A Study On The Mechanism Of ADAMTS1 Regulating Granulosa Cells Function And Oocyte Quality In PCOS Patients

BackgroundPolycystic ovary syndrome(PCOS)is a common cause of female infertility.It affects multiple systems of the body and is often accompanied by many complications,including menstrual disorders,infertility,hairy,acne,obesity and metabolism syndrome.PCOS patients have a significantly increased risk of type 2 diabetes and cardiovascular disease.The typical clinical manifestations of patients with PCOS are polycystic ovary,hyperandrogenemia,and anovulation or rare ovulation.These three clinical manifestations are also used for the diagnosis of PCOS.Studies have shown that the pathogenesis of PCOS may be affected by genetics and lifestyle,but the specific mechanism is unclear and needs further exploration.In addition,due to abnormal ovarian function in patients with PCOS,the oocyte quality and embryonic development potential during in vitro fertilization-embryo transfer(IVF-ET)are often lower than that of patients with normal menstrual cycles.Therefore,the underlying mechanisms that regulate ovarian function and the follicular development microenvironment in patients with PCOS need to be further studied in order to obtain a more efficient diagnosis and treatment plan.A disintegrin and metalloproteinase with thrombospondin motifs(ADAMTS)is an extracellular metalloproteinase family,with a total of 19 members.It is widely expressed in different human organs,and plays an important role in the pathophysiology of multiple systems such as tissue development and reconstruction,tumor migration,arthritis,and hypertension.Among them,AD AMTS 1 is most closely related to the regulation of ovarian function.In the ovary,ADAMTS1 mainly mediates angiogenesis and the dynamic shaping of the extracellular matrix.AD AMTS 1 has been found to mediate the morphogenesis and functional maintenance of the ovary,and plays an important role in oocyte development,ovulation,luteinization,and ovarian angiogenesis.Changes in expression levels are also closely involved in diseases such as PCOS,premature ovarian failure(POF),and ovarian tumor.At the same time,the substrate of AD AMTS 1 such as versican(VCAN)and aggrecan(ACAN)is also closely related to the above processes.Previous studies have confirmed the correlation between AD AMTS 1 and the occurrence of PCOS,but due to the lack of mechanism research,it is still unclear how AD AMTS1 participates in the occurrence and development of PCOS and affects the oocyte development of PCOS patients.PurposeTo clarify the expression of ADAMTS1 in granulosa cells of patients with PCOS and the mechanism underlying the oocyte quality and early embryonic development potential of PCOS patients.MethodsPCOS patients(n=48)and ovulatory controls(n=44)were recruited.Polymerase chain reaction(PCR)and western blot(WB)were used to detect AD AMTS 1 in granulosa cells;Immunofluorescence were used to explore the location of ADAMST1 in granulosa cells and early embryos in different stages;The mRNA level of marker genes(AREG,HAS2,COX2,HLA-G,VCAN,THBS1,and PTX3),which are crucial during the oocyte and embryonic development,was assessed after silencing ADAMTS1;CCK-8 assay was performed to evaluate cell proliferation;Electrochemical luminescence was used to detect estradiol(E2)secretion.Results1.The mRNA and protein level of ADA MTS1 in granulosa cells of PCOS patients were significantly increased as compared to ovulatory controls.2.The mRNA level of ADAMTS1 was significantly positively correlated to the oocyte maturation rate and good-quality embryo rate of PCOS patients.3.ADAMTS1 was mainly expressed in the cytoplasm in both granulosa cells and early developmental embryos.4.In primary granulosa cells,the expression of AREG,HAS2,COX2,and HLA-G was significantly reduced after silencing ADAMTS1,while that of VCAN,THBS1,and PTX3 was increased.5.Down-regulating AD AMTS 1 inhibited the function of granulosa cells in cell proliferation and E2 secretion;also,the expression of Bcl2,Bax,Bcl-XL and key genes of E2 synthesis such as CYP11A1 and CYP19A1 was significantly downregulated.ConclusionBoth mRNA and protein level of ADAMTS1 in granulosa cells from patients with PCOS were significantly increased,and the mRNA level was positively correlated with the oocyte maturation rate and the good-embryo rate.The further research found that AD AMTS1 is expressed in all stages of early embryo,and its expression gradually decreases as the embryo develops,suggesting that it may be involved in the regulation of embryonic development.In addition,silencing ADAMTS1 not only affects the proliferation of granulosa cells and estradiol secretion,but also the expression of marker genes indicating the oocyte quality and embryonic development potential,which suggests that AD AMTS 1 may regulate oocyte development of PCOS patients by above two ways.