**5. Hyperandrogenism is a key factor mediating increases in blood pressure in women with PCOS**

Increased BP remains the leading risk factor for death globally, accounting for 10.4 million deaths per year [36]. There is a sexual dimorphic relationship between androgens and BP in women compared to men. Free androgen index

(FAI), a measure of bioavailable androgens, is positively correlated with systolic and diastolic blood pressure [37]. A recent meta-analysis reported a greater risk of developing hypertension in reproductive age women with PCOS [38]. In contrast, epidemiological studies have shown an inverse relationship between androgens and BP in men, and this association persisted after adjusting for age and body mass index [39]. The mechanisms behind the elevated BP in response to hyperandrogenism in women remain unclear. Some of the possible mechanisms involved in androgen-mediated regulation of BP are discussed below.

## **6. Potential mechanisms that could mediate increases in blood pressure in PCOS**

The kidneys are a key regulator of long-term BP control and body fluid homeostasis in the body. The renin-angiotensin-aldosterone system (RAAS) plays a major role in several forms of hypertension, and it is composed of the classical and non-classical arms with opposite physiological effects [40, 41]. The classical RAS pathway starts with angiotensinogen (AGTN) which is enzymatically cleaved to Angiotensin I (ANG I) by renin. ANG I is then cleaved by angiotensin I converting enzyme (ACE) to Angiotensin II (ANG II), which binds to the ANG II receptor type 1 (AT1R) and ANG II type 2 receptor (AT2R). High levels of ANG II had been related to metabolic disorders. The non-classical pathway, the angiotensin I converting enzyme 2 (ACE2) reduces Ang II levels by transforming it in Angiotensin (1–7) (ANG (1–7)), which also can be generated from ANG I passing through Angiotensin (1–9) (ANG (1–9)) by the action of the endopeptidases: prolylendopeptidase and neutral endopeptidase. The main known biological effects of ANG (1–7) are associated with Mas receptor activation, causing an improvement in metabolic syndrome, obesity, and hypertension.

The rate-limiting step of the RAS is the conversion of AGTN to ANG I by renin [42]. Women with PCOS have hyperreninemia [43], and blockade of the AT1R is effective in decreasing their BP [44]. We and others have shown that androgens stimulate the synthesis of intrarenal AGTN in male and female rats [45–48]. In the kidney, the AR is highly expressed in proximal tubule cells [49], glomerular endothelial cells [50], and podocytes [51]. Moreover, the enzymes involved in androgen biosynthesis are expressed and active in the kidney [52]. In experimental models of hypertension, androgens can shift the pressure-natriuresis curve to the right, promoting sodium reabsorption [53, 54]. Androgens could also directly increase sodium reabsorption via upregulation of epithelial sodium channel (ENaC) α, β, and γ subunits expression [55]. Renal medullary blood flow and the sensitivity of the pressure-natriuresis response are regulated by various paracrine and humoral factors known to play an important role in the control of renal function and BP. Those regulatory factors include ANG II, kinins, prostaglandins, atrial natriuretic peptide (ANP), and nitric oxide (NO). Whether changes in the renal medulla microcirculation play a role in mediating the increases in BP under excess of androgens in women with PCOS remains unclear.

Plasma ACE2 activity is low in healthy subjects but elevated in patients with cardiovascular risk factors or cardiovascular disease. Hypertensive men have a high higher level of plasma ACE2 compared to women [56]. The role of increased levels of plasmatic ACE2 in men remains unknown. It has been reported that male mice have higher renal ACE2 mRNA and protein expression, as well as higher ACE2 activity, than their female counterparts [57, 58]. However, the effect of androgen excess in the non-classical RAS pathway is still not well understood. Androgens downregulate AT2R expression levels in the aorta, *in vivo*, and *ex vivo* [59]. ACE2 is

#### *Androgens and Cardiovascular Risk Factors in Polycystic Ovary Syndrome DOI: http://dx.doi.org/10.5772/intechopen.96005*

the receptor for SARS-CoV and SARS-CoV-2 [60, 61]. SARS-CoV and probably SARS-CoV-2, Spike protein binding to the ACE2 receptor causes its downregulation through internalization [62]. SARS-CoV causes an imbalance in ACE/ACE2 and consequently ANG II/ANG (1–7) that leads to lung injury [62]. Men have suffered a higher rate of severity and mortality from COVID-19 [63]. Whether such sex difference in COVID-19 outcomes is due to ACE2 expression modulation by androgens remains unknown. Further research is needed to elucidate whether and how androgens modulate the non-classical RAS pathway in PCOS and how its regulation could impact COVID-19 outcomes in this population.

