Prenatal Androgen Excess Induces Multigenerational Effects on Female and Male Descendants.

Background: It is still unelucidated how hormonal alterations affect developing organisms and their descendants. Particularly, the effects of androgen levels are of clinical relevance as they are usually high in women with Polycystic Ovary Syndrome (PCOS). Moreover, it is still unknown how androgens may affect males' health and their descendants. Objectives: We aimed to evaluate the multigenerational effect of prenatal androgen excess until a second generation at early developmental stages considering both maternal and paternal effects. Design And Methods: This is an animal model study. Female rats (F0) were exposed to androgens during pregnancy by injections of 1 mg of testosterone to obtain prenatally hyperandrogenized (PH) animals (F1), leading to a well-known animal model that resembles PCOS features. A control (C) group was obtained by vehicle injections. The PH-F1 animals were crossed with C males (m) or females (f) and C animals were also mated, thus obtaining 3 different mating groups: Cf × Cm, PHf × Cm, Cf × PHm and their offspring (F2). Results: F1-PHf presented altered glucose metabolism and lipid profile compared to F1-C females. In addition, F1-PHf showed an increased time to mating with control males compared to the C group. At gestational day 14, we found alterations in glucose and total cholesterol serum levels and in the placental size of the pregnant F1-PHf and Cf mated to F1-PHm. The F2 offspring resulting from F1-PH mothers or fathers showed alterations in their growth, size, and glucose metabolism up to early post-natal development in a sex-dependent manner, being the females born to F1-PHf the most affected ones. Conclusion: androgen exposure during intrauterine life leads to programing effects in females and males that affect offspring health in a sex-dependent manner, at least up-to a second generation. In addition, this study suggests paternally mediated effects on the F2 offspring development.

Role of Hormones During Gestation and Early Development: Pathways Involved in Developmental Programming.

Accumulating evidence suggests that an altered maternal milieu and environmental insults during the intrauterine and perinatal periods of life affect the developing organism, leading to detrimental long-term outcomes and often to adult pathologies through programming effects. Hormones, together with growth factors, play critical roles in the regulation of maternal-fetal and maternal-neonate interfaces, and alterations in any of them may lead to programming effects on the developing organism. In this chapter, we will review the role of sex steroids, thyroid hormones, and insulin-like growth factors, as crucial factors involved in physiological processes during pregnancy and lactation, and their role in developmental programming effects during fetal and early neonatal life. Also, we will consider epidemiological evidence and data from animal models of altered maternal hormonal environments and focus on the role of different tissues in the establishment of maternal and fetus/infant interaction. Finally, we will identify unresolved questions and discuss potential future research directions.

Intrauterine androgen exposure impairs gonadal adipose tissue functions of adult female rats.

Prenatal androgen exposure induces fetal programming leading to alterations in offspring health and phenotypes that resemble those seen in women with Polycystic Ovary Syndrome. It has been described that prenatal androgenization affects the reproductive axis and leads to metabolic and endocrine disorders. Adipose tissue plays a crucial role in all these functions and is susceptible to programming effects. Particularly, gonadal adipose tissue is involved in reproductive functions, so dysfunctions in this tissue could be related to fertility alterations. We aimed to investigate the extent to which prenatal hyperandrogenization is able to alter the functionality of gonadal adipose tissue in female adult rats, including lipid metabolism, adipokines expression, and de novo synthesis of steroids. Pregnant rats were treated with 1 mg of testosterone from day 16 to day 19 of pregnancy, and female offspring were followed until 90 days of age, when they were euthanized. The prenatally hyperandrogenized (PH) female offspring displayed two phenotypes: irregular ovulatory (PHiov) and anovulatory (PHanov). Regarding lipid metabolism, both PH groups displayed disruptions in the main lipid pathways with altered levels of triglyceride and increased lipid peroxidation levels. In addition, we found that Peroxisome Proliferator-Activated Receptors (PPARs) alpha protein expression was decreased in both PH phenotypes (p < 0.05), but no changes were found in PPARγ protein levels. Furthermore, regarding adipokines, no changes were found in Leptin and Adiponectin protein levels, but Chemerin protein levels were decreased in the PHiov group (p < 0.05). Regarding de novo synthesis of steroids, the PHanov group showed increased protein levels of Cyp17a1 and Cyp19, while the PHiov group only showed decreased protein levels of Cyp19 (p < 0.05). These results suggest that prenatal androgen exposure affects females' gonadal adipose tissue in adulthood, disturbing different lipid pathways, Chemerin expression, and de novo synthesis of steroids.

Polycystic ovary syndrome in Latin American populations: What is known and what remains unresolved.

