Molecular Mechanisms in the Etiology of Polycystic Ovary Syndrome (PCOS): A Multifaceted Hypothesis Towards the Disease with Potential Therapeutics

Abstract

Among the premenopausal women, Polycystic Ovary Syndrome (PCOS) is the most prevalent endocrinopathy affecting the reproductive system and metabolic rhythms leading to disrupted menstrual cycle. Being heterogeneous in nature it is characterized by complex symptomology of oligomennorhoea, excess of androgens triggering masculine phenotypic appearance and/or multiple follicular ovaries. The etiology of this complex disorder remains somewhat doubtful and the researchers hypothesize multisystem links in the pathogenesis of this disease. In this review, we attempt to present several hypotheses that tend to contribute to the etiology of PCOS. Metabolic inflexibility, aberrant pattern of gonadotropin signaling along with the evolutionary, genetic and environmental factors have been discussed. Considered a lifelong endocrinological implication, no universal treatment is available for PCOS so far however; multiple drug therapy is often advised along with simple life style intervention is mainly advised to manage its cardinal symptoms. Here we aimed to present a summarized view of pathophysiological links of PCOS with potential therapeutic strategies.

Introduction

Polycystic ovary syndrome (PCOS) is a gender specific disorder and is documented as the most common endocrine disorder-affecting women of childbearing age across the globe. It typically involves impairment in the reproduction and estrogen controlled organs in females. Rotterdam criteria hold the most accepted and classical diagnostic criteria by world scientific and health regulatories for defining the disease [1]. PCOS represents broad spectrum of symptoms, which chiefly involves hyperandrogenism (HA), anovulation and or polycystic ovarian morphology (PCOM) [1, 2]. The consequent features mainly result in the hirsutism, acne, androgenic alopecia and or raised serum level free androgens, clitoromegaly (enlargement of clitoris), seborrhea (oily skin) and acanthosisnigricans (dark patches near the skin creases) [3]. Moreover, constellation of metabolic impairment such as insulin resistance (I.R), increased serum level triglycerides, obesity, glucose intolerance, increased risk of developing type 2 diabetes mellitus (T2D), obstructive sleep apnea, hypertension and cardiovascular disease associated comorbidities can be encountered by PCOS women in later period of their life [4–6]. It has been observed that PCOS women suffers from psychological disorders like low self-esteem, anxiety, depression possibly because of negative body image perception eventually further degrading their quality of life [7, 8]. PCOS was first reported by Stien and Leventhal in 1935 as a condition associated with infertility, amenorrhoea, hirsutism and enlarged ovaries [9]. However, till then it remained as the most controversial medical disorder to be studied and diagnosed. No definite diagnostic test can categorically identifies this disease, however, the varying presence of the three defining elements i.e., clinical or biochemical hyperandrogenism, disrupted menstrual cycles (anovulation) and polymorphic ovaries stands considerable [10]. Broadly, speaking three different diagnostic criteria have been put forward from time to time to define the disease state [11] (Table 1). Recently, in 2018 International evidence-based guideline for the assessment and management of PCOS approved Rotterdam criteria for defining the disease [12]. PCOS in the adolescents is not well outlined. However, the epidemiologists and researchers diagnose the disorder based upon well-defined phenotypic features, which mainly include Oligoamenorrhoea / amenorrhoea for atleast 2 years after menarche, ovarian volume > 10cm³, increased serum level androgens and progressive hirsutism as per modified Ferriman-Gallwey scale [13, 14].

Table 1.

Diagnostic criteria and corresponding prevalence rates of PCOS

Diagnostic criteria	Parameters	Diseases prevalence rate (%)	References
NIH (1990)	HA, OD	5–10	[205, 19, 206]
Rotterdam (2003)	HA, OD and PCOM (at least 2 parameters out of 3)	2–21	[205, 19, 206]
AE and PCOS society (2006)	HA defining feature and (OD or PCOM)	2–17	[205, 19, 206]
NIH (2012)	HA, OD and PCOM (at least 2 parameters out of 3) with specific phenotypes Phenotype A: HA + OD + PCOM Phenotype B: HA + OD Phenotype C: HA + PCOM Phenotype D: OD + PCOM	2–21	[205, 19, 206]

NIH National Institutes of Health, HA hyperandrogenism, OD ovarian dysfunction, PCOM polycystic ovarian morphology, AE Androgen Excess-PCOS Society

Despite being so common, PCOS presents an enigmatic state, as the etiologies of this disease remain undefined. Evidences from the scientific data suggest that PCOS is multivariative in nature [15]. The molecular pathomechanism of PCOS chiefly revolves around the underlying hormonal imbalance, impairing the normal ovarian environment leading to disrupted menstrual cycles, cutaneous manifestation and defective metabolic rhythms [5]. To underpin its etiological roots, numerous hypotheses have been suggested from time to time and are being continuously evolving with new research [16]. Although much has been clarified on its pathogenesis, diagnostic criteria and treatment options; it is a noteworthy health condition with no exclusive cure. Currently this disorder continuously evade us, therefore it is important to underpin its pathophysiological links which ultimately can pave for the mechanism based direct intervention to tackle this disease state.

