells (Aphanizomenón Flos Aquae). Dr. Garber has made a significant contribution to Stem cell Research. Actually he is involved in Adipose Stem Cell application.

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Miguel G. Garber, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative Medicine

Chikara Furusawa, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014

Oscillatory protein expression dynamics generates robust and irreversible differentiation of stem cellsChikara FurusawaQuantitative Biology Center (QBiC), Japan

A systems-level understanding of cell differentiation is important for developmental biology. Some of the basic questions concerning such systems-level understanding include: What characteristics in a cellular state distinguish multi-potent

stem cells from differentiated cells? How are developmental processes robust to molecular noise in spite of their complexity? Following the progress in the analysis of molecular mechanisms for cell differentiations, the time is ripe to answer the above questions to unveil nature of differentiation from stem cells.

In this study, using a dynamical system modeling, we performed simulations of the developmental process using small gene regulatory networks, and screened those that could generate cell type diversity through cell-cell interactions. We found that stem cells that both proliferated and differentiated always exhibited oscillatory expression dynamics, and the differentiation frequency of such stem cells was regulated, resulting in a robust number distribution. Based on the result of computer simulations, we propose a hypothesis for the mechanism of stem cell differentiation, in which the expression levels of some genes in multipotent stem cells exhibit temporal oscillation, and itinerate over several sub-states. As development progresses, each of these quasi-stable sub-states is modified and stabilized, leading to differentiated cell types.

Importantly, this hypothesis can explain the roles and mechanism of the recently observed dynamic heterogeneity and oscillatory behavior in cellular states of stem cells, and it can predict the regulatory motifs responsible for the dynamic differentiation process. These discussions promote a system-level understanding of multicellular development and provide a basis for clinical application of stem cells.

BiographyChikara Furusawa has completed his PhD at the age of 28 years from University of Tokyo and postdoctoral studies from Center of Developmental Biology, RIKEN. He is a team reader at Quantitative Biology Center, RIKEN. He has published more than 60 papers in reputed journals

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Govindan Rangarajan, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014

Critical thresholds in cell fate determinationGovindan RangarajanIndian Institute of Science, India

It is well known that cell fate depends on a variety of complex interacting factors. Nevertheless it is important to have simple mathematical models that capture the essential features involved in cell fate determination. We hypothesize that there are

critical thresholds for expression levels of key genes (such as Myc) involved in the process. Below the critical threshold, the cell has a particular phenotype (say, normal) and above the threshold, the cell has a different phenotype (say, cancer or stem cell). Using tools from nonlinear dynamics we show that the above assumption can lead to reversible changes in cell phenotype. In particular, we implement the threshold in terms of coupling coefficients of nonlinear oscillators in the cell and we demonstrate that these oscillators undergo a bifurcation leading to oscillator death (and consequent change in cell fate) once the threshold is crossed.

BiographyGovindan Rangarajan obtained his PhD from University of Maryland, College Park, USA. He was a Staff Scientist at the Lawrence Berkeley National Laboratory, USA before moving to the Indian Institute of Science, Bangalore. He is currently a Professor there. His research interests include nonlinear dynamics, computational neuroscience and mathematical biology.

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Jianhua Xing, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014

A cascade of bistable switches controls TGF-β-induced epithelial to mesenchymal transitionJianhua XingVirginia Tech, USA

Epithelial to mesenchymal transition (EMT) is essential for cell plasticity during development and plays important roles in cancer progression. Two mathematical models proposed different mechanisms on the regulation of TGF-β1 induced

EMT. One model predicted that two bistable switches, governed by SNAIL1/miR-34 and ZEB/miR-200 double-negative feedback loops respectively, lead to epithelial, partial EMT, and mesenchymal phenotypes. Another model argued that the three phenotypes came from a tertiary switch formed by the ZEB/miR-200 loop. In this work we quantitatively measured the dynamics of TGF-β1 induced MCF10A EMT. We identified three cell subpopulations during EMT using flow cytometry. Our measurements at RNA and protein levels verified that the temporal and steady state dynamics show a two-step behavior, and can be well represented by sequential Markovian transition among the three phenotypes, reversible transition between epithetial and partial EMT phenotypes, and irreversible transition between partial EMT and mesenchymal phenotypes. Furthermore, SNAIL1 shows both unimodal and bimodal distributions under different TGF-β1 concentration, indicating the bistable behavior of SNAIL1. These experimental results quantitatively confirmed that a cascade of two binary switches governs TGF-β1 induced MCF10A EMT.

BiographyJianhua Xing has completed his PhD in theoretical chemistry from UC Berkeley, and did postdoctoral studies also at UC Berkeley. After a short period stay at Lawrence Livermore National Laboratory. He joined Virginia Tech in the department of Biological Sciences, and is affiliated to the physics department as well. The current research in Xing’s lab focuses on combined computational and experimental quantitative biology studies of regulation mechanisms of cell phenotypic transitions, especially the epithelial-to-mesenchymal transition, at transcription, translational, and epigenetic levels.

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Paul H. Lerou, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014

Exploring human pluritotent stem cell heterogeneity using a multi-scale imaging and informatics pipelinePaul H. LerouHarvard Medical School, USA

Understanding how human pluripotent stem cells (hPSCs) make cell fate decisions is central to both normal embryological development as well as therapeutic stem cell applications, such as generating uniform differentiated cell populations from

induced pluripotent stem cells (iPSCs). Since human embryonic stem cells were first isolated, various directed-differentiation protocols have been developed by empirically adding or removing inductive signals to the differentiating cell population in order to progressively enrich specific cell subsets that will yield the cell of interest. This approach fails to achieve the degree of spatiotemporal regulation found in vivo and as a result, current directed differentiation protocols are inefficient and highly variable. Due to complex tissue-level effects, population-based assay are inadequate to fully understand the pluripotent state, let alone differentiation. This complexity is multi-factorial: the cell population is heterogeneous in morphology and gene expression, and the cellular microenvironment is multidimensional. We have developed an in situ multi-scale imaging system that captures high content single-cell information of hPSCs across long length scales. This platform integrates an automated robotic microscopy system with an analysis pipeline that allows us to study relationships between cellular location within a colony and single-cellular properties, long-range interactions between different cell phenotypes, and between colony-wide features such as size, shape, and texture, and the properties of their constituent cells. In order to understand genetic network dynamics in hPCs in situ across long length scales we are developing computational algorithms to integrate and analyze single cell single-molecule mRNA FISH data into this multi-scale imaging and informatics pipeline.

BiographyPaul H. Lerou completed his MD degree from Jefferson Medical College and subsequently trained in pediatrics and neonatology at Boston Children’s Hospital, Harvard Medical School (HMS). He did his postdoctoral work in the laboratory of George Q. Daley on human pluripotent stem cell derivation. He is currently an attending neonatologist at Brigham & Women’s Hospital and assistant professor in pediatrics at HMS. His lab studies fundamental pluripotent stem cell biology and aims to apply this to the development of improved in vitro models of human disease using induced pluripotent stem cells.

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Tissue Science & Regenerative MedicineShaping the microenvironment for physiological cultivation of human adult stem cellsCornelia Kasper1, Verena Charwat1, Anne Neumann2, Maike Keck3 and Peter Ert11University of Natural Resources and Life Science (BOKU), Austria2Austrian Institute of Technology, Austria3Medical University Vienna, Austria

Although our knowledge about the in vivo microenvironment of stem cells (the biological niche) is steadily increasing, “standard” cell cultivation is still performed under not physiologically relevant conditions. Usually cells are cultivated

under ambient oxygen conditions in static 2D cultures on plastic surfaces. Since cellular responses are highly dependent on the complex interplay between cells and their microenvironment, it is important to develop more physiologically relevant cultivation systems for in vitro cellular studies. For specialized stem cell cultivation already several bioreactor systems are available. These systems have the advantage that they provide medium perfusion which enables better oxygen and pH control as well as continuous nutrient supply and waste removal. Furthermore dynamic cultivation allows mechanical stimulation, which is an important cue for stem cell differentiation. A great variety of specialized bioreactors ranging from miniaturized to large-scale systems on 2D surfaces and within 3D tissue-like matrices with and without integrated biosensors has been developed.

The lecture will provide an insight into dynamic cultivation of human adult stem cells in specialized bioreactor systems. A general introduction will be focused on the unique characteristics of dynamic stem cell cultivation and the differences to standard cultivation procedures. The potential and benefits of dynamic cultivation will be highlighted, but also risks, potential pitfalls and limitations of will be discussed. Different dynamic systems for stem cells expansion and differentiation will be introduced. The presented bioreactor types will range from miniaturized microfluidic set ups to clinical scale reactors and include 2D cultures as well as 3D tissue-like structures.

BiographyCornelia Kasper completed her PhD in 1998 from Leibniz University of Hannover (Germany) and her habilitation in 2007 at the Institute for Technical Chemistry at the Leibniz University of Hannover. She was appointed as full University Professor for “Biopharmaceutical Production and Technology” at University of Natural Resources and Life Science (BOKU) in Vienna (Austria) at the Department of Biotechnology. She has published more than 80 papers in reputed journals and several book chapters and is editor within the series “Advances in Biochemical Engineering and Biotechnology” (Springer) of several volumes covering actual areas in Tissue Engineering and Stem Cell Research. She is also reviewer for many distinguished journal within the field of biotechnology/bioprocess and bioreactor design and enablelingtechnolgies for stem cell cultivation.