Polycystic ovary syndrome (PCOS) is one of the main endocrine and reproductive disorders affecting women in their reproductive age. The syndrome is considered a multifactorial pathology. Therefore, genetic susceptibility and environmental factors contribute to PCOS development and phenotypic manifestation. Ethnicity and socioeconomic factors influence the development of PCOS and could affect the possibility of its diagnosis. Latin America is a unique case of study because of the heterogeneity within the region, complex socioeconomic status, and the mixed ancestry found in these populations. Up-to-date, most studies have focused on developed countries' populations, and there is a lack of evidence regarding Latin-American countries. We propose to review the state of the art of PCOS knowledge regarding Latin American populations, including the metabolic and reproductive aspects of the syndrome and the different influencing factors, and suggest future directions to deepen the study of PCOS.

Fetal programming by androgen excess impairs liver lipid content and PPARg expression in adult rats.

It is known that prenatal hyperandrogenization induces alterations since early stages of life, contributing to the development of polycystic ovary syndrome affecting the reproductive axis and the metabolic status, thus promoting others associated disorders, such as dyslipidemia, insulin resistance, liver dysfunction, and even steatosis. In this study, we aimed to evaluate the effect of fetal programming by androgen excess on the hepatic lipid content and metabolic mediators at adult life. Pregnant rats were hyperandrogenized with daily subcutaneous injections of 1 mg of free testosterone from days 16 to 19 of pregnancy. The prenatally hyperandrogenized (PH) female offspring displayed two phenotypes: irregular ovulatory phenotype (PHiov) and anovulatory phenotype (PHanov), with different metabolic and endocrine features. We evaluated the liver lipid content and the main aspect of the balance between fatty acid (FA) synthesis and oxidation. We investigated the status of the peroxisomal proliferator-activated receptors (PPARs) alpha and gamma, which act as lipid mediators, and the adipokine chemerin, one marker of liver alterations. We found that prenatal hyperandrogenization altered the liver lipid profile with increased FAs levels in the PHanov phenotype and decreased cholesterol content in the PHiov phenotype. FA metabolism was also disturbed, including decreased mRNA and protein PPARgamma levels and impaired gene expression of the main enzymes involved in lipid metabolism. Moreover, we found low chemerin protein levels in both PH phenotypes. In conclusion, these data suggest that prenatal hyperandrogenization exerts a negative effect on the liver and alters lipid content and metabolic mediators' expression at adult age.

Telomere Length Differently Associated to Obesity and Hyperandrogenism in Women With Polycystic Ovary Syndrome.

Background: Polycystic Ovary Syndrome (PCOS) often present metabolic disorders and hyperandrogenism (HA), facts that may influence the telomere length (TL). Aims: To compare the absolute TL (aTL) between women with PCOS and control women, and their association with the presence of obesity and HA parameters. Materials and methods: The PCOS group included 170 unrelated women outpatients and the control group, 64 unrelated donor women. Anthropometric, biochemical-clinical parameters and androgen profile were determined. The PCOS patients were divided accordingly to the presence of obesity and androgenic condition. The aTL was determined from peripheral blood leukocytes by Real Time quantitative PCR. Results: Women with PCOS exhibited a significantly longer aTL than controls after age adjustment (p=0.001). A stepwise multivariate linear regression in PCOS women, showed that WC (waist circumference) contributed negatively (b=-0.17) while testosterone levels contributed positively (b=7.24) to aTL. The non-Obese PCOS (noOB-PCOS) presented the longest aTL when compared to controls (p=0.001). Meanwhile, the aTL was significantly higher in the hyperandrogenic PCOS phenotype (HA-PCOS) than in the controls (p=0.001) and non hyperandrogenic PCOS phenotype (NHA-PCOS) (p=0.04). Interestingly, when considering obesity and HA parameters in PCOS, HA exerts the major effect over the aTL as non-obese HA exhibited the lengthiest aTL (23.9 ± 13.13 Kbp). Conversely, the obese NHA patients showed the shortest aTL (16.5 ± 10.59 Kbp). Conclusions: Whilst a shorter aTL could be related to the presence of obesity, a longer aTL would be associated with HA phenotype. These findings suggest a balance between the effect produced by the different metabolic and hormonal components, in PCOS women.

Lipid Metabolism and Relevant Disorders to Female Reproductive Health.

