Intrasplenic transplantation of allogeneic hepatocytes modified by BCL-2 gene protects rats from acute liver failure.
BACKGROUND
Apoptosis of donor hepatocytes may be induced by recipient cytotoxic T lymphocytes (CTLs) during acute rejection, representing a major impediment for these cell transplants. Because the mechanisms of transplanted hepatocyte loss involve Fas-mediated pathways, BCL-2 genetic modification may protect liver cells. In the present study, we further investigated whether BCL-2 transfer into transplanted liver cells rendered them resistant to Fas ligand-induced apoptosis, and protected rats from acute liver failure.


METHODS
Hepatocytes isolated from Sprague-Dawley rats were infected with an adenovirus vector encoding human BCL-2 gene (AdCMVBCL-2) or a control AdCMVLacZ vector. Forty-eight hours later, cells challenged with recombinant Fas ligand (rhsFasL) were assayed for apoptosis using TUNEL staining and caspase 3 activity. Other cells were transplanted into the spleens of Wistar rats with a 90% hepatectomy 12 hours later.


RESULTS
Western blot analysis and RT-PCR confirmed the expression of hBcl-2 in AdCMVhBcl-2-infected hepatocytes. Recombinant FasL produced a dose-dependent increase in TUNEL-positive percentage and caspase-3 activity in uninfected hepatocytes, but did not influence these features in AdCMVhBcl-2-infected cells. On challenge with 90% hepatectomy, the survival of Wistar rats receiving transplantation of AdCMVhBCL-2-infected hepatocytes was significantly prolonged compared with the controls.


CONCLUSIONS
Adenovirus-mediated BCL-2 gene transfer protects transplanted hepatocytes from Fas-mediated cytolysis, thus holding promise for a new avenue of acute liver failure treatment.
Wang
J
J
Department of Surgery, Sun-Yat-Sen Memorial Hospital, Guangzhou, China.
Li
W
W
Min
J
J
Ou
Q
Q
Chen
J
J
Song
E
E
D000256:Adenoviridae; D000818:Animals; D030801:Animals, Genetically Modified; D017209:Apoptosis / Q000187:drug effects; D053222:Fas Ligand Protein; D019254:Genes, bcl-2; D005822:Genetic Vectors; D022781:Hepatocytes / Q000637:transplantation*; D006801:Humans; D017114:Liver Failure, Acute / Q000517:prevention & control*; D008562:Membrane Glycoproteins / Q000494:pharmacology; D023421:Models, Animal; D051381:Rats; D017207:Rats, Sprague-Dawley; D013154:Spleen; D014162:Transfection
C505661:FASLG protein, human; D053222:Fas Ligand Protein; C505662:Faslg protein, rat; D008562:Membrane Glycoproteins