Simultaneous transplantation of co-cultured Simultaneous transplantation of co-cultured

mesenchymal stem cells and skeletal myoblasts mesenchymal stem cells and skeletal myoblasts

improves ventricular function in Chagas diseaseimproves ventricular function in Chagas disease

Luiz César Guarita-Souza MD – PhDLuiz César Guarita-Souza MD – PhD

PUCPR - Experimental Laboratory of Cell Culture Institute of PUCPR - Experimental Laboratory of Cell Culture Institute of Biological and Health SciencesBiological and Health Sciences Curitiba-Pr-BrazilCuritiba-Pr-Brazil

IntroductionIntroductionSkeletal Myoblasts cells diferentiationSkeletal Myoblasts cells diferentiation

Myocardial Scar

Skeletal Fibers

Guarita-Souza LC, Int J Cardiol. 2006 Aug 28;111(3):423-9

NeoangiogenesisNeoangiogenesis

IntroductionIntroductionMesenchymal Stem cells diferentiationMesenchymal Stem cells diferentiation

Guarita-Souza LC, Transplant Proc. 2006Jul-Aug;38(6):1953-4

SkeletalMyoblast

MesenchymalStem Cells

Angio-muscularRegeneration

+

IntroductionIntroduction Myocardial Cell TherapyMyocardial Cell Therapy

Arq. Bras. Cardiol.2004, OctOct;83(4):294-9

The Association of Myoblast and Mesenchymal Stem Cells Transplantation,

on Post Infarct Ventricular Dysfunction

Guarita-Souza LC; Carvalho KAT, Rebelatto C; Senegaglia A, Myiague N; Furuta M; Francisco J, Olandowski M,

Hansen P.

IntroductionIntroduction Myocardial Cell TherapyMyocardial Cell Therapy

Cell Therapy in Myocardial InfarctionCell Therapy in Myocardial Infarction

1.1. SegmentarSegmentar

2.2. Myocardial Scar and IschemicMyocardial Scar and Ischemic

BothBoth Cell Therapy in a Cardiomyopathy non-Cell Therapy in a Cardiomyopathy non-

ischemic ?ischemic ?

IntroductionIntroduction Cell TherapyCell Therapy

Evaluated the histological and functional effects of

simultaneous autologous transplantation of co-cultured

mesenchymal and skeletal myoblasts cells in an

experimental model of dilated cardiomyopathy due to

Chagas disease.

ObjectiveObjective

IntroductionIntroduction Chagas Disease FisiopathologyChagas Disease Fisiopathology

TransmissionTransmission Triatoma InfestansTriatoma Infestans: :

5 to 14 days:sains and 5 to 14 days:sains and symptomssymptoms

Transfusional: Transfusional: 30 to 40 days symptoms30 to 40 days symptoms

Chronic form: after 10 yearsChronic form: after 10 years

Immunologic Activity:Immunologic Activity: Humoral and Cellular Immunity Humoral and Cellular Immunity

Triatoma Infestans

“Barbeiro”

Forms of DiseaseForms of Disease AcuteAcute IndeterminatedIndeterminated 60% 60% CardiacCardiac 30% 30%

after 10-20yafter 10-20y DigestiveDigestive BothBoth CongenitalCongenital

Trypanossoma Cruzi

Myocardial Infection

IntroductionIntroduction Chagas Disease FisiopathologyChagas Disease Fisiopathology

Chronic myocarditeChronic myocardite

Chronic inflamatory processChronic inflamatory process

diffuse fibrosisdiffuse fibrosis

Impairment of microcirculationImpairment of microcirculation

Ischemic myocardiumIschemic myocardium

HE, 200x

IntroductionIntroduction Chagas Disease PathologyChagas Disease Pathology

Chronic inflamatory process

Myocardial scar

MethodsMethodsProtocol of ResearchProtocol of Research

Eco 1MEco 1M

AnimalsAnimalsinoculationinoculation TXTX

HistologyHistology

EcoEcoBaselineBaseline

7 days7 days14 of days14 of days

cultivecultive8 M 8 M

MuscularMuscular

BiopsyBiopsy

Bone MarrowBone Marrow

AspiartionAspiartion

1 Month1 Month

T CruziT Cruzi 150.000 forms150.000 forms

Cell Therapy in MyocardiumCell Therapy in MyocardiumProtocol of Cell CultureProtocol of Cell Culture

