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1.
Cell Mol Life Sci ; 81(1): 329, 2024 Aug 01.
Article in English | MEDLINE | ID: mdl-39090270

ABSTRACT

Decidualisation of the endometrium is a key event in early pregnancy, which enables embryo implantation. Importantly, the molecular processes impairing decidualisation in obese mothers are yet to be characterised. We hypothesise that impaired decidualisation in obese mice is mediated by the upregulation of leptin modulators, the suppressor of cytokine signalling 3 (SOCS3) and the protein tyrosine phosphatase non-receptor type 2 (PTPN2), together with the disruption of progesterone (P4)-signal transducer and activator of transcription (STAT3) signalling. After feeding mice with chow diet (CD) or high-fat diet (HFD) for 16 weeks, we confirmed the downregulation of P4 and oestradiol (E2) steroid receptors in decidua from embryonic day (E) 6.5 and decreased proliferation of stromal cells from HFD. In vitro decidualised mouse endometrial stromal cells (MESCs) and E6.5 deciduas from the HFD showed decreased expression of decidualisation markers, followed by the upregulation of SOCS3 and PTPN2 and decreased phosphorylation of STAT3. In vivo and in vitro leptin treatment of mice and MESCs mimicked the results observed in the obese model. The downregulation of Socs3 and Ptpn2 after siRNA transfection of MESCs from HFD mice restored the expression level of decidualisation markers. Finally, DIO mice placentas from E18.5 showed decreased labyrinth development and vascularisation and fetal growth restricted embryos. The present study revealed major defects in decidualisation in obese mice, characterised by altered uterine response to E2 and P4 steroid signalling. Importantly, altered hormonal response was associated with increased expression of leptin signalling modulators SOCS3 and PTPN2. Elevated levels of SOCS3 and PTPN2 were shown to molecularly affect decidualisation in obese mice, potentially disrupting the STAT3-PR regulatory molecular hub.


Subject(s)
Decidua , Fetal Growth Retardation , Leptin , Mice, Obese , Placenta , Protein Tyrosine Phosphatase, Non-Receptor Type 2 , STAT3 Transcription Factor , Signal Transduction , Suppressor of Cytokine Signaling 3 Protein , Animals , Female , Suppressor of Cytokine Signaling 3 Protein/metabolism , Suppressor of Cytokine Signaling 3 Protein/genetics , Pregnancy , Leptin/metabolism , Decidua/metabolism , Decidua/pathology , Protein Tyrosine Phosphatase, Non-Receptor Type 2/metabolism , Protein Tyrosine Phosphatase, Non-Receptor Type 2/genetics , Mice , Placenta/metabolism , STAT3 Transcription Factor/metabolism , Fetal Growth Retardation/metabolism , Fetal Growth Retardation/pathology , Obesity/metabolism , Obesity/pathology , Progesterone/metabolism , Mice, Inbred C57BL , Diet, High-Fat/adverse effects , Stromal Cells/metabolism
2.
BMC Endocr Disord ; 24(1): 135, 2024 Aug 01.
Article in English | MEDLINE | ID: mdl-39090659

ABSTRACT

BACKGROUND: Prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) is higher in men than in women. Hormonal and genetic causes may account for the sex differences in MASLD. Current human in vitro liver models do not sufficiently take the influence of biological sex and sex hormones into consideration. METHODS: Primary human hepatocytes (PHHs) were isolated from liver specimen of female and male donors and cultured with sex hormones (17ß-estradiol, testosterone and progesterone) for up to 72 h. mRNA expression levels of 8 hepatic lipid metabolism genes were analyzed by RT-qPCR. Sex hormones and their metabolites were determined in cell culture supernatants by LC-MS analyses. RESULTS: A sex-specific expression was observed for LDLR (low density lipoprotein receptor) with higher mRNA levels in male than female PHHs. All three sex hormones were metabolized by PHHs and the effects of hormones on gene expression levels varied depending on hepatocyte sex. Only in female PHHs, 17ß-estradiol treatment affected expression levels of PPARA (peroxisome proliferator-activated receptor alpha), LIPC (hepatic lipase) and APOL2 (apolipoprotein L2). Further changes in mRNA levels of female PHHs were observed for ABCA1 (ATP-binding cassette, sub-family A, member 1) after testosterone and for ABCA1, APOA5 (apolipoprotein A-V) and PPARA after progesterone treatment. Only the male PHHs showed changing mRNA levels for LDLR after 17ß-estradiol and for APOA5 after testosterone treatment. CONCLUSIONS: Male and female PHHs showed differences in their expression levels of hepatic lipid metabolism genes and their responsiveness towards sex hormones. Thus, cellular sex should be considered, especially when investigating the pathophysiological mechanisms of MASLD.


