Search Results (13)

Background/Objectives: Platelet-derived growth factor receptor alpha (PDGFRα) is a receptor involved in cell growth and differentiation, with unclear roles in ovarian tissues and potential interactions with KCNK3 (potassium two-pore domain channel subfamily K member 3), a member of the two-pore domain K⁺ channel involved in cellular homeostasis. This study aims to map PDGFRα expression across mouse tissues and to explore its co-expression with KCNK3 in the ovary. Methods: We visualized PDGFRα expression using RNA-seq data from the genotype-tissue expression (GTEx) BodyMAP across 54 human tissues and Cap Analysis of Gene Expression (CAGE) data for various mouse tissues. In PDGFRα^EGFP mice expressing EGFP in PDGFRα⁺ cells, histological and fluorescence imaging were used to assess ovarian expression. Immunohistochemistry determined the co-localization of PDGFRα and KCNK3, and qPCR quantified their mRNA levels in the ovary, oviduct, and uterus. Results: PDGFRα showed high expression in human and mouse female reproductive tissues, particularly the ovary. In the PDGFRα^EGFP mouse model, PDGFRα was primarily found in the thecal layer and stromal cells, not in granulosa cells or oocytes. Immunohistochemistry indicated that 90.2 ± 8.7% of PDGFRα⁺ cells expressed KCNK3 in the ovarian stroma. qPCR revealed lower PDGFRα and KCNK3 expression in the ovary compared to the oviduct and uterus. Conclusions: This study shows that PDGFRα is predominantly expressed in ovarian stromal and theca cells and is highly co-localized with KCNK3, suggesting a potential role for PDGFRα⁺ cells in ionic regulation and their possible involvement in follicular development and ovarian physiology. Full article

Olive flounder (Paralichthys olivaceus) is a commercially important fish species in Japan, China, and the Republic of Korea. Despite numerous attempts to improve productivity, there have been no studies of in vitro germline stem cell (GSC) culture in this species. Here, olive flounder testicular and ovarian cell lines (OFT and OFO, respectively) were established and characterized. RT-PCR demonstrated that OFT and OFO expressed several gonadal somatic cell markers, including wt1 and fgf2, but lacked expression of germ cell markers, such as vasa, nanos2, and scp3. In addition, SNP analysis revealed that OFT originated from XY male P. olivaceus and OFO originated from XX female P. olivaceus. These results suggest that OFT was composed of Sertoli cells and OFO was composed of granulosa cells and theca cells. Finally, coculture of OFT or OFO with enriched male P. olivaceus GSCs isolated from the top 20% and 20–30% Percoll density gradient layers showed that GSCs were attached on both cell lines. In conclusion, we established P. olivaceus testicular and ovarian cell lines, which were expected to use for development of an in vitro GSC culture system. Full article

Red light (RL) has been observed to enhance egg production in pigeons, yet the underlying histological characteristics and molecular mechanisms remain less understood. This study included fifty-four pigeons to assess follicular histology, reproductive hormones, and ovarian transcriptomics on the third day of the laying interval under RL and white light (WL). The results showed that the granulosa cell layer was significantly thicker under RL (p < 0.05), whereas the theca cell and connective tissue layers showed no significant differences (p > 0.05). Higher plasma estradiol (E₂) levels were recorded in the RL group (p < 0.05), as well as follicle stimulating hormone (FSH), although progesterone (P₄) levels were higher under WL (p < 0.05). Moreover, P₄ concentrations in follicle yolk significantly decreased under RL (p < 0.01), with higher FSH and E₂ levels in F1 yolk and similar increases in SF1 yolk (p < 0.01). Transcriptomic analysis revealed 4991 differentially expressed genes in the pigeon ovary. The protein–protein interaction network highlighted genes like HSD11B1, VEGFD, WNT6, SMAD6, and LGR5 as potential contributors to hierarchical follicle selection under RL. This research provides new insights into the molecular basis by which RL may promote hierarchical follicle selection and improve egg production in pigeons. Full article