Burden of PCOS

PCOS presents a sharp rise with respect to its prevalence [17, 18]. Understanding its prevalence and phenotype, two important considerations should be taken care of; first geographic variation and second is ethnicity [19]. The overall picture is quiet variable; reason being the various diagnostic criteria employed to validate this disorder [20, 21]. It is reported that more than, 68–75% of PCOS cases remain undiagnosed adding to the ambiguity with respect to its overall prevalence [20, 22]. The first study to determine its prevalence among the reproductive aged women of U.S was carried out by Aziz and colleagues and they documented its prevalence as 4–6.6% as per the NIH, 1990 criteria [23, 24]. Talking about the overall prevalence of this disorder, it is estimated that about 21% of U.S female population are diagnosed with it [24]. Global prevalence of PCOS is not consistent, so far it is generally taken as 5–20%, and changes as per the diagnostic criteria employed [18, 25] (Table 1). Recently Liu et al., [26] reported the global burden of PCOS and associated disability-adjusted life-years (DALYs) between the year 2007–2017 and communicated 1.55 million age-standardized incident cases of PCOS, and 0.43 million associated DALYs. Furthermore, there has been an increase of 1.45% and 1.91% for age-standardized incidence rate of PCOS and DALYs respectively. The study further added that revealed that the incidence rate with respect to the age-standardized PCOS incidence and DALYs was highest in Tropical Latin America compared to southern Latin America, North Africa, and the Middle East at regional level owing to their poor health system and reduced female awareness programs [27]. At national level, the China and the Ethiopia showed the highest increase in the incidence rates of PCOS and Afghanistan observed the lowest incidence rates probably due to the fragile socioeconomic condition of the nation and unawareness among women [26]. The prevalence of PCOS within European population using the data from Global Burden of Disease Study 2016 estimated the overall prevalence rate as 276.4 cases per 100,000 (95% UI: 207.8–363.2) with the highest rates observed in Czech Republic (460.6 per 100,000; 95% UI: 346.2–602.1) and the lowest in Sweden (34.1 per 100,000; 95% UI: 24.59–45.77) [28]. Recent study conducted by et al. communicated the pooled prevalence of PCOS in India was close to 10% using Rotterdam's criteria and AES criteria, while it was 5.8% using the NIH criteria [29]. Sharma et al. studied the overall the prevalence of various menstrual issues related to PCOS women in the Malwa region of Punjab, India and confirmed that 60.61% of women suffered menstrual dysregulation with dysmenorrhea accounting for the majority of issues (50.64%). The study further furnished that the only 3.30% subjects were aware about PCOS [30]. Undoubtedly, women with PCOS have higher lifetime risks of T2DM, metabolic disorders, and metabolic syndrome than weight-matched controls due to their higher rates of insulin resistance. Studies have revealed that 10% of women with PCOS have Type 2 Diabetes Mellitus (T2D), 70% have dyslipidaemia, there is a 5- to tenfold greater chance of developing obstructive sleep apnea, and the incidence of metabolic syndrome ranges from 34 to 46% [31, 32]. Study by Long et al., concluded that the overall prevalence of PCOS in women with T2DM was approximately 21%. It further confirmed that the prevalence of PCOS among women with T2DM was highest in Oceania, followed by Europe and Asia, and was lowest in North America [33]. Assessing the economical burden of this disease (initial diagnosis and treatment of menstrual dysfunction, infertility, and hirsutism) it estimates approximately $3.7 billion annually in 2020 USD [34].

Pathogenesis and Etiology of PCOS

PCOS is marked by abnormal cellular pathways regulating steroid synthesis, body’s metabolic processes and proper ovarian function. In the normal cycling ovaries, follicles are present at developmental stages. The immature follicle known as primodial follicle develops into preovulatory follicle and the process is termed as folliculogenesis [35]. There are finite numbers of follicles present in each ovary containing single oocyte. By the end of folliculogenesis, primodial follicle develops into antral follicle, major source of ovarian steroids. Upon the stimulation of luteinizing hormone (LH) antral follicle, releases the mature oocyte, resulting in the ovulation and what remains of antral follicle after ovulation under goes luteinization forming corpus luteum (important source of progesterone) necessary for the maintenance of pregnancy. Disruption in the major pathways governing the folliculogenesis, steroidogenesis or ovulation consequently results in the adverse conditions like amenorrhea, ovarian dysfunction, infertility, hyperandrogenism or disrupted ovarian morphology.