BACKGROUND: Lipids are essential components of cells that participate in metabolic and endocrine regulation and reproductive functions. The main organs where lipid regulation takes place are the liver and adipose tissue. Besides, when each tissue- specific action cannot be exerted, it could lead to several endocrine-metabolic disorders closely related to PCOS, such as non-alcoholic fatty liver disease (NAFLD) and obesity. OBJECTIVE: This work aims to discuss the impact of lipid alterations on metabolic and reproductive health. Therefore, this review focus on the importance of carrying out an integrated study of the molecular pathways affected in PCOS for developing target therapies. RESULTS: Lipids play a major role in PCOS pathogenesis. In this regard, failures in lipid regulation, synthesis, and/or homeostasis contribute to metabolic and reproductive abnormalities, such as those seen in PCOS. Several lipid pathways and regulators are altered in this pathology, leading to dysfunctions that worsen reproductive functions. Therefore, there are several treatments to manage dyslipidemias. Non-pharmacological therapies are considered a first-line treatment being the pharmacological treatments a second-line option. CONCLUSION: The best treatment to improve the lipid profile is lifestyle intervention, a combination of hypocaloric diet and exercise. Regarding pharmacological therapies, a combination of fibrate and statins would be the most recommended drugs. Still, in PCOS women, treatment with metformin or TZDs not only modulates the lipid metabolism, but also improves ovulation. In addition, metformin with lifestyle interventions has positive effects on the metabolic and reproductive features of PCOS patients.

Prenatal testosterone exposure induces insulin resistance, uterine oxidative stress and pro-inflammatory status in rats.

Prenatal androgen excess is considered one of the main causes of the development of polycystic ovary syndrome. In this study, we investigated the effect of prenatal hyperandrogenization (PH) on the physiology of the adult uterine tissue using a murine model of fetal programming caused by androgen excess in adult female rats. Pregnant rats were hyperandrogenized with testosterone and female offspring were studied when adult. Our results showed that PH leads to hyperglycemia and hyperinsulinemia. Consequently, PH developed insulin resistance and a systemic inflammatory state reflected by increased C-reactive protein. In the uterine tissue, levels of PPAR gamma-an important metabolic sensor in the endometrium-were found to be impaired. Moreover, PH induced a pro-inflammatory and an unbalanced oxidative state in the uterus reflected by increased COX-2, lipid peroxidation, and NF-κB. In summary, our results revealed that PH leads to a compromised metabolic state likely consequence of fetal reprogramming.

Prenatal androgen exposure affects ovarian lipid metabolism and steroid biosynthesis in rats.

Prenatal androgen exposure affects reproductive functions and has been proposed as an underlying cause of polycystic ovary syndrome (PCOS). In this study, we aimed to investigate the impact of prenatal androgen exposure on ovarian lipid metabolism and to deepen our understanding of steroidogenesis regulation during adulthood. Pregnant rats were hyperandrogenized with testosterone and female offspring were studied when adult. This treatment leads to two different phenotypes: irregular ovulatory and anovulatory animals. Our results showed that prenatally hyperandrogenized (PH) animals displayed altered lipid and hormonal profile together with alterations in steroidogenesis and ovarian lipid metabolism. Moreover, PH animals showed alterations in the PPARg system, impaired mRNA levels of cholesterol receptors (Ldlr and Srb1) and decreased expression of the rate-limiting enzyme of de novo cholesterol production (Hmgcr). Anovulatory PH animals presented an increase of ovarian cholesteryl esters levels and lipid peroxidation index. Together with alterations in cholesterol metabolism, we found an impairment of the steroidogenic pathway in PH animals in a phenotype-specific manner. Regarding fatty acid metabolism, our results showed, in PH animals, an altered expression of Srebp1 and Atgl, which are involved in fatty acid metabolism and triglycerides hydrolysis, respectively. In conclusion, fatty acid and cholesterol metabolism, which are key players in steroidogenesis acting as a source of energy and substrate for steroid production, were affected in animals exposed to androgens during gestation. These results suggest that prenatal androgen exposure leads to long-term effects that affect ovary lipid metabolism and ovarian steroid formation from the very first steps.

Prenatally androgenized female rats develop uterine hyperplasia when adult.

Prenatal hyperandrogenization (PH) is hypothesized as one of the main factors contributing to the development of polycystic ovary syndrome (PCOS). In this study, we aimed to investigate the impact of prenatal exposure to androgen excess on the uterus when animals reach their adulthood. We found that PH altered the morphology of the uteri that show a hyperplastic morphology with increased total uterine thickness as well as luminal epithelium thickness, with both enhanced and altered distribution of glands as compared with controls. Morphological alterations were associated with an unbalanced homeostasis as assessed by the expression of regulators of cell cycle progression and cell death dynamics. PH also causes disturbances in the cell cycle of the uterine tissue and dysregulates cell death and survival pathways leading to the development of uterine hyperplasia. These findings suggest that PH may have a deleterious effect on the uterus.

