Inhibitory effect of antisense human telomerase RNA(hTR) gene on hepatocellular carcinoma cells / 中国肿瘤生物治疗杂志

ABSTRACT

Objective: To study the inhibitory effect of retrovirus-mediated antisense human telomerase RNA (hTR) gene on hepatocelluar carcinoma, so as to explore an effective way to inhibit telomrerase activity in the treatment of hepatocellular carcinoma. Methods: Sense and antisense hTR gene were transfected into the packaging cell line PT67 by electroporation, and the stablely transfected cells were selected with G418. The recombinant retroviral supernatant was collected and transfected into HepG2 cells. After G418 selection, PCR was used to verify the integration of the hTR gene. Cell growth curves were drawn using MTT assay and the expression of PCNA was determined by immunofluorescence. TRAP-PCR-ELISA was adopted to detect the telomerase activity; cell cycle and apoptosis were evaluated by flow cytometry (FCM).Results: The expression of hTR gene could be amplified in HepG2-hTR-EcoRⅠ and HepG2-hTR-BamHⅠ cells, but not in untransfected HepG2 cells. The antisense hTR complementary to the template region of telomerase inhibited growth and proliferation of HepG2 cells. The expression of neutrophil proliferating cell nuclear antigen (PCNA) was decreased. Telomerase activity in the antisense hTR-treated group was (2.31?0.16),which was significantly lower than those of the other 2 groups(P