Autophagy favors survival of corpora lutea during the long-lasting pregnancy of the South American plains vizcacha, Lagostomus maximus (Rodentia, Caviomorpha).

The corpus luteum (CL) is a transient endocrine gland that plays a crucial role in establishing and maintaining pregnancy. Although autophagy and apoptosis have been suggested as cooperative mechanisms, their interaction within the CL of pregnant mammals has not been thoroughly investigated. To understand the collaborative function of autophagy and apoptosis in the CL, we analyzed both mechanisms during pregnancy in the South American plains vizcacha, Lagostomus maximus. This rodent undergoes a decline in progesterone levels during mid-gestation, a reactivation of the hypothalamus-hypophysis-gonadal axis, and the incorporation of new functional secondary CL. Our analysis of autophagy markers BECLIN 1 (BECN1), SEQUESTOSOME1 (SQSTM1), Microtubule-associated protein light chain 3 (LC3B), and lysosomal-associated membrane protein 1 (LAMP1) and anti- and pro-apoptotic markers BCL2 and ACTIVE CASPASE 3 (A-C3) revealed interactive behaviors between both processes. Healthy primary and secondary CL exhibited positive expression of BECN1, SQSTM1, LC3B, and LAMP1, while regressed CL displayed enhanced expression of these autophagy markers along with nuclear A-C3. Transmission electron microscopy revealed a significant formation of autophagic vesicles in regressed CL during full-term pregnancy, whereas healthy CL exhibited a low number of autophagy vesicles. The co-localization between LC3B and SQSTM1 and LC3B with LAMP1 was observed in both healthy and regressed CL during pregnancy, while co-localization of BECN1 and BCL2 was only detected in healthy CL. LC3B and ACTIVE CASPASE 3 co-localization were detected in a subset of luteal cells within the regressing CL. We propose that autophagy could act as a survival mechanism in the CL, allowing the pregnancy to progress until full-term, while also serving as a mechanism to eliminate remnants of regressed CL, thereby providing the necessary space for subsequent follicular maturation.

Corpus luteum presence in the bovine ovary increase intrafollicular progesterone concentration: consequences in follicular cells gene expression and follicular fluid small extracellular vesicles miRNA contents.

BACKGROUND: It is well described that circulating progesterone (P4) plays a key role in several reproductive events such as oocyte maturation. However, during diestrus, when circulating P4 is at the highest concentrations, little is known about its local impact on the follicular cells such as intrafollicular P4 concentration due to corpus luteum (CL) presence within the same ovary. Based on that, our hypothesis is that the CL presence in the ovary during diestrus alters intrafollicular P4 concentrations, oocyte competence acquisition, follicular cells gene expression, and small extracellular vesicles (sEVs) miRNAs contents. RESULTS: P4 hormonal analysis revealed that ipsilateral to the CL follicular fluid (iFF) presented higher P4 concentration compared to contralateral follicular fluid (cFF). Furthermore, oocyte maturation and miRNA biogenesis pathways transcripts (ADAMTS-1 and AGO2, respectively) were increased in cumulus and granulosa cells of iFF, respectively. Nevertheless, a RT-PCR screening of 382 miRNAs showed that three miRNAs were upregulated and two exclusively expressed in sEVs from iFF and are predicted to regulate cell communication pathways. Similarly, seven miRNAs were higher and two exclusively expressed from cFF sEVs and are predicted to modulate proliferation signaling pathways. CONCLUSION: In conclusion, intrafollicular P4 concentration is influenced by the presence of the CL and modulates biological processes related to follicular cell development and oocyte competence, which may influence the oocyte quality. Altogether, these results are crucial to improve our knowledge about the follicular microenvironment involved in oocyte competence acquisition.

Phoenixin-14 as a novel direct regulator of porcine luteal cell functions†.