In recent years, vitamin D₃ has been revealed as an important regulator of reproductive processes in humans and livestock; however, its role in the female reproductive system of poultry is poorly known. The aim of this study was to examine vitamin D₃ receptor (VDR and PDIA3) and metabolic enzyme (1α-hydroxylase and 24-hydroxylase) mRNA transcript and protein abundances, and protein localization within the hen ovary, oviductal shell gland, pituitary, liver, and kidney. We demonstrated, for the first time, the patterns of the relative mRNA and protein abundances of examined molecules in the ovary, dependent on follicle development and the layer of follicle wall, as well as in other examined organs. Immunohistochemically, PDIA3, 1α-hydroxylase, and 24-hydroxylase are localized in follicular theca and granulosa layers, luminal epithelium and tubular glands of the shell gland, pituitary, liver, and kidney. These results indicate that reproductive tissues have both receptors, VDR, primarily involved in genomic action, and PDIA3, probably participating in the rapid, non-genomic effect of vitamin D₃. The finding of 1α-hydroxylase and 24-hydroxylase expression indicates that the reproductive system of chickens has the potential for vitamin D₃ synthesis and inactivation, and may suggest that locally produced vitamin D₃ can be considered as a significant factor in the orchestration of ovarian and shell gland function in hens. These results provide a new insight into the potential mechanisms of vitamin D₃ action and metabolism in the chicken ovary and oviduct. Full article

This study sought to examine the ovarian cellular and stromal components of the zebrafish (Danio rerio) throughout the spawning season using light and electron microscopic tools. The ovaries of zebrafish showed oocytes in all stages of follicular development and degeneration (atresia). Six stages of oogenesis were demonstrated: oogonia, early oocytes, late oocytes, vacuolated follicles, the yolk globule stage (vitellogenesis), and mature follicles. The SOX9 protein was expressed in the ooplasm of the primary and previtellogenic oocytes and the theca cell layer of the mature follicles. Myostatin was expressed in the granulosa and theca cells. Many stem cells in the ovarian stroma expressed myostatin and SOX9. During the spawning season, the EM results indicated that the zona radiata increased in thickness and was crossed perpendicularly by pore canals that contained processes from both oocytes and zona granulosa. The granulosa cells contained many mitochondria, rER, sER, and vesicles. Meanwhile, the thecal layer consisted of fibroblast-like cells. Atretic follicles could be demonstrated that involved both oocytes and their follicular walls. Several types of cells were distinguished in the ovarian stroma, including mast cells, telocytes, lymphocytes, fibroblasts, endocrine cells, macrophages, adipocytes, dendritic cells, and steroidogenic (stromal) cells. The ovary of the zebrafish serves as a model to investigate follicular development. Full article

1α,25-Dihydroxyvitamin D₃ (VitD₃) is the active form of vitamin D, and it regulates gene expression and protein synthesis in mammalian follicle development. However, the function of VitD₃ in the follicular development of layers remains unclear. This study investigated, through in vivo and in vitro experiments, the effects of VitD₃ on follicle development and steroid hormone biosynthesis in young layers. In vivo, ninety 18-week-old Hy-Line Brown laying hens were randomly divided into three groups for different treatments of VitD₃ (0, 10, and 100 μg/kg). VitD₃ supplementation promoted follicle development, increasing the number of small yellow follicles (SYFs) and large yellow follicles (LYFs) and the thickness of the granulosa layer (GL) of SYFs. Transcriptome analysis revealed that VitD₃ supplementation altered gene expression in the ovarian steroidogenesis, cholesterol metabolism, and glycerolipid metabolism signaling pathways. Steroid hormone-targeted metabolomics profiling identified 20 steroid hormones altered by VitD₃ treatment, with 5 being significantly different among the groups. In vitro, it was found that VitD₃ increased cell proliferation, promoted cell-cycle progression, regulated the expression of cell-cycle-related genes, and inhibited the apoptosis of granulosa cells from pre-hierarchical follicles (phGCs) and theca cells from prehierarchical follicles (phTCs). In addition, the steroid hormone biosynthesis-related genes, estradiol (E2) and progesterone (P4) concentrations, and vitamin D receptor (VDR) expression level was significantly altered by VitD₃. Our findings identified that VitD₃ altered the gene expression related to steroid metabolism and the production of testosterone, estradiol, and progesterone in the pre-hierarchical follicles (PHFs), resulting in positive effects on poultry follicular development. Full article

