Abstract
Purpose
The purpose of this study was to investigate whether the follicle-stimulating hormone receptor (FSHR) gene p. Thr307Ala (c.919A>G, rs6165) and p. Asn680Ser (c.2039A>G, rs6166) polymorphisms are associated with susceptibility to polycystic ovary syndrome (PCOS).
Methods
Genotyping was performed in 377 women with PCOS and 388 age-matched controls. Difference in the genotype distribution was assessed using a Fisher’s exact or chi-square test, and continuous variables were compared using a Student’s t test. To evaluate the association between the presence of PCOS status and SNP, logistic regression analyses were performed.
Results
Linkage disequilibrium between the two polymorphisms was approximately complete (r 2 = 99%). The genotype distributions of the PCOS group significantly differed from those of the control group (Thr/Thr, Thr/Ala, and Ala/Ala frequencies were 38.5, 46.7, and 14.9% for the PCOS group and 46.6, 45.4, and 8.0% for the controls, respectively, P = .005; Asn/Asn, Asn/Ser, and Ser/Ser frequencies were 39.5, 47.2, and 13.3% for the PCOS group and 46.4, 45.4, and 8.2% for the controls, respectively, P = .035). Using the wild-type genotypes as the references, the odds ratios that a woman has PCOS were 2.23 (95% confidence intervals 1.38–3.68) for the Ala/Ala genotype, 1.87 (95% confidence intervals 1.14–3.06) for the Ser/Ser genotype, and 1.96 (95% confidence intervals 1.19–3.24) for the homozygous variant combination (Ser/Ser-Ala/Ala). However, there were no significant differences in serum hormonal, ovarian, and metabolic markers according to each genotype.
Conclusions
Findings of this study suggest a significant association between FSHR gene p. Thr307Ala or p. Asn680Ser coding sequence change and PCOS. The variant homozygote genotype results in a higher risk of PCOS.
Electronic supplementary material
The online version of this article (doi:10.1007/s10815-017-0953-z) contains supplementary material, which is available to authorized users.
Keywords: FSH receptor gene, Polycystic ovary syndrome, Polymorphism
Introduction
Polycystic ovary syndrome (PCOS) is a common endocrine disorder in women of reproductive age with a prevalence that ranges from 11.9 to 19.9% by the Rotterdam criteria [1, 2]. Chronic anovulation and hyperandrogenism are cardinal features of this syndrome, and obesity and insulin resistance are also common. Thus, diabetes, metabolic syndrome, and other cardiovascular disease risk factors are known to be increased in women with PCOS, and a risk of developing type 2 diabetes (DM) is approximately seven times greater in these women than unaffected women [3]. Despite considerable detrimental impact on women’s reproductive and metabolic health, PCOS etiology remains poorly understood.
Follicle-stimulating hormone (FSH) plays an important role in follicle development, oocyte maturation, and steroidogenesis. FSH exerts its effects via a specific receptor (FSHR) which is located on the granulosa cells of the ovary. The FSHR gene contains 10 exons and 9 introns, and is located at chromosome 2p21. Variants of the FSHR are known to be very rare [4, 5]; however, two polymorphisms, p. Thr307Ala (c.919A>G, rs6165) and p. Asn680Ser (c.2039A>G, rs6166), have been identified in exon 10 [6]. Earlier studies showed that the p. Asn680Ser variant allele was associated with higher basal FSH levels although within the normal range, with longer menstrual cycles, and with a higher ovarian threshold to ovulation induction, suggesting a difference in receptor sensitivity [7–11].
Although the etiology of PCOS has not been elucidated, numerous studies have suggested that genetic factors play important roles in its etiology and pathogenesis. Based on the knowledge that PCOS is characterized by failure of follicular growth, several studies have extensively investigated the association between FSHR p. Thr307Ala or p. Asn680Ser coding sequence changes and PCOS with inconsistent and controversial results [8, 11–24]. In this study, we compared the genotype distribution of FSHR gene p. Thr307Ala and p. Asn680Ser polymorphisms between PCOS patients and controls, and investigated whether serum hormonal, ovarian, and metabolic markers were affected according to these polymorphisms.