Phoenixin is a neuropeptide with a well-established role in the central regulation of reproductive processes; however, knowledge regarding its role in the ovary is limited. One of the main active phoenixin isoforms is phoenixin-14, which acts through G protein-coupled receptor 173. Our research hypothesis was that phoenixin-14 is expressed in porcine corpus luteum and exerts luteotropic action by affecting the endocrine function of luteal cells through G protein-coupled receptor 173 and protein kinase signaling. Luteal cells were cultured to investigate the effect of phoenixin-14 (1-1000 nM) on endocrine function. We showed that phoenixin-14 and G protein-coupled receptor 173 are produced locally in porcine corpus luteum and their levels change during the estrous cycle. We detected phoenixin-14 immunostaining in the cytoplasm and G protein-coupled receptor 173 in the cell membrane. Plasma phoenixin levels were highest during the early luteal phase. Interestingly, insulin, luteinizing hormone, progesterone, and prostaglandins decreased phoenixin-14 levels in luteal cells. Phoenixin-14 increased progesterone, estradiol, and prostaglandin E2 secretion, but decreased prostaglandin F2α, upregulated the expression of steroidogenic enzymes, and downregulated receptors for luteinizing hormone and prostaglandin. Also, phoenixin-14 increased the expression of G protein-coupled receptor 173 and the phosphorylation of extracellular signal-regulated kinase 1/2, protein kinase B, inhibited the phosphorylation of protein kinase A, and had mixed effect on AMP-activated protein kinase alpha and protein kinase C. G protein-coupled receptor 173 and extracellular signal-regulated kinase 1/2 mediated the effect of phoenixin-14 on endocrine function of luteal cells. Our results suggest that phoenixin is produced by porcine luteal cells and can be a new regulator of their function.

Prostaglandins as local regulators of ovarian physiology in ruminants.

Prostaglandins are synthesized from arachidonic acid through the catalytic activities of cyclooxygenase, while the production of different prostaglandin types, prostaglandin F2 alpha (PGF) and prostaglandin E2 (PGE), are regulated by specific prostaglandin synthases (PGFS and PGES). Prostaglandin ligands (PGF and PGE) bind to specific high-affinity receptors and initiate biologically distinct signalling pathways. In the ovaries, prostaglandins are known to be important endocrine regulators of female reproduction, in addition to maintaining local function through autocrine and/or paracrine effect. Many research groups in different animal species have already identified a variety of factors and molecular mechanisms that are responsible for the regulation of prostaglandin functions. In addition, prostaglandins stimulate their intrafollicular and intraluteal production via the pathway of prostaglandin self-regulation in the ovary. Therefore, the objective of the review article is to discuss recent findings about local regulation patterns of prostaglandin ligands PGF and PGE during different physiological stages of ovarian function in domestic ruminants, especially in bovine. In conclusion, the discussed local regulation mechanisms of prostaglandins in the ovary may stimulate further research activities in different methodological approaches, especially during final follicle maturation and ovulation, as well as corpus luteum formation and function.

PGF2alpha Inhibits 20alpha-HSD Expression by Suppressing CK1alpha-induced ERK and SP1 Activation in the Corpus Luteum of Pregnant Mice.

Prostaglandin F2α (PGF2α) is a luteolytic hormone that promotes parturition in mammals at the end of pregnancy by reducing progesterone secretion from the corpus luteum (CL). In rodents and primates, PGF2α rapidly converts progesterone to 20α-hydroxyprogesterone (20α-OHP) by promoting 20α-hydroxysteroid dehydrogenase (20α-HSD) expression. However, the specific mechanism of 20α-HSD regulation by PGF2α remains unclear. Casein Kinase 1α (CK1α) is a CK1 family member that regulates a variety of physiological functions, including reproductive development. Here, we investigated the effects of CK1α on pregnancy in female mice. Our experiments showed that CK1α is expressed in mouse CL, and its inhibition enhanced progesterone metabolism, decreased progesterone levels, and affected mouse embryo implantation. Further, CK1α mediated the effect of PGF2α on 20α-HSD in mouse luteal cells in vitro. Our results are the first to show that CK1α affects the 20α-HSD mRNA level by affecting the ERK signalling pathway to regulate the expression of the transcription factor SP1. These findings improve our understanding of PGF2α regulation of 20α-HSD.