Myo-inositol is a natural polyol, the most abundant among the nine possible structural isomers available in living organisms. Inositol confers some distinctive traits that allow for a striking distinction between prokaryotes and eukaryotes, the basic clusters into which organisms are partitioned. Inositol cooperates in numerous biological functions where the polyol participates or by furnishing the fundamental backbone of several related derived metabolites, mostly obtained through the sequential addition of phosphate groups (inositol phosphates, phosphoinositides, and pyrophosphates). Overall myo-inositol and its phosphate metabolites display an entangled network, which is involved in the core of the biochemical processes governing critical transitions inside cells. Noticeably, experimental data have shown that myo-inositol and its most relevant epimer D-chiro-inositol are both necessary to permit a faithful transduction of insulin and of other molecular factors. This improves the complete breakdown of glucose through the citric acid cycle, especially in glucose-greedy tissues, such as the ovary. In particular, while D-chiro-inositol promotes androgen synthesis in the theca layer and down-regulates aromatase and estrogen expression in granulosa cells, myo-inositol strengthens aromatase and FSH receptor expression. Inositol effects on glucose metabolism and steroid hormone synthesis represent an intriguing area of investigation, as recent results have demonstrated that inositol-related metabolites dramatically modulate the expression of several genes. Conversely, treatments including myo-inositol and its isomers have proven to be effective in the management and symptomatic relief of a number of diseases associated with the endocrine function of the ovary, namely polycystic ovarian syndrome. Full article

MiRNAs have been found to be involved in the regulation of ovarian function as important post-transcriptional regulators, including regulators of follicular development, steroidogenesis, cell atresia, and even the development of ovarian cancer. In this study, we evaluated the regulatory role of gga-miR-449b-5p in follicular growth and steroid synthesis in ovarian granulosa cells (GCs) of laying hens through qRT-PCR, ELISAs, western blotting and dual-luciferase reporter assays, which have been described in our previous study. We demonstrated that gga-miR-449b-5p was widely expressed in granulosa and theca layers of the different-sized follicles, especially in the granulosa layer. The gga-miR-449b-5p had no significant effect on the proliferation of GCs, but could significantly regulate the expression of key steroidogenesis-related genes (StAR and CYP19A1) (p < 0.01) and the secretion of P4 and E2 (p < 0.01 and p < 0.05). Further research showed that gga-miR-449b-5p could target IGF2BP3 and downregulate the mRNA and protein expression of IGF2BP3 (p < 0.05). Therefore, this study suggests that gga-miR-449b-5p is a potent regulator of the synthesis of steroid hormones in GCs by targeting the expression of IGF2BP3 and may contribute to a better understanding of the role of functional miRNAs in laying hen ovarian development. Full article

The development of an adequate blood vessel network is crucial for the accomplishment of ovarian follicle growth and ovulation, which is necessary to support the proliferative and endocrine functions of the follicular cells. Although the Vascular Endothelial Growth Factor (VEGF) through gonadotropins guides ovarian angiogenesis, the role exerted by the switch on of Progesterone (P₄) during the periovulatory phase remains to be clarified. The present research aimed to investigate in vivo VEGF-mediated mechanisms by inducing the development of periovulatory follicles using a pharmacologically validated synchronization treatment carried out in presence or absence of P₄ receptor antagonist RU486. Spatio-temporal expression profiles of VEGF, FLT1, and FLK1 receptors and the two major MAPK/ERKs and PI3K/AKT downstream pathways were analyzed on granulosa and on theca compartment. For the first time, the results demonstrated that in vivo administration of P₄ antagonist RU486 inhibits follicular VEGF receptors’ signaling mainly acting on the theca layer by downregulating the activation of ERKs and AKTs. Under the effect of RU486, periovulatory follicles’ microarchitecture did not move towards the periovulatory stage. The present evidence provides new insights on P₄ in vivo biological effects in driving vascular and tissue remodeling during the periovulatory phase. Full article

The ovarian follicle is the basic functional unit of the ovary, comprising theca cells and granulosa cells (GCs). Two different types of GCs, mural GCs and cumulus cells (CCs), serve different functions during folliculogenesis. Mural GCs produce oestrogen during the follicular phase and progesterone after ovulation, while CCs surround the oocyte tightly and form the cumulus oophurus and corona radiata inner cell layer. CCs are also engaged in bi-directional metabolite exchange with the oocyte, as they form gap-junctions, which are crucial for both the oocyte’s proper maturation and GC proliferation. However, the function of both GCs and CCs is dependent on proper follicular angiogenesis. Aside from participating in complex molecular interplay with the oocyte, the ovarian follicular cells exhibit stem-like properties, characteristic of mesenchymal stem cells (MSCs). Both GCs and CCs remain under the influence of various miRNAs, and some of them may contribute to polycystic ovary syndrome (PCOS) or premature ovarian insufficiency (POI) occurrence. Considering increasing female fertility problems worldwide, it is of interest to develop new strategies enhancing assisted reproductive techniques. Therefore, it is important to carefully consider GCs as ovarian stem cells in terms of the cellular features and molecular pathways involved in their development and interactions as well as outline their possible application in translational medicine. Full article