Broadly, speaking androgens are mainly produced in the ovaries and in the adrenal glands under the stimulation of their tropic hormones luteinizing hormone and adrenocorticotropic hormone (LH and ACTH respectively). Undoubtly, modest androgen excess can disrupt the major pathways associated with the negative feedback mechanism in relation to female sex hormone [36] (Fig. 1). The steroidogenic pathway in the ovaries have been described by two-cell, two-gonadotropin theory [37]. As per this theory, ovarian theca cells and granulosa cells are the major sites involved together in the biosynthesis of all the sex steroids. Cholesterol acts as the major precursor in the synthesis process. It is recruited in the theca cells via lipoprotein receptors or is synthesized de novo. Furthermore, steroidogenic acute regulatory protein (StAR) delivers it to the mitochondria [38] where cytochrome P450 cholesterol side-chain cleavage (cyp450scc; CYP11a1) results in its conversion to pregnenolone [39]. Then 17a-hydroxylase-17, 20-desmolase (CYP17a1 or CYP45017α) catalyze the conversion of pregnenolone to dehydroepiandrosterone (DHEA) and 3β-hydroxysteroid dehydrogenase (HSD3B) to progesterone [40]. DHEA and progesterone is further converted to androstenedione by HSD3B and CYP17a1 respectively [41]. It is pertinent to mention here that CYP17a1 enzyme activity depends on the LH stimulation in a dose-dependent manner [36]. Furthermore, androstenedione is the major precursor for testosterone and estrogen synthesis in ovaries [42]. It is either converted to testosterone via 17β-hydroxysteroid dehydrogenase (HSD17B) in the theca cells or gets diffused to granulosa cells where it is either converted directly to estrone by aromatase (CYP450arom; CYP19A1) [41] or again to testosterone by HSD17B from which it is finally converted to 17β- estradiol mainly by aromatase (CYP450arom; CYP19A1) [43–45]. The androgen synthesis and conversion to estrogens needs to be properly coordinated for the optimized ovarian function, which seems to be disturbed in PCOS women. Simplifying the abovementioned pathway, it can be concluded that the androgens act as obligate substrate for the synthesis of estrogen as well as plays important role in the follicular development [46, 47]. The rate limiting enzyme for the synthesis of androgens is CYP17a1, which on the other hand is dependent upon LH [48] in a dose-dependent manner. LH acts on theca cells via its receptors and induces the production androgens; concomitantly, FSH acts on granulosa cells via its receptors and converts the androgens formed in theca cells into estrogens (estradiol) via aromatase (CYP19A1). The transcription of CYP19A1 is known to be promoted by FSH and hyperandrogenic follicular environment is reported to be hazardous factor leading to the down-regulation of aromatase. Conversely, in PCOS women imbalance in the hypothalamic–pituitary–ovarian axis, results in the impaired androgen synthesis and folliculogenesis. An increase in the hypothalamic GnRH favors the synthesis of β subunit of LH over β subunit of FSH, leading in the production of LH over FSH (increased LH/FSH ratio), hallmark feature of PCOS [49]. Also, increased LH surge results in the follicular arrest at preantral and antral stages leading to hyperplasia of theca cells and the development of multiple follicles along the periphery of ovary [50]. Of note, it can be generalized that the over activity of the steroidogeneic cascade and LH/FSH surge may trigger the ovarian hyperesponsivness, ultimately leading to hyperandrogenism, disrupted ovarian morphology and anovulation.

Fig. 1.

Two-cell, two-gonadotropin theory depicting ovarian sterodgenesis; theca cells produces androgens in response to LH which are later converted into estrogens in granulosa cells under the action of FSH, governed by the series of various enzymes. The androgen synthesis and conversion to estrogens needs to be properly coordinated for the optimized ovarian function, which seems to be disturbed in PCOS women resulting in hyperandrogenism and anovulation [39]

The major ethiopathological hypotheses that have been proposed mainly include:

• 2.1Evolutionary hypothesis
• 2.2Genetic hypothesis
• 2.3Insulin hypothesis
• 2.4Environmental hypothesis
• 2.5Neuroendocrine hypothesis

Evolutionary Hypothesis

An evolutionary hypothesis suggest, that during past times humans were subjected to extreme physical activities also they have to with stand the challenging and stressful environmental conditions. The diet constituents were rich in proteins and carbohydrates with lesser fat content. Also the long periods of famine and disease epidemics might have favored in the development of thrifty genotypes and phenotypes [47]. However, with the scientific revolution and modern lifestyle the life expectancy has dramatically increased. There is no threat of famine and majority of countries have unrestricted accessibility to food with sedentary lifestyle, the need of thrifty genotype and phenotype are no longer beneficial resulting in the dramatic increase in the prevalence of obesity, T2DM and may also have contributed to the development of PCOS [47, 51].

Genetic Hypothesis

Elucidating the genetic basis of PCOS is quiet challenging task. The PCOS presents constellation of symptoms therefore the genes associated with major pathways mainly, steroidogenesis, I.R (I.R), inflammation, glucose homeostasis and those associated with the proper functioning of ovaries could be linked [52, 53]. The familial aggregation of this syndrome also validates its genetic roots [54, 55]. Number of candidate gene association studies has been carried out in relation to PCOS. Walterworth in 1997, studied the role of insulin gene, INS in the pathogenesis of PCOS in revealed its role in the familial transmission of the syndrome [56]. The role of insulin receptor gene in PCOS was demonstrated by Taylor 1992 [57] and the results showed its role in hyper insulinemia and I.R. the role of fat and obesity associated gene (FTO) and childhood obesity are strongly associated particularly in the European population and thus could be possibly linked with PCOS [58]. The association between the genetic mutation in FTO gene and increased risk of PCOS was studied by [59] and they concluded that the SNP rs9939609 is linked withincreased risk of PCOS and can be attributed to increased adiposity. The sex hormone binding globulin (HSBG) is a glycoprotein and specifically binds with the androgens thus regulates their bioavailability. The penta repeat nucleotide polymorphism (TAAAA)n was observed with decreased serum level concentration of HSBG thus marking the increased serum free androgens and consequently increased the risk of PCOS [60]. The GWAS study of DENND1A gene (rs10818854, rs2479106, and rs10986105 SNPs) have been reported to be associated to the high risk of PCOS in Chinese and mixed Asian ancestry respectively [61–63]. Whereas, SNP rs9696009 have been linked with PCOS in European ancestry [64]. SNPs in the INS-VNTR, which are linked to the development of T2DM, were also reported to implicate in PCOS related traits [63, 65]. Shi et al. reported the SNP rs2059807 in INSR associated with PCOS [61], which was further confirmed by Feng et al., [65], who furnished multiple SNP’s association with PCOS development. Studies conducted by by Chen et al., and Shi et al., discovered important FSHR gene loci (rs2268361 and rs2349415) were associated with PCOS and could be related to FSH and LH imbalances and PCOM morphology [61, 62]. The locus THADA represent another intriguing candidate gene in the aetiology of PCOS that has been reproduced across various ancestries [66, 67]. A study conducted by Tian et al., 2020 communicated the role of rs13429458 in THADA, rs10818854 in DENND1A, rs2059807 in INSR and rs4784165 in TOX3a significantly associated in the pathogenesis of PCOS chiefly via disturbing the metabolic rhythms [68]. Several other studies reported the role of GATA4/NEIL2 (rs804279), ZBTB16 (also known as PLZF), MAPRE1 (involved in adipogenesis) may mediate ovarian angiogenesis and follicle development [64, 69]. The variants at the ERBB4, YAP1, C9orf3, DENND1A, RAB5B and ZBTB16 loci were strongly associated with OD and PCOM, and therefore, might be more important for links to menstrual cycle regularity and fertility [64]. The detailed summary of candidate gene studies along with the GWAS studies in relation to PCOS is presented in Tables 2, 3, and 4 respectively.