Immunoexpression of vascular endothelial growth factor and vWF-regulating angiogenesis in cyclic corpus luteum of Indian buffalo.

The present study was conducted to localize the immunoexpression of VEGF-A (Vascular Endothelial Growth Factor) and von Willebrand factor (vWF) in corpora lutea of healthy buffaloes (24) collected from local slaughterhouses. CL collected were categorized into early (stage I, 1-5 days, n = 6), mid (stage II, 6-11 days, n = 6), late luteal phase (stage III, 12 to 16 days, n = 6) and regressing phase (stage IV, 17 to 20 days, n = 6). The percent positive immunostaining for VEGF-A was significantly (p < 0.05) higher in mid-luteal phase than the other three stages of CL. However, it was higher in early luteal phase as well indicated intense angiogenesis in both early and mid-luteal phases. The number of capillary endothelium expressing vWF was significantly (p < 0.05) highest in mid-luteal phase among all the phases. However, in late luteal phase, the percent area positive for VEGF-A immunostaining was reduced but it was significantly (p < 0.05) higher than corpus albicans phase. Thus, in regressing phase or corpus albicans, it was lowest and reduced considerably. However, in late luteal phase, the number of capillaries with vWF immunoexpression reduced significantly (p < 0.05) but it was lowest in corpus albicans phase. Therefore, the immunotaining pattern for VEGF-A and vWF concluded that there was a spositive linear correlation between the two, that is, as the VEGF-A expression was increased, the number of vWF positive capillaries also increased and vice versa. The VEGF-A expressed by the luteal parenchyma in different stages of development and regression of corpus luteum was thus observed to be involved in promoting the angiogenesis and luteal cell proliferation as supported by vWF expressed by endothelium of proliferating capillaries in buffalo corpus luteum throughout the estrous cycle.

Differential expression of angiogenesis-related genes in goat uterus and corpus luteum during pregnancy.

Vascularization and the control of luteal and endometrial development are regulated by hypoxia-inducible factors (HIFs) and vascular endothelial growth factor (VEGF) during pregnancy. In this study, the mRNA and protein expression levels of HIFs (HIF1A, HIF2A and HIF3A) and VEGF in goat uterine and ovarian tissues during various stages of pregnancy were evaluated. A total of 42 Hair goats were used and were allocated into six groups, namely embryo-positive (G1), early pregnancy (G2), mid-term pregnancy (G3), late pregnancy (G4), oocyte-positive group (G5) and diestrus group (G6). The mRNA expression of the examined genes was evaluated by RT-qPCR, and protein expression was evaluated by immunohistochemistry (IHC). In caruncles, HIF1A mRNA expression was greater in G1, G2 and G4 than in G3 (p < .05). HIF1A and HIF2A expression was greater in G1 than in G5 (p < .05). In cotyledons, HIF1A, HIF2A and HIF3A mRNA expression was greater in G2 and G3 compared to G4 (p < .05). In luteal tissue, HIF1A mRNA expression was greater in G1 and G2 than in G3 and G4 (p < .05). In the immunohistochemical examination, HIF1A, HIF2A, HIF3A and VEGF immunoreactions were detected in uterine and luteal tissues. Findings suggest that HIFs and VEGF are involved in the regulation of ovarian functions as well as the processes of implantation and placentation.

ERK1/2-SOX9/FOXL2 axis regulates ovarian steroidogenesis and favors the follicular-luteal transition.