Our research teams' studies focus on the cardiometabolic complications associated with androgen excess in female rats. The hyperandrogenic female (HAF) rat, an animal experimental model of PCOS, is generated by the chronic administration of the non-aromatizable androgen DHT. This model exhibits upregulation of intrarenal angiotensinogen and ACE mRNA expression, which are associated with a ~ 10 mmHg increase in BP compared to control female rats [46]. When HAF rats are treated with enalapril, an ACE inhibitor, their BP is lowered to values comparable to that of control rats, suggesting that the RAAS activation has a role in mediating androgens' effect on BP [64]. The stimulatory effect of androgens upon the intrarenal RAAS persisted after discontinuation of androgen exposure in female rats, suggesting a cardiometabolic androgenic memory in female rats. Interestingly, in the kidney medulla, AGTN and AT1R were still elevated after six months of DHT withdrawal [65]. AT1R blockers or ACE inhibitors are widely used as antihypertensive drugs. Women should be advised about the potential teratogenic and fetotoxic risks of ACE inhibitors or AT1R blockers if they become pregnant. ACE inhibitors and AT1R blockers use in the first trimester of pregnancy may not present significant risks for malformations in live births but a high risk of miscarriage [66]. Novel and tissue-selective RAAS inhibitors that do not cross the placental barrier are warranted to ameliorate the increases in BP in women with PCOS in the future.

In the US, a frequent finding in PCOS patients is an increase in body mass index (BMI), with up to 80% being either overweight or obese [67]. There is a strong link between adiposity and hypertension, with multiple mechanisms being suggested [68]. Hypertrophy of adipocytes is associated with local hypoxia, leading to increased oxidative stress and inflammatory cytokines, followed by capillary rarefaction [69, 70]. These processes can lead to a positive feedback loop, ultimately releasing more inflammatory cytokines and reactive oxidative species into the systemic circulation. Chronic inflammation can ultimately lead to increased BP. In HAF rats, there is increased fat mass and BP coupled with increased plasma tumor necrosis factor-alpha and renal mRNA expression of NADPH oxidase 4 [46, 71]. Increased adiposity is also associated with increased circulating adipokines, such as leptin [69]. Chronic hyperleptinemia is known to stimulate the sympathetic nervous system [72], which could lead to vasoconstriction. It has been reported that in women with PCOS, leptin levels can be elevated [73]. Furthermore, using heart rate variability as a measure of autonomic dysfunction, women with PCOS have increased sympathetic activity compared to control women matched for body mass index, systolic and diastolic BP [74]. Additionally, leptin is linked to activation of the RAAS via the renal sympathetic nervous system [72]. All the findings mentioned above suggest that BP control is complex and depends on multiples pathways in women with PCOS.

Endothelial dysfunction refers to the impaired function in the endothelium, the inner lining of blood vessels, which could lead to inappropriate vasoconstriction and atherosclerosis [75]. Endothelial dysfunction frequently occurs under chronic inflammation conditions or high oxidative stress, which interfere with the nitric oxide production needed for vasodilation [76]. Interestingly, this occurs not just in obese females but also in lean females with PCOS. A recent study found that normotensive lean females with PCOS, even without insulin resistance, had increased endothelial dysfunction compared to controls [77]**.** One of the major vascular oxygen-derived free radicals is superoxide anion. Superoxide is routinely scavenged by superoxide dismutase (SOD). Superoxide can also combine with nitric oxide (NO), which results in quenching of NO and, theoretically, can induce vasoconstriction. There is an interaction between NO and oxidative stress to maintain endothelial function. We previously showed that an intact NO system is necessary for antioxidants to elicit a BP-lowering effect [78]. Furthermore, Huirliman and colleagues demonstrated that the presence of endothelial dysfunction and IR develops in pair-fed DHT-treated female rats, suggesting an obesity-independent mechanism [79]. Increased endothelial dysfunction has also been found in transgender men compared to cisgender women matched for body mass index [80], suggesting a broader link between endothelial dysfunction and female androgen excess in addition to women with PCOS. Another study with lean females with PCOS also found that they had decreased plasma total antioxidant status [81]. This reduced ability to handle oxidative stress can contribute to the endothelial dysfunction in hyperandrogenic females.

Cardiovascular diseases are the leading cause of death in women. Furthermore, there have been an overall decline in CVD mortality over the past 40 years; however, the mortality in younger women has plateaued since 2000 [82]. Increases in BP is a primary cardiovascular risk factor. The carotid artery intima-media thickness (cIMT) has emerged as an important surrogate marker of target organ damage in hypertensive heart disease. A recent prospective cross-sectional study in PCOS women showed that cIMT was significantly increased in women with PCOS compared to controls, and this increase was independent of BMI, age, and smoking status [83]. Consequenly, the cIMT could be used to determine the cardiovascular risk profile in women with PCOS.

In summary, hyperandrogenemia in females has multiple mechanisms of causing increased BP and impaired vascular function. Pharmacological agents that target multiple pathways could constitute effective therapeutic agents to be used in women with PCOS.