Table 2.

Candidate gene studies and the associated pathways involved in the pathogenesis of PCOS

Associated pathway	Candidate gene	References
Insulin secretion and metabolism	INS (Insulin gene)	[56, 207]
	INSR (Insulin receptor gene)	[57, 208]
	CAPN10	[209]
	IRS1 (Insulin receptor substrate 1)	[210]
Fat metabolism	FTO gene	[58, 59]
	MC4R	[211]
	ADIPOQ	[212]
Steroidogenic pathway	CYP11a	[213, 214]
	CYP17	[215, 216]
	CYP19	[217]
	CYP21	[218]
	INSL3 (Insulin-like factor 3)	[219]
	HSD17B5 (17β-hydroxysteroid dehydrogenase type 5)	[220]
Regulation of gonadotropin	FSHR	[221]
	FSHβ	[222]
	AMHR	[223]
	LH	[224]
	AMH	[225]
Androgen bioavailability	SHBG	[60, 226]
Inflammation	TNF-alpha	[227]
	IL-6 gene	[228]
Vitamin D	VDR	[229]

Table 3.

Genome-Wide Association Studies (GWAS) and the SNPs identified in relation to Polycystic Ovary Syndrome (PCOS

Nearest gene	Gene locus	SNPs	References
THADA	2p21	rs12468394	[62, 64, 68]
		rs13429458
		rs12478601
		rs7563201
DENND1A	9q33.3	rs10818854	[62, 64, 68, 230]
		rs2479106
		rs10986105
		rs9696009
LHCGR, STON1, GTF2A1L	2p16.3	rs13405728	[64, 230]
FSHR	2p16.3	rs2349415	[61]
		rs2268361
C9orf3	9q22.32	rs4385527	[61, 62]
		rs3802457
		rs10818854
		rs10986105
		rs10993397
YAP1	11q22.1	rs1894116	[61, 230, 231]
		rs11225154
RAB5B, SUOX	12q13.2	rs705702	[61, 64]
HMGA2	12q14.3	rs2272046	[61, 232]
TOX3	16q12.1	rs4784165	[61, 64, 68]
		rs8043701
INSR	19p13.3	rs2059807	[61, 68]
		rs1799817
SUMO1P1	20q13.2	rs6022786	[61, 233]
ERBB4	2q.34	rs2178575	[61]
IRF1/RAD50	5q31.1	rs13164856	[64]
PEX5L	3q26.33	rs7652876	[223]
ERBB3/RAB5B	12q13.2	rs2271194	[64]
HIPK3	11p13	rs4755571	[223]
MAPRE1	20q11.21	rs853854	[64]
GATA4, NEIL2	8p32.1	rs804279	[64]
GYS2		rs7485509	[234]
		rs10841843
		rs6487237

Table 4.

Genome-wide significant associations with PCOS subtypes

Subtype	Gene	Locus	SNPs	References
Reproductive subtype	PRDM2/KAZN	1p36.21	rs78025940	[235]
	IQCA1	2q37.3	rs76182733	[235]
	BMPR1B/ UNC5C	4q22.3	rs17023134	[235]
	CDH10	5p14.2–p14.1	rs7735176	[235]
Metabolic subtype	KCNH7/FIGN	2q24.2–q24.3	rs55762028	[235]
Intermediate subtype	FSHB	11p14.1	rs10835638	[235]

Insulin Hypothesis

Insulin is mainly secreted by the beta cells of pancreas. Its role is well established in ovarian growth and in the modulation of androgens by ovaries [70]. PCOS women are often hyperinsulinemic (50–70% of PCOS patients) [71], thus representing a unique characteristic of this disorder (Fig. 1). I.R simply involves the reduced sensitivity to the normal insulin levels resulting in the irregular glucose metabolism primarily in skeletal muscle and adipose tissue [72]. Studies have revealed that I.R promotes PCOS by disrupting two major pathways- phosphatidylinositol 3-kinase (PI-3 K)/Akt pathway and the mitogen-activated protein kinase (MAPK) pathway [73–75] [Fig. 2]. Different studies have supported the fact that the cross talk between insulin and LH results in the over expression of StAR and Cyp-17 genes in the theca cells thus establishing their potent link in governing the androgen biosynthesis [76]. I.R is a result of disrupted signaling pathways involved in activation of GLUT4 transporter [77]. In trophoblastic cells of humans, I.R results in the decreased expression of sex hormone-binding globulin (SHBG) consequently affecting the expressions of Insulin Receptor sensitive 1 (IRS1), Insulin Receptor Sensitive 2 (IRS2), PI3Kp85 and finally GLUT4 [78] suggesting its active role in inducing PI3K/Akt pathway-mediated systemic I.R. In addition to this, more insulin levels tend to diminish the synthesis of SHBG so to reduce the binding with testosterone thereby increasing the bioavailability of the serum androgens [79]. We can conclude that in PCOS women I.R and hyperandrogenism can be regarded as “partners in crime” in delivering the pathogenesis [Fig. 2]. Burghen et al. [80] firstly described the role of insulin in the ovarian function and proposed a link between hyperinsulinemia and hyperandrogenism. Till then various studies have explained the possible role of I.R in the pathogenesis of PCOS [73, 81]. Thus, I.R and compensatory hyperinsulinism can induce hyperandrogenism in number of ways as:

• Insulin acts as cogonadotropin on ovaries thus, contributing to the excess androgens [73, 82].