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Cornelia Kasper et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineRegulation of metastasis by endogenous metaboliteSudhakar YakkantiCell Signalling & Metabolism Centre, USA

Metastasis is frequently deadlier than the original tumor-ultimately, reducing the risk or occurrence of metastasis could effectively cure or at least manage human cancer. My laboratory has carried out research that could significantly contribute

to the control of malignant progression, involving the use of endogenous metabolite that is released from the extracellular matrix, which is both an endogenous angiogenesis inhibitor and anti-metastatic molecule. The signaling mechanism(s) underlying the influence of these metabolites on regulation of tumor angiogenesis is known, where as regulation of tumor metastasis are not yet known. We identified that one of the endogenous metabolite binds to different cell surface integrins, inhibits different cellular signaling in a manner distinct from that of other metabolites studied to date. Treatment of endothelial cells with this metabolite specifically inhibiting κ-elastin mediated phosphorylation of FAK, Akt, mTOR and PI-3K signaling. In additiondifferent in-vitro and in-vivo studies, we found that this metabolite possibly binding to Laminin D-III and D-IV domains and inhibits Laminin degradation by the matrix metalloprotease-14 (MMP-14), and thereby reduces the generation of different sized Laminin peptides that can bind to the EGF receptor and promote cancer metastasis, in addition to its integrin(s) mediated signaling. Our findings suggest that this metabolite interacting with different cell surface integrins and cross talking with othermolecules and inhibiting tumor angiogenesis and tumor metastasis both in-vitro and in-vivo.

BiographySudhakar Yakkanti, Former Associate Director at Center for Cancer & Metabolism, Bioscience Division, California, San Fransisco. He did his postdoctoral training at Harvard Medical, Boston USA. He received Indian President’s fellowships, YCS, Michael A. O’Connor Young Investigator Awards from FAMRI and Mayo Clinic.He has more than 40 research articles including Science, JCI, Blood, PNAS etc. His research focuses on cell signaling and tumor metastasis, which is supported by NIH and foundation grants. He was served as grant reviewer for DT study section. He is serving as an Editor-in-Chief and also honored as keynote speaker and session chair for many conferences.

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Sudhakar Yakkanti, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineNew BEST - biomaterials enhanced simulation testMichael GasikAalto University, Finland

This work presents the development of new in vitro testing method for biomaterials in the closest to hostile conditions. The objective of this BEST - biomaterials enhanced simulation test - is to provide maximal possible yet realistic control and

monitoring of chemical, biological, cytological etc. reactions, and leading to decrease of animal testing. Recently no combined in vitro solution exists capable of answering the demands and needs of all stakeholders at reasonable costs, speed and safety. New challenges require more consistent and holistic approaches to ensure reliability and safety of the implants including those with ATMP, as well as primary cultures based TE/RM applications.

Here the design of the BEST methodology and test equipment is shown for the case of load-bearing implants such as orthopaedic and dental ones. The demonstration includes based porous coated biomaterials at different conditions showing the preferential potential for bone, cartilage or fibrous tissue formation. The testing is supported by time- and frequency-domains simulation with models in silico.

The results show the importance of proper application of relevant testing parameters vs. traditionally used protocols, leading also to reduction of specimens and faster screening of new materials formulations with new method.

BiographyMichael Gasik’s scientific interests are mainly in biomaterials for hard tissue replacements (orthopaedic, dental) and CATMP. Multifunctional bioresorbable biomaterials with biofilm inhibition and new alternative testing methods (3R) are of a particular interest. MC member (Finland) in COST MP1005 "From Nano to Macro Biomaterials and applications to stem cells regenerative orthopaedic and dental medicine". Member of the Finnish board of Japanese Society for Promotion of Science. Honorary member of Technet Alliance. Participant in biomaterials activities such as BioTiNet and EFORT. Rapporteur for EU framework programs, ERA-NET and joint undertakings. Reviewer of national research programs in several countries and selected scientific journals.

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Michael Gasik, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineA successful approach to rapidly scale up microcarrier-based expansions of human mesenchymal stem cells for allogeneic cell therapiesValentin Jossen1, Stephan Kaiser1, Ann Siehoff2, Christian van den Bos2, Dieter Eibl1 and Regine Eibl11Zurich University of Applied Sciences, Switzerland2Lonza Cologne GmbH, Germany

Human mesenchymal stem cells (hMSCs) isolated from adipose tissue and bone marrow represent often used cell types in many pre- and clinical studies running today. This can be explained by their high therapeutic potential resulting from

their differentiation capability, immunosuppressive, immune regulating, migrating and trophic properties. In the majority of clinical studies with adipose tissue- and bone marrow-derived hMSCs the treatment of orthopaedic and cardiac diseases is focused on autologous and in special cases on allogeneic cell therapies. However, the required number of therapeutic hMSCs for such allogeneic treatments is in the range of trillions per year. Due to the high cell quantities, alternatives to the 2‐dimensional planar cultivation systems (stacked plate systems) typically being applied to propagate hMSCs and suitable scale-up strategies of the expansion procedure are stringently required.

The talk will present a successful scale‐up Approach based on microcarriers and stirred cultivation systems with cultivation containers intended for one‐time usage. This approach, combining knowledge in bioengineering and cell biology allowed direct transfer of process and cultivation conditions from 100 mL upto 35L and 50L culture volume as shown for adipose tissue- and bone marrow-derived hMSCs. It presupposes extensive screening studies performed in spinner flasks in order to establish the optimal culture medium-microcarrier type-combination, the optimal medium composition and the optimal operational conditions. For rapid process scale-up, experimental and numerical investigations have to be realized in order to predict the optimum stirrer speed and fluid flow pattern, the suspension criteria (NS1 and NS1U), and to keep the shear stress low. In so doing, we achieved one of the highest peak cell densities (0.7-0.8•∙106 cells mL-1) reported to date for hMSCs expanded on microcarriersat L‐scale under low-serum conditions.

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Valentin Jossen et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineSpontaneous fusion of breast cancer cells with mesenchymal stem cells: Support for a long-standing hypothesis on cancer metastasisBrenda M. OgleUniversity of Minnesota, USA

Ninety percent of cancer-related deaths are due to secondary tumors, or metastases, that form at sites far removed from the primary tumor. To successfully relocate in the body, a tumor cell must acquire transient properties that

enable dissemination, followed by the reestablishment of the original primary phenotype at a distant site. Exactly how this is accomplished is yet unclear and therefore reliable treatments are lacking. Here we test a long-standing, but technically challenging, hypothesis that spontaneous fusion of tumor cells with cells of stromal lineage give rise to hybrid cells capable of dissemination and new tumor growth. Here we analyzed the ability of breast cancer cells to spontaneously fuse with mesenchymal stem cells. We used breast cancer cells with different degrees of aggressiveness (T47D which is non metastatic, and MDA-MB-231 which is metastatic) and normal breast epithelial cells MCF10a. To enable these studies, we have developed a powerful tool for the detection of fusion products in vitro that utilizes bimolecular fluorescence complementation. Using this tool, we found that MSCs fuse spontaneously with all breast epithelial cell types. Interestingly, this fusion occurs to a greater extent between MSCs and the breast cancer cells T47D and MDA-MB-231 (P< 0.05) compared to normal breast epithelium suggesting that cancer cells more readily fuse with MSCs. In addition, we found that hypoxia stimulates a significant increase in fusion between MSCs and non-metastatic breast cancer cells T47D (P<0.05) which is intriguing given the hypoxic nature of the tumor microenvironment relative to healthy tissue. And finally, hybrids are more migratory than breast cancer cells both in terms of accumulated distance and velocity. These results suggest that cell fusion might contribute to the ability of cancer cells to disseminate from the primary tumor site and perhaps metastasize. Future follow-on studies may uncover new strategies for cancer treatment and/or preventing metastatic spread.

[email protected]

Brenda M. Ogle, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineDevelopment of a bio-inspired blood factory for personalised healthcareAthanasios (Sakis) MantalarisImperial College London, UK

Personalized medicine is a medical model that proposes the customization of healthcare, with decisions and practices being tailored to the individual patient by use of patient-specific information (initially genetics information) and/or the

application of patient-specific cell-based therapies.

The BioBlood project (an ERC Advanced Investigator Award) aims to deliver personalised healthcare through a “step change” in the clinical field of haemato-oncology. BioBlood represents an engineered bio-inspired integrated experimental/modelling platform for normal and abnormal haematopoiesis that receives disease & patient input (patient primary cells & patient/disease-specific data) and will produce cellular (red blood cell product) and drug (optimal drug treatment) therapies as its output. Herein, we will present the experimental platform, which is a novel three-domensional hollow fibre bioreactor capable of culturing normal and abnormal haematopoietic cells in the absence of exogenous growth factors by mimicking the structure and function of the bone marrow, alongside a population balance model (PBM) that is able to capture cellular heterogenity and in particular leukaemia heterogenity. The PBM, which is able to extract patient- and disease-specific information is linked to a pharmacokinetic/pharmacodynamic (PD/PD) model, which is the used to optimise chemotherapy treatment in a personalised manner.

BiographyAthanasios (Sakis) Mantalaris is Professor of BioSystems Engineering in the Department of Chemical Engineering at Imperial College London. He received his PhD (2000) in Chemical Engineering from the University of Rochester. His expertise is in modelling of biological systems and bioprocesses with a focus on mammalian cell culture systems, stem cell bioprocessing, and tissue engineering. He has published over 150 original manuscripts, co-edited one book, and holds several patents with several more pending. He has received several awards including the Junior Moulton Award for best paper by the Institute of Chemical Engineers (IChemE) in 2004. In 2012, he was elected Fellow of the American Institute for Medical & Biological Engineering and in 2013 he was awarded a European Research Council (ERC) Advanced Award.

[email protected]

Athanasios (Sakis) Mantalaris, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineOur experience with autologous stem cell applications in chronic non healing ulcers of lower limbSribatsa Kumar MahapatraV.S.S. Medical College, India

Chronic non healing ulcers of lower limb due to diabetes, sickle cell disease, peripheral vascular disease, Hansen’s disease, Burns and venous ulcers, which are refractory to conventional treatment are tried with autologous bone marrow derived

mesenchymal stem cell therapy. Ulcer healing was then observed with control group of similar type of ulcers where only conventional ulcer therapy was tried. Progress of ulcer healing was then observed by appearance of good quality granulation tissue, marginal epithelial migration, ulcer contraction in stem cell applied ulcers in contrast to control group.