Estradiol and progesterone are the primary sex steroids produced by the ovary. Upon luteinizing hormone surge, estradiol-producing granulosa cells convert into progesterone-producing cells and eventually become large luteal cells of the corpus luteum. Signaling pathways and transcription factors involved in the cessation of estradiol and simultaneous stimulation of progesterone production in granulosa cells are not clearly understood. Here, we decipher that phosphorylated ERK1/2 regulates granulosa cell steroidogenesis by inhibiting estradiol and inducing progesterone production. Down-regulation of transcription factor FOXL2 and up-regulation of SOX9 by ERK underpin its differential steroidogenic function. Interestingly, the incidence of SOX9 is largely uncovered in ovarian cells and is found to regulate FOXL2 along with CYP19A1 and STAR genes, encoding rate-limiting enzymes of steroidogenesis, in cultured granulosa cells. We propose that the novel ERK1/2-SOX9/FOXL2 axis in granulosa cells is a critical regulator of ovarian steroidogenesis and may be considered when addressing pathophysiologies associated with inappropriate steroid production and infertility in humans and animals.

Adropin may regulate corpus luteum formation and its function in adult mouse ovary.

BACKGROUND: Adropin, a unique peptide hormone, has been associated with the regulation of several physiological processes, including glucose homeostasis, fatty acid metabolism, and neovascularization. However, its possible role in ovarian function is not understood. Our objective was to examine the expression of adropin and its putative receptor, GPR19, in the ovaries of mice at various phases of the estrous cycle. METHODS: Immunohistochemistry and western blot analysis were performed to explore the localization and changes in expression of adropin and GPR19 in the ovaries during different phases of the estrous cycle in mice. Hormonal assays were performed with ELISA. An in vitro study was performed to examine the direct effect of adropin (10, 100 ng/ml) on ovarian function. RESULTS: A western blot study showed that adropin and GPR19 proteins were maximum during the estrus phase of the estrous cycle. Interestingly, adropin and GPR19 displayed intense immunoreactivity in granulosa cells of large antral follicles and corpus luteum. This suggested the possible involvement of adropin in corpus luteum formation. Adropin treatment stimulated progesterone synthesis by increasing GPR19, StAR, CYP11A1, and 3ß-HSD expressions, while it decreased estrogen synthesis by inhibiting 17ß-HSD and aromatase protein expressions. Moreover, adropin treatment upregulated the cell cycle arrest-CDK inhibitor 1B (p27kip1), pERK1/2, and angiogenic protein (EG VEGF) that are involved in the process of luteinization. CONCLUSIONS: Adropin GPR19 signaling promotes the synthesis of progesterone and upregulates the expression of p27kip1, EG VEGF, and erk1/2, resulting in cell cycle arrest and neovascularization, which ultimately leads to corpus luteum formation.

Luteal Lipid Droplets: A Novel Platform for Steroid Synthesis.

Progesterone is an essential steroid hormone that is required to initiate and maintain pregnancy in mammals and serves as a metabolic intermediate in the synthesis of endogenously produced steroids, including sex hormones and corticosteroids. Steroidogenic luteal cells of the corpus luteum have the tremendous capacity to synthesize progesterone. These specialized cells are highly enriched with lipid droplets that store lipid substrate, which can be used for the synthesis of steroids. We recently reported that hormone-stimulated progesterone synthesis by luteal cells requires protein kinase A-dependent mobilization of cholesterol substrate from lipid droplets to mitochondria. We hypothesize that luteal lipid droplets are enriched with steroidogenic enzymes and facilitate the synthesis of steroids in the corpus luteum. In the present study, we analyzed the lipid droplet proteome, conducted the first proteomic analysis of lipid droplets under acute cyclic adenosine monophosphate (cAMP)-stimulated conditions, and determined how specific lipid droplet proteins affect steroidogenesis. Steroidogenic enzymes, cytochrome P450 family 11 subfamily A member 1 and 3 beta-hydroxysteroid dehydrogenase (HSD3B), were highly abundant on lipid droplets of the bovine corpus luteum. High-resolution confocal microscopy confirmed the presence of active HSD3B on the surface of luteal lipid droplets. We report that luteal lipid droplets have the capacity to synthesize progesterone from pregnenolone. Lastly, we analyzed the lipid droplet proteome following acute stimulation with cAMP analog, 8-Br-cAMP, and report increased association of HSD3B with luteal lipid droplets following stimulation. These findings provide novel insights into the role of luteal lipid droplets in steroid synthesis.

