Dataset Information

Identification of a New Theca/Interstitial Cell-Specific Gene and Its Biological Role in Growth of Mouse Ovarian Follicles at the Gonadotropin-Independent Stage.

ABSTRACT: Theca/interstitial cells are responsible for the growth and maturation of ovarian follicles. However, little is known about the theca/interstitial cell-specific genes and their functions. In this study, we explored transcriptomes of theca/interstitial cells by RNA-seq, and the novel biological roles of a theca cell marker, asporin (Aspn)/periodontal ligament-associated protein 1 (PLAP-1). RNA-seq detected 432 and 62 genes expressed specifically in theca/interstitial cells and granulosa cells isolated from 3-weeks old mouse ovaries. Gene ontology analysis demonstrated that these genes were largely categorized into four major groups: extracellular matrix organization-related terms, chemotaxis-related terms, the angiogenesis-related terms, and morphogenesis-related terms. In situ hybridization demonstrated that the newly detected representative gene, Aspn/PLAP-1, was detected specifically in the outer layer of theca cells in contrast with the expression of the basal lamina-specific gene, Nidgen-1. Intriguingly, an Aspn/PLAP-1 antibody completely arrested the growth of secondary follicles that is the gonadotropin-independent follicle developmental stage. Furthermore, transforming growth factor-? (TGF-?)-triggered signaling was induced by the Aspn/PLAP-1 antibody treatment, which is consistent with the inhibitory effect of Aspn/PLAP-1 on TGF-?. Altogether, these results suggest that theca cells are classified into subpopulations on the basis of new marker genes and their biological functions, and provide evidence that Aspn/PLAP-1 is expressed exclusively in the outer layer of theca cells and plays a pivotal role in the growth of secondary follicles via downregulation of the canonical TGF-? signaling cascade.

SUBMITTER: Aoyama M

PROVIDER: S-EPMC6702446 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Identification of a New Theca/Interstitial Cell-Specific Gene and Its Biological Role in Growth of Mouse Ovarian Follicles at the Gonadotropin-Independent Stage.

Aoyama Masato M Shiraishi Akira A Matsubara Shin S Horie Kaoru K Osugi Tomohiro T Kawada Tsuyoshi T Yasuda Keiko K Satake Honoo H

Frontiers in endocrinology 20190814

Theca/interstitial cells are responsible for the growth and maturation of ovarian follicles. However, little is known about the theca/interstitial cell-specific genes and their functions. In this study, we explored transcriptomes of theca/interstitial cells by RNA-seq, and the novel biological roles of a theca cell marker, asporin (Aspn)/periodontal ligament-associated protein 1 (PLAP-1). RNA-seq detected 432 and 62 genes expressed specifically in theca/interstitial cells and granulosa cells iso ...[more]

PMID: 31474939

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Project description:The theca interna layer of the ovarian follicle forms during the antral stage of follicle development and lies adjacent to and directly outside the follicular basal lamina. It supplies androgens and communicates with the granulosa cells and the oocyte by extracellular signaling. To better understand developmental changes in the theca interna, we undertook transcriptome profiling of the theca interna from small (3-5 mm, n = 10) and large (9-12 mm, n = 5) healthy antral bovine follicles, representing a calculated >7-fold increase in the amount of thecal tissue. Principal Component Analysis and hierarchical classification of the signal intensity plots for the arrays showed no clustering of the theca interna samples into groups depending on follicle size or subcategories of small follicles. From the over 23,000 probe sets analysed, only 76 were differentially expressed between large and small healthy follicles. Some of the differentially expressed genes were associated with processes such as myoblast differentiation, protein ubiquitination, nitric oxide and transforming growth factor β signaling. The most significant pathway affected from our analyses was found to be Wnt signaling, which was suppressed in large follicles via down-regulation of WNT2B and up-regulation of the inhibitor FRZB. These changes in the transcriptional profile could have been due to changes in cellular function or alternatively since the theca interna is composed of a number of different cell types it could have been due to any systematic change in the volume density of any particular cell type. However, our study suggests that the transcriptional profile of the theca interna is relatively stable during antral follicle development unlike that of granulosa cells observed previously. Thus both the cellular composition and cellular behavior of the theca interna and its contribution to follicular development appear to be relatively constant throughout the follicle growth phase examined.