Subject(s)
Gonadal Steroid Hormones , Hepatocytes , Lipid Metabolism , Humans , Male , Female , Hepatocytes/metabolism , Hepatocytes/drug effects , Lipid Metabolism/genetics , Lipid Metabolism/drug effects , Gonadal Steroid Hormones/pharmacology , Gonadal Steroid Hormones/metabolism , Cells, Cultured , Middle Aged , Testosterone/pharmacology , Testosterone/metabolism , Estradiol/pharmacology , Adult , Progesterone/pharmacology , Progesterone/metabolism , Sex Factors
3.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 55(3): 596-604, 2024 May 20.
Article in Chinese | MEDLINE | ID: mdl-38948288

ABSTRACT

Objective: This study aims to analyze the relationship between reproductive tract microecological changes, metabolic differences, and pregnancy outcomes at different time points in the frozen-thawed embryo transfer cycle while patients are undergoing hormone replacement therapy, which will be a breakthrough point for improving outcomes. Methods: A total of 20 women undergoing frozen-thawed single blastocyst transfer for the first time at the Reproductive Medicine Center of Fujian Maternal and Child Health Hospital between July 2022 and January 2023 were recruited for this study. Their vaginal and cervical secretions were collected for 16S rRNA sequencing and non-targeted metabolomics analysis on days 2-5 of menstruation, day 7 after estrogen replacement therapy started, the day when progesterone was added, and the day of transplantation. The subjects were divided into different groups according to their clinical pregnancy status and the sequencing results were analyzed using bioinformatics methods. Results: 1) The alpha-diversity index of the vaginal and cervical microbiota was higher on days 2-5 of menstruation (P<0.01), but did not differ significantly on day 7 after oral estrogen replacement therapy started, the day of progesterone administration, and the day of transplantation (P≥0.1). 2) Both the pregnant group and the non-pregnant group showed a variety of microorganisms and metabolites with significant differences in the lower reproductive tract at different time points. 3) Microbial analysis at different time points showed that there were significant differences in vaginal flora, including Peptoniphilus, Enterocloster, Finegoldia, Klebsiella, Anaerobutyricum, Agathobaculum, Sporanaerobacter, Bilophila, Prevotella, and Anaerococcus in the pregnant group (P<0.05). 4) Metabolite analysis at different time points showed that there were significant differences in 3-hydroxybenzoic acid, linatine, (R)-amphetamine, hydroxychloroquine, and L-altarate in the vaginal secretions of the pregnant group (P<0.05), and that there were significant differences in isocitric acid, quassin, citrinin, and 12(R)-HETE in the cervical secretions (P<0.05). 5) Metabolite analysis at different time points showed that, in the non-pregnant group, there were significant differences in linatine, decanoyl-L-carnitine, aspartame, sphingosine, and hydroxychloroquine in the vaginal secretions (P<0.05), and the isocitric acid, quassin, ctrinin, and 12(R)-HETE in the cervical secretions (P<0.05). 6) Combined microbiome and metabolomics analysis showed that certain metabolites were significantly associated with microbial communities, especially Klebsiella. Conclusions: Significant differences in the microbiota genera and metabolites at different time points were found during the frozen-embryo transfer cycle of hormone replacement therapy, which may be used as potential biomarkers to predict pregnancy outcomes of embryo transfer.


Subject(s)
Embryo Transfer , Microbiota , Pregnancy Outcome , Progesterone , Vagina , Humans , Female , Pregnancy , Embryo Transfer/methods , Vagina/microbiology , Progesterone/metabolism , Adult , Cryopreservation , RNA, Ribosomal, 16S/genetics , Cervix Uteri/metabolism
4.
Proc Biol Sci ; 291(2027): 20232712, 2024 Aug.
Article in English | MEDLINE | ID: mdl-39043247

ABSTRACT

Although men's attraction to women's body odour has been suggested to vary over the ovulatory cycle, peaking around the fertile window, we still lack methodologically robust evidence corroborating this effect. Further, the chemical underpinnings of male preference for the odour of ovulating women remain unknown. Here, we combined perceptual and chemical analyses to investigate the axillary odour of naturally cycling women over 10 days, covering the gradual change in fertility across the ovulatory cycle with a focus on fertile days. The fertile state was confirmed by urinary ovulation tests as well as salivary oestradiol and progesterone levels. Men rated the scent of unfamiliar women, resembling a first encounter. We used multivariate analyses to relate variation in both odour ratings and chemical composition to female conception probability, temporal distance to ovulation and ovarian hormone levels. Our results provide no evidence that males prefer the odour of fertile women. Furthermore, the volatile analysis indicated no link between axillary odour composition and current fertility status. Together, our results showed no convincing support for a chemical fertility cue in women's axillary odour, questioning the presence of olfactory fertility information that is recognizable during first encounters in modern humans.


Subject(s)
Odorants , Ovulation , Humans , Female , Odorants/analysis , Male , Adult , Ovulation/physiology , Young Adult , Fertility , Progesterone/metabolism , Olfactory Perception , Estradiol/metabolism
5.
Nat Commun ; 15(1): 5619, 2024 Jul 04.
Article in English | MEDLINE | ID: mdl-38965213

ABSTRACT

The sigma-1 receptor (σ1R) is a non-opioid membrane receptor, which responds to a diverse array of synthetic ligands to exert various pharmacological effects. Meanwhile, candidates for endogenous ligands of σ1R have also been identified. However, how endogenous ligands bind to σ1R remains unknown. Here, we present crystal structures of σ1R from Xenopus laevis (xlσ1R) bound to two endogenous neurosteroid ligands, progesterone (a putative antagonist) and dehydroepiandrosterone sulfate (DHEAS) (a putative agonist), at 2.15-3.09 Å resolutions. Both neurosteroids bind to a similar location in xlσ1R mainly through hydrophobic interactions, but surprisingly, with opposite binding orientations. DHEAS also forms hydrogen bonds with xlσ1R, whereas progesterone interacts indirectly with the receptor through water molecules near the binding site. Binding analyses are consistent with the xlσ1R-neurosteroid complex structures. Furthermore, molecular dynamics simulations and structural data reveal a potential water entry pathway. Our results provide insight into binding of two endogenous neurosteroid ligands to σ1R.