Progesterone signaling in the regulation of luteal steroidogenesis.

The corpus luteum is the major source of progesterone, the essential hormone for female reproductive function. While progesterone activity has been the subject of extensive research for decades, characterization of non-canonical progesterone receptor/signaling pathways provided a new perspective for understanding the complex signal transduction mechanisms exploited by the progesterone hormone. Deciphering these mechanisms has significant implications in the management of luteal phase disorders and early pregnancy complications. The purpose of this review is to highlight the complex mechanisms through which progesterone-induced signaling mediates luteal granulosa cell activity in the corpus luteum. Here, we review the literature and discuss the up-to-date evidence on how paracrine and autocrine effects of progesterone regulate luteal steroidogenic activity. We also review the limitations of the published data and highlight future research priorities.

Basic fibroblast growth factor induces proliferation and collagen production by fibroblasts derived from the bovine corpus luteum†.

Cyclic regression of the ovarian corpus luteum, the endocrine gland responsible for progesterone production, involves rapid matrix remodeling. Despite fibroblasts in other systems being known for producing and maintaining extracellular matrix, little is known about fibroblasts in the functional or regressing corpus luteum. Vast transcriptomic changes occur in the regressing corpus luteum, among which are reduced levels of vascular endothelial growth factor A (VEGFA) and increased expression of fibroblast growth factor 2 (FGF2) after 4 and 12 h of induced regression, when progesterone is declining and the microvasculature is destabilizing. We hypothesized that FGF2 activates luteal fibroblasts. Analysis of transcriptomic changes during induced luteal regression revealed elevations in markers of fibroblast activation and fibrosis, including fibroblast activation protein (FAP), serpin family E member 1 (SERPINE1), and secreted phosphoprotein 1 (SPP1). To test our hypothesis, we treated bovine luteal fibroblasts with FGF2 to measure downstream signaling, type 1 collagen production, and proliferation. We observed rapid and robust phosphorylation of various signaling pathways involved in proliferation, such as ERK, AKT, and STAT1. From our longer-term treatments, we determined that FGF2 has a concentration-dependent collagen-inducing effect, and that FGF2 acts as a mitogen for luteal fibroblasts. FGF2-induced proliferation was greatly blunted by inhibition of AKT or STAT1 signaling. Our results suggest that luteal fibroblasts are responsive to factors that are released by the regressing bovine corpus luteum, an insight into the contribution of fibroblasts to the microenvironment in the regressing corpus luteum.

Prostaglandin F2α regulates mitochondrial dynamics and mitophagy in the bovine corpus luteum.

Prostaglandins are arachidonic acid-derived lipid mediators involved in numerous physiological and pathological processes. PGF2α analogues are therapeutically used for regulating mammalian reproductive cycles and blood pressure, inducing term labor, and treating ocular disorders. PGF2α exerts effects via activation of calcium and PKC signaling, however, little is known about the cellular events imposed by PGF2α signaling. Here, we explored the early effects of PGF2α on mitochondrial dynamics and mitophagy in the bovine corpus luteum employing relevant and well characterized in vivo and in vitro approaches. We identified PKC/ERK and AMPK as critical protein kinases essential for activation of mitochondrial fission proteins, DRP1 and MFF. Furthermore, we report that PGF2α elicits increased intracellular reactive oxygen species and promotes receptor-mediated activation of PINK-Parkin mitophagy. These findings place the mitochondrium as a novel target in response to luteolytic mediator, PGF2α. Understanding intracellular processes occurring during early luteolysis may serve as a target for improving fertility.

Resistin as a new player in the regulation of porcine corpus luteum luteolysis: in vitro effect on proliferation/viability, apoptosis and autophagy.