Fetal programming by androgen excess in rats affects ovarian fuel sensors and steroidogenesis.

Fetal programming by androgen excess is hypothesized as one of the main factors contributing to the development of polycystic ovary syndrome (PCOS). PCOS is more than a reproductive disorder, as women with PCOS also show metabolic and other endocrine alterations. Since both ovarian and reproductive functions depend on energy balance, the alterations in metabolism may be related to reproductive alterations. The present study aimed to evaluate the effect of androgen excess during prenatal life on ovarian fuel sensors and its consequences on steroidogenesis. To this end, pregnant rats were hyperandrogenized with testosterone and the following parameters were evaluated in their female offspring: follicular development, PPARG levels, adipokines (including leptin, adiponectin, and chemerin as ovarian fuel sensors), serum gonadotropins (LH and FSH), the mRNA of their ovarian receptors, and the expression of steroidogenic mediators. At 60 days of age, the prenatally hyperandrogenized (PH) female offspring displayed both an irregular ovulatory phenotype and an anovulatory phenotype with altered follicular development and the presence of cysts. Both PH groups showed altered levels of both proteins and mRNA of PPARG and a different expression pattern of the adipokines studied. Although serum gonadotropins were not impaired, there were alterations in the mRNA levels of their ovarian receptors. The steroidogenic mediators Star, Cyp11a1, Cyp17a1, and Cyp19a1 were altered differently in each of the PH groups. We concluded that androgen excess during prenatal life leads to developmental programming effects that affect ovarian fuel sensors and steroidogenesis in a phenotype-specific way.

Effects of in utero androgen excess and metformin treatment on hepatic functions.

This study aimed to evaluate the role of prenatal hyperandrogenization in liver functions and the extent of metformin as treatment. Pregnant rats were hyperandrogenized with subcutaneous testosterone (1mg/rat) between 16 and 19 of pregnancy. Prenatally hyperandrogenized (PH) female offspring displayed, at the adult life, two phenotypes; a PH irregular ovulatory phenotype (PHiov) and a PH anovulatory (PHanov) phenotype. From day 70 to the moment of sacrifice (90 days of age), 50% of the animals of each group received a daily oral dose of 50 mg/kg of metformin. We found that both PH phenotypes displayed a hepatic disruptions of insulin and glucose pathway and that metformin treatment reversed some of these alterations in a specific-phenotype manner. Our findings show, for the first time, that androgen excess in utero promotes hepatic dysfunctions and that metformin treatment is able to specifically reverse those hepatic alterations and sheds light on the possible mechanisms of metformin action.

Metformin improves ovarian insulin signaling alterations caused by fetal programming.

Insulin resistance is the decreased ability of insulin to mediate metabolic actions. In the ovary, insulin controls ovulation and oocyte quality. Alterations in ovarian insulin signaling pathway could compromise ovarian physiology. Here, we aimed to investigate the effects of fetal programming on ovarian insulin signaling and evaluate the effect of metformin treatment. Pregnant rats were hyperandrogenized with testosterone and female offspring born to those dams were employed; at adulthood, prenatally hyperandrogenized (PH) offspring presented two phenotypes: irregular ovulatory (PHiov) and anovulatory (PHanov). Half of each group was orally treated with metformin. Metformin treatment improved the estrous cyclicity in both PH groups. Both PH groups showed low mRNA levels of IR, IRS1 and Glut4. IRS2 was decreased only in PHanov. Metformin upregulated the mRNA levels of some of the mediators studied. Protein expression of IR, IRS1/2 and GLUT4 was decreased in both PH groups. In PHiov, metformin restored the expression of all the mediators, whereas, in PHanov, metformin restored only that of IR and IRS1/2. IRS1 phosphorylation was measured in tyrosine residues, which activates the pathway, and in serine residues, which impairs insulin action. PHiov presented high IRS1 phosphorylation on tyrosine and serine residues, whereas PHanov showed high serine phosphorylation and low tyrosine phosphorylation. Metformin treatment lowered serine phosphorylation only in PHanov rats. Our results suggest that PHanov rats have a defective insulin action, partially restored with metformin. PHiov rats had less severe alterations, and metformin treatment was more effective in this phenotype.

Developmental programming of the female neuroendocrine system by steroids.