80 animals were inoculated - 80 animals were inoculated - 200 gramas200 gramas 21.25%21.25% - 17 animals included - 17 animals included - FE < 37% FE < 37%

Co-culture group: 9 animals Co-culture group: 9 animals mesenchymal stem cellsmesenchymal stem cellsskeletal myoblast cellsskeletal myoblast cells0.30 ml - 0.30 ml - 5,4x106 8,0x106 cells

Control group : 8 animalsControl group : 8 animalssaline - 0.30 mlsaline - 0.30 ml

Material and MethodMaterial and Method

Material and MethodMaterial and MethodEchocardiographic studyEchocardiographic study

Bi-Dimensional Bi-Dimensional Echocardiographic studyEchocardiographic study

HP/SONOS 5500- Linear Transduction of 15MHzHP/SONOS 5500- Linear Transduction of 15MHz

After eigth months of After eigth months of

inoculationinoculation

One month after One month after

transplantationtransplantation

Parameters AnalysisParameters Analysis LVEDV (ml)LVEDV (ml)

LVESV (ml)LVESV (ml)

LVEF (%)LVEF (%)

ResultsResults Echocardiographic studyEchocardiographic study

VariableVariable ControlControl Co-cultureCo-culture

LVEF% preLVEF% pre 36,74 ± 3,6336,74 ± 3,63 30,10 ± 5,7130,10 ± 5,71

LVEF% posLVEF% pos 37,42 ± 6,6637,42 ± 6,66 51,76 ± 6,6351,76 ± 6,63

pp 0,76840,7684 <0,0001<0,0001

Mena±sd

LVEF%

LVE

F%

0

10

20

30

40

50

60

70

Baseline 1-month Baseline 1-month

Control Co-culturep=0,7684 p<0,0001

ResultsResultsLVEF(%)LVEF(%)

(*) Paired Student’s test p<0,05

VariableVariable ControlControl Co-cultureCo-culture

LVESV preLVESV pre 0,44 ± 0,080,44 ± 0,08 0,56 ± 0,050,56 ± 0,05

LVESV posLVESV pos 0,46 ± 0,120,46 ± 0,12 0,32 ± 0,090,32 ± 0,09

pp 0,65230,6523 <0,0001<0,0001

Mean±sd

LVESV

LVE

SV

0,0

0,1

0,2

0,3

0,4

0,5

0,6

0,7

Baseline 1-month Baseline 1-month

Control Co-culturep=0,6523 p<0,0001

ResultsResults LVESV (ml)LVESV (ml)

(*) Paired Student’s test p<0,05

VariableVariable ControlControl Co-cultureCo-culture

LVEDV preLVEDV pre 0,69 ± 0,110,69 ± 0,11 0,82 ± 0,070,82 ± 0,07

LVEDV posLVEDV pos 0,73 ± 0,140,73 ± 0,14 0,65 ± 0,140,65 ± 0,14

pp 0,63110,6311 0,00040,0004

Mean±sd

LVEDV

LVE

DV

0,45

0,55

0,65

0,75

0,85

0,95

Baseline 1-month Baseline 1-month

Control Co-culturep=0,6311 p=0,0004

ResultsResults LVEDV (ml)LVEDV (ml)

(*) Paired Student’s test p<0,05

ResultsResults PathologyPathology

HE, 200X

Control group Treat group

ResultsResultsPhatologyPhatology

ResultsResultsPhatologyPhatology

Gomery Trichrome Treat group

200X 400X 400X

ResultsResults Phatology / Phatology / ImmunostainedImmunostained

Fast Miosin, 40x

Control Treat group

The co-transplant of mesenchymal stem cell and skeletal The co-transplant of mesenchymal stem cell and skeletal

myoblasts cell is functionally effective in the Chagas disease myoblasts cell is functionally effective in the Chagas disease

ventricular dysfunction.ventricular dysfunction.

ConclusionConclusion