Materials and methods
Subjects
Three hundred seventy-seven premenopausal women were enrolled as PCOS diagnosis. PCOS diagnosis was based on the 2003 Rotterdam consensus meeting guidelines [25]. Oligomenorrhea was defined as fewer than eight periods per year or cycles longer than 35 days, and amenorrhea was defined as the absence of menstruation for more than 3 months without pregnancy. Clinical hyperandrogenism was defined by a modified Ferriman and Gallwey score (mF-G score) of 6 or greater in our population [26]. Biochemical hyperandrogenism was defined by an elevation of serum androgen levels beyond the 95% confidence limits measured in 89 ovulatory, non-hirsute controls in our population who did not have a PCO based on ultrasonography (total testosterone [T] >0.68 ng/ml, free T > 1.72 pg/ml, free androgen index [FAI] >5.36) [27]. All PCOS patients were screened to exclude hyperprolactinemia and thyroid dysfunction. Serum 17-hydroxyprogesterone (OHP) was also measured in all PCOS patients. If a patient’s serum 17-OHP level was greater than 2 ng/ml, a repeat test was performed during the early morning follicular phase. Patients who showed continuous elevation of 17-OHP were excluded from the study group.
A total of 388 premenopausal women were matched with patients based on age (± 1 year). These women visited the healthcare center of our hospital as part of a group checkup for work and lacked specific health problems. All controls had regular (21–35-day) menstrual cycles and mF-G scores below 6. They all received a transvaginal or transrectal pelvic ultrasound examination to evaluate ovarian morphology, and were excluded if a polycystic ovarian morphology was identified. Neither the cases nor the controls had taken hormonal medications, including oral contraceptives, for at least 3 months. The review board for human research of the Seoul National University Hospital approved this project (IRB No. 1005-050-319), and written informed consent was obtained from all individual participants included in the study.
Clinical and biochemical measurements
Clinical variables such as body weight, height, waist circumference, and blood pressure (BP) were assessed, and body mass index (BMI) was calculated as weight (kg) divided by the square of height (m2).
Basal gonadotropin hormone levels were measured in all PCOS subjects, including serum luteinizing hormone (LH), FSH, and estradiol (E2) levels. All PCOS women were evaluated for serum total T, free T, 17-OHP, dehydroepiandrosterone sulfate (DHEAS), and sex hormone-binding globulin (SHBG) using radiomimunoassay (Siemens, Los Angeles, CA, USA). Plasma insulin levels were measured using a commercial kit (BioSource Europe S.A., Belgium). FAI was calculated as (total T / SHBG) × 100, and the values for T were converted from nanograms per milliliter to nanomoles per liter using the following index provided by the manufacturer: 1 ng/ml = 3.467 nmol/l. Fasting and 2-h glucose and insulin levels were evaluated by a 75-g glucose tolerance test to assess glucose intolerance and insulin resistance in PCOS patients. The homeostatic model for insulin resistance (HOMA-IR) was calculated as glucose (mg/dl) × insulin (μU/ml) / 405, and HOMAβcell (%) was calculated as follows: (20 × fasting insulin) / (fasting glucose − 3.5). Serum cholesterol, triglyceride (TG), high-density lipoprotein (HDL) cholesterol, and low-density lipoprotein (LDL) cholesterol levels were measured using a 200FR system (Toshiba, Tokyo, Japan). In most of the controls, serum total T, free T, SHBG, fasting glucose and insulin, total cholesterol, TG, HDL cholesterol, and LDL cholesterol levels were also measured. The intra- and inter-assay coefficients of variation were 4.0–11.0 and 5.9–11.0% for total testosterone, 4.0–17 and 8.0–18.3% for free T, 5.0–7.1 and 5.0–11.0% for 17-OHP, 3.8–5.1 and 6.3–11.0% for DHEAS, 2.8–5.3 and 7.9–8.5% for SHBG, and 1.6–2.2 and 6.1–6.5% for plasma insulin, respectively.
Genomic DNA analysis
Genomic DNA was extracted from peripheral blood samples with the Wizard DNA Purification Kit (Promega, Madison, WI). Allelic discrimination was done using the MGB-NFQ primer/probe TaqMan assay on the ABI Prism 7500 Real-Time PCR System (Applied Biosystems, Foster City, CA, USA). Primer pairs and probes for FSHR p.Thr307Ala and FSHR p. Asn680Ser were Assays-on-Demand, C___2676873_30 (Applied Biosystems), and Assays-on-Demand, C___2676874_10 (Applied Biosystems), respectively. The PCR mixture consisted of 10 μl of TaqMan Universal PCR Master Mix 2× (Applied Biosystems) and 25 ng DNA. The PCR cycling conditions consisted of one 2-min cycle at 50 °C, and one 10-min cycle at 95 °C, followed by 40 cycles at 95 °C for 15 s and 60 °C for 1 min. We used distilled water as a negative PCR control in each amplification.