Developmental programming refers to processes that occur during early life that may have long-term consequences, modulating adult health and disease. Complex diseases, such as diabetes, cancer and cardiovascular disease, have a high prevalence in different populations, are multifactorial, and may have a strong environmental component. The environment interacts with organisms, affecting their behaviour, morphology and physiology. This interaction may induce permanent or long-term changes, and organisms may be more susceptible to environmental factors during certain developmental stages, such as the prenatal and early postnatal periods. Several factors have been identified as responsible for inducing the reprogramming of various reproductive and nonreproductive tissues. Among them, both natural and synthetic steroids, such as endocrine disruptors, are known to have either detrimental or positive effects on organisms depending on the dose of exposure, stage of development and biological sexual background. The present review focuses on the action of steroids and endocrine disruptors as agents involved in developmental programming and on their modulation and effects on female neuroendocrine functions.

Prenatal hyperandrogenism induces alterations that affect liver lipid metabolism.

Prenatal hyperandrogenism is hypothesized as one of the main factors contributing to the development of polycystic ovary syndrome (PCOS). PCOS patients have high risk of developing fatty liver and steatosis. This study aimed to evaluate the role of prenatal hyperandrogenism in liver lipid metabolism and fatty liver development. Pregnant rats were hyperandrogenized with testosterone. At pubertal age, the prenatally hyperandrogenized (PH) female offspring displayed both ovulatory (PHov) and anovulatory (PHanov) phenotypes that mimic human PCOS features. We evaluated hepatic transferases, liver lipid content, the balance between lipogenesis and fatty acid oxidation pathway, oxidant/antioxidant balance and proinflammatory status. We also evaluated the general metabolic status through growth rate curve, basal glucose and insulin levels, glucose tolerance test, HOMA-IR index and serum lipid profile. Although neither PH group showed signs of liver lipid content, the lipogenesis and fatty oxidation pathways were altered. The PH groups also showed impaired oxidant/antioxidant balance, a decrease in the proinflammatory pathway (measured by prostaglandin E2 and cyclooxygenase-2 levels), decreased glucose tolerance, imbalance of circulating lipids and increased risk of metabolic syndrome. We conclude that prenatal hyperandrogenism generates both PHov and PHanov phenotypes with signs of liver alterations, imbalance in lipid metabolism and increased risk of developing metabolic syndrome. The anovulatory phenotype showed more alterations in liver lipogenesis and a more impaired balance of insulin and glucose metabolism, being more susceptible to the development of steatosis.

Nonalcoholic fatty liver disease in children and adolescents - Relationship with Polycystic Ovary Syndrome.

Nonalcoholic fatty liver disease (NAFLD) is defined as the accumulation of triglycerides (TGs) within hepatocytes exceeding 5 % of liver weight. NAFLD is a spectrum of pathological processes from nonalcoholic fatty liver or simple steatosis to nonalcoholic steatohepatitis (NASH), fibrosis, cirrhosis, and hepatocellular carcinoma. As NAFLD induces metabolic syndrome (MS), then, NAFLD is associated with insulin resistance (IR), type 2 diabetes mellitus (T2DM), hypertension and even Polycystic Ovary Syndrome (PCOS). Because it is well established that patients carrying gene mutations also develop NAFLD in the absence of IR, the genetic predisposition to NAFLD is also discussed. Little is known about the diagnosis and treatment of NAFLD in children and adolescents and the lack of non-invasive diagnostic tools in these populations is a major problem faced by physicians. The present review aims to discuss recent findings of NAFLD in children and adolescents and, considering the features in common with PCOS, we also discuss their relationship.

The biology of the peroxisome proliferator-activated receptor system in the female reproductive tract.

Fuel sensors such as glucose, insulin or leptin, are known to be directly involved in the regulation of fertility at each level of the hypothalamic-pituitary-gonadal axis. The discovery of the peroxisome proliferator-activated receptor (PPAR) family of transcription factors has revealed the link between lipid/glucose availability and long-term metabolic adaptation. By binding to specific regions of DNA in heterodimers with the retinoid X receptors (RXRs), the members of the PPAR family (α, ß/δ, γ) are able to regulate the gene expressions of several key regulators of energy homeostasis including several glucose regulators (glucose transporters, insulin receptor, substrate insulin receptor, etc), and also metabolic and endocrine pathways like lipogenesis, steroidogenesis, ovulation, oocyte maturation, maintenance of the corpus luteum, nitric oxide system, several proteases and plasminogen activator among others. All the three PPAR isoforms are expressed in different tissues of the female reproductive tract and regulate gametogenesis, ovulation, corpus luteum regression and the implantation process among others. The present review discusses the mechanisms involved in PPAR activation focusing on endogenous and synthetic ligands of PPAR not only in physiological but also in pathological conditions (such as polycystic ovary syndrome, pathologies of implantation process, chronic anovulation, etc).