Polycystic ovarian syndrome (PCOS) is a common
endocrine disorder associated with characteristic
features including hyperandrogenemia, insulin resistance
and obesity, which profoundly impact a
woman’s reproductive life (
This study was approved by the Ethics Committee of the University of Hamedan, Iran, and informed written consents were obtained from participants.
This was a randomized clinical trial study. Twenty-
eight infertile patients (male or female factor)
diagnosed with PCOS according to the Rotterdam
ESHRE/ASRM criteria (
S1=1.4 standard deviation of body mass index (BMI) in PCOS patients before treatment.
S2=1.4 standard deviation of BMI in PCOS patients after treatment.
M1=30.65 mean of BMI in PCOS patients before treatment. and
M2=28.75 mean of BMI in PCOS patients after treatment.
S1, S2, M1 and M2 were determined according to
literature reviews. Inclusion criteria PCOS was
defined as the presence of polycystic ovaries on
transvaginal ultrasound scan (TVS); more than 12
cysts (2-9 mm in diameter in one plane in at least
one ovary) and increased stroma, usually combined
with increased ovarian volume >10 ml; clinical or
biochemical hyperandrogenism with at least one
of the following symptoms: oligomenorrhea or
amenorrhea; and clinical manifestations of hyperandrogenism
such as hirsutism and acne, Anovulation
was defined as the presence of amenorrhea or
oligomenorrhea (cycle length greater than 35 days)
Exclusion criteria included concurrent hormone therapy within the previous six weeks, any chronic disease that interfered with the absorption, distribution, metabolism or excretion of metformin and the presence of renal or liver disease. Patients with significant systemic disease were excluded. Smokers, those taking sex hormones or drugs effecting insulin secretion, clomiphene citrate, intense physical activity, as well as the loss of 3 kg of body weight two months prior to study entry were excluded.
Weight, height, and waist and hip circumferences
were measured. Because of the impact of body fat
distribution on androgen levels and glucose metabolism,
waist-to-hip ratios (WHR) were measured.
Waist circumference was determined as the
minimum value between the iliac crest and the lateral
costal margin, whereas hip circumference was
determined as the maximum value over the buttocks.
The cut-off point for high WHR for women
was set at 0.80. Body weight was measured using
analogue scales in light clothing; height was
measured barefoot using a stadiometre. BMI (kg/
m2) was calculated to assess obesity and WHR assessed
body fat distribution. Obesity was defined
as BMI ≥30 and overweight as BMI from 25 to
Ovarian volume was calculated for each ovary using
the formula for a prolate ellipsoid: π/6 × (D1 ×
D2 × D3), where D1-D3 represent the maximum
diameter in the transverse, antero-posterior and
longitudinal axes (
Venous blood samples were collected from all patients after 12 hours of overnight fasting. Samples were centrifuged immediately and serum was stored at -20°C until assayed for total testosterone, estradiol, 17-α-OH progesterone (17OHP), luteinizing hormone (LH), follicle stimulating hormone (FSH), estradiol, testosterone, 17-α-OH progesterone (17OHP), dehydroepiandrosterone sulfate (DHEAS), C-reactive protein (CRP), Hcy, lipid profiles and fasting blood sugar (FBS).
All patients received 1500 mg metformin per day (500 mg, three times daily) for three months. All women were urged to maintain the same diet as before treatment and were checked monthly. No severe side effects were reported during the study. After three months of treatment, patients were reevaluated clinically, biochemically and hormonally. All measurements were performed using the ChemWell® Analyzer, unless otherwise stated. FBS (mg/dl) was determined by the glucose oxidase color method (Glucose GOD-PAP). Total cholesterol (mg/dl) was determined by enzymatic photometric (CHOD-PAP) precipitation of low density lipoprotein (LDL), very low density lipoprotein (VLDL) and chylomicrons. High density lipoprotein (HDL) (mg/dl) was measured by magnesiumphosphotungstate precipitation, and precipitation of LDL, VLDL and chylomicrons. LDL-C (mg/dl) levels were calculated by the Friedewald formula. Total testosterone (ng/dl), 17OHP (ng/ml), estradiol (pg/ml), LH (mlu/ml), FSH (mlu/ml), DHEAS (μg/ml), CRP (mg/l) and Hcy (μmol/l) were measured by enzyme-linked immunosorbent assay (ELISA).
Statistics were calculated by the SPSS version 16.0 with student’s paired t-test. Significance was set at p<0.05. The correlations between mean ovarian volume with androgen levels, BMI, triglyceride (TG), LDL, CRP, Hcy and WHR were tested by applying Pearson's correlation coefficient using bivariate analysis.
There were 28 PCOS patients with a mean age 25.67 ± 8.54 years who participated in this study. In patients, the following PCOS signs and symptoms were noted: hirsutism (75%), acne (50%) and acantosis nigricans (39.3%). Regular menstruation was seen in 7 (25%) patients, 18 (64.3%) had oligomenorrhea and 3 (10.7%) had amenorrhea. After treatment, 17 women (65.38%) had regular menstrual cycles.