Statistical analysis
Power calculations were performed using the QUANTO v.1.2.4 (log-additive model) software (http://hydra.usc.edu/gxe/). A 12% population prevalence of PCOS was assumed, and minor allele frequency was obtained from other Korean studies [1, 18]. Based on the sample size (377 patients with PCOS and 388 controls), the power to detect an allelic odds ratio of 1.5 at an α value of 0.05 was 83.2%. Possible deviations from the normality of the data distribution were tested by visual inspection of quantile-normal plots or the Shapiro-Wilk test of normality. If variables did not follow Gaussian distributions, normal distributions were achieved by natural logarithmic or square root transformation. In these cases, data were shown as geometric means with 95% confidence intervals (95% CI). Hormonal and other metabolic parameters were compared using either Student’s t test or analysis of variance (ANOVA). The differences in genotype distribution and allele frequency were tested using a Fisher’s exact or chi-square test. Genotype distribution was also examined for significant departures from the Hardy-Weinberg equilibrium by a goodness-of-fit chi-square test. To evaluate the association between the presence of PCOS status and SNP, logistic regression analyses were performed using the wild-type homozygote as the reference category. All data analyses were performed using the Statistical Package for the Social Sciences software (version 22.0, IBM SPSS, NY, USA), and a haplotype analysis was performed using Haploview software (version 4.2; Broad Institute). Significance was accepted for two-sided P values <.05.
Results
The clinical and endocrine characteristics of the PCOS patients and controls are shown in Table 1. By definition, there were significant differences in hirsutism score and serum androgen levels between women with PCOS and controls. Women with PCOS were more obese than controls, and as expected, there were also significant differences in metabolic parameters between the two groups except HDL cholesterol and fasting glucose levels.
Table 1.
Clinical and biochemical features of PCOS patients and age-matched controls
| PCOS (n = 377) | Controls (n = 388) | P value | |
|---|---|---|---|
| Age (years)a | 28.5 ± 4.9 | 28.5 ± 4.9 | .955 |
| Body mass index (kg/m2)a | 22.2 ± 4.0 | 20.1 ± 2.3 | <.001 |
| Waist circumference (cm)a | 74.1 ± 10.6 | 74.4 ± 6.8 | .615 |
| Hirsutism scoreb | 5 (0–22) | 0 (0–5) | <.001 |
| Total testosterone (ng/ml)c | 0.32 (0.30, 0.34) | 0.25 (0.24, 0.26) (n = 210) | .002 |
| Free testosterone (pg/ml)c | 1.0 (0.9, 1.1) | 0.6 (0.5, 0.7) (n = 210) | .028 |
| Sex hormone-binding globulin (nmol/l)c | 39.0 (34.6, 43.4) | 64.3 (60.5, 68.1) (n = 197) | <.001 |
| Free androgen indexc | 3.2 (2.9, 3.5) | 1.5 (1.3, 1.7) (n = 197) | <.001 |
| Luteinizing hormone (mIU/ml)c | 7.3 (6.7, 8.0) | Not available | |
| Follicle-stimulating hormone (mIU/ml)c | 4.6 (4.3, 4.9) | Not available | |
| Estradiol (pg/ml)c | 46.1 (42.1, 50.5) | Not available | |
| Fasting plasma glucose (mg/dl)a | 88.3 ± 13.1 | 87.3 ± 8.2 | .240 |
| Fasting insulin (μU/ml)c | 8.5 (6.6, 10.4) | 6.2 (4.5, 7.9) | <.001 |
| Homeostatic model assessment for insulin resistancec | 1.86 (1.52, 2.20) | 1.32 (1.24, 1.40) | <.001 |
| Homeostatic model assessmentβcell (%)c | 131.8 (130.0, 133.6) | 97.7 (95.6, 99.8) | <.001 |
| Systolic blood pressure (mmHg)a | 108.4 ± 15.7 | 104.5 ± 14.4 | .001 |
| Diastolic blood pressure (mmHg)a | 69.6 ± 10.8 | 65.7 ± 10.8 | <.001 |
| Total cholesterol (mg/dl)a | 180.6 ± 31.1 | 173.3 ± 26.9 | .002 |
| Triglyceride (mg/dl)c | 75.9 (74.2, 77.6) | 67.6 (66.2, 69.0) | .001 |
| High-density lipoprotein cholesterol (mg/dl)a | 63.1 ± 14.9 | 63.9 ± 13.9 | .513 |