BiographySribatsa Kumar Mahapatra completed his M.B.B.S with Honors from M.K.C.G. Medical College, Berhampur(gm), Odissa, India, in 1977, later got his M.S. in general surgery from post graduate Institute of Medical Education and Research, Chandigarh, India in 1982 and obtained his F.R.C.S. from royal college of surgeons of Edinburgh in1994. Presently he is working as Professor and Head of the Dept. of General Surgery at V.S.S. Medical College, Sambalpur, Odissa, with teaching, treatment and research activity in a busy 1200 bed hospital. He has published many papers on cell therapy in national and international journals.

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Sribatsa Kumar Mahapatra, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineOxygen controlled bioprocessing of pluripotent stem cellsFarlan VeraitchUniversity College London, UK

Stem cells can self-renew in vitro whilst retaining their ability to differentiate into multiple adult cell types. These properties suggest that stem cells will have a number of potential applications including the generation of adult tissue for regenerative

medicine, drug discovery, drug development and whole cell delivery of gene therapies. One of the major technical challenges will be the development of scalable, cost effective, reproducible and safe whole bioprocesses. Farlan Veraitch's talk will focus on his research group's work the use of oxygen tension for the control of both the expansion and differentiation of pluripotent stem cells. In particular the talk will focus on recent findings showing how controlling oxygen tension during the exapnsion step can “prime” pluripotent stem cells to differentiated into specific lineages.

BiographyFarlan Veraitch gained his PhD from the University of Birminham where his research focused on the optimisation of mammalian cell culture processes. He then moved to UCL where he worked as a Post Doctoral Research Assistant on the automation of embryonic stem cell processing. Since gaining his Letureship, and subsequent Senior Lectureship, Farlan has helped to establish the UCL’s Cell Therapy Bioprocessing programme which has been applying ultra scale-down, bioprocess modelling and a ‘whole bioprocess’ vision to the development of robust stem cell production processes.

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Farlan Veraitch, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThymic tissue regeneration using natural scaffoldsIleana Bortolomai1, Monica Sandri2, Tiziano Di Tomaso2, Marco Catucci3, Angelo Lombardo1, Silvia Panseri2, Luigi Naldini1,4, Georg A. Hollander3,5, Anna Tampieri2, Anna Villa1,6 and Marita Bosticardo1

1San Raffaele Scientific Institute, Italy2Institute of Science and Technology for Ceramics (ISTEC), Italy3University of Basel, Switzerland4Vita-Salute San Raffaele University, Italy5University of Oxford, UK6UOS Milano, Italy

Thymus transplantation has great clinical potential, but the shortage of transplant donors limits the progress of this therapy. Creation of a bioengineered thymus, where the cellular component is autologous, will overcome two major obstacles in

transplantation: the lack of organs and the toxicity arising from lifelong immunosuppression. Here, we show how the induction of ectopic Oct4 in adult TEC could allow the expansion of thymic epithelial progenitor-like cells (pTECs) in 2D and 3D culture systems, retaining their capability to differentiate into mature TECs and potentially form a functional thymus in vivo. In order to obtain bipotent TECs precursors that could differentiate into both cortical and medullary TECs we transduced freshly isolated TECs with a lentiviral vector (LV) that allows for ectopic expression of Oct4. Through LV transduction we were able to achieve high levels of Oct4 expression in adult TECs. Moreover, we demonstrated that ectopic expression of Oct4 is sufficient to promote the de-differentiation of primary mature TECs into pTECs, inducing the expression of the progenitor markers, such as Thy1.2, Sca1 and CD44, and decreasing the expression of maturation markers, such as MHCII. To improve thymic organogenesis and restore its functions both in vitro and in vivo, we exploited a 3D collagen type I scaffolds, mimicking the thymus structure, for the culture of TECs. Our results showed that collagen type I scaffolds allow the efficient expansion of pTECs. pTECs grown on scaffolds displayed a high proliferation rate without losing their developmental potential. To induce pTECs maturation and thymic regeneration, we are now evaluating a LV-based strategy for Oct4 inducible expression that allows us to switch off Oct4 expression and control pTECs differentiation. We will then perform a morphological and phenotypical characterization of TECs cultured on scaffolds and evaluate their ability in restoring thymic function both in vitro and in vivo.

BiographyIleana Bortolomai has completed her PhD with the Open University of London at the IRCCS Istituto dei Tumori di Milano in 2011. At Yale University School of Medicine she completed her first postdoctoral studies at Dr.Santin lab where she was involved in evaluating the targeting potential of CPE functionalized PLGA-NPs for gene therapy against cervical cancer cells. In 2014 she has joined the group of Dr. Anna Villa at Istituto San Raffaele Telethon (TIGET) where she is involved in the project of thymus regeneration, as innovative research on gene transfer and cell transplantation for successful clinical application for genetic diseases.

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Ileana Bortolomai et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineChanges in gene expression, protein content and morphology of chondrocytes cultured on a 3D random positioning machine and 2D rotating clinostatGanna AleshchevaOtto-von-Guericke University, Germany

Chondrocytes are the only cell type found in human cartilage consisting of proteoglycans and type II collagen. Several studies on chondrocytes cultured either in space or on a ground-based facility for simulation of microgravity revealed

that these cells are very resistant to adverse effects and stress induced by altered gravity. Tissue engineering of chondrocytes is a new strategy for cartilage regeneration. Using a three-dimensional Random Positioning Machine and a 2D rotating clinostat, devices designed to simulate microgravity on Earth, we investigated the early effects of microgravity exposure on human chondrocytes of six different donors after 30 min, 2h, 4h, 16h, and 24h and compared the results with the corresponding static controls cultured under normal gravity conditions. As little as 30 min of exposure resulted in increased expression of several genes responsible for cell motility, structure and integrity (betaactin); control of cell growth, cell proliferation, cell differentiation and apoptosis; and cytoskeletal components such as microtubules (beta-tubulin) and intermediate filaments (vimentin). After 4 hours disruptions in the vimentin network were detected. These changes were less dramatic after 16 hours, when human chondrocytes appeared to reorganize their cytoskeleton. However, the gene expression and protein content of TGF-β1 was enhanced for 24h. Based on the results achieved, we suggest that chondrocytes exposed to simulated microgravity seem to change their extracellular matrix production behavior while they rearrange their cytoskeletal proteins prior to forming three-dimensional aggregates.

BiographyGanna Aleshcheva has completed her study of biotechnology at the Technical University to Berlin (Berlin, Germany) and Zurich University of Applied Sciences (Waedenswil, Switzerland) in 2012. Afterwards, she started her PhD about “Tissue Engineering of cartilage” at the Otto-von-Guericke University (Magdeburg, Germany) and Aarhus University (Aarhus, Denmark) as a member of Space Life Sciences Research School (SpaceLife) of the German Aerospace Center and Helmholtz Association.

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Ganna Aleshcheva, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineElectrospun nanofibers based on natural biomaterials for regenerative medicineHadi Hajiali, Maria Summa, Rosalia Bertorelli, Athanassia Athanassiou and Elisa MeleItalian Institute of Technology, Italy

In tissue regeneration, it is fundamental that the used scaffolds mimic the structure and the functions of the extracellular matrix. Common scaffolding methodologies, such as solvent casting, particulate leaching, gas foaming, and freeze drying,

difficulty satisfy this requirement. On the contrary, the electrospinning technique has demonstrated its ability in fabricating nanofibrous scaffolds able to duplicate the native tissue structure. Electrospinning is a versatile method to create fibres having a diameter can range from 5 nm to more than 1 µm. Electrospun webs provide scaffolds with a wide range of mechanical properties (from plastic to elastic), optimized porosity and topographical cues. This results in improved cellular response and it is often translated into accelerated tissue healing and regeneration. Various synthetic and natural biodegradable polymers have been electrospun for producing nanofibrous substrates with the desired mechanical and chemical properties. Moreover, drugs, growth factors and other active molecules can be added inside the nanofibers. We demonstrated the fabrication of composite nanofibers based on polymers derived from natural sources (such as alginate, cellulose, and chitosan) and functionalized with active agents. The antimicrobial activity, cytotoxicity and inflammatory response of the realized fibrous scaffolds were investigated in-vitro and in-vivo. The obtained results show that they are promising architectures for regenerative medicine and wound care medical devices.

BiographyHadi Hajiali completed his M.Sc. degree in Biomedical Engineering (Biomaterials) from Iran University of Science and Technology in 2010. He has consistently ranked the first student of his class in B.Sc. and M.Sc. courses. His Master’s dissertation was “Synthesize Nanocomposite Scaffolds and Examine their Properties for Bone Tissue Engineering”. Moreover, he joined Tissue Engineering and Nanomedicine Research Center at Shahid Beheshti University of Medical Sciences to work on the projects which were related to regenerative medicine. He is currently a PhD student in Bioengineering (Bionanotechnology) at University of Genova and Italian Institute of Technology (IIT). His PhD project is “Smart-Nanostructured Scaffolds for Regenerative Medicine”. The results of his research projects have been published as several papers.

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Hadi Hajiali et al, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineA robust HIV-1 viral load detection assay optimized for Indian sub type C specific strains and resource limiting settingParth Shah3, Arpan Acharya1, Salil Vaniawala2, Parth Shah3, Harsh Parekh4, Rabindra Nath Misra5, Minal Wani1 and Pratap N Mukhopadhyaya4

1Dr. D. Y. Patil Biotechnology & Bioinformatics Institute, India2SN Genelab, India3Supratech Micropath Laboratory & Research Institute, India4Interdisciplinary Science, Technology and Research Academy, India5Padmashree Dr. D. Y. Patil Medical College & Research Centre, India

Human Immunodeficiency Virus Type 1 (HIV-1) viral load testing at regular intervals is an integral component of disease management in AIDS patients. The need in countries like India is an assay that is not only economical but efficient and

highly specific for HIV-1 sub type C virus. This study reports a SYBR Green-based HIV-1 real time PCR assay for viral load testing and is designed for enhanced specificity towards HIV-1 sub type C viruses prevalent in India. Linear regression of the observed and reference concentration of standards used in this study generated a correlation coefficient of 0.998 (p<0.001). Lower limit of detection of the test protocol was 50 copies/ml of plasma. The assay demonstrated 100% specificity when tested with negative control sera. The Spearman coefficient of the reported assay with an US-FDA approved, kit was found to be 0.997. No significant difference in viral load was detected when the SYBR Green based assay was used to test infected plasma stored at -200C and room temperature for 7 days respectively (Wilcoxon signed rank test, p=0.105). The HIV-1 viral load assay reported in this study was found to be robust, reliable, economical and effective in resource limited settings such as those existing in India. PCR probes specially designed from HIV-1 Subtype C-specific nucleotide sequences originating from India imparted specificity towards such isolates and demonstrated superior results when compared to two similar commercial assays widely used in India.