• It can act on adrenal glands and facilitates its further secretion [83].

• It affects the LH pulse frequency thus disturbing the delicate balance and driving increased pituitary LH secretion more than FSH secretion, thereby inducing hyperandrogenism and PCOM [84].

• I.R can suppress the production of SHBG, which intrun can further exaggerate with the insulin related signaling pathways and favors hyperandrogenism.

Fig. 2.

Depiction of the possible mechanism involved in triggering the hyperandogenism via defective insulin signaling pathway. Increased serine phosphorylation of IRS-1 hampers the activation of PI3K leading to reduced uptake of glucose by GLUT4 favoring I.R. I.R interferes with the SHBG, acts on theca cells and provokes LH excess to aggravate hyperandrogenism. On the other hand, the picture becomes further exaggerated when HA downregulates the aromatase activity leading to follicular atresia. MAPK-ERK1/2 impairment inhibits GCs proliferation resulting in follicular atresia [73–75]

The abovementioned picture becomes more exacerbated in PCOS obese women because of adiposity. The increased phosphorylation of the serine residue of the insulin receptor substrate-1 molecule (IRS-1^{ser 312}) has been witnessed in the obese PCOS women, consequently leading to impaired Insulin receptor signaling (impaired regulation of IRS-1 and PI3K activity) leading to hyperinsulinemia, favoring hyperandrogenism [85]. Additionally, downregulation of MAPK-ERK1/2 caused enhanced rate of apoptosis and diminished proliferation of granulosa cells in PCOS women. Studies suggested that disruption in mTORC1 autophagy pathway results in mitochondrial impairment and reduction in uptake of glucose, leading to I.R in skeletal muscles due to hyperandrogenism [86, 87]. The Vitamin D also plays a significant role in governing the insulin signaling pathway in PCOS women however, the deficiency of the former modifies the entire picture. Vitamin D deficiency triggers IR [88]. Vitamin D also serves very important precursor for the adipose cells development and is involved in the activation of enzyme system important for lipid and carbohydrate metabolism thus playing an important role in glucose metabolism [89]. In addition, the genetic variation in the MTNR1B gene has been witnessed in PCOS women resulting in the impairment in insulin secretion thus indicating its possible role in PCOS and IR cross talk [90]. Furthermore the role of insulin like growth factors 1&2, their receptors and ILGFBPs have been well studied for their role in antral follicle development [91, 92]. Studies have revealed that hyperinsulinemia can alter synthesis of IGF- binding proteins thus increase the levels of free IGF-1[93]. IGF-1 directly affects the miRNA-323-3p regulation [94] consequently leading to accelerated granulose cell apoptosis via IGF-1, thus obstructs follicular development [94]. Moreover, the role of vascular endothelial growth factor (VEGF) has also been documented in the pathogenesis of PCOS [95]. However, more research is needed to simplify how I.R actually alters the follicular development and hinders the ovulation.

Environmental Hypothesis

There has been growing evidences in the recent years that environmental toxins could contribute to the pathogenesis of PCOS. In the current industrial era, the indiscriminate use of chemicals has significantly affected the health status and has potentially altered the reproductive health by directly interfering with the normal ovarian morphology as well as function. Researches throughout the world are taking pains to understand the possible link between environmental toxins and pathophysiology of PCOS. The most well studied environmental risk factor in the pathogenesis of PCOS includes Endocrine disrupting chemicals (EDCs). EDCs are the group of heterogeneous group of compounds containing a phenolic group in their chemical structure making them to mimic the body’s endogenous hormones (e.g., steroid hormones, and thyroid hormones) eventually interfering with their receptors trigging altered molecular mechanism. Not only this, these chemicals alter the delivery of natural blood borne hormones by mingling with their transport proteins (like sex hormone binding globulin) consequently disturbing body’s homeostasis and impairing with developmental, metabolic and reproduction processes [96, 97]. However, the toxicity of such chemicals is determined by the time of exposure (fetal, adulthood), variety and concentration as it is likely that the cocktail of chemicals might result in more complex effects on the individual’s health [98]. More than 800 chemicals are documented as EDC’s and some of them possesses lipophilic properties can bioaccumlate in the adipose tissue (pesticides, POPs) leading to more adverse health effects [99, 100]. Humans are consuming packed foods in plastic containers, using cosmetics, plastic baby bottles, sanitary napkins, diapers, thermal papers, PVC pipes; these are found to be loaded with plasticizers which is considered to be the chief source of EDCs (BPA, BPF, phthalates). Also the pesticides are studied as the persistent toxicants with similar properties [101, 102]. EDC’s can lead to the disruption of various pathways potentially leading to PCOS and its consequences (Fig. 3). Some of the ways are summarized as under:

• Environmental toxins chiefly EDC’s can directly affect the hormonal homeostasis thus leading to anovulation and reproductive dysfunction. EDC’S can either directly target ovaries or indirectly impact the HPO axis (hypothalamic pituitary ovarian axis) [103, 104] leading to hormonal disruption. BPA possess weak affinity towards variety of receptors (classical NRs α and β) [105], non-classical membrane ERs [106] thus favoring hormonal destabilization. Furthermore, studies have revealed that the expression of aromatase and E2 synthesis in the granulosa cells was downregulated in PCOS women vis-à-vis with higher levels of BPA in their follicular fluid sighting role of such toxins in disrupting the intrafollicular environment [107].