| Low-density lipoprotein cholesterol (mg/dl)a | 101.3 ± 26.7 | 94.8 ± 21.8 | .006 |
| Hemoglobin A1c (%)a | 5.53 ± 0.253 | 5.43 ± 0.26 | <.001 |
P values are indicated for the differences in groups as analyzed by Student’s t test
aData are shown as means ± SD
bMedian (range)
cGeometric mean and 95% CI
The genotype distribution in the control group was in the Hardy-Weinberg equilibrium, and a linkage disequilibrium (LD) between p. Asn680Ser and p. Thr307Ala polymorphisms was approximately complete (r 2 = 99%). The genotype distributions of the PCOS group significantly differed from those of the control group (Asn/Asn, Asn/Ser, and Ser/Ser frequencies were 39.5, 47.2, and 13.3% for the PCOS group and 46.4, 45.4, and 8.2% for the control group, respectively, P = .035; Thr/Thr, Thr/Ala, and Ala/Ala frequencies were 38.5, 46.7, and 14.9% for the PCOS group and 46.6, 45.4, and 8.0% for the control group, respectively, P = .005) (Table 2). Using the wild-type genotype as the reference, the odds that a woman has PCOS was 1.87 (95% CIs 1.14–3.06) times higher if she has a Ser/Ser genotype and 2.23 (95% CIs 1.38–3.68) times higher if she has an Ala/Ala genotype (Table 2). We additionally evaluated distributions of p. Asn680Ser and p. Thr307Ala genotype combinations, and found significant differences between the women with PCOS and the controls (Supplementary Table 1). The Asn/Asn-Thr/Thr, Asn/Ser-Thr/Ala, and Ser/Ser-Ala/Ala genotype combinations made up almost the whole subject pool (366/377 in women with PCOS and 387/388 in controls). Distributions of these three major genotype combinations significantly differed between the PCOS patients and controls (Asn/Asn-Thr/Thr, Asn/Ser-Thr/Ala, and Ser/Ser-Ala/Ala frequencies were 38.5, 45.6, and 13.0% for the PCOS group and 46.4, 45.4, and 8.0% for the controls, respectively, P = .026) (Table 2). Using the wild-type genotype combination (Asn/Asn-Thr/Thr) as the reference, the odds that a woman has PCOS was 1.96 (95% CIs 1.19–3.24) times higher if she has a homozygous variant combination (Ser/Ser-Ala/Ala).
Table 2.
Genotype distributions of the p. Asn680Ser and p. Thr307Ala polymorphisms in the follicle-stimulating hormone receptor gene in PCOS patients and controls
| Genotype | PCOS (n = 377) | Control (n = 388) | P valuea | Odds ratio (95% CIs) | P valueb |
|---|---|---|---|---|---|
| p. Asn680Ser | |||||
| Asn/Asn (%) | 149 (39.5%) | 180 (46.4%) | .035 | ||
| Asn/Ser (%) | 178 (47.2%) | 176 (45.4%) | 1.21 (0.89–1.63) | .219 | |
| Ser/Ser (%) | 50 (13.3%) | 32 (8.2%) | 1.87 (1.14–3.06) | .013 | |
| p. Thr307Ala | |||||
| Thr/Thr (%) | 145 (38.5%) | 181 (46.6%) | .005 | ||
| Thr/Ala (%) | 176 (46.7%) | 176 (45.4%) | 1.25 (0.92–1.69) | .150 | |
| Ala/Ala (%) | 56 (14.9%) | 31 (8.0%) | 2.23 (1.38–3.68) | .001 | |
| Genotype combination | |||||
| Asn/Asn-Thr/Thr (%) | 145 (38.5%) | 180 (46.4%) | .026 | ||
| Asn/Ser-Thr/Ala (%) | 172 (45.6%) | 176 (45.4%) | 1.21 (0.90–1.64) | .212 | |
| Ser/Ser-Ala/Ala (%) | 49 (13.0%) | 31 (8.0%) | 1.96 (1.19–3.24) | .008 | |
CI confidence interval, PCOS polycystic ovary syndrome
aEvaluated by chi-square test in comparison with the control group
bEvaluated by logistic regression models using the wild-type Asn/Asn genotype, Thr/Thr genotype, or Asn/Asn-Thr/Thr genotype combination as the reference
For further analysis, subjects were divided according to genotype, and we assessed whether serum hormonal, ovarian, and metabolic markers were affected by the p. Asn680Ser or p. Thr307Ala genotypes (Table 3). However, we could not find any significant differences in these parameters that were correlated with genotype in women with PCOS (nor in controls, data not shown).
