Study On The Effect Of GRP78 On Autophagy And Apoptosis In Ovarian Carcinoma

ObjectiveThe purpose of this study is to investigate the effect and mechanisms of GRP78 on autophagy and apoptosis in ovarian carcinoma, to explore its impact on the growth and sensitivity to cisplatin in ovarian cancer cells. It may provide new ideas for treatment and novel methods to overcome cisplatin resistance in ovarian carcinoma.MethodsThe human ovarian cancer cell line SKOV3 were cultured in vitro. The GRP78 regulator BAPTA-AM and A23187 were used to down-regulate and up-regulate the expression levels of GRP78, respectively. The expressions of Beclin1, Bcl-2 and CHOP m RNA and their protein levels were detected by RT-PCR and Western Blot. The autophagy levels were observed by GFP-LC3-II fluorescence staining and the apoptosis rates of cells were analyzed by the flow cytometry. MTT was used to detect the influences on cell growth and sensitivity to cisplatin of SKOV3.Results1.The effect of GRP78 on the m RNA expressions of Belin1, Bcl-2 and CHOP.After decreased and increased the GPR78 expression levels of ovarian cancer cell SKOV3 by the GRP78 regulator BAPTA-AM and A23187, the m RNA expressions of Beclin1 were(0.8605 ? 0.05478) and(0.6537 ? 0.06500), respectively, compared with the control group whose was(0.7724 ? 0.02871) and the differences were statistically significant(t=2.768, P<0.05; t=2.893, P<0.05). The m RNA expressions of Bcl-2 were(0.7136 ? 0.03230) and(0.9075 ? 0.04730), respectively, compared with the control group whose was(0.8450 ? 0.01785) and the differences were statistically significant(t=3.334, P<0.05; t=3.920, P<0.05). The m RNA expressions of CHOP were(0.8108 ? 0.00380) and(0.6195 ? 0.06148), respectively, compared with the control group whose was(0.7116 ? 0.07689) and the differences were statistically significant(t=3.141, P<0.05; t=2.877, P<0.05).2.The effect of GRP78 on the protein expressions of Belin1, Bcl-2 and CHOP.After decreased and increased the GPR78 expression levels of ovarian cancer cell SKOV3 by the GRP78 regulator BAPTA-AM and A23187, the protein expressions of Beclin1 were(0.9521 ? 0.02238) and(0.6375 ? 0.02489), respectively, compared with the control group whose was(0.7890 ? 0.08349) and the differences were statistically significant(t=3.268, P<0.05; t=3.012, P<0.05). The protein expressions of Bcl-2 were(0.3854 ? 0.03231) and(0.5956 ? 0.02902), respectively, compared with the control group whose was(0.4918 ? 0.03551) and the differences were statistically significant(t=3.810, P<0.05; t=3.248, P<0.05). The protein expressions of CHOP were(0.6813 ? 0.09462) and(0.4313 ? 0.09534), respectively, compared with the control group whose was(0.5306 ? 0.00110) and the differences were statistically significant(t=3.404, P<0.05; t=3.222, P<0.05).2.The effect of GRP78 on autophagy in SKOV3.After decreased and increased the GPR78 expression levels of ovarian cancer cell SKOV3 by the GRP78 regulator BAPTA-AM and A23187, the autophagy fluorescence of SKOV3 were(706.25 ? 117.258) and(473.17 ? 127.903), respectively, compared with the control group whose was(594.58 ? 126.232) and the differences were statistically significant(t=5.381, P<0.05; t=4.380, P<0.05).3.The effect of GRP78 on apoptosis analyzed by flow cytometry in SKOV3.After treated by BAPTA-AM, the apoptosis rate of SKOV3 was 27.42%, which was higher than the control group whose was 19.57%. After treated by A23187, the apoptosis rate of SKOV3 was 12.21%, which was lower than the control group whose was 19.57%.4.The effect of GRP78 on the sensitivity to cisplatin in SKOV3.The IC50 of SKOV3 to cisplatin was(3.02 ? 0.624) μg/ml. After treated by BAPTA-AM, the IC50 was(2.00 ? 0.172) μg/ml and the sensitivity of SKOV3 to cisplatin increasd by 33.78%. The difference was statistically significant compared with the control group(t=17.787, P<0.05). And after treated by A23187, the IC50 was(4.91?2.515) μg/ml and the sensitivity of SKOV3 to cisplatin decreasd by 62.58%. The difference was statistically significant compared with the control group(t=29.516, P<0.05).Conclusions1.GRP78 could regulate autophagy and apoptosis of ovarian cancer cells by regulating the expressions of Beclin1, Bcl-2 and CHOP, thereby affecting the sensitivity to cisplatin in ovarian carcinoma. Regulating the expression of GRP78 could be a new method for the treatment and improvement of the sensitivity to casplatin in ovarian carcinoma.2.GPR78 may play an important role in cisplatin resistance in ovarian carcinoma by regulating autophagy and apoptosis pathways.