The formation and luteolysis of the corpus luteum (CL) is strictly controlled by many factors. Imbalance between proliferation and apoptosis processes leads to deficiency of the luteal phase and infertility. Our previous study showed resistin expression in porcine luteal cells and an inhibitory effect on progesterone synthesis. Thus, the aim of the present study was to examine the in vitro effect of resistin on the proliferation/viability, apoptosis and autophagy of porcine luteal cells as well as the involvement of mitogen-activated kinase (MAP3/1), protein kinase B (AKT) and signal transducer and activator of transcription 3 (STAT3) in these processes. Porcine luteal cells were incubated with resistin (0.1-10 ng/mL) for 24-72 h and viability was assessed using the alamarBlue or 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Then, the time-dependent effect of resistin on mRNA and protein expression of proliferating cell nuclear antigen (PCNA), caspase 3, BCL2-like protein 4 (BAX), B-cell lymphoma 2 (BCL2), beclin1, microtubule-associated protein 1A/1B-light chain 3 (LC3) and lysosomal-associated membrane protein 1 (LAMP1) was measured by real-time polymerase chain reaction (PCR) and immunoblotting, respectively. We found that resistin enhanced luteal cell viability with no effect on caspase 3 mRNA and protein, increased the BAX/BCL2 mRNA and protein ratio and significantly stimulated the initiation of autophagy, which promotes the maintenance of CL function rather than its regression. Additionally, using pharmacological inhibitors of MAP3/1 (PD98059), AKT (LY294002) and STAT3 (AG490), we observed that the effect of resistin was reversed to the control level in viability and, by influence, MAP3/1 and STAT3 in autophagy. Taken together, our results suggest that resistin, in addition to its well-known effect on granulosa cell function has direct influence on CL luteolysis and the formation and maintenance of luteal cell function.

The involvement of let-7 in hCG-induced progesterone synthesis via regulating p27Kip1 and p21Cip1 expression.

Progesterone is essential in females to maintain a regular menstrual cycle and pregnancy. The luteinizing hormone (LH) surge induces the luteinization of granulosa cells and thecal cells to form the corpus luteum, which is responsible for progesterone synthesis. However, the specific mechanism of how hCG, the analog of LH, regulates progesterone synthesis has yet to be fully discovered. In this study, we found that progesterone level was increased in adult wild-type pregnant mice 2 and 7 days post-coitum, along with a decrease in let-7 expression compared with the estrus stage. Besides, the let-7 expression was negatively correlated with progesterone level in post-delivery day 23 wild-type female mice after being injected with PMSG and hCG. Then, using let-7 transgenic mice and a human granulosa cell line, we found that overexpression of let-7 antagonized progesterone level via targeting p27Kip1 and p21Cip1 and steroidogenic acute regulatory protein (StAR) expression, which is a rate-limiting enzyme in progesterone synthesis. Furthermore, hCG suppressed let-7 expression by stimulating the MAPK pathway. This study elucidated the role of microRNA let-7 in regulating hCG-induced progesterone production and provided new insights into its role in clinical application.

Role of intraluteal and intrauterine prostaglandin signaling in LH-induced luteolysis in pregnant rats.