Subject(s)
Dehydroepiandrosterone Sulfate , Molecular Dynamics Simulation , Progesterone , Receptors, sigma , Sigma-1 Receptor , Xenopus laevis , Receptors, sigma/metabolism , Receptors, sigma/chemistry , Animals , Ligands , Binding Sites , Progesterone/metabolism , Progesterone/chemistry , Dehydroepiandrosterone Sulfate/metabolism , Dehydroepiandrosterone Sulfate/chemistry , Protein Binding , Crystallography, X-Ray , Neurosteroids/metabolism , Neurosteroids/chemistry , Hydrogen Bonding , Hydrophobic and Hydrophilic Interactions
6.
FASEB J ; 38(14): e23839, 2024 Jul 31.
Article in English | MEDLINE | ID: mdl-39037418

ABSTRACT

During early pregnancy in mice, the establishment of uterine receptivity and endometrial decidualization require the extensive proliferation and differentiation of endometrial epithelial cells or stromal cells. Pin1 has been suggested to act as a molecular 'timer' of the cell cycle and is involved in the regulation of cellular proliferation and differentiation by binding many cell-cycle regulatory proteins. However, its physiological role during early pregnancy is still not fully understood. Here, we employed immunohistochemistry to determine the spatiotemporal pattern of Pin1 expression during early pregnancy. We found that Pin1 was mainly localized in subluminal stromal cells on day 4, in the decidual zone on days 5 to 8 of pregnancy and in artificial decidualization. Using a uterine stromal cell culture system, we found that progesterone, but not estrogen, induced the expression of Pin1 in a progesterone receptor-dependent manner. Inhibition of Pin1 in the uterus leads to impaired embryo implantation and decidualization in mice. Notably, a decrease in Pin1 activation affected the functional execution of several implantation- or decidualization-related factors. These findings provide new evidence for a previously unknown function of Pin1 in mediating embryo implantation and decidualization during successful pregnancy establishment and maintenance.


Subject(s)
Decidua , Embryo Implantation , NIMA-Interacting Peptidylprolyl Isomerase , Uterus , Animals , Female , NIMA-Interacting Peptidylprolyl Isomerase/metabolism , NIMA-Interacting Peptidylprolyl Isomerase/genetics , Embryo Implantation/physiology , Mice , Pregnancy , Decidua/metabolism , Decidua/cytology , Uterus/metabolism , Uterus/cytology , Progesterone/metabolism , Stromal Cells/metabolism , Receptors, Progesterone/metabolism , Cells, Cultured , Endometrium/metabolism , Endometrium/cytology
7.
J Pineal Res ; 76(2): e12936, 2024 Mar.
Article in English | MEDLINE | ID: mdl-39041348

ABSTRACT

Women typically sleep and wake earlier than men and have been shown to have earlier circadian timing relative to the light/dark cycle that synchronizes the clock. A potential mechanism for earlier timing in women is an altered response of the circadian system to evening light. We characterized individual-level dose-response curves for light-induced melatonin suppression using a within-subjects protocol. Fifty-six participants (29 women, 27 men; aged 18-30 years) were exposed to a range of light illuminances (10, 30, 50, 100, 200, 400, and 2000 lux) using melatonin suppression relative to a dim control (<1 lux) as a marker of light sensitivity. Women were free from hormonal contraception. To examine the potential influence of sex hormones, estradiol and progesterone was examined in women and testosterone was examined in a subset of men. Menstrual phase was monitored using self-reports and estradiol and progesterone levels. Women exhibited significantly greater melatonin suppression than men under the 400-lux and 2000-lux conditions, but not under lower light conditions (10-200 lux). Light sensitivity did not differ by menstrual phase, nor was it associated with levels of estradiol, progesterone, or testosterone, suggesting the sex differences in light sensitivity were not acutely driven by circulating levels of sex hormones. These results suggest that sex differences in circadian timing are not due to differences in the response to dim/moderate light exposures typically experienced in the evening. The finding of increased bright light sensitivity in women suggests that sex differences in circadian timing could plausibly instead be driven by a greater sensitivity to phase-advancing effects of bright morning light.


Subject(s)
Circadian Rhythm , Light , Melatonin , Humans , Female , Adult , Circadian Rhythm/physiology , Adolescent , Young Adult , Male , Melatonin/metabolism , Estradiol/blood , Progesterone/blood , Progesterone/metabolism , Testosterone/blood , Menstrual Cycle/physiology
8.
J Membr Biol ; 257(3-4): 215-230, 2024 Aug.
Article in English | MEDLINE | ID: mdl-38970681

ABSTRACT

Progesterone (P4) acts as a key conserved signalling molecule in vertebrate reproduction. P4 is especially important for mature sperm physiology and subsequent reproductive success. "CatSpermasome", a multi-unit molecular complex, has been suggested to be the main if not the only P4-responsive atypical Ca2+-ion channel present in mature sperm. Altogether, here we analyse the protein sequences of CatSper1-4 from more than 500 vertebrates ranging from early fishes to humans. CatSper1 becomes longer in mammals due to sequence gain mainly at the N-terminus. Overall the conservation of full-length CatSper1-4 as well as the individual TM regions remain low. The lipid-water-interface residues (i.e. a 5 amino acid stretch sequence present on both sides of each TM region) also remain highly diverged. No specific patterns of amino acid distributions were observed. The total frequency of positively charged, negatively charged or their ratios do not follow in any specific pattern. Similarly, the frequency of total hydrophobic, total hydrophilic residues or even their ratios remain random and do not follow any specific pattern. We noted that the CatSper1-4 genes are missing in amphibians and the CatSper1 gene is missing in birds. The high variability of CatSper1-4 and gene-loss in certain clades indicate that the "CatSpermasome" is not the only P4-responsive ion channel. Data indicate that the molecular evolution of CatSper is mostly guided by diverse hydrophobic ligands rather than only P4. The comparative data also suggest possibilities of other Ca2+-channel/s in vertebrate sperm that can also respond to P4.


