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Retrovirus-mediated multidrug resistance gene (MDR1) overexpression inhibits chemotherapy-induced toxicity of granulosa cells
Salih, Sana M.
Article from Journal - ilmiah internasional
Fertility and Sterility (keterangan: ada di ClinicalKey) vol. 95 no. 04 (Mar. 2011)
premature ovarian failure
protection from chemotherapy toxicity
1 (dapat dipinjam: 0)
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Objective To protect granulosa cells from chemotherapy-induced toxicity by retrovirus-mediated multidrug resistance (MDR1) gene transfection. Design Laboratory study. Setting Academic research laboratory in a university hospital. Patient(s) None. Intervention(s) KK15 immortalized murine granulosa cell line transiently transduced with sf91m3 retrovirus vector carrying MDR1 complementary DNA that encodes P-glycoprtoein (P-gp); transduced cells selected with colchicine and treated with doxorubicin or paclitaxel for 24-72 hours; expression and function of MDR1 and the messenger RNA (mRNA) expression of selected steroidogenesis enzymes evaluated by flow cytometry, cell viability assays, Western blot, and reverse-transcriptase polymerase chain reaction (RT-PCR). Main Outcome Measure(s) Viability of sf91m3-transduced KK15 cells after treatment with doxorubicin and paclitaxel. Result(s) The sf91m3-transduced KK15 demonstrated high expression of biologically active MDR1, as shown by flow cytometry analysis and immunoblotting using P-gp monoclonal antibody and Rhodamine 123 efflux assays. The sf91m3-transduced KK15 exhibited statistically significant resistance to toxicity of 10 µM paclitaxel. The MDR1-transduced KK15 cells were also protected from doxorubicin toxicity (10 nM to 2.5 µM), as shown by cell viability assay. Both flow cytometry and cell viability assays showed that the protection of KK15 from doxorubicin toxicity was lost at 5 µM of doxorubicin; equivalent to 500 times LD50. The sf91m3-transduced KK15 showed normal mRNA expression of a panel of selected steroidogenesis enzymes. Conclusion(s) Retroviral gene delivery of human MDR1 inhibited chemotherapy-induced granulosa cell toxicity and offered chemoprotection in an in vitro model.
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