[Effect of human hepatocyte growth factor gene-modified bone marrow mesenchymal stem cells on immunological rejection after allograft liver transplantation in rats].

OBJECTIVE: To study the effect of recombinant lentiviral vector mediated human hepatocyte growth factor (hHGF) gene-modified bone marrow mesenchymal stem cells (BMSCs) on the immunological rejection after allograft liver transplantation in rats, and to reveal the mechanism of immune tolerance.

METHODS: Eight male Sprague Dawley (SD) rats of clean grade (aged 3 to 4 weeks, weighing 75-85 g) were selected for the isolation and culture of BMSCs; 64 adult male SD rats of clean grade (weighing 200-250 g) were used as donors; and 64 adult male Wistar rats of clean grade (weighing 230-280 g) were used as receptors. After establishing a stable model of rat allogeneic liver transplantation, 1 mL saline, 2 x10(6)/mL of BMSCs 1 mL, 2 x 106/mL of BMSCs/green fluorescent protein 1 mL, and 2 x 10(6)/mL of BMSCs/hHGF 1 mL were injected via the portal vein in groups A, B, C, and D respectively. Then the survival time of the rats was observed. The hepatic function was determined and the histological observation of the liver was performed. The hHGF mRNA expression was detected by RT-PCR, the level of cytokine including hHGF, interleukin 2 (IL-2), IL-4, IL-10, and interferon gamma (IFN-gamma) by ELISA assay, the level of apoptosis by TUNEL method, and the expression level of proliferating cell nuclear antigen (PCNA) by immunohistochemical method.

RESULTS: The survival time of group D was significantly higher than that of groups A, B, and C (P < 0.01); the survival time of groups B and C was significantly higher than that of group A (P < 0.01), but there was no significant difference between group B and group C (P > 0.05). RT-PCR demonstrated the transcription of hHGF mRNA in the grafts of group D; the serum cytokine hHGF reached to (6.2 +/- 1.0) ng/mL. Compared with groups B and C, group D exhibited significant inhibitory effect, significantly improved liver function, and showed mild acute rejection. In addition, the levels of cytokine IL-2 and IFN-gamma decreased; the levels of cytokine IL-4 and IL-10 increased; the level of apoptosis reduced; and the expression level of PCNA increased. Except for the expression of IL-4 (P > 0.05), there were significant differences in the other indexes between group D and groups B, C (P < 0.05).

CONCLUSION: BMSCs/hHGF implanting to rat liver allograft via portal vein can induce immune tolerance. Compared with injection of BMSCs alone, BMSCs/hHGF treatment can alleviate acute rejection and prolong the survival time significantly. The immunosuppressive effect of BMSCs/hHGF is correlated with Th2 shifts up of Th1/Th2 shift, reduced apoptosis, promoted liver regeneration.