• EDC’s can interfere with the steroidogenic pathways in multiple ways. A vast literature supports that BPA as well as phthalate interferes agonistically or antagonistically with the steroid binding proteins, steroid receptors or somehow modulates steroidogenic enzymes in the ovaries. BPA modulates the expression of various steroidogenic enzymes chiefly CYP17a1 and Cyp11a1, involved in the synthesis of androgens [107]. Additionally EDC’s may also interfere with the androgen catabolism thereby increasing the total free androgen index in PCOS women [108, 109]. This on the other hand downregulates the activity of hepatic UGT enzymes, key phase II players involved in the metabolism and clearance of BPA [110], thus exacerbating this vicious cycle in PCOS women.

• EDC’s can alter the metabolic pathways as well which is been already entangled among the PCOS women thus making them more vulnerable. Obesity, I.R and compensatory hyperinsulinemia witnessed in majority of PCOS women; EDC’s can find their way in disturbing these delicate metabolic pathways. EDC’s are regarded as obesogens as they promote fat storage [111]. Animal models have revealed that EDC’s like mono-isobutyl phthalate, tributyltin and BPA act via various receptors (PPARγand ER) thus modulating different molecular pathways involved in adipogenisis and accumulation of lipids [112–114]. The role of various EDC’s in inducing hyperinsulinimia has been well documented in various toxicological studies [115]. EDC’s can directly alter the glucose metabolism [116] or indirectly may decrease the secretion of adiponectin which may later on lead to I.R [117].

• Apart from plasticizers (BPA, phthalate) number of air pollutants have been studied for their role in causing the defects related to reproductive health [118]. Some of these are enlisted as EDC’s (PHA, heavy metals- Pb, Cu, Zn) [118, 119]. Studies suggest that these chemicals may induce compromised oxidative state or can lead to DNA abrasion leading to epigenetic modifications [120]. PCOS has been linked to the decreased antioxidant concentration and is termed as an oxidative state [121]. Oxidative stress developed in response to the excess ROS production have been seen to cause impaired folliculogenesis [122]. Elevated levels of oxidative stress biomarkers were reported from PCOS women indicating their role in diseases development [123]. A study conducted by Lin et al., [124] concluded that the women exposed to air pollutants chiefly- SO₂, NO₂, NO and PM_2.5 are at high risk in developing PCOS. Sun and colleagues studied the role of particulate matter (< 2.5 μm; PM_2.5) in diet-induced I.R, adipose inflammation, and visceral adiposity in C57BL/6 mice. They communicated that the inflammatory state because of oxidative stress and Tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), E-selectin, intracellular adhesion molecule-1 (ICAM-1), plasminogen activator inhibitor-1, and resistin were main culprits in resulting this compromised metabolic state. Another study carried out by González et al., [125] revealed that the proinflmmatory response could stimulate the ovarian theca cells for the over production of androgens leading to PCOS. Thus, we can conclude that the inflammatory condition in response to the oxidative stress may result in the creation of hostile redox environment favoring metabolic inflexibility and hyperandrogenism consequently leading to PCOS progression [102, 126]. Furthermore, defective menstrual and ovarian functions were reported in relation to the numerous organic pollutants such as Tetrachlorodibenzo-p-dioxin, Dichlorodiphenyltrichloroethane, polychlorinated biphenyl (TCDD, DDT, and PCBs) [127–129].

Fig. 3.

Potential impact of environmental toxins on female reproductive health linked to PCOS phenotypes and adverse health effects

