Asscociation Study Of Mutation Of CYP17 Gene And LH-β Gene To Polycystic Ovarian Syndrome In Chinese Women

Background and Objective Polycystic ovarian syndrome (PCOS) is the most common endocrinologic disorder in women of reproductive age. It is the most common cause of chronic unovulatory infertility and hyperandrogenism. This syndrome is characterized by diverse clinical symptoms and complicated endocrine features. Hyperandrogenism and abnormity of luteinizing hormone are the primary endocrine features of PCOS and can lead to diverse clinical symptoms and bad ending. Up to date, the certain cause of PCOS is still unknown. PCOS shows significant familial aggregation and studies reveals that the hereditary factors play an important role in the pathogenesis of PCOS. The clinical symptoms and biochemical features in PCOS appear to be heterogeneity. Many studies pointed out that PCOS may be explained by the interaction of a few key genes and tiny effect genes with genetic susceptibility and environmental factors, and the key gene is likely different between different ethics and families. There were many different investigations of genes of PCOS abroad, but the results were not uniform between different studies. There were few studies on the causative gene of PCOS in our country.We chose the candidate genes associated with androgen synthesis CYP17 and that of encoding LH- β to study, considering the endocrinal characteristics such as hyperandrogenism and high level of LH. The purpose of this study is to investigate the relationship between the two genes and PCOS, and to search the causative genesof PCOS so as to provide theoretic base to prevent and treat PCOS.Methods Blood samples were collected from the peripheral vessels of 118 patients with PCOS and 106 controls. Genomic DNA was isolated from peripheral blood collected above. Polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) were employed to detect the mutation of CYP17 gene and LH- (3 gene. First, fragments of CYP17 gene , exon 2 and exon 3 regions of LH-|3 gene were amplified by PCR. Then the PCR product of CYP17 gene was digested with restriction endonuclease MspAl I, PCR product of exon 2 of LH- β gene was digested with Nco I, Fok I and PCR product of exon 3 was digested with EcoO109 I. Finally, the digested product was subjected to electrophoresis on a gel containing 2% agarose. The serum sexual hormones were determined by ELISA. The relationship of mutation of CYP17 and LH-β gene and sexual hormone levels of patients with PCOS was analysed.Analysis was performed using the SPSS10.0 software package. Statistical analysis of differences in imitative rate, genotype frequency and allele frequency were assessed by using the x~2 test. Differences in serum sexual hormone levels and BMI were compared by using one-way ANOVA t test, α =0.05 was considered statistically significant. Results1. The T—C substitution of -34bp of the promotor of CYP17 gene created a site of restriction endonuclease (MspAl I) resulting in two alleles Al and A2. The prevalence rate of CYP17 genotype A1A1, A1A2, A2A2 were 10.2%, 55.9%, 33.9% in PCOS and 19.8%, 51.9%, 28.3% in control group, respectively. The frequencies of Al and A2 alleles were 38.1%, 61.9% in PCOS group and 45.8%, 54.2% in control group, respectively. Neither the genotype distributions nor the allele frequencies were statistically different between PCOS group and control group, P>0.05.2. In PCOS group, the testosterone levels of the genotype A2A2 group (1.52± 0.50μg/L) were significantly higher than that of A1A2 (1.30±0.85μg/L) andAlAl(1.00±0.66ug/L) groups (PO.05). The testosterone levels in PCOS patients with T-*C substitution (1.43±0.67ug/L) were significantly higher than that without T-*C substitution(1.00±0.66|ig/L), PO.05.3. We found two mutations [T986GG (Trp8) to C986GG (Arg8) and AT1008C (He15) to AC1008C (Thr15) ] in exon 2 of LH-P gene in Chinese women after digestion with Nco I and Fok I. This two mutations were linkage occurrence. Mutation rate was 10.2% in PCOS and 16.0% in controls. Neither the genotype distributions nor the allele frequencies were significantly different between PCOS group and control group. Sexual hormne levels were not statistically different between mutation group and non-mutation group, P>0.05.4. Mutation [G15°2GT^AI502GT (Gly102—Ser102)] in exon 3 of LH-p gene existed inChinese women after digestion with EcoO109 I. Mutation rate in PCOS (37.7%) was significantly higher than that in controls (18.9%) (x 2=9.284, PO.005). The frequencies of G and A alleles were 81.4%, 18.6% in the PCOS group and 90.6%, 9.4% in the control group, respectively. There were statistical difference between the two groups, ( x 2=7.736, PO.01).5. The levels of testosterone and luteining hormone were higher in PCOS with A allele than that with G allele, P<0.05. This difference was not exist in controls. Conclusions1. The T-* C substitution of-34bp of CYP17 gene might be correlated with the high testosterone levels of PCOS, and CYP17 was one of the susceptive genes of PCOS.2. There were two missence mutations[T986GG (Trp8) to C986GG (Arg8) and AT1008C(He15 )to AC1008C (Thr15) ] in exon 2 of LH-p gene in Chinese. They were linkae occurence. But these mutations were not correlated with the pathogenesis of PCOS.3. We found the mutation G1502GT to A1502GT (Gly102to Ser102) in exon 3 of LH-P gene in Chinese. This mutation occured more frequently in PCOS than that in controls, and might have effect on levels of serum testosterone and luteininghormone. This result clued that LH-0 gene might be one of the primary causative genes of PCOS and be worthy of further studying.