Table 3.
Hormonal and metabolic variables according to the p. Asn680Ser and p. Thr307Ala polymorphisms in the follicle-stimulating hormone receptor gene in women with polycystic ovary syndrome
| Asn/Asn (n = 149) | Asn/Ser (n = 178) | Ser/Ser (n = 50) | P a | Thr/Thr (n = 145) | Thr/Ala (n = 176) | Ala/Ala (n = 56) | P a | |
|---|---|---|---|---|---|---|---|---|
| Age (years)a | 26.1 ± 1.3 | 26.9 ± 0.9 | 27.2 ± 1.4 | .398 | 26.5 ± 1.4 | 26.3 ± 0.9 | 26.4 ± 1.4 | .203 |
| Body mass index (kg/m2)a | 22.2 ± 4.2 | 22.0 ± 3.9 | 22.4 ± 3.8 | .913 | 22.2 ± 4.1 | 22.0 ± 3.9 | 22.5 ± 3.7 | .721 |
| Total testosterone (ng/ml)b | 0.32 (0.28, 0.36) | 0.36 (0.33, 0.39) | 0.33 (0.29, 0.37) | .141 | 0.32 (0.28, 0.36) | 0.36 (0.33, 0.39) | 0.34 (0.29, 0.39) | .129 |
| Free testosterone (pg/ml)b | 1.0 (0.8, 1.2) | 1.1 (1.0, 1.2) | 1.1 (0.9, 1.3) | .068 | 1.0 (0.8, 1.2) | 1.1 (1.0, 1.2) | 1.1 (0.9, 1.3) | .067 |
| Sex hormone binding globulin (nmol/l)b | 34.6 (29.6, 40.0) | 34.1 (30.7, 37.7) | 37.1 (31.2, 43.6) | .336 | 34.5 (29.7, 39.3) | 34.1 (30.7, 37.7) | 37.2 (31.3, 43.7) | .595 |
| Free androgen indexb | 3.37 (2.72, 4.18) | 3.83 (3.32, 4.41) | 3.12 (2.45, 3.96) | .313 | 3.38 (2.74, 4.02) | 3.83 (3.32, 4.41) | 3.14 (2.47, 3.81) | .734 |
| Total cholesterol (mg/dl)a | 181.9 ± 33.8 | 176.9 ± 29.2 | 189.7 ± 26.7 | .079 | 181.3 ± 33.9 | 177.7 ± 29.6 | 188.2 ± 28.7 | .176 |
| Triglycerides (mg/ml)b | 75.4 (65.5, 87.8) | 70.9 (64.0, 79.2) | 82.4 (72.0, 89.3) | .569 | 75.5 (65.5, 85.5) | 71.4 (64.0, 78.8) | 82.5 (72.1, 92.9) | .527 |
| High density lipoprotein cholesterol (mg/ml)a | 62.3 ± 14.7 | 63.4 ± 15.6 | 63.4 ± 14.5 | .790 | 62.1 ± 14.2 | 63.5 ± 15.6 | 63.4 ± 15.2 | .695 |
| Low density lipoprotein cholesterol (mg/ml)a | 101.6 ± 28.0 | 98.7 ± 24.6 | 108.7 ± 28.5 | .188 | 102.8 ± 29.5 | 100.6 ± 25.5 | 105.7 ± 26.9 | .628 |
| Fasting plasma glucose (mg/dl)a | 90.0 ± 17.8 | 87.0 ± 8.8 | 87.8 ± 8.5 | .153 | 90.1 ± 17.8 | 87.9 ± 8.9 | 87.9 ± 8.7 | .156 |
| Fasting insulin (μU/ml)b | 9.2 (7.9, 10.7) | 10.0 (9.1, 11.0) | 9.5 (8.4, 10.9) | .768 | 9.2 (7.9, 10.7) | 10.0 (9.1, 11.0) | 9.6 (8.4, 10.8) | .502 |
| Homeostatic model assessment for insulin resistanceb | 2.04 (1.74, 2.39) | 2.20 (1.98, 2.44) | 1.99 (1.73, 2.30) | .758 | 2.05 (1.74, 2.36) | 2.20 (1.98, 2.44) | 2.19 (1.73, 2.65) | .995 |
| Homeostatic model assessmentβcell (%)b | 144.7 (142.8, 146.6) | 147.7 (145.7, 149.7) | 159.1 (157.3, 160.9) | .458 | 144.7 (142.8, 146.6) | 158.1 (145.6, 170.6) | 159.8 (157.3, 162.3) | .195 |