BiographyParth Shah has completed his MBBS (Medicine) from the Gujarat University in India at the age of 24. He has trained at the Brigham and Womens hospital in Molecular Genetics and has been working as a Research Scientist for Infectious diseases and Oncogenic Markers on Next Generation Sequencing at Supratech Micropath Labs (CAP and NABL accredited) for the last 1 year.

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Parth Shah et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineAllogenic mesenchymal stem cells for the treatment of equine degenerative joint diseaseCharlotte Beerts, Sarah Broeckx, Marc Suls and Jan H. SpaasGlobal Stem Cell Technology, Belgium

Degenerative joint disease (DJD) is a major cause of reduced athletic function and retirement in equine performers. However, conventional therapies are merely aimed at alleviating the symptoms or enhancing clinical recovery for a short

period of time, whereas regenerative medicine aims at long-term clinical improvement and pain relief of the affected joints. Indeed, equine (n=33) as well as human (n=12) studies confirm a clinical improvement of patients with knee injuries after intra-articular mesenchymal stem cell (MSC) injection. Furthermore, due to many interspecies similarities, the horse might be considered as a valuable animal model to evaluate human orthopedic therapies. For all the aforementioned reasons, MSC studies in equine joint pathologies are of therapeutic interest to the scientific community.

In the present study, peripheral blood (PB) from one donor horse was used to isolate MSCs and platelet-rich plasma (PRP) for combined use. Indeed, it has been confirmed that PRP increases the cell viability of chondrocytes, enhances the migration and chondrogenesis of MSCs and improves the clinical outcome of MSC therapy in osteoarthritic joints. However, it has been reported that the clinical effect of PRP alone is significant inferior to the MSC-PRP combination. Therefore, patients with naturally occurring DJD of stifle, fetlock, pastern and coffin joints were included in the study and randomly treated with MSC-PRP or chondrogenic induced MSCs-PRP. Short-and long-term follow-up consisted of a clinical evaluation and scores were given for failure of return to work (0), rehabilitation (1), return to work (2) and return to previous level (3).

In conclusion, the present study reports a considerable, yet joint-specific enhanced clinical improvement when using chondrogenic induced MSCs for the treatment of DJD in horses.

BiographyCharlotte Beerts graduated as a Doctor in Veterinary Medicine in June 2012 from the University of Liège. She received the Pfizer Scientific Prize for an outstanding Master Thesis about the current aspects of Equine Infectious Anemia in Europe. In August 2012 she started an internship in the Equine Clinic Equitom in Belgium. Recently she joined the scientific team of Global Stem Cell Technology (GST) to take part in the further research on stem cell technologies for the treatment of musculoskeletal and metabolic disorders in horses and to turn her passion into her carrier.

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Charlotte Beerts et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineStrategy optimization of the combination of 3D extruded-based poly (ɛ-caprolactone) scaffolds with mesenchymal stem cells towards cartilage regenerationCarla Sofia Monteiro de Moura1, Claudia Lobato da Silva2, Paulo Jorge Bartolo1, 3 and Frederico Castelo Ferreira2

1Polytechnic Institute of Leiria, Portugal2University of Lisbon, Portugal3University of Manchester, UK

Hyaline cartilage is a critical tissue to normal articular function. Natural restoring of this tissue in patients with rheumatoid arthritis, osteoarthritis or traumas is extremely limited considering its low regenerative capacity. These conditions cause

an enormous constraint to the daily routine and, consequently, lower quality of life and they are managed, according with severity, through the use of pharmaceutics, surgery, transplant or prosthetic procedures. Tissue Engineering (TE) emerges as a new promising tool to provide long lasting regenerating solutions. In this work we used the combination of 3D extruded-based scaffolds made of poly (e-caprolactone) (PCL) and human bone marrow-derived mesenchymal stem cells (MSC) to provide initial higher cell densities, followed by differentiation in chondrocytes. The aim of this study is to understand the influence in cell behavior of scaffolds manufactured by layer-by-layer extrusion with a pore size (190 – 390 µm) and fiber alignments (0-45º and 0-90º), as well as of two different atmospheric conditions, normoxia (21% O2) and hypoxia (5% O2). Results obtained showed that pore size and fiber alignment do not pose a limitation for cell adhesion and proliferation within the range of pore sizes studied. However, cell distribution and dimension of chondrocyte aggregates strongly depend on fiber alignment. The results obtained in this study point out that higher chondrocyte population is obtained, when previously to differentiation stage, MSC are expanded in the PCL under hypoxia condition, rather than normoxia.

BiographyCarla Moura is a PhD student of the MIT-Portugal Program in Bioengineering. Her work is done in collaboration between Centre for Rapid and Sustainable Product Development (CDRsp), Polytechnic Institute of Leiria and Institute of Biotechnology and Bioengineering (IBB), University of Lisbon. This research is sponsored by the Portuguese Foundation for Science and Technology through MIT Portugal Program, Bioengineering Systems Focus Area a PhD grant SFRH/BD/73970/2010 and an Investigator FCT research contract IF/00442/2012.

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Carla Sofia Monteiro de Moura et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThe use of Keratinocytes: Things we should keep in mind!Lars-Peter KamolzMedical University of Graz, Austria

Background: Cultivation of keratinocytes and their clinical application was an essential step towards the development of new treatment concepts for patients suffering from severe burns and chronic cutaneous wounds.

The aim of this review is to give a current overview of keratinocyte cultivation and keratinocyte application under experimental and clinical conditions as well as to discuss their limitations, complications and future perspectives.

Methods: Pubmed and Medline was systematically searched for correlative publications.

Results: Promising at first, but over time its limitations became evident: demanding infrastructural requirements, high costs, lack of “ex-vitro” stability, additional requirement for dermal support, and the absence of other skin cell types or appendages have limited the introduction in daily clinical routine. Nonetheless, continuous efforts have been made in the past decades in order to improve the application of keratinocytes. Novel techniques of keratinocyte harvest and cultivation have simplified the clinical application, improved stability, and consequent outcomes.

Conclusions: The herein presented efforts despite their drawbacks during the development process and resultant clinical outcomes of keratinocyte culturing bear a solid foundation for the future. The ultimate challenge however remains, bringing any of these efforts into clinical practice while avoiding the failures of the past.

BiographyLars-Peter Kamolz is Professor for Plastic, Aesthetic and Reconstructive Surgery and Head of the Division of Plastic, Aesthetic and Reconstructive Surgery, Department of Surgery, Medical University of Graz. Furthermore, he is the Head of the Research Unit for Tissue Regeneration, Repair and Reconstruction at the Medical University of Graz.

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Lars-Peter Kamolz, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineNeural competent cells of adult human dermis belong to the Schwann lineageAnder Izeta1, Adrian Perez-San Vicente1, Usue Etxaniz1, Nuria Gago-Lopez2, Mario Garcia-Dominguez3, Ariane Aduriz4, Haizea Iribar1, Haritz Irizar1, Maider Munoz-Culla1, Virginia Perez-Lopez1, Izaskun Burgoa1, Ainara Vallejo1, Paz Lopez- Mato4, Angel G. Martin5, Olatz Leis5, David Otaegui1 and Robb MacLellan2

1Instituto Biodonostia, Spain2University of Washington, USA3CABIMER, Spain4Inbiomed Foundation, Spain5StemTek Therapeutics, Spain

Resident neural precursor cells (NPCs) have been reported for a number of adult tissues. Understanding their physiological function or, alternatively, their activation after tissue damage or in vitro manipulation remains an unsolved issue. Here

we investigated the source of human dermal NPCs in the adult. By following an unbiased, comprehensive approach of cell surface marker screening, cell separation, transcriptomic characterization and in vivo fate analyses, we found that p75NTR+ precursors of human foreskin can be ascribed to the Schwann (CD56+) and perivascular (CD56-) cell lineages. Moreover, neural differentiation potential was restricted to the p75NTR+CD56+ Schwann cells and correlated with Sox2 expression levels. Loss and gain of function experiments demonstrated that Sox2 levels dictate neural competence in dermal precursors and thus Sox2 is a major determinant of cell fate also in this system. Double positive NPCs were similarly obtained from human cardiospheres, indicating that this phenomenon might be widespread and underlie stromal NPCs previously described in diverse tissues.

BiographyAnder Izeta completed B.Sc. from Navarra University (1994) and PhD in Biology by Universidad Autonoma of Madrid (2000). The aim of his lab is to elucidate the ontogeny, expansion and differentiation capacity of precursor cells in mouse and human dermis, with the ultimate purpose of facilitating their therapeutic use through generation of tissue engineered constructs and help clarify their possible contribution to carcinogenic processes or other pathologies. Specifically, they are studying human skin-derived precursors (SKPs) and other dermal stem cells and their relationship to wound healing and aging, as well as their use in tissue engineering. They also do clinical wound healing research.

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Ander Izeta et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineBupivacaine crystal deposits after long-term epidural infusionIngrid BalgaUniversity Leipzig, Germany

The case of a 45-year-old male patient (body weight 52 kg, height 1.61 m) with a locally invasive gastric carcinoma infiltrating into the retroperitoneal space is reported. Because of severe cancer pain a tunnelled thoracic epidural catheter (EC) was

placed at thoracic spinal level 7/8 and a local anesthetic (LA) mixture of bupivacaine 0.25% and morphine 0.005% was infused continuously at 6 ml h−1. To optimize pain therapy the concentration was doubled (bupivacaine 0.5%, morphine 0.01%) 3 months later but the infusion rate was reduced to 3 ml h−1 thus the total daily dose did not change. The patient died 6 months after initiation of the epidural analgesia from the underlying disease. The total amount of bupivacaine infused was 69 g and of morphine 1.37 g. The patient never reported any neurological complications. The autopsy revealed large white crystalline deposits in the thoracic epidural space which were identified as bupivacaine base by infrared spectrometry. Morphine could not be detected. A histological examination showed unreactive fatty tissue necrosis within the crystalline deposits but nerve tissue could not be identified. It is concluded that the bupivacaine crystalline deposits arose due to precipitation but the clinical significance with regard to sensory level and neuraxial tissue toxicity is unknown.