We hypothesized that the creation of a 3-dimensional ovarian follicle, with embedded granulosa and theca cells, would better mimic the environment necessary to support early oocytes, both structurally and hormonally. Using a microfluidic system with controlled flow rates, 3-dimensional two-layer (core and shell) capsules were created. The core consists of murine granulosa cells in 0.8 mg/mL collagen + 0.05% alginate, while the shell is composed of murine theca cells suspended in 2% alginate. Somatic cell viability tests and hormonal assessments (estradiol, progesterone, and androstenedione) were performed on days 1, 6, 13, 20, and 27. Confocal microscopy confirmed appropriate compartmentalization of fluorescently-labeled murine granulosa cells to the inner capsule and theca cells to the outer shell. Greater than 78% of cells present in capsules were alive up to 27 days after collection. Artificially constructed ovarian follicles exhibited intact endocrine function as evidenced by the production of estradiol, progesterone, and androstenedione. Oocytes from primary and early secondary follicles were successfully encapsulated, which maintained size and cellular compartmentalization. This novel microfluidic system successfully encapsulated oocytes from primary and secondary follicles, recapitulating the two-compartment system necessary for the development of the mammalian oocyte. Importantly, this microfluidic system can be easily adapted for sterile, high throughput applications. Full article

Lipopolysaccharide (LPS) from gram-negative bacteria was found to be involved in the decrease in laying performance in goose flocks with high stocking density during summer months. LPS injection delayed the increase in the laying rate and altered hierarchical follicle morphology. While there is evidence that LPS exerts suppressive effects on goose reproduction, the time course effects of LPS on the hypothalamus-pituitary-ovary (HPG) axis remain elusive. In this study, we investigated the expression of genes in the HPG axis and the plasma gonadotrophin hormone concentrations in breeding geese at 0, 6, 12, 24, and 36 h after intravenous injection with LPS. The results showed that LPS treatment enhanced and suppressed expression of hypothalamic gonadotropin-inhibiting hormone (GnIH) and gonadotrophin-releasing hormone (GnRH) mRNA, respectively, and similar effects were observed on the mRNA expression of their receptors, GnIHR and GnRHR, in the pituitary. LPS treatment transiently increased follicle FSHβ mRNA expression at 12 h and exerted no significant effect on LHβ mRNA expression in the pituitary. Regardless of the expression of FSHβ and LHβ, plasma follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations were significantly increased during 24–36 h after LPS treatment. In the ovary, StAR and Cyp11a1 were mainly expressed in the granulosa layer (GL) of hierarchical follicles, while Cyp17a1 and Cyp19a1 were mainly expressed in white follicles (WFs) and yellowish follicles (YFs), and to a lesser extent in the theca layer (TL). After LPS treatment, the mRNA levels of Cyp11a1 in the GLs, Cyp17a1 in the WFs and TL, and Cyp19a1 in the WFs, YFs, and TL were significantly decreased. However, LPS treatment transiently upregulated StAR expression at 12 h. These results indicate that the exposure of laying geese to LPS may impair the HPG axis and disturb ovarian steroidogenesis. Our research provides new insights into reproductive dysfunction caused by LPS and the immune challenge in birds. Full article

Development of ovarian follicles requires interactions between granulosa cells, theca cells, and oocytes. Multiple transcription levels are involved but information about the role of noncoding RNAs, especially circular RNAs (circRNAs), is lacking. Here, we used RNA sequencing to profile circRNAs and mRNAs in theca cells from three types of follicle: small yellow follicles (SYF), the smallest hierarchical follicles (F6), and the largest hierarchical follicles (F1). Using bioinformatics analysis, we identified a total of 14,502 circRNAs in all theca cells, with 5622 widely distributed in all stages of development. Differential expression analysis suggested that some genes display differential isoforms during follicular development. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed enrichment of both differentially expressed circRNAs and mRNAs in pathways associated with reproduction, including the TGF-β signaling pathway, oocyte meiosis, and vascular smooth muscle contraction. Our study provides the first visual information about circRNAs and mRNAs in theca cells during follicle development in chickens and adds to the growing body of knowledge about theca cells. Full article