| 2-h post-load plasma glucose (mg/dl)a | 108.2 ± 40.7 | 103.5 ± 28.0 | 107.7 ± 25.8 | .611 | 108.2 ± 41.2 | 103.7 ± 27.9 | 106.6 ± 25.5 | .636 |
| 2-h post-load plasma insulin (μU/ml)b | 41.7 (39.2, 44.2) | 36.3 (34.1, 38.5) | 46.8 (44.3, 49.3) | .313 | 41.7 (39.2, 44.2) | 36.3 (34.1, 38.5) | 46.8 (44.3, 49.3) | .329 |
| Hemoglobin A1ca | 5.53 ± 0.26 | 5.55 ± 0.24 | 5.53 ± 0.24 | .861 | 5.54 ± 0.25 | 5.55 ± 0.24 | 5.54 ± 0.31 | .859 |
| Lt. ovarian volumeb | 6.26 (4.37, 8.15) | 5.75 (3.92, 7.58) | 4.81 (2.99, 6.63) | .085 | 6.45 (4.51, 8.40) | 5.84 (3.97, 7.71) | 4.98 (3.14, 6.82) | .090 |
| Lt. ovarian follicle numbera | 14.7 ± 5.4 | 14.9 ± 6.2 | 14.6 ± 5.4 | .947 | 14.9 ± 5.6 | 15.1 ± 6.2 | 14.7 ± 5.4 | .896 |
| Rt. ovarian volumeb | 7.67 (5.92, 9.42) | 7.57 (5.78, 9.36) | 6.24 (4.27, 8.21) | .913 | 7.69 (5.93, 9.45) | 7.54 (5.84, 9.24) | 6.40 (4.63, 8.17) | .398 |
| Rt. ovarian follicle numbera | 16.7 ± 6.1 | 17.0 ± 7.0 | 15.5 ± 6.7 | .402 | 16.8 ± 6.3 | 16.9 ± 6.7 | 15.7 ± 6.7 | .431 |
| Luteinizing hormoneb | 6.88 (4.83, 8.93) | 8.03 (6.07, 9.99) | 9.40 (7.21, 11.59) | .080 | 6.88 (4.84, 8.92) | 7.99 (6.05, 9.93) | 9.41 (7.21, 11.61) | .081 |
| Follicle stimulating hormonea | 4.93 ± 2.58 | 4.94 ± 2.25 | 5.54 ± 2.55 | .391 | 4.81 ± 2.37 | 5.07 ± 2.37 | 5.35 ± 2.56 | .512 |
| Estradiolb | 43.0 (40.6, 45.4) | 44.7 (42.5, 46.6) | 43.4 (41.3, 45.5) | .932 | 43.1 (40.6, 45.6) | 44.9 (42.4, 47.4) | 44.4 (41.3, 47.5) | .608 |
P values are indicated for the differences in groups as analyzed by ANOVA
aData are shown as means ± standard deviations
bData are shown as geometric means and 95% confidence intervals
Discussion
The aim of the present study was to evaluate the association between FSHR gene p. Thr307Ala or p. Asn680Ser coding sequence polymorphisms and PCOS in Korean women. According to our data, a nearly complete LD between p. Thr307Ala and p. Asn680Ser polymorphism exists, and the FSHR p. Asn680Ser or p. Thr307Ala genotype distribution of the PCOS group was significantly different from that of the control group. Using the wild-type genotypes as the references, the odds ratios that a woman has PCOS were 1.87 (95% CIs 1.14–3.06) for the Ser/Ser genotype, 2.23 (95% CIs 1.38–3.68) for the Ala/Ala genotype, and 1.96 (95% CIs 1.19–3.24) for the homozygous variant combination (Ser/Ser-Ala/Ala). However, in women with PCOS, there were no significant differences in the serum levels of hormones, biochemical variables, or ovarian markers associated with any genotype.