Actually, only two human cases with complications of bupivacaine crystalline deposits after long-term application of local anesthetics are known.

BiographyIngrid Balga has completed her Dr. med. at the age of 27 years from Bern University, Switzerland. She worked as an anesthesiologist (main topics: adult and pediatric anesthesia, pain medicine) at Kantonsspital Lucerne, Switzerland and actually specialises as an anesthesiologist for cardiac anesthesia, Heart Centre Leipzig, Germany. She has published several papers with the topics of pain medicine, pediatric postoperative quality management and hematology.

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Ingrid Balga, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineCardiac telocyte therapy for infarcted myocardiumCai DongqingJi Nan University, China

Regeneration of damaged myocardium is still a big challenge in clinic. Stem cell therapy has shed light to regenerate the ischemic myocardium. However, low survival rate of transplanted stem cells, very low terminal differentiation of

transplanted stem cells, and serious fibrosis of infarcted zone limit therapeutic effects of stem cell to achieve functional and structural regeneration of ischemic myocardium. Recent studies have shown that supporting niche cells which are consisted in cardiac unit in myocardium might play an important role in regeneration of myocardium. A novel interstitial cell, named as telocyte, has been identified recently in heart interstitium. Recently we reported that cardiac telocyte (CTs) network in myocardium was impaired during myocardial infarction (MI). In addition, transplantation of CTs in both infarcted and border zone of myocardium simultaneously was able to decrease the infarct size and improve the myocardial function. Our up-to-date study further revealed that the midterm therapeutic effects (decrease of infarct size and improve myocardial function) of CTs transplantation for MI was maintained till 14 weeks. The cellular mechanism of therapeutic effects using CTs for MI was attributed to increasing of angiogenesis, improving CT network reconstruction and decreasing fibrosis in ischemic myocardium. The finding of our study suggested that therapeutic effects using CTs was able to maintained at least for 14 weeks. The CT therapy would be considered as one of the potential novel cell therapies for MI. The CTs have high potential to be applied to regenerate MI used alone or tandem stem cells.

BiographyCai Dongqing has completed his MD in Guangzhou Medical College; 1987, his PhD was done in The Chinese University of Hong Kong; 2000, Postdoctoral Associate in Weill Medical College of Cornell University [U.S.A.]; 2000-2003. He is a Professor and Director for Key Laboratory for Regenerative Medicine, Ministry of Education, Ji Nan University and also being Professor and Director for Department of Developmental and Regenerative Biology, Ji Nan University. His scientific interests are 1) Aging and microenvironment in regeneration of myocardial infarction (MI); 2) Cardiac vascular specific targeting and therapy (stem cell and therapeutic angiogenesis) for MI; 3) Aging and regeneration of Tissue & Organ. Grant: 2003-present: 863, International collaboration grant of Ministry of Science & Technology, five NSFC-grants, two Key grant of GDZRKXJJ and other five Guangdong and Guangzhou government grants. He has published 40 SCI papers included: JCMM, Proteomics, Am J Physiol Heart Circ Physiol and Physiological Genomics etc.

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Cai Dongqing, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineRegenerative potential of outer root sheath melanocytes in tissue graftsMarie SchneiderUniversity of Leipzig, Germany

Our group at the Translational Centre for Regenerative Medicine in Leipzig developed a method for melanocyte cultivation from the Outer Root Sheath(ORS) by expanding stem cells, precursors and dedicated cells and differentiating them

into melanocytes. Based on this methodology, we are developing an autologous, transplantation-based, causal treatment of depigmentation disorders such as Vitiligo. In the scope of cultivation, we are tracking shifts in gene expression profiles, from pluripotency in undifferentiated cells to melanotic status in late differentiation stages of the ORS melanocyte culture.

For the purposes of melanocyte transplantation, a biocompatible, biodegradable graft carrier is needed. In order to identify candidates for biocompatible as well as mechanically stable niche, we are working on a palette of scaffolds, testing function and development of 3D cell cultures compared to 2D adherent culture.

Melanocytes were cultivated from plucked hair follicles by the means of an optimized explant method by Savkovic et al. Differentiated cells were characterized for expression of melanocyte markers and melanin production. ORS melanocytes were seeded and cultivated on different types of scaffolds– Polycaprolactone (PCL) microspunscaffold, decellularized bovine collagen backbone called Collagen Cell Carrier (CCC), as well as hydrogel produced of shortened, chemically cross-linked bovine collagen chains named Collagel – for a cultivation period of one week. Expression of melanocyte markers was assessed by immuno-fluorescence and gene expression was analyzed by the means of quantitative real-time PCR.

Gene expression of the ORS cells differentiating into melanocytes shifted from pluripotency-like profiles in early cultivation passages to melanocyte-like profiles in late passages. These results corroborate the hypothesis that the ORS melanocytes descend from the ORS stem cell and progenitor sub-pool and possess a high developmental potential within the follicle and in early passages of primary culture. ORS melanocytes showed correct melanocyte marker expression and displayed melanotic features in adherent cultures, which were potentiated on 3D scaffolds, especially on Collagel scaffolds. Their architecture and mechanical stability promote melanocyte features mimicking the native structure of the extracellular matrix, which makes them exceptionally good candidates for biomimetic graft carriers.

BiographyMarie Schneider has completed his Doctor in Translational Centre for Regenerative Medicine in 2013 and Master Thesis, in TRM Leipzig (Group Dr.

Savkovic) on “Gene expression and melanotic properties of the hair follicle outer root sheath melanocytes in varying culture conditions”. He is a Full Professor in the University of Leipzig, Gemany. The Bachelor Thesis was done in year 2011 Fraunhofer IZI Leipzig under the supervision of Dr. Lehmann. The High School Degree was done in Abitur during 2008.

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Marie Schneider, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThe impact of histological re-evaluation of tissue engineered startegies in articular cartilage repair -Results of human applicationsChristoph BrochhausenUniversity Medical Centre Mainz, Germany

Several tissue engineering based strategies to treat articular cartilage defects – such as autologous chondrocytes transplantation (ACT) or matrix assited autologous chondrocyte transplantation (MACT) - are already in clinical use. However, none of

them provides optimal results with view to regeneration of hyaline cartilage. An innovative new strategy to treat articular cartilage lesions is the implantation of autologous chondrocytic spheroids. We present histological re-evaluations of second-look biopsies of MACT patients and for the first time such of autologous chondrocytic spheroid application. Biopsies were taken from 6 patients treated with Standardized synthesized MACT-constructs or autologous chondrocytic spheroids. The Second-look biopsies were taken between four and sixteen months after implantation during a second arthroscopy, which was indicated independent of the (ACT). Biopsies (n=2) were fixed in buffered formalin and prepared according to standard methods for conventional histological (HE, alcian-blue) and immunohistochemical (collagen II, aggrecan) examination followed by semiquantitative evaluation. Biopsies from MACT-constructs showed fibrous tissue without relevant collagen II or aggrecan expression. Biopsies from spheroid based autologous chondrocyte transplantation showed typical articular cartilage architecture with expression of collagen II and aggrecan. To our knowledge this study is the first histomorphological evaluation of scaffold-free spheroid based ACT in humans. The heterogeneous results of our re-evaluations demonstrated the need for a systematic biopsy-collection and their combination with clinical data to provide an evident evaluation of the used tissue engineering based strategies to treat cartilage defects. Furthermore, such analyses could give an innovative input in further developments of the guided tissue engineering of cartilage defects.

BiographyChristoph Brochhausen is Chief Consultant Pathologist and Head of Electron Microscopy at the Institute of Pathology at the University Medical Centre Mainz as well as Group Leader of the Cartilage Tissue Engineering Group at the REPAIR-lap. His research-work was awarded by national and international awards. He has published more than 80 papers in reputed journals and serving as an editorial board member of some reputed journals.

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Christoph Brochhausen, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineCritique in vitro analysis of platelet concentration effect on cells involved in in vivo wound healingVincenza DoloUniversity of L'Aquila, Italy

Over time the role of platelet derivatives (firstly Platelet Rich Plasma and Platelet Gel) in wound healing processes has become more clear; its clinical use has been investigated for several pathologies, ranging from chronic skin and soft

tissue ulcerations treatments to several surgical applications (oral-, maxillofacial-, orthopedic and trauma-, plastic-surgery). Despite this, their use has progressed in an “empirical” fashion, only sustained by individual doctors’ experiences but not by experimental data. Thus, the need of strict in vitro experiments to possibly identify the parameters that should be used in clinical applications to obtain satisfactory results in wound healing. In our opinion, among more useful parameters, the concentration of platelet seems to be very important even if it is, usually, ignored. That being so, our purpose was to evaluate the in vitro effect of different platelet concentrations on biological processes used by cells during the in vivo wound healing. Several cell types were assessed, taking into account their role during wound healing processes; pro-angiogenic response was analyzed in human endothelial cells treated with increasing concentrations of platelet per microliter, and ability to restore extracellular matrix lost after tissue damage was evaluated in human fibroblasts and human tenocytes subjected to the same treatments. For all cell lines assessed, our data highlighted that exists a specific optimal concentration for each process and each cell but, in any case, excessively high concentrations (over 1.500.000 platelet/μl) have an inhibitory effect on the wound healing processes and could be, therefore, counterproductive in clinical applications.

BiographyVincenza Dolo has completed his PhD in Cellular Biology from Palermo University and postdoctoral studies from L’Aquila University School of Medicine. She is the Director of Post-graduate School of Clinical Pathology, and Chairman of Master Degree in Health Professions Sciences. She has published more than 67 papers in reputed journals.

