Regulation Of The HIF-1α/VEGF Pathway In Ovarian Granulosa Cells Of PCOS

ObjectivePolycystic ovary syndrome(PCOS)is a common endocrine disease and metabolic disorder syndrome in women of childbearing age.Up to now,the pathogenesis of PCOS is unclear.At present,most studies showed that cell apoptosis and inflammatory response may be involved in the occurrence and development of PCOS.Gene associated with retinoid interferon induced mortality 19(GRIM-19)is one of formic acid and interferon inducing cell apoptosis related gene families.It worked through the interaction with multiple cytokines involved in regulating cell apoptosis and inflammation related pathophysiological process.It has been found that Grim-19 is related to oocyte maturation and early embryonic development,but its role in PCOS has not been reported.In this study,the effect of Grim-19 in ovarian granulosa cells of PCOS patients on HIF-1α/VEGF pathway was studied,and the possible molecular regulatory mechanism was explored,hoping to reveal the possible role and mechanism of GRIM-19 in the occurrence and development of PCOS,and to provide a new idea for the diagnosis and treatment of PCOS.MethodsOvarian granulocytes from PCOS patients and patients with normal ovarian function were selected as the research objects to study the effects of GRIM-19 in ovarian granulocytes on HIF-1α and VEGF in PCOS patients.KGN cell line with hydrogen peroxide solution(H2O2)was used as study group,and H2O2 groups with different concentrations(0,0.1,0.3 and 0.5mM)were set to establish oxidative stress model,and reactive oxygen species(ROS)scavenging agent N-acetyl-L-cysteine(NAC)was used as the control group,and the required concentration was selected by detecting the expression levels of GRIM-19,HIF-la and VEGF.The expression of GRIM-19 and HIF-1α genes in KGN cells was down-regulated by small interfering RNA(siRNA)technique,and the mRNA and protein expression levels of GRIM-19,HIF-lα and VEGF were compared with negative control(NC)group.ResultsAfter Grim-19 was down-regulated,mRNA expression of Grim-19,HIF-1α and VEGF in treated KGN cells was detected by RT-qPCR.In the down-regulated GRIM-19 group,the mRNA expression level of GRIM-19 was significantly lower than that of NC control group(P=0.00).Meanwhile,the mRNA expression of HIF-la was significantly higher than that of NC control group(P=0.01),and the mRNA expression of VEGF was significantly higher than that of NC control group(P=0.01).WB detected the protein expression in KGN cells after Grim-19 down-regulation:the protein expression of Grim-19 in the down-regulated GRIM-19 group was significantly lower than that in the NC control group(P=0.01).Meanwhile,the protein expression of HIF-la was significantly higher than that of NC control group(P=0.00),and the protein expression of VEGF was significantly higher than that of NC control group(P=0.02).After down-regulation of HIF-1α,mRNA expressions of Grim-19,HIF-1α and VEGF in treated KGN cells were detected by RT-qPCR.The mRNA expression of HIF-1α in down-regulated HIF-la group was significantly lower than that in NC control group(P=0.04).The mRNA expression level of Grim-19 was not significantly different from that of NC control group(P=0.90),and the mRNA expression level of VEGF was significantly lower than that of NC control group(P=0.02).The protein expression of HIF-la in down-regulated HIF-la group was significantly lower than that in NC control group(P=0.00).Meanwhile,the protein expression level of Grim-19 was not significantly different from that of NC control group(P=0.14),and the protein expression level of VEGF was significantly lower than that of NC control group(P=0.00).Conclusion(1)The expression of Grim-19 was low in PCOS ovarian granulosa cells,while the expression of HIF-lα and VEGF were high.(2)Under ROS stimulation,Grim-19 expression was inhibited,but HIF-1α expression was promoted.(3)The reduction of Grim-19 increased VEGF expression by acting on HIF-1α.