Subject(s)
Calcium Channels , Progesterone , Spermatozoa , Male , Animals , Spermatozoa/metabolism , Calcium Channels/metabolism , Calcium Channels/genetics , Calcium Channels/chemistry , Progesterone/metabolism , Humans , Vertebrates/genetics , Vertebrates/metabolism , Amino Acid Sequence , Conserved Sequence
9.
Reprod Fertil ; 5(3)2024 Jul 01.
Article in English | MEDLINE | ID: mdl-38990713

ABSTRACT

Abstract: Reactive oxygen species (ROS) are a by-product of the activity of cytochrome P450 steroidogenic enzymes. Antioxidant enzymes protect against ROS damage. To identify if any particular antioxidant enzyme is used to protect against ROS produced by granulosa cells as follicles enlarge and produce oestradiol, we measured in the bovine granulosa cells the expression of two steroidogenic enzymes (CYP11A1, CYP19A1), important for progesterone and oestradiol production. We also measured the expression of the members (FDXR, FDX1, POR) of their electron transport chains (ETC). We measured antioxidant enzymes (GPXs 1-8, CAT, SODs 1 and 2, PRDXs 1-6, GSR, TXN, TXNRDs 1-3). Since selenium is an active component of GPXs, the selenium-uptake receptors (LRPs 2 and 8) were measured. Only the selenium-dependent GPX1 showed the same increase in expression as the steroidogenic enzymes did with increasing follicle size. GPX4 and PRDX2/6 decreased with follicle size, whereas SOD1/2, CAT, GSR, and TXNRD3 were lowest at the intermediate sizes. The other antioxidant enzymes were unchanged or expressed at low levels. The expression of the selenium-uptake receptor LRP8 also increased significantly with follicle size. Correlation analysis revealed statistically significant and strongly positive correlations of the steroidogenic enzymes and their ETCs with both GPX1 and LRP8. These results demonstrate a relationship between the expression of genes involved in steroidogenesis and selenium-containing antioxidant defence mechanisms. They suggest that during the late stages of folliculogenesis, granulosa cells are dependent on sufficient expression of GPX1 and the selenium transporter LRP8 to counteract increasing ROS levels caused by the production of steroid hormones. Lay summary: In the ovary, eggs are housed in follicles which contain the cells that produce oestrogen in the days leading up to ovulation of the egg. Oestrogen is produced by the action of enzymes. However, some of these enzymes also produce by-products called reactive oxygen species (ROS). These are harmful to eggs. Fortunately, cells have protective antioxidant enzymes that can neutralise ROS. This study was interested in which particular antioxidant enzyme(s) might be involved in neutralising the ROS in follicle cells. It was found that only one antioxidant enzyme, GPX1, appeared to be co-regulated with the enzymes that produce oestrogen and progesterone in the follicular cells. GPX1 contains the essential mineral selenium. In summary, this study has identified which antioxidant appears to be involved in neutralising ROS in the days leading to ovulation. It highlights the importance of selenium in the diet.


Subject(s)
Glutathione Peroxidase GPX1 , Glutathione Peroxidase , Granulosa Cells , Female , Granulosa Cells/metabolism , Animals , Cattle , Glutathione Peroxidase/metabolism , Glutathione Peroxidase/genetics , Selenium/metabolism , Antioxidants/metabolism , Aromatase/metabolism , Aromatase/genetics , Cholesterol Side-Chain Cleavage Enzyme/metabolism , Cholesterol Side-Chain Cleavage Enzyme/genetics , Progesterone/metabolism , Reactive Oxygen Species/metabolism , Estradiol/metabolism , Ovarian Follicle/metabolism
10.
Cell Mol Life Sci ; 81(1): 324, 2024 Jul 30.
Article in English | MEDLINE | ID: mdl-39080028

ABSTRACT

Polycystic ovary syndrome (PCOS) is a complex common endocrine disorder affecting women of reproductive age. Ovulatory dysfunction is recognized as a primary infertile factor, however, even when ovulation is medically induced and restored, PCOS patients continue to experience reduced cumulative pregnancy rates and a higher spontaneous miscarriage rate. Hyperandrogenism, a hallmark feature of PCOS, affects ovarian folliculogenesis, endometrial receptivity, and the establishment and maintenance of pregnancy. Decidualization denotes the transformation that the stromal compart of the endometrium must undergo to accommodate pregnancy, driven by the rising progesterone levels and local cAMP production. However, studies on the impact of hyperandrogenism on decidualization are limited. In this study, we observed that primary endometrial stromal cells from women with PCOS exhibit abnormal responses to progesterone during in vitro decidualization. A high concentration of testosterone inhibits human endometrial stromal cells (HESCs) decidualization. RNA-Seq analysis demonstrated that pyruvate dehydrogenase kinase 4 (PDK4) expression was significantly lower in the endometrium of PCOS patients with hyperandrogenism compared to those without hyperandrogenism. We also characterized that the expression of PDK4 is elevated in the endometrium stroma at the mid-secretory phase. Artificial decidualization could enhance PDK4 expression, while downregulation of PDK4 leads to abnormal decidualization both in vivo and in vitro. Mechanistically, testosterone excess inhibits IGFBP1 and PRL expression, followed by phosphorylating of AMPK that stimulates PDK4 expression. Based on co-immunoprecipitation analysis, we observed an interaction between SIRT1 and PDK4, promoting glycolysis to facilitate decidualization. Restrain of AR activation resumes the AMPK/SIRT1/PDK4 pathway suppressed by testosterone excess, indicating that testosterone primarily acts on decidualization through AR stimulation. Androgen excess in the endometrium inhibits decidualization by disrupting the AMPK/SIRT1/PDK4 signaling pathway. These data demonstrate the critical roles of endometrial PDK4 in regulating decidualization and provide valuable information for understanding the underlying mechanism during decidualization.