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Vincenza Dolo, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThe roles of autophagy and apoptosis by spermidine in burn wound progression in ratsDaryousch ParviziMedical University of Graz, Austria

Introduction: Spermidine is a naturally occurring polyamine involved in multiple biological processes, including DNA metabolism, autophagy and aging. Spermidine induces autophagy in cultured yeast and mammalian cells, as well as in nematodes and flies. Genetic inactivation of genes essential for autophagy abolishes the life span-prolonging effect of spermidine in yeast, nematodes and flies. These findings complement expanding evidence that autophagy mediates cytoprotection against a variety of noxious agents and can confer longevity when induced at the whole-organism level. In an acute burn injury the zone of stasis is initially vital but may progress to coagulation necrosis with time. In this experimental study, we hypothesized that apoptosis, autophagy induced by spermidine plays a role in burn wound progression.

Material and Methods: Healthy adult male Sprague Dawley rats (300–350 g, n = 40) were provided by the Institute for Biomedical Research, Medical University of Graz, Austria. A well-described and reliable “scald burn model with dressing protector” was employed to evaluate the roles of autophagy and apoptosis by spermidine in burn wound progression 5 days post-burn in a rat model. In two groups (each 8 rats) spermidine was injected intraperitoneally in two different concentrations thirty minutes and 48h after burn injury. Furthermore two groups (each 8 rats) were treated by a silicone foam dressing, where spermidine was injected in two different concentrations, over the burnt area. Animals in the control group (n= 8) were given the same amount of saline without spermidine. The animals were euthanized five days after burn injury and burn depth was assessed by hematoxylin and eosin staining.

Results: In both treated groups (intraperitoneally and topical) a significantly higher rate of inflammation with granulation tissue in the deep dermal and muscular layer than that of the control group was found. Furthermore lower burn wound progression, by less swelling collagen and edema in the dermal and muscular layer, was seen in the spermidine groups than in the control group.

Conclusion: Histological results of this study confirm that spermidine treatment has a statistically significant benefit in burn wound progression and survival of the stasis zone in an acute burn injury.

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Daryousch Parvizi, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineA combinatorial relative mass value evaluation of bioactive factors in degenerative cervical and lumbar discs: New indications for gene therapeutic approaches of spinal disc degenerationDemissew S. MernInnsbruck Medical University, Austria

Degenerative intervertebral disc disease is common after middle age and can cause loss of disc height with painful nerve impingement, bone and joint inflammation. Despite the clinical importance of these problems, the pathology of cervical

disc degeneration were studied merely from a morphologic view point using magnetic resonance imaging (MRI), without addressing the issue of biological treatment approaches. Moreover, a wide range of endogenously expressed bioactive factors in degenerative cervical and lumbar discs were not investigated. Nucleus pulposus (NP) cells play a central role in intervertebral disc maintenance by organizing the expression of anabolic, catabolic, anti-catabolic and inflammatory cytokines that affect the extracellular matrix. Intervertebral disc degeneration is associated with imbalances of these factors, resulting in a catabolic inflammatory metabolism. Although degenerative lumbar discs have been targeted by different biological treatment approaches, the quantities of disc cells and the concentrations of gene therapeutic factors used in animal models fluctuate extremely. These indicate lack of experimentally acquired data regarding disc cell proliferation and levels of therapeutic targets, which are vital for rational gene therapeutic approaches. Therefore, we analyzed proliferation rates of degenerative NP cells and their endogenous expression levels of anabolic, catabolic, anti-catabolic and inflammatory matrix proteins.

Grades of intervertebral disc degeneration were determined by preoperative MRI. Then grade III, IV and V disc tissues were isolated from 78 patients operated due to spinal disc herniation (63 lumbar, 15 cervical, mean age 56 / range 29 - 84 years). NP cells were cultured for four weeks with low-glucose in collagen I scaffold. Their proliferation rates were analyzed using 3-(4, 5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide. The protein expression levels of 28 therapeutic targets were analyzed using enzyme-linked immunosorbent assay.

During progressive grades of degeneration NP cell proliferation rates remained similar in all groups, independent of gender, age and grades of degeneration. Significantly decreased aggrecan and collagen II expressions (P < 0.0001) were accompanied by accumulations of selective catabolic and inflammatory cytokines ADAMTS-4, ADAMTS-5, MMP-3, IL-1β and IL-1R combined with low expression level of the anti-catabolic factor TIMP-3 (P < 0.0001). However, the concentration of the anabolic factors BMP-2, BMP-4, BMP-6, BMP-7, IGF-1, TGF-β1 and TGF-β3 remained below the minimal detectable quantities.

Hence, gene therapeutic interventions regulating relevant bioactive factors identified in this work might contribute to the development of regenerative treatment approaches and inhibit inflammatory catabolism of intervertebral discs.

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Demissew S. Mern, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineNanostructured electrically active scaffolds for neural and muscle regenerationRob M. I. Kapsa1,2, Anita F. Quigley1,2, Joselito M. Razal1, Javad Foroughi1, Magdalena Kita1,2, Catriona Sinclair1,2, Rohoullah Jalili1, Raquel Ovalle-Robles3, Simon Moulton1, Ray H. Baughman3 and Gordon G. Wallace1

1University of Wollongong, Australia2St. Vincent’s Hospital, Australia3University of Texas at Dallas, USA

Neural and muscle regeneration requires platforms that facilitate the guidance and maintenance of differentiated cells. Various laboratories have been investigating the use of micro and nanostructured polymer scaffolds for control of cell

growth, cell guidance and tissue regeneration. In this presentation, we describe our investigations into the effects of orientated nanostructured conducting polymers on muscle and neural cell behavior, as well as the effects of electrical stimulation applied through these scaffolds.

Conducting nanostructured platforms were synthesized from aligned Multi Walled carbon NanoTube (MWNT) aerogel sheets deposited on gold coated mylar, as previously described. Conductive polypyrrole doped with para-toluenesulphonic acid (PPy-pTS) was deposited over the aligned carbon nanotube aerogel sheets, to create nanostructured PPy-pTS surfaces as required. DRG neural explants, PC12 cells or primary murine myoblasts were then grown on the scaffolds under differentiating conditions. Electrical stimulation (bipolar waveform) was applied for 8 hours per day for three days to analyse the effects of electrical stimulation on PC12 and myoblast behavior.

DRG neural explants and PC12s grown and differentiated on these substrates demonstrated a high degree of alignment, dependent on the thickness of the PPy-pTS overlay. Similarly, myoblasts and myotubes exhibited a high degree of alignment on nanostructured surfaces, this effect was also dissipated with increasing thickness of the PPy-pTS overlay. Electrical stimulation of myoblasts on the scaffolds resulted in a small but significant increase in myoblast fusion and cell density. In addition, PC12s electrically stimulated on these platforms exhibited an increased number of longer neurites, suggesting that electrical stimulation applied through conducting polymer platforms can increase neurite outgrowth, as previously observed in primary neural explants.

These studies demonstrate that nanostructured conducting polymer platforms can be used to influence neural and skeletal muscle differentiation ex vivo. In addition, these platforms can influence myofibre and neurite orientation in a manner reflecting the in vivo architecture of the parent tissue. Such platforms have applications in controlling the regeneration of skeletal muscle and neural tissues in vivo and for the integration of bionic devices designed to facilitate tissue regeneration and function.

BiographyRob M. I. Kapsa completed his PhD in 1996 at the University of Melbourne Dept Medicine, St Vincent’s Hospital (Melbourne). He is Program Director for the Bionics Platform of the ARC Centre for Electromaterials Science (ACES) and concurrent Principal Scientist and Head of Research Neurosciences at St Vincent’s Hospital in Melbourne. He has published 85 peer-reviewed manuscripts including 2 book chapters and one book in the areas of muscle biochemistry, genetics, gene therapy and muscle and nerve engineering. His work is focused on the development of autologous cell-based regenerative therapies for hereditary nerve and muscle disease.

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Rob M. I. Kapsa et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineAmniotic derived progenitor cells in different animal species in view of cell therapy applicationsAnna Lange-ConsiglioLarge Animal Hospital University of Milan, Italy

The human amnion membrane is a source of mesenchymal stem cells (AMCs). For the first time, AMCs in horse, cow, dog and cat were characterized by immunocytochemical studies which showed the expression of specific embryonic markers

(TRA-1-60, SSEA-3, SSEA-4 and Oct-4) and by molecular studies which proved the positivity to CD105, CD73, CD90, CD29, CD166, and CD44, confirming the stemness of these cells. AMCs showed high proliferative capacity for prominent telomerase activity and high differentiate potential toward osteogenic, adipogenic, chondrogenic and neurogenic lineages (mesodermic and ectodermic).

Moreover, equine AMCs were compared to bone marrow-derived cells (BM-MSCs) demonstrating, in vitro, higher proliferative and differentiative potential and, in vivo, lower rate (4%) of re-injury when allogenic cryopreserved AMCs were, for the first time, transplanted in spontaneous tendon injuries in horses compared to fresh autologous BM-MSCs group (23.08%). Moreover, horse AMCs have immunomodulatory capacity demonstrated by their ability to inhibit PBMC proliferation not only when cultured in cell–cell contact with responder cells but also when separated from them by a transwell membrane. This ability could be attributed to soluble factors released by AMCs. This hypothesis was further supported by the inhibition of the PBMC proliferation exerted by the conditioned medium secreted by AMCs (AMC-CM). When AMC-CM was used to treat spontaneous horse tendon and ligament injuries, regenerative/reparative responses were achieved. Our outcomes demonstrated that the AMCs transplantation results in improved tendon healing and that the AMC-CM could be a novel therapeutic biological cell-free product in spontaneous tendon and ligament diseases.

BiographyAnna Lange-Consiglio is a Doctor in Animal Science. She completed her PhD from the Veterinary School of the University of Milan and she is in charge of the Reproduction Laboratory at the Large Animal Hospital of University of Milan. Her field of research is focused on stem cells-based regenerative medicine and reproductive biotechnologies, as in vitro fertilization and cloning, with the purpose of in vivo and in vitro studies and clinical trials. Lange-Consiglio collaborates with Italian and foreign research groups. She has published more than 40 papers in international journals with impact factor and peer review.

