European Journal of Cardiovascular Medicine

Polycystic ovarian syndrome (PCOS) is a multi-aetiological and heterogeneous endocrine disorder that affects approximately 6 to 12% of women of reproductive age worldwide.1,2 PCOS is a common endocrinopathy characterized by menstrual irregularities, obesity, hirsutism, acne, insulin resistance, hyperandrogenism, anovulatory infertility and multiple ovarian cysts on ultrasonography.3,4 Dyslipidemia and obesity are very common with a prevalence of approximately 93.3% in women with PCOS. Obesity, atherogenic dyslipidemia particularly hyper triglyceridemia, increased levels of LDL & VLDL and decreased level of HDL are common metabolic abnormalities associated with PCOS.5 Dyslipidemia, diabetes, obesity, central body fat distribution, hyperandrogenemia, insulin resistance and glucose intolerance are all potent cardiovascular risk factors that tend to cluster in women with polycystic ovary syndrome.6

PCOS is also called as Stein-Leventhal syndrome as it was first reported in modern medical literature by Stein and Leventhal in 1935. Stein-Leventhal syndrome is the clinical manifestation of PCOS typically associated with anovulation and infertility with the classical triad of oligomenorrhea or amenorrhea, hirsutism and obesity.7 Accumulating evidence shows increased prevalence of cardiovascular disease (CVD) in women with PCOS than their non-PCOS counterparts. Futhermore, women with PCOS have a clustering of factors, such as obesity, dyslipidemia, insulin resistance, atherosclerotic plaque formation and type 2 DM predisposing them to cardiovascular risk.8,9

Proinflammatory markers like hs-CRP provide an adjunctive method for global assessment of cardiovascular risk.10 High plasma level of Lp (a) represent an independent risk factor for cardiovascular events, linked to an increased risk of myocardial infarction, stroke and coronary heart disease (CHD). Moreover, elevated plasma Lp (a) levels and lipid profile levels may contribute to the development of atherosclerosis and predisposes to increased cardiovascular risk in women with PCOS.11,12 Plasminogen activator inhibitor-1 (PAI-1) is an well-established risk factor for CVD and elevated plasma concentration of PAI-1 seem to be linked to an increased risk of thrombotic vascular events and associated with abdominal obesity, metabolic syndrome, insulin resistance and type-2 diabetes mellitus.13-15 The present study was conducted to evaluate lipid profile and serum biomarkers; hs-CRP, Lp-a and PAI-1 activity in women with PCOS and their role in the assessment of cardiovascular risk.

All women in the age group of 18 to 35 years who attended OPD in the Department of Obstetrics and Gynecology with the primary complaints of menstrual irregularities (amenorrhea or oligomenorrhea) and/or hirsutism with or without infertility were evaluated for PCOS and womendiagnosed with PCOS (n=100) were enrolled for the study. PCOS confirmation was done by high resolution ultrasonography in the follicular phase as per the Rotterdam revised criteria which required the presence of at least two out of three of the following features: (1) oligo- and/or anovulation (2) clinical and/or biochemical evidence of hyperandrogenism and (3) ultrasonographic findings of polycystic ovaries with exclusion of other known disorders of hyperandrogenism.16 Selection of PCOS cases and healthy controls were done by simple random sampling. One hundred (n=100) age-matched healthy women were taken as controls as they did not have PCOS, or any other signs of hyperandrogenism and had regular menstrual cycles. It was a prospective case-control study performed in the Department of Biochemistry and Obstetrics & Gynecology at SGT University, Gurugram, Haryana, India. All the PCOS cases were included as per the Rotterdam revised criteria. The patients with known cases of diabetes mellitus, hypertension, severe insulin resistance, CVD, congenital adrenal hyperplasia, androgen secreting tumors, Cushing’s syndrome, history of smoking and/or alcohol intake, androgenic/anabolic drug use or abuse, hperprolacteinemia, and thyroid dysfunction were excluded from the study. The size of the ovaries, volume, morphology and the number & size of the follicles were noted. Detailed history with special reference to menstrual irregularity, hirsutism, alopecia, infertility, voice change, weight gain, presence of acne and obstetric history was noted.

Anthropometric Parameters

Standard anthropometric data including height, weight, waist circumference (WC) and hip circumference (HC), weremeasured. BMI was calculated using the equation(body weight in kilograms divided by body height in meterssquared; kg/m2)11. The waist-to-hip ratio (WHR)wasmeasuredusing dressmakers tape, taking care that it was applied horizontally.Waist circumference is middlecircumference between the iliac crest and the lateral coastal margin and hip circumference is maximum circumference around the buttocks posteriorly and indicated anteriorly by the symphysis pubis.

Sample Collection and Analysis

After 12 hours overnight fasting, approximately 5 ml of venous blood samples were collected in plain tubes (for estimations of lipid profile, hormonal profile& serum biomarkers) and in the tube containing sodium fluoride and Oxalate (for plasma Glucose estimation).Serum was separated and preserved at -80ºC for subsequent analyses. Estimations of fasting plasma glucose, serum total cholesterol, triglycerides and HDL concentrations were assayed by using commercial kits available for standard photometric methods in fully automatedERBA XL (EM-200) Biochemistry analyzer.LDL was calculated by using Fredrickson Friedewald’s formula.17 Serum luteinizing hormone (LH), follicle stimulating hormone (FSH), prolactin (PRL)and thyroid stimulating hormone (TSH) were assayed by chemi luminescent immunoassay (CLIA)using Siemen’s Advia Centaur CP kit. Quality controls were used to check the accuracy and precision of the analyzer, reagents and assay results.The study was approved by the Institutional Ethical Committee (IEC) and an informed written consent was obtained from all the participants at the study entry after apprising them the nature and objectives of the study.

