UNIVERSITÉ DE GENÈVE

FACULTÉ DE MÉDECINE

Département de Gériatrie
Section de Médecine Clinique
Laboratoire de Biologie du Vieillissement

Thérapie cellulaire de l'insuffisance cardiaque dans un modèle d'infarctus du myocarde chez le rat

Thèse
présentée à la faculté de Médecine de l'Université de Genève
pour obtenir le grade de Docteur en Médecine

par

Qing HE

(de Shangai / Chine)

sous la direction du
Pr Karl-Heinz Krause

et sous la supervision du
Dr Marisa Jaconi

Thèse n° Méd. 10358

Genève, 2004


      To my husband Ke-zheng and my daughter Yun-Yun
with all my love


BSA
Bovine serum albumin
CCM
Cellular cardiomyoplasty
CHF
Congestive heart failure
CGR8
Center Genome Research 8
CsA
Cyclosporin A
Cx43
Connexin 43, gap-junction protein
EB
Embryoid Body
EBFP
Enhanced Blue Fluorescent Protein
EG
Embryonic germ
EGFP
Enhanced Cyan Fluorescent Protein
ESC
Embryonic stem cells
FACS
Fluorescence -assisted cell sorting
FS
Fractional Shorting
GFP
Green Fluorescent Protein
H&E
Hematoxylin and eosin
LIF
Leukemia Inhibitory Factor
LVEF
Left Ventricular Ejection Fraction
LVEDD
Left Ventricular End-Diastolic Diameter
LVESD
Left Ventricular End-Systolic Diameter
MEF
Murine Embryonic Fibroblast
MHC
Myosin Heavy Chain
MI
Myocardial infarction
MSC
Mesenchymal stem cells
NIH
National Institute of Health
PCNA
Proliferating Cell Nuclear Antigen
TGF-b
Transforming Growth Factor b-1

      Congestive heart failure (CHF) is a major cause of cardiovascular morbidity and mortality in developed countries. Treatment consists basically of long-standing symptomatic drug therapy, effective percutaneous and surgical revascularization. The only causal treatment at the end-stage of CHF is heart transplantation. However, the shortage of donor hearts the complications of immunosuppression, the failure of grafted organs and, not at least, the advanced age of patients suffering from CHF limit the utility of cardiac transplantation significantly. In recent years the idea emerged that cells rather than whole organs can be used to replace damaged tissues.

      Two different cell types displaying these qualities have been identified so far: 1) embryonic stem cells (ESC), and 2) adult somatic stem cells. ESC are pluripotent cells derived from the inner cell mass of the blastocyst. They have the ability to differentiate into virtually all kinds of cell types, a capacity that becomes progressively restricted with development.

      Can ESC be used as the source of cells for the therapy of CHF? Could a failing heart instruct the engrafted cells to become cardiac cells in situ, or is it first necessary to differentiate them in culture into cardiomyocytes?

      To address these fundamental questions, our research focused on the intracardiac implantation of undifferentiated murine ESC into a rat model of myocardial infarction caused by coronary artery ligature. We studied the possibility of cell survival and differentiation in situ. In particular we address the necessity of using immunosuppression when implanting non-autologous cells. Specifically, we investigated :

  1. The influence of cell implantation as a function of time after MI (acute (1 week) versus chronic phase (4 weeks));
  2. Different period after cell implantation (1 versus 4 weeks of engraftment);
  3. Different types of ESC, feeder cell dependent and LIF (Leukemia Inhibitory Factor) dependent;
  4. The effect of immunosuppression treatment using CsA (Cyclosporin A) on cell fate.

      The results show that under the circumstance of immunosuppression by CsA, the implantation of different types of undifferentiated ESC inevitably leads to teratoma formation. The occurrence increased over time from 20% at 1 week to 100% 4 weeks after implantation. The teratoma formation was not influenced by cardiac injury (MI). In the absence of immunosuppression (CsA), the injected ESC could not be retrieved. We conclude that it is necessary to use immunosuppression when grafting undifferentiated murine ESC to avoid cell rejection. Even though the use ESC as a source of cell therapy for CHF remain a promising idea, there is still a long way to go from basic research to the clinic application. Careful attention must be paid to numerous aspects of stem cell therapy to avoid deleterious effects.


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