Neuroendocrine Hypothesis

PCOS considered to be an ovarian dysfunction but accumulating various clinical and pre-clinical evidence indicates the role of master gland “pituitary-gland”, located in the fore brain as the major suspect in the pathophysiology of the disease [130–132]. Numerous findings have suggested that any alteration in the specific neuronal network in the brain controlling fertility can results in the downstream ovarian dysfunction. Gonadotropin Releasing Hormone (GnRH) neuronal network is comprised of the small population of GnRH neurons. This portion is the fertility governing circuit scattered mainly in the fore brain and around glial cells, transmitting normal hormonal, metabolic and stress related signals to the reproductive axis [133]. GnRH neurons stimulate the release of tropical peptide hormone GnRH, consequently governs the release of two important gonadotropins; LH and FSH from the anterior pituitary. GnRH is released in a pulsatile manner and the frequency of the pulses determines which gonadotropin is secreted. A faster GnRH pulse favors the release of LH on contrary a weak pulse drives FSH from the master gland [134, 135]. Of note, the balanced LH/FSH ratio downstream regulates important ovarian events typically ssteroidogenesis and folliculogenesis. The ovarian cycle comprises of three basic phases, (1) Follicular phase, (2) Ovulatory phase and (3) Leutal phase. There exists a balanced neuroendocrine loop between the ovaries and hypothalamus pituitary axis governing the release of GnRH, LH and FSH. Over most of the ovarian cycle, the estradiol and progesterone send a negative feedback to the GnRH neuron activity favoring FSH surge and thus regulating the folliculogenesis. A positive feedback is mediated during the mid follicular phase driving GnRH/LH surge triggering ovulation. This delicate loop is somewhat disrupted in PCOS women resulting in the ovarian dysregulation. It is well recognized that increased LH frequency favors the hyperproduction of androgens by the theca cells leading to hyperandrogenism [136, 137]. Moreover, the decreased FSH frequency does not allow the follicular development thus arresting the folliculogenesis eventually resulting in the formation of tiny follicular cysts and anovulation [136, 138] [Fig. 4. Studies have showed that more than 95th percentile of the elevated LH at a single time measurement has been witnessed in 75% of PCOS women [139]. Scientists worldwide are proposing various mechanisms to explain this neuroendocrine impairment of PCOS. The upstream neuron network mostly regulates GnRH activity [140]. Karolina et al., [141] reported that the kisspeptin network plays significant role in reproductive processes. Kisspeptin directly regulates GnRH function via its receptors, located on the GnRH neurons. Administration of kisspeptin can directly regulate the LH secretion [142]. Studies have revealed that the invitro kisspeptin exposure can increase the firing rate of GnRH and stimulates the secretion of GnRH in hypothalamic explants [143, 144]. In addition, to underpin its role in the release of LH, it was observed that using Kisspeptin antagonist the pulsatile release of GnRH was affected consequently affecting the LH/FSH pulse which was further demonstrated by number of studies [145, 146]. Taking the advantage of optogenetics and chemogenetics techniques, the arcuate nucleus population of kisspeptin neurons (ARNKISS) has been identified as the extrinsic GnRH pulse generator [147, 148]. Studies have further confirmed that the inactivating mutations in kiss genes or its receptors resulted in hypogonadotropic hypogonadism and delayed puberty [149, 150], whist its activating mutations was related to precocious puberty [151, 152], suggesting that kisspeptin modulates GnRH pulsatility. It is pertinent to mention here that the classical neurotransmitter GABA is seen to play a significant role in the maintaining proper steroid hormone feedback [153]. The role of these neuronal population in GnRH/LH pulse patterns and PCOS pathophysiology have also been reported in several studies [154–158]. In addition, several animal models revealed that the exposure to the exogenous androgens or anti-Müllerian hormone (AMH) during the various developmental stages may induce PCOS like symptoms latter on [159–162]. The prenatal animal model (PNA) supports the fact that the prenatal exposure to the excessive androgens results in the PCOS-like neuroendocrine dysfunction [132, 161, 163, 164]

Fig. 4.

Neuroendocrine circuit disruption leading to PCOS

Potential Drugs and Therapeutics

PCOS is turning out to be monstrous in nature with increasing prevalence in both developing and developed countries. Unfortunately, no universal treatment is currently available to cure it completely but the current treatments are mostly symptom oriented, thus managing this disorder seems to be the only promising approaches available [165]. The present strategy of the researchers and pharmacologist is to target I.R, hyperandrogenism and oligomenorrehea. The long-term management to check the associated commorbities should include life style counselling to prevent obesity, T2DM, CVD or other metabolic complication [166].

Managing Hyperandrogenism

More than 80% of PCOS women suffer from hyperandrogenism i.e., hyperproduction of androgens leading to the elevated levels of circulating testosterone and androsterodione It represents to be the most burdensome characteristics as it triggers masculine phenotypic appearance besides having health consequences. Cholesterol serves as the chief precursor for the synthesis of androgens via series of various enzymatic reactions. The upreguation of various steroidogenic enzymes chiefly CYP17a1, Cyp11a1, 3βHSD, AKRIC1 are seen to be the major hyperandrogenic culprits in PCOS women [36, 167]. Thus, the simple approach that can be manifested to manage hyperandrogenism is to attenuate the expression of aforementioned enzymes. Moreover studies have confirmed that the downregulation of the expression of GnRH can lessen down LH pluse thus balancing LH/FSH ratio [165]; proving helpful in managing hyperandrogenism. A variety of drugs mainly oral contraceptive pills (OCPs) and anti-androgens are advised to PCOS women who are not attempting to conceive [168]. Anti-androgenic drugs mainly eflornithine, finastride can prove teratogenic [169, 170] therefore a combined therapy of OCPs with neutral or anti-progestin and/or anti-androgens (androgen receptors blockers) namely spironolactone, cyproterone acetate or flutamide acting as antagonists of androgen receptors proves highly effective for hirsutism [168]. Dopamine and dopamine agonists have long been known to decrease females' production of LH [171]. Bromocriptine, a dopamine agonist, may be helpful for PCOS patients by lowering LH hypersecretion. For acne retinoids[172], and for alopecia minoxidil [173] proves to be drug of choice. Cosmetic techniques are also advised along with the oral drugs depending upon the severity and patient’s requirement. For managing hirsutism waxing, bleaching, shaving, plucking and laser treatment are employed [168]. For acne dermocosmetics [174], light therapy [175] or cosmetic surgery can be helpful. Alopecia can be resolved by hair transplantation [176] or inducing growth factors from platelet rich plasma [177]. Statins are the most recent therapies that have been incorporated into PCOS treatment methods. They are helpful in the treatment of PCOS because they decrease the synthesis of sex hormones, alleviate dyslipidemia, and lessen the development of ovarian androgen by preventing the creation of androgen by thecal cells[171, 178]. Studies have shown that women with PCOS specifically have higher levels of bad bacteria (Escherichia and Shigella) and lower levels of good bacteria (Lactobacilli and Bifidobacteria) [179, 180]. The metabolic profile in PCOS has been demonstrated to benefit from probiotic administration [181, 182]. When compared to placebo, PCOS patients showed a statistically significant decrease in weight, BMI, plasma glucose, serum insulin levels, and a reduction in androgen levels [181, 183, 184]