Subject(s)
AMP-Activated Protein Kinases , Endometrium , Polycystic Ovary Syndrome , Sirtuin 1 , Stromal Cells , Humans , Female , Polycystic Ovary Syndrome/metabolism , Polycystic Ovary Syndrome/pathology , Stromal Cells/metabolism , Stromal Cells/pathology , Stromal Cells/drug effects , Sirtuin 1/metabolism , Sirtuin 1/genetics , Endometrium/metabolism , Endometrium/pathology , Endometrium/drug effects , AMP-Activated Protein Kinases/metabolism , Adult , Hyperandrogenism/metabolism , Hyperandrogenism/pathology , Decidua/metabolism , Decidua/pathology , Testosterone/metabolism , Testosterone/pharmacology , Androgens/pharmacology , Androgens/metabolism , Progesterone/metabolism , Progesterone/pharmacology , Pyruvate Dehydrogenase Acetyl-Transferring Kinase/metabolism , Pyruvate Dehydrogenase Acetyl-Transferring Kinase/genetics , Signal Transduction/drug effects
11.
Mol Cell Biol ; 44(8): 316-333, 2024.
Article in English | MEDLINE | ID: mdl-39014976

ABSTRACT

Pregnancy involving intricate tissue transformations governed by the progesterone hormone (P4). P4 signaling via P4 receptors (PRs) is vital for endometrial receptivity, decidualization, myometrial quiescence, and labor initiation. This study explored the role of TCF23 as a downstream target of PR during pregnancy. TCF23 was found to be expressed in female reproductive organs, predominantly in uterine stromal and smooth muscle cells. Tcf23 expression was high during midgestation and was specifically regulated by P4, but not estrogen. The Tcf23 knockout (KO) mouse was generated and analyzed. Female KO mice aged 4-6 months exhibited subfertility, reduced litter size, and defective parturition. Uterine histology revealed disrupted myometrial structure, altered collagen organization, and disarrayed smooth muscle sheets at the conceptus sites of KO mice. RNA-Seq analysis of KO myometrium revealed dysregulation of genes associated with cell adhesion and extracellular matrix organization. TCF23 potentially modulates TCF12 activity to mediate cell-cell adhesion and matrix modulation in smooth muscle cells. Overall, TCF23 deficiency leads to impaired myometrial remodeling, causing parturition delay and fetal demise. This study sheds light on the critical role of TCF23 as a dowstream mediator of PR in uterine remodeling, reflecting the importance of cell-cell communication and matrix dynamics in myometrial activation and parturition.


Subject(s)
Mice, Knockout , Myometrium , Parturition , Animals , Female , Parturition/metabolism , Parturition/genetics , Parturition/physiology , Mice , Pregnancy , Myometrium/metabolism , Progesterone/metabolism , Uterus/metabolism , Myocytes, Smooth Muscle/metabolism , Receptors, Progesterone/metabolism , Receptors, Progesterone/genetics , Mice, Inbred C57BL , Litter Size
12.
Front Biosci (Landmark Ed) ; 29(6): 238, 2024 Jun 25.
Article in English | MEDLINE | ID: mdl-38940045

ABSTRACT

BACKGROUND: Hormone receptors exert their function through binding with their ligands, which results in cellular signaling activation mediated by genomic or non-genomic mechanisms. The intrinsic molecular communication of tick Rhipicephalus microplus and its host Bos taurus comprises an endocrine regulation involving hormones. In the present study, we performed a molecular and in silico analysis of a Membrane Associated Progesterone Receptor in R. microplus (RmMAPRC). METHODS: The RmMAPRC protein sequence was analyzed with bioinformatics tools, and its structure was characterized by three-dimensional (3D) modeling and molecular docking. A semi-quantitative reverse transcription and polymerase chain reaction (sqRT-PCR) assessed the RmMAPRC gene presence and relative expression in tick organs and embryonic cells. RESULTS: RmMAPRC relative expression in salivary glands, ovaries, and embryonic cells showed overexpression of 3%, 13%, and 24%, respectively. Bioinformatic analysis revealed that RmMAPRC corresponded to a Progesterone Receptor Membrane Component 1 (RmPGRMC1) of ~23.7 kDa, with an N-terminal transmembrane domain and a C-terminal Cytochrome b5-like heme/steroid binding domain. The docking results suggest that RmPGRMC1 could bind to progesterone (P4), some progestins, and P4 antagonists. The phylogenetic reconstruction showed that Rhipicephalus spp. MAPRC receptors were clustered in a clade that includes R. appendiculatus, R. sanguineus, and R. microplus (RmMAPRC), and mammals and helminths MAPRC receptors clustered in two separated clades away from ticks. CONCLUSIONS: The presence of RmPGRMC1 highlights the importance of transregulation as a conserved adaptive mechanism that has succeeded for arthropod parasites, making it a target for tick control.