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Anna Lange-Consiglio, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineOsteo differentiation of intact amniotic membrane and outcome for tissue engineering Florelle GindrauxUniversity Hospital of Besancon, France

The human amniotic membrane (hAM) is known to have a good potential for tissue regeneration, as a scaffold containing mesenchymal stromal cells (MSC) and growth factors, with low immunogenicity, anti-microbial, anti-inflammatory, anti-

fibrotic and analgesic properties. Several case reports relate its use for oral and maxillofacial surgery (vestibuloplasty; guided bone regeneration and buccal defects).

Based on previous findings reporting the possibility to osteodifferentiate whole tissue (Lindenmair et al. 2010), we aimed to use hAM as a bioactive membrane for bone repair. In this study, we sought to confirm its in vitro osteogenic potential and to observe the effects of the culture on cell function and phenotype. At the same time, we aimed to develop simple and rapid phenotypic and functional controls, which could be easily implemented in a tissue bank, in order to qualify cultured hAM. Finally, we evaluated the in vivo biocompatibility of cultured hAM.

Results showed that hAM could be osteo differentiated in vitro by an osteogenic medium and presented an apparently osteocyte phenotype. Moreover, the culture, especially in this condition, led to structural alterations of epithelium related to modification of cell function, very probably beneficial for bone repair. We report that phenotype could be easily determined by von Kossa staining and that cultured, especially osteo differentiated, hAM did not involve in vivo allogenic reaction, testifying to its good biocompatibility.

BiographyFlorelle Gindraux graduated with Masters (in 2001) and a PhD degree (2007) from the University of Franche-Comté (Besançon, France). She performed a post-doctoral internship for 2 years as R&D Project manager with TBF (Tissue Bank of France) in Mions-Lyon, France, where she developed her Ph.D. project. i.e. the development of an advanced therapeutic medicinal product for bone repair using Mesenchymal Stem Cells (MSCs). She is specialized in bone repair (for orthopaedic and maxillo-facial surgeries) and more generally for other tissue repair (cartilage, ligament, tendon, dental pulp, teeth, skin). For the last 6 years, she has been working at the University Hospital of Besançon and the University of Franche-Comté where she combines R&D with clinical research on MSCs (from perinatal tissue) and biomaterials for tissue regeneration. Florelle Gindraux is currently EORS (European Orthopaedic Research Society) Ambassador for France and is co-organizing the next meeting to be held in Nantes in July 2014. She has authored 35 international publications (6 Pubmed cited); 14 Book chapters; 2 Books; as well as 27 international & 34 national communications (as presenting author). Florelle Gindraux has also obtained 3 awards for her previous work.

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Florelle Gindraux, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineCollagen plus elastin dermal substituteLars-Peter KamolzMedical University of Graz, Austria

The gold standard for the coverage of full-thickness skin defects are autologous skin grafts. However, poor skin quality and scar contracture are well-known problems in functional, highly strained regions. The use of dermal substitutes is an

appropriate way to minimise scar contraction and, thereby, to optimise the quality of the reconstructed skin. The aim of this presentation is to present our long time experiences with a collagen-elastin matrix, Matriderm. All patients (n=70) with full-thickness skin defects were treated with the dermal substitute, Matriderm, and a skin graft. The take rate of the matrix-and-skin graft was 96%. Long-term follow-up revealed an overall Vancouver scar scale of 1.7. No limitation concerning function were observed.

This matrix represents a viable alternative to other types of defect coverage and should therefore be considered in the treatment of skin injuries, especially in very delicate regions such as the joint regions. The possibility of performing a one-stage procedure is supposed to be a major advantage in comparison to a two-stage procedure.

BiographyLars-Peter Kamolz is Professor for Plastic, Aesthetic and Reconstructive Surgery and Head of the Division of Plastic, Aesthetic and Reconstructive Surgery, Department of Surgery, Medical University of Graz. Furthermore, he is the Head of the Research Unit for Tissue Regeneration, Repair and Reconstruction at the Medical University of Graz.

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Lars-Peter Kamolz, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineThe Polycomb group protein L3MBTL1 represses a smad5-mediated hematopoietic transcriptional program in human pluripotent stem cellsFabiana PernaMolecular Pharmacology and Chemistry Program, USA

Epigenetic regulation of key transcriptional programs is a critical mechanism of hematopoietic development and, aberrant epigenetic patterns and mutations frequently occur in hematologic malignancies perturbing hematopoietic stem cells. We

demonstrate that the Polycomb protein L3MBTL1, which is monoallelically deleted in 20q- myeloid malignancies, represses the ability of stem cells to drive hematopoietic-specific transcriptional programs by regulating the expression of smad5 and impairing its recruitment to target regulatory regions. Indeed, knock-down of L3MBTL1 promotes the development of hematopoiesis and impairs neural cell fate in human pluripotent stem cells. Furthermore, we found that regulation of smad5 targets by L3MBTL1 also occurs in mature hematopoietic cell populations, affecting erythroid differentiation. Altogether, these findings define an epigenetic mechanism underlying priming of hematopoietic-specific transcriptional networks and provide rationale for development of therapeutic approaches for patients with anemia.

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Fabiana Perna, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineCord blood banking. Understanding the critical quality attributes (CQAs) for cell product development and manufactureAndreea Iftimia ManderAnthony Nolan Cell Therapy Center, UK

Cord blood banks are a potential source of cellular material for the emerging cell based therapy industry. Umbilical cord blood (UCB) is now a well-recognized source of hematopoietic stem cells and it has a widespread use for transplantation.

It has several advantages over bone marrow or peripheral blood, including increased tolerance for Human Leukocyte Antigen mismatches, decreased incidence of graft-versus-host disease (GVHD), and easy availability. These advantages make UCB a desirable cell therapy treatment in the growing need of suitable grafts for patients with hematologic malignancies or underlying bone marrow or metabolic defects. It is also important to note that a significant population of patients, specifically minority groups, lack a suitable HLA-matched bone marrow, or peripheral stem cell donor, consequently UCB is able to fill this gap as an important source of allografts for such groups. Umbilical cord tissue (UCT) banking is also increasing in suchfacilities as a source of mesenchymal stem cells for future therapeutic use. However, earlyhandling of the placenta and umbilical cord is relatively uncontrolled due to the clinical demands of the birth environment and subsequent transport logistics. It is therefore necessary to develop extractionmethods that are robust to real world operating conditions, rather than idealised operation. Also in a recent study performed by our cord blood bank with the help of 50 members of the World Marrow Donor Association (WMDA), a gap in theevaluation of the critical quality attributes (CQAs) of a cord blood unit (CBU) used for transplantation was identified. The results of this study emphasize that a better understanding and communication of UCB parameters and their limitations needs to be established between the Cord Blood Providers Group (CBPG) and the Cord Blood Selectors Group (CBSG). This would allow the CBPG to set parameters that can be translated into standardised testing methods, with proven cross centre reproducibility, and with long standing correlations to engraftment.

BiographyAndreea Iftimia-Mander is an interdisciplinary scientist with a wide range of expertise in the fields of regenerative medicine and chemical engineering, organic synthesis. She has completed her PhD at the Centre for Biological Engineering, Loughborough University, Loughborough, UK, in the field of Regenerative Medicine. Currently she is the Clinical Processing Manager for Anthony Nolan Cell Therapy Centre in Nottingham, UK.

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Andreea Iftimia Mander, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineIn situ characterization of tissue constructs using a multimodal optical imaging/spectroscopy approachNicusor IftimiaPhysical Sciences, Inc., USA

Currently, most of the engineered tissue constructs are characterized at the scaffold level for biocompatibility, toxicity, porosity for cell infiltration, mechanical strength, compliance, toxicity, etc., using partially or totally invasive and

destructive procedures. Additionally, two-dimensional basic imaging is often performed to determine cell maturation and growth. However, more precise spatial and temporal assessment of the growing 3D tissue is required during in vitro culture and after implantation. Unfortunately, tissue-specific analysis is complicated by the complexity of the cellular environment. Several cell types can be utilized in the engineered construct, targeted to perform multiple functions. As an example, muscle progenitor cells differentiate to form myofibers in a soft tissue construct. Co-seeded fibroblasts express soluble factors to maintain muscle viability, enhance differentiation, and produce extracellular matrix critical to tissue function. Endothelial cells and supporting stromal cells are co-seeded to form extensive endothelial-lined networks which readily connect with host blood vessels upon implantation. These constructs are typically cultured in vitro under electrical or dynamic stimulation. Although the 2D cell cultures can be characterized with a simple microscope working in the reflectance or fluorescence mode, imaging of cells in 3D is significantly more challenging. Cells must be fluorescently labeled and few compounds are designed for live cell imaging. Fluorescent dyes or fluorophore-linked antibodies are usually toxic and require permeable cell membranes for cell entry. If multiple cell types are used, they must be separately labeled prior to seeding. Cells are frequently genetically modified to express a fluorescent protein. Collagen-based biological scaffolds and many polymeric scaffolds are strongly auto fluorescent, making the cells and the supporting scaffold difficult to distinguish.

Physical Sciences Inc. (PSI), in collaboration with the Center for Regenerative Medicine at the Massachusetts General Hospital have preliminarily evaluated the use combined high-resolution Scanning Optical Coherence Microscopy (SOCM) and Raman spectroscopy for real-time three-dimensional quantitative assessment of growing ex vivo engineered soft tissue construct noninvasively in order to assess the most important parameters needed to determine its readiness for transplantation: viability, microstructure, vascularization, and maturity. The main benefit of our approach is that it provides very useful and comprehensive data sets in real time noninvasively, while no part of the manufactured tissue is destroyed. SOCM provides cross-sectional and three-dimensional structural maps of the tissue at the micron scale to depths of at least 1.5 mm, and thus to accurately assess the morphology of the tissue. It combines the depth sectioning capabilities of Optical Coherence Tomography (OCT) with the lateral sectioning capability of confocal microscopy (CM). Raman spectroscopy on the other side provides details about the biochemical composition of the tissue, allowing for determining its viability status before implantation.