Statistical Analysis

Statistical analysis was performed using the Statistical Package for the Social Sciences (SPSS) version 24.0 for Windows (SPSS, Inc., Chicago, IL). The normality of distribution of all the variables was checked by using Kolmogorov–Smirnov test. The quantitative data were expressed as mean ± standard deviation (SD). Unpaired Student’s t-test was used to compare the values (PCOS vs. Controls) and Pearson’s correlation coefficient was used to elucidatethe relationship between the variables. At a confidence interval of 95%, p<0.05 were considered statistically significant.

The age group of the PCOS patients ranges from 18 to 35 years. The anthropometric parameters of the study population calculated were height, weight, BMI, waist circumference, hip circumference and WHR as shown in table 1.

Table 1: Demographic features and anthropometric parameters (mean ± SD) of the study population.

Abbreviations: BMI: body mass index; PCOS: polycystic ovary syndrome; SD: standard deviation; WHR: waist-to-hip ratio. (p** value less than 0.001 is considered highly significant).

We have evaluated both PCOS cases and the controls for the fasting blood sugar, lipid profile, serum TSH, LH, FSH, PRL and results are shown in table 2 and figure 1. The mean levels of serum total cholesterol, triglycerides, LDL and TC/HDL ratio were significantly higher (p<0.001) in PCOS cases than controls. Whereas, the mean level of serum HDL cholesterol was significantly decreased (p<0.001) in PCOS cases than controls. The mean TSH, LH, FSH, LH/FSH ratio & prolactin were significantly higher (p<0.001) in PCOS cases than the controls.

Table 2: Comparison of biochemical and hormonal profiles (mean ± SD) among PCOS patients and controls.

Abbreviations: FBS: fasting blood sugar, TC: total cholesterol, TG: triglyceride, HDL: high-density lipoprotein; LDL:low-density lipoprotein; VLDL: very low-density lipoprotein; TSH: thyroid stimulating hormone; LH: luteininzing hormone; FSH: follicle stimulating hormone; PRL: prolactin.

*p-value less than 0.05 is considered significant.

**p-value less than 0.001 is considered highly significant.

We have also evaluated cardiovascular risk markers in both PCOS cases and the controls. A significant increase was observed in the level of serum biomarkers such as high sensitivity C-reactive protein (hs-CRP), lipoprotein-a (Lp-a), and plasminogen activator inhibitor type-1 antigen (PAI-1) in PCOS patients (p<0.001) compared to the healthy subjects as shown in table 3.

Table 3: comparison of cardiovascular risk markers among PCOS patients and controls.

Abbreviations: hs-CRP: high sensitivity c-reactive protein; Lp-a: lipoprotein a; PAI-1: plasminogen activator inhibitor-1.

*p-value less than 0.05 is considered significant.

**p-value less than 0.001 is considered highly significant.

In the present study an increased level of Fasting blood sugar in PCOS patients (p=0.003) was found indicating abnormal glucose metabolism and is supported by the existing studies.18, 19 Our study showed dyslipidemia associated with PCOS that has been characterized by significant increase in the mean level of lipid parameters (TC, TG, LDL-C, VLDL-C, LDLC/HDL-C ratio and TC/HDL-C ratio) (p<0.001) and significant decrease in HDL-C level in PCOS patients compared to the healthy subjects. These findings were supported by the study conducted by Legro RS et al., Diamanti-Kandarakis E et al. and Rocha MP et al. Decreased HDL-C has been reported previously in more than two thirds of PCOS patients and more than one thirds of the PCOS patients are associated with hypertriglyceridemia.20-22

All these are known to be cardiovascular disease risk factors and were identified in a considerable number of young women. In addition, obesity and altered lipid parameters may be regarded as an underlying risk factor for atherosclerosis and coronary artery disease.6 LDL-C level was significantly elevated in PCOS patients (p<0.001) of this study when compared to controls. Elevated plasma levels of Lp-a have been associated with the increased risk of CHD in PCOS patients. The mean level of Lp-a in our study was increased in women with PCOS (p<0.001) than the controls. Our findings are in concordance with the previous studies.2,12,23 Conversely, no significant difference in the baseline characteristics of Lp-a between PCOS patients and controls have also been reported.24 The mean level of PAI-1 in our study was increased in women with PCOS as compared with the age-matched healthy controls (p<0.001). In addition to this, it has been reported that significantly increased level of PAI-1 antigen is associated with cardiovascular morbidity in PCOS patients independent of obesity.25 These findings are in concordance with the previous reports in PCOS patients.2,22,25 Furthermore, increased PAI-1 level in PCOS patients are also correlated with insulin resistance and obesity (BMI).46.

Our study revealed the derangements in anthropometric parameters, lipid profile, hormonal levels & cardiovascular risk markers in PCOS patients suggesting the high prevalence of hypothyroidism, increased risk of hyperlipidemia, type-2 diabetes mellitus and coronary artery disease. Identification of hypothyroidism, biochemical and endocrine derangements would help us to treat and prevent future complications in PCOS women. Furthermore, the evaluation of serum biomarkers such as; hs-CRP, Lp-a and PAI-1 routinely in PCOS patients may have diagnostic role in the early detection of metabolic abnormalities and endocrine derangements and timely management of co-morbid diabetes and cardiovascular disease in PCOS females.

Acknowledgment

We sincerely thank all the faculies and staffs of the Departments of Biochemistry and Obstetrics & Gynecology, SGT University, Gurugram, Haryana, India.We also acknowledge the contribution of the statistician and healthcare staff involved in patient care and data collection for their constant support & help. Finally yet importantly, special thanks to our patients without whom this study would never have been possible.

Conflicts of Interest

The authors have no conflicts of interest to declare.

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