Subject(s)
Progesterone , Receptors, Progesterone , Rhipicephalus , Animals , Rhipicephalus/metabolism , Rhipicephalus/genetics , Receptors, Progesterone/metabolism , Receptors, Progesterone/genetics , Progesterone/metabolism , Cattle , Molecular Docking Simulation , Host-Parasite Interactions , Female , Amino Acid Sequence , Protein Binding , Phylogeny
13.
Res Vet Sci ; 175: 105321, 2024 Aug.
Article in English | MEDLINE | ID: mdl-38843689

ABSTRACT

The aim of the present study was to determine the effects of the adipokines progranulin and omentin on the basic functions of feline ovarian cells. For this purpose, we investigated the effects of the addition of progranulin and omentin (0, 0.1, 1, or 10 ng/ml) on the proliferation (accumulation of PCNA and cyclin B1), apoptosis (accumulation of Bax and caspase 3) and progesterone release of cultured feline ovarian granulosa cells by quantitative immunocytochemistry and enzyme-linked immunosorbent assays (ELISAs). Both progranulin and omentin increased cell proliferation and decreased apoptosis. Both progranulin and omentin promoted progesterone release. The present findings demonstrate that the adipokines progranulin and omentin can directly regulate basic feline ovarian cell functions.


Subject(s)
Apoptosis , Cell Proliferation , Granulosa Cells , Animals , Female , Cats , Granulosa Cells/metabolism , Granulosa Cells/drug effects , Apoptosis/drug effects , Cell Proliferation/drug effects , Progesterone/metabolism , Progesterone/pharmacology , Progranulins/metabolism , Cytokines/metabolism , Cells, Cultured , Lectins/metabolism , Lectins/pharmacology
14.
FASEB J ; 38(13): e23744, 2024 Jul 15.
Article in English | MEDLINE | ID: mdl-38885031

ABSTRACT

The hypothalamic-pituitary-gonadal axis (HPG) is the key neuroendocrine axis involved in reproductive regulation. Brain and muscle ARNT-like protein 1 (Bmal1) participates in regulating the metabolism of various endocrine hormones. However, the regulation of Bmal1 on HPG and female fertility is unclear. This study aims to explore the regulation of female reproduction by Bmal1 via the HPG axis in mice. Bmal1-knockout (Ko) mice were generated using the CRISPR/Cas9 technology. The structure, function, and estrous cycle of ovarian in Bmal1 Ko female mice were measured. The key genes and proteins of the HPG axis involved in regulating female reproduction were examined through transcriptome analysis and then verified by RT-PCR, immunohistochemistry, and western blot. Furthermore, the fertility of female mice was detected after intervening prolactin (PRL) and progesterone (Pg) in Bmal1 ko mice. The number of offspring and ovarian weight were significantly lower in Bmal1-Ko mice than in wild-type (Wt) mice. In Bmal1-Ko mice, ovarian cells were arranged loosely and irregularly, and the total number of follicles was significantly reduced. No corpus luteum was found in the ovaries. Vaginal smears revealed that Bmal1-Ko mice had an irregular estrus cycle. In Bmal1-Ko mice, Star expression was decreased, PRL and luteinizing hormone (LH) levels were increased, and dopamine (DA) and Pg levels were decreased. Inhibition of PRL partially recovered the estrous cycle, corpus luteum formation, and Star expression in the ovaries. Pg supplementation promoted embryo implantation in Bmal1-Ko female mice. Bmal1 Ko increases serum PRL levels in female mice likely by reducing DA levels, thus affecting luteal formation, resulting in decreased Star expression and Pg production, hindering female reproduction. Inhibition of PRL or restoration of Pg can partially restore reproductive capacity in female Bmal1-Ko mice. Thus, Bmal1 may regulate female reproduction via the HPG axis in mice, suggesting that Bmal1 is a potential target to treat female infertility.


Subject(s)
ARNTL Transcription Factors , Hypothalamo-Hypophyseal System , Ovary , Reproduction , Animals , Female , Mice , ARNTL Transcription Factors/metabolism , ARNTL Transcription Factors/genetics , Estrous Cycle , Fertility , Hypothalamo-Hypophyseal System/metabolism , Hypothalamo-Hypophyseal System/physiology , Mice, Inbred C57BL , Mice, Knockout , Ovary/metabolism , Progesterone/metabolism , Prolactin/metabolism
15.
Proc Natl Acad Sci U S A ; 121(25): e2400601121, 2024 Jun 18.
Article in English | MEDLINE | ID: mdl-38861608