BiographyNicusor Iftimia is a Pr. Scientist at Physical Sciences, Inc- USA. His major research interests include disease diagnosis, regenerative medicine, and therapy guidance. He applies various optical imaging and spectroscopy modalities to buid complex instrumentation that can be used in the above-mentioned areas. He is the author of over 70 peer-review papers, numerous conference presentations, several book chapters, and the editor of two books. He also serves as reviewer and editorial board member on several peer-review journals, such as Molecular Imaging and Dynamics, International Journal of Optics, Journal of Medical Engineering, Journal of Optoelectronics and Advanced Materials, etc.

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Nicusor Iftimia, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineMultimodal multi component polymer conduits for repair of nerve injuryRob M.I. KapsaUniversity of Wollongong, Australia

Injury to nerve tissue in the peripheral nervous system (PNS) results in long-term impairment of limb function, dysaesthesia and pain, often with associated psychological effects. Minor injuries can regenerate without intervention and short gaps

can be repaired by end joining, however larger or more severe injuries commonly require autogenous nerve grafts which often result in suboptimal clinical outcomes. The challenges that persist with nerve repair have resulted in development of synthetic nerve guides from non-neural biological tissues and polymers to improve the prognosis for repair of damaged nerves. This study describes the design and fabrication of a nerve regeneration conduit, synthesised using polylactic acid and poly (lactic-co-glycolic) acid co-polymers, in addition to aneurotrophin-enriched hydrogel. The conduit was used to promote repair of transected sciatic nerve in rats over a period of 4 weeks. Over this period, it was observed that over-grooming and self-mutilation (autotomy) of the limb implanted with the conduit was significantly reduced in rats implanted with the full-configuration conduit compared to rats implanted with conduits containing only an alginate hydrogel. This indicates return of some feeling to the limb via the fully-configured conduit. Immuno histo chemical analysis of the implanted conduits removed from the rats after the implantation period confirmed the presence of myelinated axons within the conduit, distal to the site of implantation, further supporting that the conduit promoted nerve repair over this period of time. This study describes the design and fabrication considerations of a novel multicomponent, multimodal bio-engineered synthetic conduit for peripheral nerve repair.

BiographyRob M.I. Kapsa completed his PhD in 1996 at the University of Melbourne Dept. Medicine, St Vincent’s Hospital (Melbourne). He is Program Director for the Bionics Platform of the ARC Centre for Electromaterials Science (ACES) and concurrent Principal Scientist and Head of Research Neurosciences at St Vincent’s Hospital in Melbourne. He has published 85 peer-reviewed manuscripts including 2 book chapters and one book in the areas of muscle biochemistry, genetics, gene therapy and muscle and nerve engineering. His work is focused on the development of autologous cell-based regenerative therapies for hereditary nerve and muscle disease.

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Rob M.I. Kapsa, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineStem cell behavior in development and regeneration: To balance or imbalanceAhmed HK El-HashashUniversity of Southern California, USA

A defining feature of stem cells is their ability to continuously maintain a stem cell population (i.e. self-renew) while generating differentiated progeny. Both embryonic and tissue-specific stem cells are faced with a uniquely difficult task:

to avoid cell cycle exit and differentiation, and to avoid uncontrolled proliferation and tumor formation. Therefore, stem cells are required to maintain a proper balance between cell gain (self-renewal) and cell loss (apoptosis or differentiation). Loss of this balance leads to severe developmental defects and adult diseases, including cancer and fibrosis, all of which are highly costly and morbid medical conditions. How stem cells walk this developmental tightrope is an extremely interesting question that is of relevance to our understanding of the processes of cell differentiation and cancer, and of the developmental diseases that result from the premature loss of stem cell pools. Molecular programs regulating the balance between the self-renewal and differentiation and the balance of apoptosis versus self-renewal/differentiation of endogenous organ-specific stem cells are likely critical both to development and to regenerating diseased and damaged tissues. Recent studies from our laboratory have identified the importance and molecular mechanisms of several transcription factors and protein phosphatases in balancing self-renewal versus differentiation or self-renewal versus apoptosis of tissue-specific stem cells. Using the lung as a model organ, we discovered that two classes of protein phosphatases (PTPs): Asp-based PTPs and non-receptor PTPs are essential for balancing self-renewal versus differentiation, as well as self-renewal versus apoptosis, respectively, of lung-specific stem cell during embryonic and adult stages. These enzymes do this by controlling the activity of the Par polarity complex and Notch1 signaling activity. Conditional genetic deletion of Asp-based PTPs in murine lungs results in loss of the balance between self-renewal and differentiation of lung-specific stem cells, leading to lung hypoplasia in mice. Conversely, conditional genetic deletion of non-receptor PTPs results in loss of the balance between self-renewal and apoptosis of these stem cells, leading to lung hypoplasia during embryogenesis or pulmonary fibrosis in adult mice. This imbalance between self-renewal and differentiation or apoptosis can be restored by intratracheal administration of these phosphatases or by genetic activation of downstream signal pathways such as Notch1, which leads to alleviations of lung hypoplasia during embryogenesis as well as reversing of pulmonary fibrosis in adult mice. These novel findings signify that PTP deficiency-mediated fibrosis is a feasible therapeutic target. They will also help in devising new therapeutic approaches to understand and eventually correct pulmonary fibrosis that is a lethal disease affecting 5 million people world-wide, with an average survival of 2-3 years following diagnosis and without an approved-treatment till now.

BiographyAhmed HK El-Hashash has completed his PhD from Manchester University, UK and postdoctoral studies from Mount Sinai School of Medicine of New York University and Children’s Hospital Los Angeles. He is currently Assistant Professor of Stem Cell Biology and Regenerative Medicine at Keck School of Medicine and Ostrow School of Dentistry of University of Southern California. He has published more than 20 papers in reputed journals and serving as an editorial board member of repute.

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Ahmed HK El-Hashash, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineConcept of cancer stem cells in oral squamous cell carcinomaGokul SridharanYMT Dental College and Hospital, India

Oral squamous cell carcinoma (OSCC) is an epithelial malignancy commonly affecting developing countries. Disease progression is due to a complex molecular mechanism resulting in accumulated genetic mutations causing uncontrolled

cell proliferation. These genetic alterations are primarily evident in somatic stem cells and recently, it has been demonstrated that a subset of these cells referred to as cancer stem cells play an important role in cancer progression.

Cancer stem cells are defined as a small subset of cells with the capability of self renewal and differentiation into the heterogenous lineages that constitute the tumor mass. These cells were found to be highly tumorigenic with capability of self renewal and behaviour akin to tumor progenitor cells. The origin of cancer stem cells could be from normal somatic stem cells, dedifferentiated tumor cells, fusion of tumor epithelial cells with hematopoitic stem cells or due to the process of epithelial-mesenchymal transition. Other minor factors giving rise to cancer stem cells could be human papilloma virus, tobacco habits, neosis and tumor microenvironment which act as stem cell niche. Identification of cancer stem cells could be done by markers such as CD44, CD133, E-cadherin; Wnt signaling pathway, Oct-4 and ABCG2. Cancer stem cells are responsible for tumor recurrence, metastasis and resistance to chemotherapy.

The concept of cancer stem cells and its association with OSCC is relatively new and detailed study is necessary for its better understanding. This review highlights the relevant findings on cancer stem cells, its origin, identification and its role in OSCC.

BiographyGokul Sridharan completed is Masters in Oral Pathology (Dentistry) in 2009 and currently working as lecturer in the department of Oral Pathology. He is currently pursuing in PhD with the study titled “Oxidative stress in leukoplakia and oral squamous cell carcinoma”. He has several publications in national and international journals and serving as an editorial board member of reputed journals.

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Gokul Sridharan, J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014






Tissue Science & Regenerative MedicineHuman serum as an alternative to fetal bovine serum in adipose tissue-derived stromal cells culturesAna Cláudia C Paula1, Thaís M M Martins1, Alessandra Zonari1, Soraia P P J Frade2, Patrícia C Angelo2 and Alfredo M Goes1

1Universidade Federal de Minas Gerais, Brazil 2Instituto Hermes Pardini, Brazil

Multiple clinical trials are underway to evaluate the use of adipose tissue-derived stromal cells (hASC) for regenerative medicine. Nevertheless, prior to its clinical application, hASC need ex vivo expansion in compliance with GMP

guidelines in order to obtain a suitable amount of cells. Fetal bovine serum (FBS) is the most widely used serum in standard culture conditions, but the use of animal origin components bears disadvantages and is not an option in clinical cell therapy due to several safety issues. So our aim was to show the effectiveness of pooled allogeneic human serum (aHS) as a supplement for hASC culturing. In this study, the hASC were expanded in DMEM supplemented with aHS and the immunophenotype and differentiation potential of the cells were evaluated. The first difference observed between FBS and aHS cultures was that hASC in aHS proliferated markedly faster than in FBS, which was confirmed by Ki-67 expression, the cumulative population doubling, and by a growth curve. To better understand this hASC expansion pattern, the transcription factors C-myc and C-fos as well as mRNA levels of hTERT, and pluripotency genes (OCT-4 and NANOG) were assessed. No significant expression was observed and the hASC cultured in aHS underwent replicative senescence with normal karyotype. These cells assayed were not able to generate a teratoma. Therefore, this study confirms and extends the benefits of rapid expansion of hASC in culture media supplemented with aHS, with the cells maintaining their phenotype and with no spontaneous cell transformation.

BiographyAna Cláudia Chagas de Paula at age of 26 years is at the end of her PhD at Laboratory of Cellular and Molecular Immunology of Federal University of Minas Gerais (Brazil). Her areas of research interest are in cell biology and immunological approaches for cell culture, mainly stem cells, as well as development of novel tissue engineering strategies for bone replacement. She has published more than 5 papers in reputed journals in the last 2 years.

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Ana Cláudia C Paula et al., J Tissue Sci Eng 2014, 5:3http://dx.doi.org/10.4172/2157-7552.S1.014

involvement of the caspase enzymes in satellite cells

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