Managing I.R

I.R appears to be the most common metabolic consequences related to PCOS. Hyperinsulinemia can pave to various secondary comorbidities in PCOS women in future of which T2DM and CVD appears to be most vulnerable. The basic strategy to manage this is life style modifications i.e. maintaining proper diet plan and physical activities. Studies have revealed that life style modifications not only improve hyperinsulinimia but also prove to be effective against hyperandrogenism in PCOS women [185]. A variety of anti-obesity drugs are advised to improve weight loss in PCOS women but their effectiveness remains to be established [186]. The bariatric surgery proves to be fruitful option for PCOS obese women [187]. The mainstay of pharmacotherapy of PCOS against hyperinsulinimia proves to be metformin, insulin sensitizer drug. It proves to be a mutihand drug which not only improves weight loss but also reduces hyperandrogenism and related symptoms [188–190]. Inositols, another insulin sensitizer drug have also proven effective in attenuating the PCOS symptoms [191, 192]. Berberine, a plant based alkaloid also showed promising results in animal based models against I.R by modulating expression of GLUT type 4 (glucose transport proteins) [193] but it needs to be further elucidated [194]. The class of anti-diabetic medications known as GLP-1 receptor agonists imitates the effects of incretin. In both diabetic and PCOS patients, GLP-1 agonists provide a dual control by lowering body fat and hyperglycemia thus showing promising results in managing I.R. [195]. DPP-4 inhibitors are seen to increase the levels of incretin in the body [196] there by regulating the normal blood glucose levels. SGLT2 inhibitors have also proven impactful in managing I.R by regulating blood glucose levels in the body through glycosuria and natriuresis. SGLT2 inhibitors increase urinary glucose excretion, weight loss and improve cardiovascular risks in patients with type 2 diabetes. Only one drug of SGLT2 inhibitors (Empagliflozin) have been used in PCOS patients with markedly effective results to manage I.R when compared to metformin [197]. Supplementing with vitamin D and calcium may have therapeutic effects on I.R, menstrual regularity, hyperandrogenism, and follicular development in women with PCOS [198, 199]. Vitamin D therapy normalizes the serum AMH levels, which may lead to improved folliculogenesis [200].

Managing Oligomennnorhea

Oligoovulation or oligomennorhea represents another common feature of PCOS. It is associated with menstrual dysfunction where ovaries seldom ovulate leading to subfertility. PCOS women experiencing oligomennorhea are at higher risk of endometrial hyperplasia, canacer and infertility [1]. Two basic approaches could be put into action based on patient’s requirement. PCOS women not attempting for pregnancy should be prescribed OCPs [201], cyclic or continuous progestin administration [201], or levonorgestrel –releasing intra utride device [202] can be employed as per the requirement and severity of her symptoms. Women expressing desire to get pregnant it is important to know the severity of oligoanovulation. If only mild menstrual dysfunction is present, proper counseling and suggestions regarding their most fertile days will suffice their desire. In contrast when the symptoms are more severe with acute menstrual dysfunction or when ovulation is unpredictable, ovulation needs to be induced [1] and accordingly clomiphene citrate or letrozole remain drug of choice [203]. Metformin may also be used as the data suggests that it can improve ovulation and pregnancy rates [204].

Conclusion and Future Perspectives

PCOS represents the most prevalent female endocrine disorder affecting the ovulation consequently leading to sub-fecundity in the industrialized population. The disease represent spectrum of symptoms thus making it difficult to understand and diagnose. In this review, we tried to discuss its major etiological factors and pathological links (Fig. 5). Plentiful researches suggests that the PCOS is no longer restricted as an ovarian diseases, rather numerous pathways—neuroendocrine, metabolic dysregulation, defective steroid state, environmental factors and genetic predisposition may induce and paves way for disease development. So taking consideration its increasing prevalence, it is need of the hour to understand the complex interplay among the potential pathways inducing HA, IR, metabolic aberration in shaping this disease. Molecular biology provides a promising approach to simplify this multifactorial paradox by exploring a common denominator that may trigger abnormal regulation of steroidogenesis, homologous desensitization to LH, folliculogenesis and metabolic dysregulation thus ultimately kick starts this vicious cycle. Meanwhile, unraveling the diverse pathological links will help to deduce a proper treatment opportunities of PCOS at early stages.

Fig. 5.

Interplay between the potential pathways that gets disrupted, eventually leading to reproductive and metabolic failure witnessed in PCOS

Abbreviations

NIH

National Institutes of Health

HA

Hyperandrogenism

OD

Ovarian dysfunction

PCOM

Polycystic ovarian morphology

AE

Androgen Excess-PCOS Society

I.R

Insulin resistance

LH

Luteinizing hormone

FSH

Follicle stimulating hormone

Author Contributions

Khair Ul Nisa and Bashir Ahmad Ganai conceived and designed this review. Khair Ul Nisa, Najeebul Tarfeen, Shahnaz Ahmad Mir, Ajaz Ahmad Waza, and Mir Bilal Ahmad collected the literature for this review. Khair Ul Nisa wrote the manuscript draft. Bashir Ahmad Ganai and Shahnaz Ahmad Mir edited this manuscript. All the authors gave final shape to this manuscript.

Funding

The work was not supported by any research grant.

Declartions

Conflict of interest

Authors declare no conflict of interest.