Apoptosis of human primary ovarian granulose cells infected with lentivirus carrying bcl-2 gene / 中国组织工程研究

ABSTRACT

BACKGROUND: Lentivirus can infect divided and undivided cells. It remains uncertain whether the lentivirus can successful y infect primary ovarian granulosa cells. OBJECTIVE: To investigate infecting ratio and cel apoptosis of lentivirus carrying bcl-2 gene in primary human ovarian granulose cells cultured in vitro. METHODS: The lentiviral vector carrying bcl-2 gene was constructed using molecular biology, and packaged into lentivirus with high titer. The resulting recombinant lentivirus carrying bcl-2 genes were then used to infect primary human ovarian granulosa cells in vitro at different multiplicity of infection, 10, 50, 100, 200, and 400. Infection efficiency and cel proliferation were observed at 24, 48, 72, and 96 hours fol owing infection. Cel apoptosis was detected by flow cytometry, and bcl-2 gene transcription was assessed using reverse transcription PCR. RESULTS AND CONCLUSION: Primary human ovarian granulosa cells adhered at 24 hours, and exhibited polygon- or fusiform-shape and colony-like growth. When multiplicity of infection was 100, cel appearance and growth remained unchanged, and infection efficiency was high, which reached the peak up to 72 hours. Moreover, the positive rate was up to 60% in granulosa cells. Lentivirus carrying bcl-2 gene could increase expression of Bcl-2 protein and inhibit apoptosis of primary ovarian granulosa cells.