Quercetin activates autophagy to protect rats ovarian granulosa cells from H2O2-induced aging and injury.

Autophagy is closely related to the aging of various organ systems, including ovaries. Quercetin has a variety of biological activities, including potential regulation of autophagy. However, whether quercetin-regulated autophagy activity affects the process of ovarian aging and injury has not been clarified yet. This study explores whether quercetin can resist H2O2-induced aging and injury of granulosa cells by regulating autophagy and its related molecular mechanisms in vitro experiments. The cell viability, endocrine function, cell aging, and apoptosis were detected to evaluate the effects of quercetin and autophagy regulators like 3-methyladenine and rapamycin. The levels of autophagy markers Atg5, Atg12, Atg16L, Lc3B II/I, and Beclin1 were determined by Western blot to assess the effects of quercetin, 3-methyladenine and rapamycin on autophagy. Our results showed quercetin resisted H2O2-induced granulosa cell aging and injury by activating protective autophagy. The treatment of 3-methyladenine and rapamycin confirmed the protective function of autophagy in H2O2-induced granulosa cells. 3-methyladenine treatment inhibited the expression of autophagy markers Atg5, Atg12, Atg16L, Lc3B II/I, and Beclin1 and abolished the positive effects on cell viability, estradiol secretion, and cell apoptosis activated by quercetin. In conclusion, quercetin activates autophagy by upregulating the expression of autophagy-related proteins to resist H2O2-induced aging and injury, which is crucial for stabilizing the function of granulosa cells under oxidative injury conditions and delaying aging. This study may explain the protective effects of quercetin on ovarian aging and injury from the perspective of regulating autophagy.

Resveratrol Protects Rat Ovarian Luteinized Granulosa Cells from H2O2-Induced Dysfunction by Activating Autophagy.

Resveratrol performs a variety of biological activities, including the potential regulation of autophagy. However, it is unclear whether resveratrol protects against luteal dysfunction and whether autophagy involves the regulation of resveratrol. This study aims to investigate whether resveratrol can regulate autophagy to resist H2O2-induced luteinized granulosa cell dysfunction in vitro. Our results showed that resveratrol can enhance cell viability, stimulate the secretion of progesterone and estradiol, and resist cell apoptosis in H2O2-induced luteinized granulosa cell dysfunction. Resveratrol can activate autophagy by stimulating the expression of autophagy-related genes at the transcriptional and translational levels and increasing the formation of autophagosomes and autophagolysosomes. Rapamycin, 3-methyladenine, and bafilomycin A1 regulated the levels of autophagy-related genes in H2O2-induced luteinized granulosa cell dysfunction and further confirmed the protective role of autophagy activated by resveratrol. In conclusion, resveratrol activates autophagy to resist H2O2-induced oxidative dysfunction, which is crucial for stabilizing the secretory function of luteinized granulosa cells and inhibiting apoptosis. This study may contribute to revealing the protective effects of resveratrol on resisting luteal dysfunction from the perspective of regulating autophagy.

Resveratrol Attenuates Hydrogen Peroxide-induced Injury of Rat Ovarian Granulosa-lutein Cells by Resisting Oxidative Stress via the SIRT1/Nrf2/ARE Signaling Pathway.

INTRODUCTION: This paper aims to reveal the molecular mechanism of resveratrol against oxidative stress and cell injury. The ovarian granulosa-lutein cell injury and apoptosis induced by oxidative stress may be responsible for female luteal phase deficiency. The antioxidant function of resveratrol has been confirmed; however, its effect on the expression of antioxidant enzymes and regulatory mechanisms in ovarian granulosa-lutein cells remains unclear. OBJECTIVE: This study aimed to investigate the role of the SIRT1/Nrf2/ARE signaling pathway in the effect of resveratrol on the hydrogen peroxide-induced injury of rat ovarian granulosa-lutein cells. METHODS: In this study, ovarian granulosa-lutein cells extracted from 3-week female SD rats were treated with 200 µM H2O2 in the presence or absence of 20 µM resveratrol. siRNA-SIRT1 and siRNA-Nrf2 were used to inhibit the expression of SIRT1 and Nrf2, respectively. Cell counting kit 8 (CCK-8), cellular morphology, progesterone secretion, and estradiol were used to evaluate cell injury. Hoechst 33258 staining was used to measure cell apoptosis. DHE staining, DCFH-DA staining, malondialdehyde content, protein carbonyl content, total antioxidant capacity and SOD viability were used to estimate the levels of oxidative stress. Western blot analysis was used to detect the levels of apoptosis-related proteins, and SIRT1/Nrf2/ARE signaling pathway-related proteins. RESULTS: The H2O2 treatment-induced rat ovarian granulosa-lutein cells injury was shown as decreased cell viability, impaired cellular morphology, and decreased levels of progesterone and estradiol. The H2O2 treatment also exacerbated cell apoptosis demonstrated as more apoptotic cells stained by Hoechst staining, decreased level of anti-apoptosis protein Bcl-2 and increased level of pro-apoptosis protein Bax. These effects of cell injury and apoptosis induced by H2O2 can be ameliorated by resveratrol. Resveratrol also alleviated oxidative stress induced by H2O2, supported by decreased superoxide anion and cellular total ROS, decreased malondialdehyde and protein carbonyl levels, and increased total antioxidant capacity and SOD viability. Western blot results demonstrated resveratrol reversed the H2O2-induced decrease in levels of antioxidant enzymes containing ARE sequences and activated SIRT1/Nrf2 pathway. Further treatment by siRNA-Nrf2 suggested resveratrol could not activate the expression of antioxidant enzymes under a condition of inhibition of Nrf2. CONCLUSION: This study demonstrates that resveratrol attenuated oxidative stress to protect H2O2-induced rat ovarian granulosa-lutein cell injury and apoptosis via SIRT1/Nrf2/ARE signaling pathway.

Melatonin Attenuates H2O2-Induced Oxidative Injury by Upregulating LncRNA NEAT1 in HT22 Hippocampal Cells.

More research is required to understand how melatonin protects neurons. The study aimed to find out if and how long non-coding RNA (lncRNA) contributes to melatonin's ability to defend the hippocampus from H2O2-induced oxidative injury. LncRNAs related to oxidative injury were predicted by bioinformatics methods. Mouse hippocampus-derived neuronal HT22 cells were treated with H2O2 with or without melatonin. Viability and apoptosis were detected by Cell Counting Kit-8 and Hoechst33258. RNA and protein levels were measured by quantitative real-time PCR, Western blot, and immunofluorescence. Bioinformatics predicted that 38 lncRNAs were associated with oxidative injury in mouse neurons. LncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) was related to H2O2-induced oxidative injury and up-regulated by melatonin in HT22 cells. The knockdown of NEAT1 exacerbated H2O2-induced oxidative injury, weakened the moderating effect of melatonin, and abolished the increasing effect of melatonin on the mRNA and protein level of Slc38a2. Taken together, melatonin attenuates H2O2-induced oxidative injury by upregulating lncRNA NEAT1, which is essential for melatonin stabilizing the mRNA and protein level of Slc38a2 for the survival of HT22 cells. The research may assist in the treatment of oxidative injury-induced hippocampal degeneration associated with aging using melatonin and its target lncRNA NEAT1.