Dataset Information

Proteomics detection of mouse embryonic 17.5 dpc cultured ovaries in vitro

ABSTRACT: In this project, the ovaries of mice at 17.5 dpc embryonic stage were collected, and KN93, an inhibitor of CAMKII, and IWP2, an inhibitor of Wnt5a, were added for in vitro culture. Proteins affected by KN93 and IWP2, which were regulated by CAMKII and Wnt5a, were detected by proteomics technology, so as to identify the target proteins regulated by CAMKII and Wnt5a. To analyze the possible mechanisms of CAMKII and Wnt5a on the development of primordia follicles in mouse ovaries.

ORGANISM(S): Mus Musculus

SUBMITTER: Guoliang Xia

PROVIDER: PXD035136 | iProX | Thu Jul 07 00:00:00 BST 2022

REPOSITORIES: iProX

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Publications

Effects and potential mechanism of Ca<sup>2+</sup>/calmodulin‑dependent protein kinase II pathway inhibitor KN93 on the development of ovarian follicle.

Yu Jianjie J Xie Xianguo X Ma Yabo Y Yang Yi Y Wang Chao C Xia Guoliang G Ding Xiangbin X Liu Xinfeng X

International journal of molecular medicine 20220805 4

Adequate regulation of the speed of follicular development has been reported to prolong the reproductive life of the ovary. The aim of the present study was to assess the potential effects and mechanism of the Ca<sup>2+</sup>/calmodulin‑dependent protein kinase II (CaMKII) pathway on the development of ovarian follicle. In the present study, the expression of CaMKII was measured in the ovary of mice at different developmental stages by immunofluorescence, confirming that CaMKII has a role in fol ...[more]

PMID: 35929517

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Project description:The oocytes found within the primordial follicles of mammalian ovaries remain quiescent for months to years until they receive the appropriate signals to undergo the primordial to primary follicle transition and initiate folliculogenesis. The molecular mechanisms and extracellular signaling factors that regulate this process remain to be fully elucidated. The current study investigates the mechanisms utilized by anti-MÃ¼llerian hormone (AMH; i.e. MÃ¼llerian inhibitory substance) to inhibit the primordial to primary follicle transition. Ovaries from 4-day-old rats were placed into organ culture and incubated in the absence or presence of AMH, either alone or in combination with known stimulators of follicle transition, including basic fibroblast growth factor (bFGF), kit ligand (KITL), or keratinocyte growth factor (KGF). Following 10 days of culture, the ovaries were sectioned, stained, and morphologically evaluated to determine the percentage of primordial versus developing follicles. As previously demonstrated, AMH treatment decreased primordial to primary follicle transition. Interestingly, AMH inhibited the stimulatory actions of KITL, bFGF, and KGF. Therefore, AMH can inhibit the basal and stimulated development of primordial follicles. To investigate the mechanism of AMH actions, the influence AMH has on the ovarian transcriptome was analyzed. AMH treatment when compared with controls was found to alter the expression of 707 genes. The overall effect of AMH exposure is to decrease the expression of stimulatory factors, increase the expression of inhibitory factors, and regulate cellular pathways (e.g. transforming growth factor beta signaling pathway) that result in the inhibition of primordial follicle development. Analysis of the regulatory factors and cellular pathways altered by AMH provides a better understanding of the molecular control of primordial follicle development. Experiment Overall Design: RNA samples from two control groups (pooled untreated cultured ovaries) are compared to two treated groups (pooled cultured ovaries treated with AMH)

Project description:Exposure to persistent organic pollutants (POPs), such as dichlorodiphenyltrichloroethane (DDT) and polychlorinated biphenyls (PCBs), has historically been linked to population collapses in wildlife. Despite international regulations, these legacy chemicals are still currently detected in women of reproductive age, and their levels correlate with reduced ovarian reserve, longer time-to-pregnancy, and higher risk of infertility. However, the specific modes of action underlying these associations remain unclear. Here, we examined the effects of five commonly occurring POPs − hexachlorobenzene (HCB), p,p'-dichlorodiphenyldichloroethylene (DDE), 2,3,3′,4,4′,5-hexachlorobiphenyl (PCB156), 2,2′,3,4,4′,5,5′-heptachlorobiphenyl (PCB180), perfluorooctane sulfonate (PFOS) − and their mixture on human ovaries in vitro. We exposed human ovarian cancer cell lines COV434, KGN, and PA1 as well as primary ovarian cells for 24 h, and ovarian tissue containing unilaminar follicles for 6 days. RNA-sequencing of samples exposed to concentrations covering epidemiologically relevant levels revealed significant gene expression changes related to central energy metabolism in the exposed cells, indicating glycolysis, oxidative phosphorylation, fatty acid metabolism, and reactive oxygen species as potential shared targets of POP exposures in ovarian cells. Alpha-enolase (ENO1), lactate dehydrogenase A (LDHA), cytochrome C oxidase subunit 4I1 (COX4I1), ATP synthase F1 subunit alpha (ATP5A), and glutathione peroxidase 4 (GPX4) were validated as targets through qPCR in additional cell culture experiments in KGN. In ovarian tissue cultures, we observed significant effects of exposure on follicle growth and atresia as well as protein expression. All POP exposures, except PCB180, decreased unilaminar follicle proportion and increased follicle atresia. Immunostaining confirmed altered expression of LDHA, ATP5A, and GPX4 in the exposed tissues. Moreover, POP exposures modified ATP production in KGN and tissue culture. In conclusion, our results demonstrate the disruption of cellular energy metabolism as a novel mode of action underlying POP-mediated interference of follicle growth in human ovaries.

Dataset Information

Proteomics detection of mouse embryonic 17.5 dpc cultured ovaries in vitro

Publications

Effects and potential mechanism of Ca<sup>2+</sup>/calmodulin‑dependent protein kinase II pathway inhibitor KN93 on the development of ovarian follicle.

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