Tissue Engineering: Scaffold MaterialsBY: ELAHEH ENTEZAR-ALMAHDI

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References:Hollander AP, Hatton PV. Biopolymer methods in tissue engineering: Springer; 2004.

Shoichet MS, Hubbell JA. Polymers for tissue engineering. Journal of Biomaterials Science,Polymer Edition. 1998;9(5):405-6.

Yang S, Leong K-F, Du Z, Chua C-K. The design of scaffolds for use in tissue engineering. Part I.Traditional factors. Tissue engineering. 2001;7(6):679-89.

Shoichet MS. Polymer scaffolds for biomaterials applications. Macromolecules. 2009;43(2):581-91.

Araci IE, Brisk P. Recent developments in microfluidic large scale integration. Current opinion inbiotechnology. 2014;25:60-8.

Coluccino L, Stagnaro P, Vassalli M, Scaglione S. Bioactive TGF-β1/HA alginate-based scaffoldsfor osteochondral tissue repair: design, realization and multilevel characterization. Journal ofapplied biomaterials & functional materials. 2016;14(1).

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Characterizing Regenerative Medicine

1.Regenerative medicine is a broad definition for innovative medical therapies that will enable thebody to repair, replace, restore and regenerate damaged or diseased cells, tissues and organs. (MayoClinic)

2.Tools and Procedures (Biofabrication or Additive Manufacturing) of Regenerative Medicine

Tissue Engineering: Tissue Repair/Replacement and Lab Grown Organs

Technologies

Stem cells

Natural and Synthetic Scaffolds

3-D Printing and Chip Technologies

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Regenerative Medicine

1. Artificial organs

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Regenerative Medicine

2. Tissue Engineering and Biomaterials

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The Beginning…Joseph Vacanti (Harvard Stem Cell Institute) And Robert Langer (MIT) 1993

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Regenerative medicine pioneerDr. Ali Khademhosseini (Wyss Institute at Harvard)

Organs in the lab

Lab on a chip

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Process of Tissue Engineering

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3 Tools for Tissue Engineering• Cells

– Living part of tissue

– Produces protein and provides function of cells

– Gives tissue reparative properties

• Scaffold

– Provides structural support and shape to construct

– Provides place for cell attachment and growth

– Usually biodegradable and biocompatible

• Cell Signaling

– Signals that tell the cell what to do

– Proteins or Mechanical Stimulation

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Stem CellsStem cells are cells that:

(1) can self-renew

(2) have the potential to differentiate along one or two lineages.

1. Totipotent Can produce all cell types

2. Pluripotent Can produce most cell types

3. Multipotent Can produce more than one cell type

4. Unipotent Can produce one cell type

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Cell SourcesAutologous: Come from the person that needs the new cells.

Allogeneic: Come from a body from the same species.

Xenogenic: Come from a different species then the organism they’re going into.

Isogenic (Syngenic): Come from identical twins.

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ScaffoldsScaffolds are 3-dimensionalmaterials constructed in order toprovide structure to a developingtissue and to allow cells to adhere,proliferate, differentiate and mostimportantly, secrete extracellularmatrix (ECM) (Leong MF. et al.,2009).

Many different materials have beeninvestigated in order to constructscaffolds such as polymers (PLA,PGA, PCL, PEG), bioactive ceramics(HA, TCP) as well as naturalpolymers (Collagen, GAGs,Chitosan).

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Scaffold Construction RequirementsScaffolds should provide void volume for vascularization, new tissue formation and remodeling so as to facilitate host tissue integration upon implantation.

Should have high porosity and have suitable pore sizes, and the pores should be interconnected.

the biomaterials should also be degradable upon implantation at a rate matching that of the new matrix production by the developing tissue.

The biomaterials used to fabricate the scaffolds need to be compatible with the cellular components of the engineered tissues and endogenous cells in host tissue.

Scaffolds provide mechanical and shape stability to the tissue defect.

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Scaffolds SynthesisNanofiber self-assembly:

Molecular self-assembly is one of the few methods for creating biomaterials with properties similar in scale and chemistry to that of the natural in vivo extracellular matrix (ECM), a crucial step toward tissue engineering of complex tissues.Moreover, these hydrogel scaffolds have shown superiority in in vivo toxicology and biocompatibility compared to traditional macro scaffolds and animal-derived materials (Cassidy JW, 2014).

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Scaffolds SynthesisTextile technologies:

These techniques include all the approaches that have been successfully employed for the preparation of non-woven meshes of different polymers. In particular, non-woven polyglycolide structures have been tested for tissue engineering applications: such fibrous structures have been found useful to grow different types of cells. The principal drawbacks are related to the difficulties in obtaining high porosity and regular pore size(Ekevall E, 2004).

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Scaffolds SynthesisFreeze- drying:

First, a synthetic polymer is dissolved into a suitable solvent (e.g. polylactic acid in dichloromethane) then water is added to the polymeric solution and the two liquids are mixed in order to obtain an emulsion. Before the two phases can separate, the emulsion is cast into a mold and quickly frozen by means of immersion into liquid nitrogen. The frozen emulsion is subsequently freeze-dried to remove the dispersed water and the solvent, thus leaving a solidified, porous polymeric structure (Haugh MG, 2010).