Luteal dysfunctions lead to fertility disorders and pregnancy complications. Normal luteal function is regulated by many factors, including luteinizing hormone (LH). The luteotropic roles of LH have been widely investigated but its role in the process of luteolysis has received little attention. LH has been shown to have luteolytic effects during pregnancy in rats and the role of intraluteal prostaglandins (PGs) in LH-mediated luteolysis has been demonstrated by others. However, the status of PG signaling in the uterus during LH-mediated luteolysis remains unexplored. In this study, we utilized the repeated LH administration (4×LH) model for luteolysis induction. We have examined the effect of LH-mediated luteolysis on the expression of genes involved in luteal/uterine PG synthesis, luteal PGF2α signaling, and uterine activation during different stages (mid and late) of pregnancy. Further, we analyzed the effect of overall PG synthesis machinery blockage on LH-mediated luteolysis during late pregnancy. Unlike the midstage of pregnancy, the expression of genes involved in PG synthesis, PGF2α signaling, and uterine activation in late-stage pregnant rats' luteal and uterine tissue increase 4×LH. Since the cAMP/PKA pathway mediates LH-mediated luteolysis, we analyzed the effect of inhibition of endogenous PG synthesis on the cAMP/PKA/CREB pathway, followed by the analysis of the expression of markers of luteolysis. Inhibition of endogenous PG synthesis did not affect the cAMP/PKA/CREB pathway. However, in the absence of endogenous PGs, luteolysis could not be activated to the full extent. Our results suggest that endogenous PGs may contribute to LH-mediated luteolysis, but this dependency on endogenous PGs is pregnancy-stage dependent. These findings advance our understanding of the molecular pathways that regulate luteolysis.

PPARß/δ Ligands Regulate Oxidative Status and Inflammatory Response in Inflamed Corpus Luteum-An In Vitro Study.

Inflammation in the female reproductive system causes serious health problems including infertility. The aim of this study was to determine the in vitro effects of peroxisome proliferator-activated receptor-beta/delta (PPARß/δ) ligands on the transcriptomic profile of the lipopolysaccharide (LPS)-stimulated pig corpus luteum (CL) in the mid-luteal phase of the estrous cycle using RNA-seq technology. The CL slices were incubated in the presence of LPS or in combination with LPS and the PPARß/δ agonist-GW0724 (1 µmol/L or 10 µmol/L) or the antagonist-GSK3787 (25 µmol/L). We identified 117 differentially expressed genes after treatment with LPS; 102 and 97 differentially expressed genes after treatment, respectively, with the PPARß/δ agonist at a concentration of 1 µmol/L or 10 µmol/L, as well as 88 after the treatment with the PPARß/δ antagonist. In addition, biochemical analyses of oxidative status were performed (total antioxidant capacity and activity of peroxidase, catalase, superoxide dismutase, and glutathione S-transferase). This study revealed that PPARß/δ agonists regulate genes involved in the inflammatory response in a dose-dependent manner. The results indicate that the lower dose of GW0724 showed an anti-inflammatory character, while the higher dose seems to be pro-inflammatory. We propose that GW0724 should be considered for further research to alleviate chronic inflammation (at the lower dose) or to support the natural immune response against pathogens (at the higher dose) in the inflamed corpus luteum.

Prokineticin 1 is a novel factor regulating porcine corpus luteum function.

Prokineticin 1 (PROK1) is a pleiotropic factor secreted by endocrine glands; however, its role has not been studied in the corpus luteum (CL) during pregnancy in any species. The present study aimed to investigate the contribution of PROK1 in regulating processes related to porcine CL function and regression: steroidogenesis, luteal cell apoptosis and viability, and angiogenesis. The luteal expression of PROK1 was greater on Days 12 and 14 of pregnancy compared to Day 9. PROK1 protein expression during pregnancy increased gradually and peaked on Day 14, when it was also significantly higher than that on Day 14 of the estrous cycle. Prokineticin receptor 1 (PROKR1) mRNA abundance increased on Days 12 and 14 of pregnancy, whereas PROKR2 elevated on Day 14 of the estrous cycle. PROK1, acting via PROKR1, stimulated the expression of genes involved in progesterone synthesis, as well as progesterone secretion by luteal tissue. PROK1-PROKR1 signaling reduced apoptosis and increased the viability of luteal cells. PROK1 acting through PROKR1 stimulated angiogenesis by increasing capillary-like structure formation by luteal endothelial cells and elevating angiogenin gene expression and VEGFA secretion by luteal tissue. Our results indicate that PROK1 regulates processes vital for maintaining luteal function during early pregnancy and the mid-luteal phase.