No patients had BP ≥130/85. Laboratory analysis showed that 22 (78.57%) had a TG _≥150mg/dl and 12 women (42.85%) had HDL levels ≤50mg/ dl, however no patients had impaired FBS. A WHR ratio ≥0.85 was seen in 4 cases (14.28%) and there was a positive correlation between mean ovarian volume and WHR (r=0.547, p=0.003).
Twenty-one patients (75%) had sonographic characteristics of polycystic ovary and 17 women (60.71%) had a mean ovarian volume greater than 10 ml.
According to BMI, there were 7 (25%) obese patients, 15 (53.57%) overweight and 6 (21.42%) who were in the normal BMI range. Positive correlations were noted between mean ovarian volume and BMI (r=0.589, p=0.001), and testosterone levels and BMI (r=0.663, p=0.000).
Anthropometric characteristics of PCOS patients before and after treatment are listed in table 1. Weight, BMI and mean ovarian volume were significantly different before and after treatment in PCOS patients, but there was no significant difference in WHR and BP. After treatment, baseline FSH was higher, however decreases were seen in LH, DHEAS, 17OHP, estradiol and testosterone levels.
The lipid profile, Hcy, FBS and CRP in PCOS patients significantly reduced after three months treatment with metformin. Hcy was more than the desired level (10 μg/ml) in 15 cases (53.57%), but not more than the upper limit of normal (5-16 μmol/l) in any of the patients. All patients had positive CRP (≥3mg/l). Twenty-one PCOS women (75%) had testosterone levels more than the 95th percentile. Table II lists the main hormonal and metabolic profile before and after metformin treatment in PCOS patients.
There was a positive correlation between reduction in ovarian volume and decreases in CRP (r=0.603, p=0.001), LDL (r=0.436, p=0.026) and Hcy levels (r=0.479, p=0.013) after three months of treatment with metformin.
The ovarian volume correlated to BMI and thus suggested a possible relationship between ultrasound findings and anthropometric characteristics. In our findings, the prevalence of obesity, high androgen, CRP, Hcy levels and existence of metabolic syndrome within patients with larger ovarian volumes was higher than in PCOS patients with normal ovarian volumes. This finding possibly confirmed an interaction between ovarian morphology and volume, and anthropometric characteristics. We hypothesized that patients with larger ovarian volumes were more insulin resistant; this would explain the higher BMI, androgen and CV risk factors. Metformin administration was associated with reduced ovarian volume and this reduction had a positive correlation with the decrease in CV risk factors.
Hyperinsulinemia stimulates the development of
antral follicles, increasing the sensitivity of granulosa
cells to FSH, thus increasing the numbers of
follicles and ovarian volume (
The results of our study, which are also in agreement
with Genazzani et al. (
Obesity is seen in 40% to 50% of women with
PCOS. This obesity is usually of the android type,
with an increased WHR (
A moderately increased total plasma Hcy concentration
is associated with an increased risk of
atherosclerosis. Elevations of Hcy can be due to
demographic, genetic, nutritional or metabolic factors.
Hyperhomocysteinemia induces sustained
injury to the arterial endothelial cell, which accelerates
the development of thrombosis and atherosclerosis
Wijeyaratne et al. declared that Yarali first reported
a significant elevation of plasma Hcy among PCOS
subjects when compared with older BMI-matched
controls. This correlated with echocardiographic
evidence of diastolic dysfunction (considered as
an early marker of CAD), plasma insulin and uric
acid; thus linking hyper homocysteinemia with
the insulin resistance of PCOS. Also they declared
that Loverro et al. reported significantly greater
plasma Hcy in a group of 35 women with PCOS
when compared with age-matched controls and
conversely, Morgante et alreported no difference in
plasma Hcy inwomen with PCOS, although their
study had fewer patients. Also they reported the results
of Schachter et al. study on 150 women with
PCOS, of whom 53.5% were insulin resistant, and
reported a significant elevation of fasting plasma
Hcy that correlated with insulin resistance (
Endothelial dysfunction in PCOS was documented
both by decreased response to vasodilation and
by the finding of increased levels of endothelin-1
in insulin-resistant PCOS patients and increased
oxidative stress markers (
Badawy et al. (
This increase in Hcy levels may be explained by
the fact that metformin affects folate and vitamin
B levels by decreasing their absorption from the
gut, which significantly increases Hcy levels (
Many PCOS patients may also have an increase in
subclinical atherosclerotic disease, as suggested by
greater carotid intima media thickness and higher
levels of coronary calcifications (
Metformin improves insulin sensitivity. Metformin
may improve ovulation and menstrual cycles, and
reduce ovarian volume. It may decrease circulating
androgen levels, thus addressing the traditional
goals of long-term treatment. Available clinical
evidence supports the use of metformin as a protective
measure against the adverse CV effects of
insulin resistance and insulin excess (
Existing data support the importance of increased
CV and metabolic risks in hyperandrogenic women
with classic features of PCOS (