ABSTRACT

The process of human parturition involves inflammation at the interface where fetal chorion trophoblast cells interact with maternal decidual stromal (DS) cells and maternal immune cells in the decidua (endometrium of pregnancy). This study tested the hypothesis that inflammation at the chorion-decidua interface (CDI) induces labor by negating the capacity for progesterone (P4) to block labor and that this is mediated by inactivation of P4 in DS cells by aldo-keto reductase family 1 member C1 (AKR1C1). In human, Rhesus macaque, and mouse CDI, AKR1C1 expression increased in association with term and preterm labor. In a human DS cell line and in explant cultures of term human fetal membranes containing the CDI, the prolabor inflammatory cytokine, interleukin-1ß (IL-1ß), and media conditioned by LPS-stimulated macrophages increased AKR1C1 expression and coordinately reduced nuclear P4 levels and P4 responsiveness. Loss of P4 responsiveness was overcome by inhibition of AKR1C1 activity, inhibition of AKR1C1 expression, and bypassing AKR1C1 activity with a P4 analog that is not metabolized by AKR1C1. Increased P4 activity in response to AKR1C1 inhibition was prevented by the P4 receptor antagonist RU486. Pharmacologic inhibition of AKR1C1 activity prevented parturition in a mouse model of inflammation-induced preterm parturition. The data suggest that inflammatory stimuli at the CDI drive labor by inducing AKR1C1-mediated P4 inactivation in DS cells and that inhibiting and/or bypassing of AKR1C1-mediated P4 inactivation is a plausible therapeutic strategy to mitigate the risk of inflammation-associated preterm birth.


Subject(s)
20-Hydroxysteroid Dehydrogenases , Decidua , Inflammation , Macaca mulatta , Parturition , Progesterone , Stromal Cells , Female , Animals , Progesterone/metabolism , Progesterone/pharmacology , Decidua/metabolism , Humans , Mice , Stromal Cells/metabolism , Pregnancy , Inflammation/metabolism , 20-Hydroxysteroid Dehydrogenases/metabolism , 20-Hydroxysteroid Dehydrogenases/genetics , Interleukin-1beta/metabolism , Chorion/metabolism
16.
Reprod Domest Anim ; 59(6): e14655, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38924202

ABSTRACT

Understanding the normal physiology of the canine mammary gland (CMG) is crucial, as it provides a foundational reference for understanding canine mammary neoplasms. The relation between the Proliferation Index (PI) indicated by Ki-67 expression, along with the Apoptotic Index (AI) determined through Caspase-3 expression during the oestrous cycle, is inadequately documented in existing literature. This study seeks to offer insights into the interplay between PI and AI in the CMG across oestrous cycle phases. An extensive investigation was conducted on a diverse case series of bitches (n = 18). Oestrous cycle stages were determined through vaginal cytology, histological examination of the reproductive tract and serum progesterone and oestradiol concentrations. The entire mammary chain was histologically examined, and proliferation and apoptosis were assessed via double immunohistochemistry employing anti-Ki-67 and Caspase-3 antibodies. PI and AI were evaluated through a systematic random sampling approach, counting a minimum of 200 cells for each cell type. There was a significantly higher PI during early dioestrus in all mammary gland components, with a greater proportion of positive cells observed in epithelial cells compared to stromal cells. The highest PI was detected in epithelial cells within the end buds. Significant differences were found in Ki-67 labelling across the cranial mammary glands. A positive and strong correlation was noted between progesterone concentration and PI in epithelial cells. The AI remained consistently low throughout the oestrous cycle, with few differences observed across histological components. Caspase-3 labelling displayed the highest positivity in caudal mammary pairs. A negative and moderate correlation was identified between progesterone concentration and AI in interlobular mesenchymal cells. This study highlights the influence of endocrine regulation on cell proliferation indices in mammary tissue, emphasizing the need to consider these hormonal variations in toxicopathological studies involving canine mammary gland.


Subject(s)
Apoptosis , Caspase 3 , Cell Proliferation , Estrous Cycle , Ki-67 Antigen , Mammary Glands, Animal , Progesterone , Animals , Female , Ki-67 Antigen/metabolism , Dogs , Apoptosis/physiology , Mammary Glands, Animal/physiology , Mammary Glands, Animal/cytology , Caspase 3/metabolism , Estrous Cycle/physiology , Progesterone/blood , Progesterone/metabolism , Estradiol/blood , Estradiol/metabolism , Epithelial Cells
17.
Reprod Domest Anim ; 59(6): e14628, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38828525

ABSTRACT

This study aimed to investigate the impact of the epidermal growth factor receptor ligands amphiregulin (AREG) and epiregulin (EREG) on the fundamental functions of feline ovarian granulosa cells. Granulosa cells isolated from feline ovaries were incubated with AREG and EREG (0, 0.1, 1 or 10 ng/mL). The effects of these growth factors on cell viability, proliferation (assessed through BrdU incorporation), nuclear apoptosis (evaluated through nuclear DNA fragmentation) and the release of progesterone and estradiol were determined using Cell Counting Kit-8 assays, BrdU analysis, TUNEL assays and ELISAs, respectively. Both AREG and EREG increased cell viability, proliferation and steroid hormone release and reduced apoptosis. The present findings suggest that these epidermal growth factor receptor ligands may serve as physiological stimulators of feline ovarian cell functions.


Subject(s)
Amphiregulin , Apoptosis , Epiregulin , Granulosa Cells , Animals , Cats , Female , Amphiregulin/metabolism , Amphiregulin/genetics , Cell Proliferation , Cell Survival , Cells, Cultured , Epiregulin/metabolism , Epiregulin/genetics , Estradiol/metabolism , Granulosa Cells/cytology , Granulosa Cells/metabolism , Progesterone/metabolism
18.
Reprod Domest Anim ; 59(6): e14643, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38877774

ABSTRACT

Progesterone has been shown to stimulate glycogen catabolism in uterine epithelial cells. Acid α-glucosidase (GAA) is an enzyme that breaks down glycogen within lysosomes. We hypothesized that progesterone may stimulate glycogenolysis in the uterine epithelium via GAA. We found that GAA was more highly expressed in the stroma on Day 1 than on Day 11. However, GAA did not appear to differ in the epithelium on Days 1 and 11. Progesterone (0-10 µM) had no effect on the levels of the full-length inactive protein (110 kDa) or the cleaved (active) peptides present inside the lysosome (70 and 76 kDa) in immortalized bovine uterine epithelial (BUTE) cells. Furthermore, the activity of GAA did not differ between the BUTE cells treated with 10 µM progesterone or control. Overall, we confirmed that GAA is present in the cow endometrium and BUTE cells. However, progesterone did not affect protein levels or enzyme activity.