Expression and localization of apelin and apelin receptor (APJ) in buffalo ovarian follicles and corpus luteum and the in-vitro effect of apelin on steroidogenesis and survival of granulosa cells.

Apelin is an adipose tissue-derived hormone with many physiological functions, including the regulation of female reproduction. It acts through an orphan G protein-coupled receptor APJ/APLNR. The present study aimed to investigate the expression of apelin and its receptor APJ in the ovarian follicles and corpus luteum (CL) and the role of apelin on steroidogenesis and cell survival. Ovarian follicles were classified into four groups based on size and estradiol (E2) level in the follicular fluid as follows: (i) F1 (4-6 mm; <0.5 ng/mL) (ii) F2 (7-9 mm; 0.5-5 ng/mL) (iii) F3 (10-13 mm; 5-40 ng/mL) and (iv) F4 (dominant/pre-ovulatory follicle) (>13 mm; >180 ng/mL). The corpora lutea (CL) were categorized into early (CL1), mid (CL2), late luteal (CL3), and regressing (CL4) CL stages. Expression of apelin increased with follicle size, with significantly greatest in the dominant or pre-ovulatory follicle (P < 0.05). Expression of APJ was greater in large and dominant follicles than in small and medium follicles (P < 0.05). In CL, the mRNA and protein abundance of apelin and apelin receptor was greater during mid (CL2) and late luteal (CL3) stages as compared to early (CL1) and regressing (CL4) stages (P < 0.05). Both the factors were localized in granulosa and theca cells of follicles and small and large luteal cells of CL. The pattern of the intensity of immunofluorescence was similar to mRNA and protein expression. Granulosa cells were cultured in vitro and treated at 1, 10, and 10 ng/mL apelin-13 either alone or in the presence of the follicle-stimulating hormone (FSH) (30 ng/mL) or insulin-like growth factor-I (IGF-I) (10 ng/mL) for 48 h. The luteal cells were treated with apelin-13 at 1, 10, and 100 ng/mL doses for 48 h. Apelin treatment at 10 and 100 ng/ml significantly (P < 0.05) increased E2 secretion, cytochrome P450 aromatase or CYP19A1 expression in GC. In luteal cells, apelin at 10 ng/mL and 100 ng/mL significantly (P < 0.05) increased progesterone (P4) secretion and HSD3B1 expression. In GCs, apelin, either alone or in combination, increased PCNA expression and inhibited CASPASE3 expression suggesting its role in cell survival. In conclusion, this study provides novel evidence for the presence of apelin and receptor APJ in ovarian follicles and corpora lutea and the stimulatory effect on E2 and P4 production and promotes GC survival in buffalo, suggesting the role of apelin in follicular and luteal functions in buffalo.

Continuous light exposure influences luteinization and luteal function of ovary in ICR mice.

With the rapid change of people's lifestyle, more childbearing couples live with irregular schedules (i.e., staying up late) and suffer from decreased fertility and abortion, which can be caused by luteal phase defect (LPD). We used continuous light-exposed mice as a model to observe whether continuous light exposure may affect luteinization and luteal function. We showed that the level of progesterone in serum reduced (p < .001), the number of corpus luteum (CL) decreased (p < .01), and the expressions of luteinization-related genes (Lhcgr, Star, Ptgfr, and Runx2), clock genes (Clock and Per1), and Mt1 were downregulated (p < .05) in the ovaries of mice exposed to continuous light, suggesting that continuous light exposure induces defects in luteinization and luteal functions. Strikingly, injection of melatonin (3 mg/kg) could improve luteal functions in continuous light-exposed mice. Moreover, we found that, after 2 h of hCG injection, the level of pERK1/2 in the ovary decreased in the continuous light group, but increased in the melatonin administration group, suggesting that melatonin can improve LPD caused by continuous light exposure through activating the ERK1/2 pathway. In summary, our data demonstrate that continuous light exposure affects ovary luteinization and luteal function, which can be rescued by melatonin.