ABSTRACT The microorganisms of the vaginal tract are critical for vaginal and reproductive health. However, the regulation of these microorganisms is not well understood. Therefore, we investigated whether different factors regulate the vaginal microbiota of healthy college-aged women (n = 26) with high temporal resolution by collecting daily self-administered vaginal swabs and using 16S rRNA sequencing for bacterial identification. As expected, vaginal microbiota clustered into five predefined community state types. Vaginal microbial diversity, stability, and Lactobacillus abundances were associated with the menstrual cycle and hormonal contraceptive use. Vaginal microbial diversity, as measured using the Shannon index, increased during menses (P < 0.001), while Lactobacillus abundances decreased (P = 0.01). The covariance of these microbial measures with previously established estradiol levels suggests that estrogens can regulate vaginal microbiota. Moreover, the use of hormonal contraceptives may alter the temporal dynamics of the vaginal microbiota and decrease Lactobacillus abundances, depending on hormonal content and release method. Interestingly, intrasample diversity was greater in participants on a vegetarian diet (P = 0.004) and among participants who exercised more (P = 0.04). These findings indicate that ovarian hormones, diet, and exercise can regulate vaginal microbial composition and stability and may impact vaginal and reproductive health. IMPORTANCE The vaginal microbiome is a critical component of women’s sexual and reproductive health, with variations in microbial composition, particularly the loss of Lactobacillus species, being implicated in gynecologic and obstetric diseases. Given that the vaginal microbiome is so crucial, why do vaginal microbial profiles vary strikingly from person to person and even change over time within the same person? In the present study, which tracked the daily vaginal microbiomes of young healthy women through different lifestyles, we found that use of a locally released progestin contraceptive, a vegetarian diet, and intense exercise appear to lead to vaginal microbiome alterations and loss of Lactobacillus species. The impact of these vaginal microbiome changes on immediate and long-term health remain to be investigated.

estrogens protect against diet-induced obesity in women and female rodents. for example, a lack of estrogens in postmenopausal women is associated with an increased risk of weight gain, cardiovascular diseases, low-grade inflammation, and cancer. Estrogens act with leptin to regulate energy homeostasis in females. Leptin-deficient mice (ob/ob) exhibit morbid obesity and insulin resistance. the gut microbiome is also critical in regulating metabolism. the present study investigates whether estrogens and leptin modulate gut microbiota in ovariectomized ob/ob (obese) or heterozygote (lean) mice fed high-fat diet (HFD) that received either 17β-Estradiol (E2) or vehicle implants. E2 attenuated weight gain in both genotypes. Moreover, both obesity (ob/ob mice) and E2 were associated with reduced gut microbial diversity. ob/ob mice exhibited lower species richness than control mice, while E2-treated mice had reduced evenness compared with vehicle mice. Regarding taxa, E2 was associated with an increased abundance of the S24-7 family, while leptin was associated with increases in coriobacteriaceae, Clostridium and Lactobacillus. Some taxa were affected by both E2 and leptin, suggesting these hormones alter gut microbiota of HFD-fed female mice. Understanding the role of E2 and leptin in regulating gut microbiota will provide important insights into hormone-dependent metabolic disorders in women. Estrogens profoundly influence energy homeostasis 1-3 , as well as reproductive physiology and behavior 4-6. Estrogens reduce food intake, attenuate body weight gain and adiposity, and increase physical activity in humans and rodents 1,2. Postmenopausal women have lower levels of circulating estrogens and an increased tendency to gain fat weight, which increases their risk for obesity, cardiovascular disease, stroke, and type 2 diabetes 7-9. Similarly, in mice on a high-fat diet (HFD), ovariectomy increases energy intake and obesity, while estradiol (E2) treatment prevents weight gain 2,10-13 , indicating that estrogens protect against HFD-induced obesity. Leptin is a peptide hormone secreted primarily by adipocytes, which acts primarily in the brain to stimulate metabolism, promote satiety, and regulate fat storage 14-16. A mutation in the ob gene that encodes leptin results in mice lacking the hormone (ob/ob) 17. While phenotypically normal at birth, ob/ob mice quickly develop obesity and diabetes 18. Additionally, ob/ob mice exhibit increased food intake and decreased physical activity, energy metabolism, and body temperature compared to lean controls, making ob/ob mice an excellent genetic model of obesity 19-21. Administering leptin to adult ob/ob mice reverses these effects by decreasing food intake, increasing energy output and decreasing circulating levels of glucose and insulin 19,22. Leptin and estrogen signaling pathways interact to influence reproduction and energy metabolism. High levels of E2 are associated with increased leptin sensitivity in both male and female rodents 23. In contrast, decreased estrogens in ovar...