FSH plays a key role in human reproduction. Based on the knowledge that PCOS is characterized by failure of follicular growth, an association between p. Thr307Ala or p. Asn680Ser polymorphism and PCOS has been extensively studied across ethnicity with conflicting results [8, 11–24]. Most studies were performed within small numbers of subjects, but two large studies could not find an association between FSHR polymorphism and PCOS. The largest one was the study by Valkenburg et al., which compared 518 Caucasian PCOS patients and 2996 controls, and the second largest one was the study by Fu et al., which compared 384 Chinese women with PCOS and 768 controls [16, 20]. Our study contained the third largest number of subjects (377 patients and 386 controls) ever reported of women with PCOS in which p. Asn680Ser polymorphism was analyzed, and our current study might offer data to the literature about the significance of FSHR polymorphism in PCOS. One interesting point was that the frequency of the Ser680Ser variant genotype (13.3% in women with PCOS and 8.2% in controls) was lower than that observed in Caucasians (22–25%). Another study performed in Italian women with PCOS also found higher frequency (30–40%) of the Ser680Ser variant genotype, suggesting the importance of ethnicity background [14].
The FSH receptor is a G-protein-coupled receptor, and has transmembrane, intracellular, and extracellular domains. The intracellular domains are encoded by exon 10, in which the p. Asn680Ser variant is located [28]. Although in vitro studies could not find different activity associated with variants of the FSH receptor [13, 29], substantial studies reported that FSHR gene polymorphism was associated with phenotype in women with PCOS. In addition, the FSHR p. Asn680Ser variant allele was associated with higher levels of FSH or clomiphene citrate resistance [11, 13, 16, 20–22, 24]. We also assessed the possibility that FSHR gene polymorphism affects ovarian markers represented by serum FSH concentrations, antral follicle count, or ovarian size, but could not find any association between genotype and phenotype. FSH levels in women with PCOS are generally within reference values, thus there is a possibility that small differences in serum FSH levels within the normal range may need larger numbers of subjects to demonstrate an association. Alternatively, the p. Asn680Ser variant might be involved only in local regulation of FSHR in women with PCOS. Either way, the mechanism by which FSHR gene polymorphism modulates the pathophysiology of PCOS needs to be determined.
Genome-wide association (GWA) studies on Chinese and European ancestry women found FSHR as PCOS susceptibility locus. Especially, the variant (rs2268361) in the FSHR gene intron region at 2p16.3 was associated with PCOS, although functional significance was not clear [30–33]. Recently, two European GWA studies reported that chromosome 11p14.1 SNP (rs11031006), which lies near the FSH β-subunit coding region, was strongly associated with PCOS diagnosis [34, 35]. Whether FSHR gene or FSH gene polymorphisms have complementarity in the PCOS pathogenesis is uncertain; these findings suggest that gonadotropin action modulation may have some roles in the PCOS pathogenesis.
In conclusion, the current study suggests a significant association between FSHR gene p. Thr307Ala or p. Asn680Ser coding sequence changes and PCOS. Since disorder in follicle development is a key feature of PCOS, FSHR polymorphism studies which might affect receptor function need to be performed continuously.
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Compliance with ethical standards
Ethical approval
All procedures performed in studies involving human participants were in accordance with ethical standards of the institutional and/or national research committees and with the 1964 Helsinki Declaration and its later amendments or comparable ethical standards.
Conflict of interest
The authors declare that they have no conflict of interest.
Funding
This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (20100009075).
Footnotes
Electronic supplementary material
The online version of this article (doi:10.1007/s10815-017-0953-z) contains supplementary material, which is available to authorized users.
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