The potential role of early-phase graft injury in induction of late-phase chemoresistance after liver transplantation

Abstract

OBJECTIVE: We aim to investigate the potential role of acute phase liver graft injury on induction of late phase chemoresistance after liver transplantation and to explore the underlying mechanism. MATERIALS AND METHODS: A rat orthotopic liver transplantation model using cirrhotic recipients was established with applying whole or small-for-size (50%) graft. The recipients were injected with a rat hepatoma cell line (MH7777) via portal vein after reperfusion. Proline tyrosine kinase (Pyk2) expression was compared between small-for-size and whole grafts, as well as the tumor developed from small and whole grafts. Pyk2-overexpressed HCC cell stable clones were established. The role of Pyk2 in cisplatin resistance was investigated by in vitro studies on HCC cell proliferation and apoptosis under cisplatin environment, and further confirmed by in vivo xenograft tumorigenesis and orthotopic nude mice models. The gene profile of Pyk2-overexpressed HCC cells was screened by cDNA microarray to explore the mechanism. RESULTS: Intragraft gene expression of Pyk2 increased gradually after transplantation (4h, 2d, 4d and 7d) in both small-for-size and whole grafts. mRNA levels of Pyk2 were significantly higher in small-for-size grafts at day 4 (55.71 vs 24.25 folds of normal liver, p=0.005) and day 7 (90.51 vs 48.5 folds, p=0.005) after transplantation. Pyk2 expression level was also significantly higher in liver tumor from small-for-size graft at day 14 (1.75 folds of whole graft, p=0.01) and day 21 (2.41 folds of whole graft, p=0.01) after operation. In vitro studies including MTT, colony formation and Annexin-V-FLOUS assay showed that overexpression of Pyk2 promoted cell proliferation and increased survival rate under cisplatin treatment. cDNA microarray screening showed that MDR1, GAGE, STAT1, CASP9 and MAP7, which related to drug resistance, were up-regulated in Pyk2-overexpressed HCC cells. The in vivo study demonstrated that suppression of Pyk2 could sensitize liver tumor to cisplatin by induction of significant necrosis and apoptosis. CONCLUSION: Pyk2 was significantly up-regulated in both of small-for-size grafts and the tumor developed from small graft after transplantation. Pyk2 overexpression in HCC cells remarkably induced cisplatin resistance. Therefore, earlyphase liver graft injury may induce late-phase chemoresistance after liver transplantation by up-regulating Pyk2.