Leptin and oestradiol have overlapping functions in energy homeostasis and fertility, and receptors for these hormones are localised in the same hypothalamic regions. Although, historically, it was assumed that mammalian adult neurogenesis was confined to the olfactory bulbs and the hippocampus, recent research has found new neurones in the male rodent hypothalamus. Furthermore, some of these new neurones are leptin-sensitive and affected by diet. In the present study, we tested the hypothesis that diet and hormonal status modulate hypothalamic neurogenesis in the adult female mouse. Adult mice were ovariectomised and implanted with capsules containing oestradiol (E2) or oil. Within each group, mice were fed a high-fat diet (HFD) or maintained on standard chow (STND). All animals were administered i.c.v. 5-bromo-2′-deoxyuridine (BrdU) for 9 days and sacrificed 34 days later after an injection of leptin to induce phosphorylation of signal transducer of activation and transcription 3 (pSTAT3). Brain tissue was immunohistochemically labelled for BrdU (newly born cells), Hu (neuronal marker) and pSTAT3 (leptin sensitive). Although mice on a HFD became obese, oestradiol protected against obesity. There was a strong interaction between diet and hormone on new cells (BrdU+) in the arcuate, ventromedial hypothalamus and dorsomedial hypothalamus. HFD increased the number of new cells, whereas E2 inhibited this effect. Conversely, E2 increased the number of new cells in mice on a STND diet in all hypothalamic regions studied. Although the total number of new leptin-sensitive neurones (BrdU-Hu-pSTAT3) found in the hypothalamus was low, HFD increased these new cells in the arcuate, whereas E2 attenuated this induction. These results suggest that adult neurogenesis in the hypothalamic neurogenic niche is modulated by diet and hormonal status and is related to energy homeostasis in female mice.

Progestins bind to the progestin receptor (PR) isoforms, PR-A and PR-B, in brain to influence development, female reproduction, anxiety, and stress. Hormone-activated PRs associate with multiple proteins to form functional complexes. In the present study, proteins from female mouse hypothalamus that associate with PR were isolated using affinity pull-down assays with glutathione S-transferase–tagged mouse PR-A and PR-B. Using complementary proteomics approaches, reverse phase protein array (RPPA) and mass spectrometry, we identified hypothalamic proteins that interact with PR in a ligand-dependent and isoform-specific manner and were confirmed by Western blot. Synaptic proteins, including synapsin-I and synapsin-II, interacted with agonist-bound PR isoforms, suggesting that both isoforms function in synaptic plasticity. In further support, synaptogyrin-III and synapsin-III associated with PR-A and PR-B, respectively. PR also interacted with kinases, including c-Src, mTOR, and MAPK1, confirming phosphorylation as an integral process in rapid effects of PR in the brain. Consistent with a role in transcriptional regulation, PR associated with transcription factors and coactivators in a ligand-specific and isoform-dependent manner. Interestingly, both PR isoforms associated with a key regulator of energy homeostasis, FoxO1, suggesting a novel role for PR in energy metabolism. Because many identified proteins in this PR interactome are synaptic proteins, we tested the hypothesis that progestins function in synaptic plasticity. Indeed, progesterone enhanced synaptic density, by increasing synapsin-I–positive synapses, in rat primary cortical neuronal cultures. This novel combination of RPPA and mass spectrometry allowed identification of PR action in synaptic remodeling and energy homeostasis and reveals unique roles for progestins in brain function and disease.

Estradiol-induced structural dimorphisms exist in the songbird brain. However, how they arise is not clear since there is a scarce distribution of ERα and lack of ERβ in song control nuclei. This suggests that other receptors are involved. The G-protein coupled membrane-bound estrogen receptor, GPR30, is a candidate but has never been investigated in songbirds. In this study, we characterized its gene and protein in the zebra finch brain. Analysis of the putative GPR30 protein sequence revealed a strong similarity to avian and mammalian homologues. Quantitative PCR indicated that the gene was elevated in the telencephalon of both sexes from posthatching day (P) 15 to P45, with a male-biased sex difference at P21 and P30. In comparison, expression at younger posthatching ages and in adults was significantly less. At P21, GRP30 protein was widespread, nonuniform, and overlapped with song control nuclei. Of particular interest, the number of immunoreactive cells was greatest in HVC and RA, but less in LMAN and Area X. Labeling in HVC was also dimorphic; with more cells present in males than in females. In parallel with the gene, by adulthood, protein expression was reduced across most brain regions. Taken together these data suggest that GPR30 may contribute to differences in song system development by mediating dimorphic responses to estrogens. In addition, the extensive protein distribution indicates that it may also have a role in general brain development in both sexes.