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Scaffolds SynthesisCAD/CAM (3D-Printing):

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Scaffolds SynthesisElectrospinning:

A highly versatile technique that can be used to produce continuous fibers fromsubmicrometer to nanometer diameters. In a typical electrospinning set-up, a solution isfed through a spinneret and a high voltage is applied to the tip. The buildup ofelectrostatic repulsion within the charged solution, causes it to eject a thin fibrousstream. A mounted collector plate or rod with an opposite or grounded charge draws inthe continuous fibers, which arrive to form a highly porous network. The primaryadvantages of this technique are its simplicity and ease of variation. At a laboratorylevel, a typical electrospinning set-up only requires a high voltage power supply (up to30 kV), a syringe, a flat tip needle and a conducting collector. For these reasons,electrospinning has become a common method of scaffold manufacture in many labs. Bymodifying variables such as the distance to collector, magnitude of applied voltage, orsolution flow rate researchers can dramatically change the overall scaffold architecture(Lannutti J, et al., 2007).

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Electrospinning

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Poly-α-hydroxy acid

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Methods: Drying

Extrusion

Knitting

Gamma sterilization

Poly-α-hydroxy acidExtensive research has been performed in developing a full range of PLGApolymers.

Both L- and DL-lactides have been used for co-polymerization.

The ratio of glycolide to lactide at different compositions allows control of thedegree of crystallinity of the polymers.

When the crystalline PGA is co-polymerized with PLA, the degree ofcrystallinity is reduced and as a result this leads to increases in rates of hydrationand hydrolysis.

In general, the higher the content of glycolide, the quicker the rate ofdegradation. However, an exception to this rule is the 50:50 ratio of PGA: PLA,which exhibits the fastest degradation.

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Fibrin microbeads (FMB)Fibrinogen exerts adhesive effects on cultured fibroblasts and other cells.Specifically, fibrin(ogen) and its various lytic fragments (e.g., FPA, FPB,fragments D and E) were shown to be chemotactic to macrophages, humanfibroblasts, and endothelial cells

Micro-carrier beads made of some plastic polymers or glass provide cells with asurface area on the order of 104 cm2/L for cell attachment, which is one order ofmagnitude larger than the area available with stack plates or multi-tray cell-culture facilities

microparticles from plasma proteins, such as albumin or fibrinogen, generallyusing glutaraldehyde to cross-link the proteins.

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Setup for producing FMB by oil emulsion method

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Natural PolymersBlends of collagen andglycosaminoglycans (GAG) have beenutilized extensively for dermalregeneration.

Chondroitin sulfate has been added tocollagen type I for dermal regenerationtemplates and aggrecan (chondroitinsulfate/dermatan sulfate/keratin sulfate)to collagen type II for articularcartilage tissue engineering

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Hyaluronan Composed of repeated disaccharide units

of D-glucuronic acid and N-acetylglucosamine

The unique properties of HA aremanifested in its mechanical function inthe synovial fluid, the vitreous humor ofthe eye, and the ability of connectivetissue to resist compressive forces, as inarticular cartilage.

Plays a fundamental role duringembryonic development and in woundhealing

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Hyaluronan scaffold for central neural tissue engineering

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CollagenIn the form of collagen sponge

Porosity, biodegradability, and biocompatibility

Can be modified using growth factors or other manipulations to promote chondrocyte

growth and cartilage matrix formation

Scaffolds made from a single collagen type or composites of two or more types

Disadvantages

Poor dimensional stability. Variability in drug release kinetics.

Poor mechanical strength.

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ChitosanIt consists of β-1-4 linked 2 amino-2-deoxygluco –pyranose moieties.

Commercially manufactured by N-deacetylationof Chitin which is obtained from Mollusc shells.

It is soluble only in acidic pH i.e. when aminogroup is protonated.

Thereby it readily adheres to bio membranes.

It is degraded mainly by Glycosidases &lysozymes.

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Chitosan ScaffoldFabrication of bulk porous chitosan scaffolds

Freezing of a chitosan-acetic acid solution

Subsequent lyophilization

Scaffold microstructure will depend on the shape of the mold used for freezingand on the freezer temperature.

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Chitosan and Collagen-chitosan scaffold

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Hydrogels as scaffoldCells are suspended within or adhered to the 3D hydrogel framework during orafter formulation as scaffolds

RGD (arginine–glycine–aspartic acid) adhesion peptide sequence. Inclusion ofthese RGD domains in hydrogels has shown improved cellular migration,proliferation, growth, and organization in tissue regeneration applications.

Cells have been shown to favorably bind to the RGD-modified hydrogelscaffolds. These cells include endothelial cells (ECs), fibroblasts, smooth musclecells (SMCs), chondrocytes and osteoblasts.

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Alginate as Scaffold

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Pure Alginate as Scaffold

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Bioactive TGF-β1/HA alginate-based scaffolds for osteochondral tissue repair

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