Project description:LH triggers the biosynthesis of androgens in the theca-interstitial (T-I) cells of ovary through the activation of a cAMP-dependent pathway. We have previously shown that LH/human chorionic gonadotropin (hCG) activates mammalian target of rapamycin complex 1 (mTORC1) signaling network, leading to cell proliferation. In the present study, we provide evidence that the LH/hCG-mediated activation of the mTORC1 signaling cascade is involved in the regulation of steroidogenic enzymes in androgen biosynthesis. Treatment with LH/hCG increased the expression of downstream targets of mTORC1, ribosomal protein S6 kinase 1, and eukaryotic initiation factor 4E as well as steroidogenic enzymes. LH/hCG-mediated stimulation of the steroidogenic enzyme mRNA was blocked by the mTORC1 inhibitor, rapamycin. This inhibitory effect was selective because rapamycin failed to block hCG-mediated increase in the expression of Star mRNA levels. Furthermore, pharmacological targeting of mTORC1 with rapamycin also blocked LH/hCG- or forskolin-induced expression of cAMP response element-binding protein (CREB) and steroidogenic enzymes (P450 side-chain cleavage enzyme, 3?-hydroxysteroid dehydrogenase type 1, and 17?-hydroxylase/17,20 lyase) but produced no effect on steroidogenic acute regulatory protein levels. These results were further confirmed by demonstrating that the knockdown of mTOR using small interfering RNA selectively abrogated the LH/hCG-induced increase in steroidogenic enzyme expression, without affecting steroidogenic acute regulatory protein expression. LH/hCG-stimulated androgen production was also blocked by rapamycin. Furthermore, the pharmacological inhibition of mTORC1 or ribosomal protein S6 kinase 1 signaling prevented the LH/hCG-induced phosphorylation of CREB. Chromatin immunoprecipitation assays revealed the association of CREB with the proximal promoter of the Cyp17a1 gene in response to hCG, and this association was reduced by rapamycin treatment. Taken together, our findings show for the first time that LH/hCG-mediated activation of androgen biosynthesis is regulated by the mTORC1 signaling pathway in T-I cells.

Project description:Background Human umbilical cord mesenchymal stem cells (hUCMSCs, retrospectively registered) have a lot of promise for treating theca interstitial cells(TICs) dysfunction in premature ovarian insufficiency (POI). The mechanisms, however, are still unknown. Methods To examine the therapeutic and find the cause, we used both in vivo cisplatin-induced POI rat model and in vitro TICs model. HUCMSCs were injected into the tail veins of POI rats in an in vivo investigation. Then, using ELISA, HE staining, TUNEL apoptosis test kit, immunohistochemistry and western blot, researchers examined hormonal levels, ovarian morphology, TICs apoptosis, NR4A1 and Cyp17a1 in response to cisplatin treatment and hUCMSCs. TICs were obtained from the ovaries of rats and treated with the cisplatin, hUCMSCs supernatant, and the antagonist of NR4A1——DIM-C-pPhOH. ELISA, immunofluorescence, flow cytometry, JC-1 labeling and western blot analysis were used to detect T levels, Cyp17a1, NR4A1, and the anti-apoptotic protein Bcl-2, as well as pro-apoptotic proteins Bax, caspase-9, caspase-3, and cytochrome C(cytc). Results We discovered that hUCMSCs restored the ovarian function, particularly TICs function based on measures of Cyp17a1 and T expression. NR4A1 was found in ovarian TICs of each group and NR4A1 expression was lower in the POI rats but higher following hUCMSCs therapy. The apoptosis of TICs generated by cisplatin was reduced after treatment with hUCMSCs. In vitro, NR4A1 was expressed in the nucleus of TICs, and NR4A1 as well as phospho-NR4A1 were decreased, following the apoptosis of TICs was emerged after cisplatin treatment. Interestingly, the localization of NR4A1 was translocated from the nucleus to the cytoplasm due to cisplatin. HUCMSCs were able to boost NR4A1 and phospho-NR4A1 expression while TICs’ apoptosis and JC-1 polymorimonomor fluorescence ratios reduced. Furthermore, Bcl-2 expression dropped following cisplatin treatment, whereas Bax, cytc, caspase-9, and caspase-3 expression rose; however, hUCMSCs treatment reduced their expression. In addition, DIM-C-pPhOH had no effect on the NR4A1 expression, but it did increase the expression of apoptosis-related factors such as Bax, cytc, caspase-9, and caspase-3, causing the apoptosis of TICs. Conclusions These data show that hUCMSCs therapy improves ovarian function in POI rats by inhibiting TICs apoptosis through regulating NR4A1 -mediated mitochondrial mechanisms. Supplementary Information The online version contains supplementary material available at 10.1186/s12958-022-00992-5.

Dataset Information

Identification of a New Theca/Interstitial Cell-Specific Gene and Its Biological Role in Growth of Mouse Ovarian Follicles at the Gonadotropin-Independent Stage.

Publications

Identification of a New Theca/Interstitial Cell-Specific Gene and Its Biological Role in Growth of Mouse Ovarian Follicles at the Gonadotropin-Independent Stage.

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