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.
BackgroundThe mechanism of human Umbilical Cord Mesenchymal Stem Cells (hUMSCs) transplantation to improve ovarian function in the rats with Premature Ovarian Insu ciency (POI) is still unclear. The aim of this study is to investigate the signal axis mechanism that is involved in the ovarian function recovery of POI rats following hUMSCs transplantation. MethodsThe rat model with POI was established by intraperitoneal injection of cisplatin. The hUMSCs were transplanted by caudal vein injection into POI rats. Hematoxylin-eosin (H&E) staining was performed to examine the morphology of rat ovarian tissue. Masson staining, Sirus red staining and immuno uorescence were used to observe the brosis extent of ovarian tissue. The levels of serum sex hormones and the expression of brosis related markers in ovarian tissues were measured by enzymelinked immunosorbent assay (ELISA). The expression of NR4A1, Phospho-NR4A1 and AMP-activated protein kinase (AMPK) signaling in rat ovarian tissues was measured by immunohistochemistry and immuno uorescence. The role of AMPK/NR4A1 signaling axis in the regulation of ovarian function recovery in POI rats following hUMSCs transplantation was further investigated by adenovirus and siRNA intervention in isolated stromal cells. ResultsThe results showed that the hUMSCs transplantation signi cantly inhibited ovarian tissue brosis and restored the ovarian function in POI rats. The level of NR4A1 and AMPK expression in ovarian tissue of POI rats after hUMSCs transplantation was signi cantly increased compared with the control group. In the cultured ovarian stromal cells, the similar results were obtained on the expression of NR4A1 and its regulation on brosis related molecular markers in Cisplatin (CDDP) damaged stromal cells following hUMSCs supernatant treatment. Both hUMSCs supernatant treatment and the addition of AMPK inhibitors increased NR4A1 expression in stromal cells. And after NR4A1 molecular intervention, brosisrelated indicators in stromal cells changed. The data suggests that the AMPK/NR4A1 signaling axis is involved in the ovarian function changes in POI rats following hUMSCs transplantation. ConclusionThe data from this study indicate that the inhibition of tissue brosis and recovery of ovarian function is regulated by AMPK/NR4A1 signaling axis in POI rats following hUMSCs transplantation.
Background The mechanism of human Umbilical Cord Mesenchymal Stem Cells (hUMSCs) transplantation to improve ovarian function in the rats with Premature Ovarian Insufficiency (POI) is still unclear. The aim of this study is to investigate the signal axis mechanism that is involved in the ovarian function recovery of POI rats following hUMSCs transplantation. Methods The rat model with POI was established by intraperitoneal injection of cisplatin. The hUMSCs were transplanted by caudal vein injection into POI rats. Hematoxylin-eosin (H&E) staining was performed to examine the morphology of rat ovarian tissue. Masson staining, Sirus red staining and immunofluorescence were used to observe the fibrosis extent of ovarian tissue. The levels of serum sex hormones and the expression of fibrosis related markers in ovarian tissues were measured by enzyme-linked immunosorbent assay (ELISA). The expression of NR4A1, Phospho-NR4A1 and AMP-activated protein kinase (AMPK) signaling in rat ovarian tissues was measured by immunohistochemistry and immunofluorescence. The role of AMPK/NR4A1 signaling axis in the regulation of ovarian function recovery in POI rats following hUMSCs transplantation was further investigated by adenovirus and siRNA intervention in isolated stromal cells. Results The results showed that the hUMSCs transplantation significantly inhibited ovarian tissue fibrosis and restored the ovarian function in POI rats. The level of NR4A1 and AMPK expression in ovarian tissue of POI rats after hUMSCs transplantation was significantly increased compared with the control group. In the cultured ovarian stromal cells, the similar results were obtained on the expression of NR4A1 and its regulation on fibrosis related molecular markers in Cisplatin (CDDP) damaged stromal cells following hUMSCs supernatant treatment. Both hUMSCs supernatant treatment and the addition of AMPK inhibitors increased NR4A1 expression in stromal cells. And after NR4A1 molecular intervention, fibrosis-related indicators in stromal cells changed. The data suggests that the AMPK/NR4A1 signaling axis is involved in the ovarian function changes in POI rats following hUMSCs transplantation. Conclusion The data from this study indicate that the inhibition of tissue fibrosis and recovery of ovarian function is regulated by AMPK/NR4A1 signaling axis in POI rats following hUMSCs transplantation.
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