Genetically engineered bone marrow-derived mesenchymal stem cells co-expressing IFN-γ and IL-10 inhibit hepatocellular carcinoma by modulating MAPK pathway.

PURPOSE: One of the major challenges in delivering cytokines for the treatment of hepatocellular carcinoma (HCC) is the mode of delivery. This study hypothesized that genetically engineered bone marrow derived mesenchymal stem cells (BMSCs) co-expressing IFN-γ and IL-10 can serve as a potential therapeutic strategy in the treatment of HCC by inhibiting cell proliferation.

METHODS: Male Sprague-Dawley rats (n=5, 200-250 g) for BMSCs isolation and Nude/SCID mice (n=35,12-20g) to develop liver cancer xenograft model were used. Mice were subcutaneously injected HepG2 cell suspension on left flank. BMSCs were genetically engineered with the recombinant lentiviral vectors expressing IFN-γ and IL-10. The experiments were performed in 5 groups (phosphate buffered saline/PBS, BMSCs, BMSC-IFN-γ, BMSC-IL-10 and BMSC-IFN-γ-IL-10) and the genetically engineered BMSCs were transplanted into HCC mice. Cell viability was measured by MTT assay followed by the evaluation of the effect of cell-cycle regulators (p21, p27, cyclin D1 and Rb). Protein expression of p38, ERK and JNK was assessed by immunohistochemistry using the cell proliferation marker Ki67.

RESULTS: The combination of two cytokines (IFN-γ and IL- 10) engineered into BMSCs resulted in a significant reduction in HepG2 cell viability (*p<0.05 vs PBS-treated and #p<0.05 vs BMSC-treated group). Significantly increased expression of cell cycle inhibitors p21 and p27 in parallel with reduced cyclin D1 expression were observed. Reduced phosphorylation of Rb demonstrated the repression of G1/S progression. BMSC-IFN-γ-IL-10 treatment significantly reduced the tumor growth at the end of 36 days compared to the group treated with PBS or BMSCs alone. This effect was accompanied with the modulation of MAPK pathway with the activation of p38 and JNK, and inactivation of ERK.

CONCLUSION: The co-expression of IFN-γ and IL-10 in BMSCs inhibits HCC in vitro and in vivo by modulating cell cycle regulators and MAPK pathway.