Subject(s)
Endometrium , Progesterone , alpha-Glucosidases , Animals , Cattle , Female , Endometrium/metabolism , Endometrium/enzymology , Progesterone/pharmacology , Progesterone/metabolism , alpha-Glucosidases/metabolism , alpha-Glucosidases/genetics , Epithelial Cells/metabolism , Glycogenolysis , Lysosomes/enzymology , Lysosomes/metabolism , Glycogen/metabolism
19.
FASEB J ; 38(11): e23710, 2024 Jun 15.
Article in English | MEDLINE | ID: mdl-38822676

ABSTRACT

Steroidogenic tissues contain cytosolic lipid droplets that are important for steroidogenesis. Perilipin 2 (PLIN2), a structural coat protein located on the surface of lipid droplets in mammalian cells, plays a crucial role in regulating lipid droplet formation and contributing to various cellular processes such as lipid storage and energy homeostasis. Herein, we examine the role that PLIN2 plays in regulating progesterone synthesis in the bovine corpus luteum. Utilizing gene array databases and Western blotting, we have delineated the expression pattern of PLIN2 throughout the follicular to luteal transition. Our findings reveal the presence of PLIN2 in both ovarian follicular and steroidogenic luteal cells, demonstrating an increase in its levels as follicular cells transition into the luteal phase. Moreover, the depletion of PLIN2 via siRNA enhanced progesterone production in small luteal cells, whereas adenovirus-mediated overexpression of both PLIN2 and Perilipin 3 (PLIN3) induced an increase in cytosolic lipid droplet accumulation and decreased hormone-induced progesterone synthesis in these cells. Lastly, in vivo administration of the luteolytic hormone prostaglandin F2α resulted in an upregulation of PLIN2 mRNA and protein expression, accompanied by a decline in serum progesterone. Our findings highlight the pivotal role of PLIN2 in regulating progesterone synthesis in the bovine corpus luteum, as supported by its dynamic expression pattern during the follicular to luteal transition and its responsiveness to luteotropic and luteolytic hormones. We suggest PLIN2 as a potential therapeutic target for modulating luteal function.


Subject(s)
Luteal Cells , Perilipin-2 , Progesterone , Animals , Female , Cattle , Progesterone/metabolism , Perilipin-2/metabolism , Perilipin-2/genetics , Luteal Cells/metabolism , Lipid Droplets/metabolism , Membrane Proteins/metabolism , Membrane Proteins/genetics , Perilipin-3/metabolism , Corpus Luteum/metabolism , Cells, Cultured
20.
J Biol Chem ; 300(7): 107484, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38897566

ABSTRACT

Progesterone (P4), acting via its nuclear receptor (PR), is critical for pregnancy maintenance by suppressing proinflammatory and contraction-associated protein (CAP)/contractile genes in the myometrium. P4/PR partially exerts these effects by tethering to NF-κB bound to their promot-ers, thereby decreasing NF-κB transcriptional activity. However, the underlying mechanisms whereby P4/PR interaction blocks proinflammatory and CAP gene expression are not fully understood. Herein, we characterized CCR-NOT transcription complex subunit 1 (CNOT1) as a corepressor that also interacts within the same chromatin complex as PR-B. In mouse myome-trium increased expression of CAP genes Oxtr and Cx43 at term coincided with a marked decline in expression and binding of CNOT1 to NF-κB-response elements within the Oxtr and Cx43 promoters. Increased CAP gene expression was accompanied by a pronounced decrease in enrichment of repressive histone marks and increase in enrichment of active histone marks to this genomic region. These changes in histone modification were associated with changes in expression of corresponding histone modifying enzymes. Myometrial tissues from P4-treated 18.5 dpc pregnant mice manifested increased Cnot1 expression at 18.5 dpc, compared to vehicle-treated controls. P4 treatment of PR-expressing hTERT-HM cells enhanced CNOT1 expression and its recruitment to PR bound NF-κB-response elements within the CX43 and OXTR promoters. Furthermore, knockdown of CNOT1 significantly increased expression of contractile genes. These novel findings suggest that decreased expression and DNA-binding of the P4/PR-regulated transcriptional corepressor CNOT1 near term and associated changes in histone modifications at the OXTR and CX43 promoters contribute to the induction of myometrial contractility leading to parturition.


Subject(s)
Myometrium , Receptors, Progesterone , Female , Animals , Pregnancy , Myometrium/metabolism , Mice , Receptors, Progesterone/metabolism , Receptors, Progesterone/genetics , Transcription Factors/metabolism , Transcription Factors/genetics , Uterine Contraction/metabolism , Uterine Contraction/genetics , NF-kappa B/metabolism , NF-kappa B/genetics , Humans , Progesterone/metabolism , Connexin 43/metabolism , Connexin 43/genetics , Promoter Regions, Genetic , Gene Expression Regulation
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