| 江雪娟,陈晓菲,俞佳,等.槲皮素对多囊卵巢综合征大鼠卵巢颗粒细胞增殖与凋亡的影响研究[J].浙江中医药大学学报,2022,46(10):1055-1062, 1069. |
| 槲皮素对多囊卵巢综合征大鼠卵巢颗粒细胞增殖与凋亡的影响研究 |
| Effect of Quercetin on Proliferation and Apoptosis of Ovarian Granulosa Cells in Rats with Polycystic Ovarian Syndrome |
| DOI：10.16466/j.issn1005-5509.2022.10.001 |
| 中文关键词: 槲皮素 多囊卵巢综合征 胰岛素抵抗 细胞活力 二甲双胍 细胞增殖 细胞凋亡 AGEs/RAGE |
| 英文关键词: quercetin polycystic ovarian syndrome insulin resistance cell viability metformin cell proliferation cell apoptosis AGEs/RAGE |
| 基金项目:杭州市卫生科技计划项目(ZD20200016、ZD20210029、OO2019011、OO2019008)；浙江省中医药科学研究基金项目(2021ZB210)；杭州市红十字会医院院内科研基金项目(HHQN2021005)；浙江省自然科学基金项目(LQ19H270005) |
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| 中文摘要: |
| [目的] 探究槲皮素(quercetin，Q)对多囊卵巢综合征(polycystic ovarian syndrome，PCOS)大鼠卵巢颗粒细胞增殖与凋亡的影响及其作用机制。[方法] 建立PCOS大鼠模型，提取培养卵巢颗粒细胞进行鉴定，并分为PCOS组、PCOS+4-氯-N-环己基-N-(苯基甲基)苯甲酰胺(N-benzyl-4-chloro-N-cyclohexylbenzamide，FZ)组、PCOS+L-Q(槲皮素0.5 μg·mL-1)组、PCOS +H-Q(槲皮素1 μg·mL-1)组、PCOS+H-Q+FZ组、PCOS+盐酸二甲双胍(metformin hydrochloride，MH)组，并取正常颗粒细胞作为对照组。以四甲基偶氮唑盐[3-(4，5)-dimethylthiahiazo(-z-y1)-3，5-di-phenytetrazoliumromide，MTT]法检测细胞活力，酶联免疫吸附试验(enzyme linked immunosorbent assay，ELISA)检测细胞上清液中雌二醇(estradiol，E2)、孕酮(progesterone，P)、促卵泡生成素(follicle-stimulating hormone，FSH)、黄体生成素(luteinizing hormone，LH)、雄激素(testosterone，T)含量，流式细胞术测定细胞凋亡率，并进行活性氧(reactive oxygen species，ROS)活性检测。实时荧光定量聚合酶链式反应(Real-time quantitative polymerase chain reaction，Real-time qPCR)检测细胞中晚期糖基化终末产物受体(receptor of advanced glycation end products，RAGE)、Bax、Bcl-2及炎性介质高迁移率族蛋白B1(high mobility group protein B1，HMGB1)的mRNA表达情况，免疫印迹法检测细胞中晚期糖基化终末产物/晚期糖基化终末产物受体(advanced glycation end products/receptor of advanced glycation end products，AGEs/RAGE)及细胞外信号调节蛋白激酶/有丝分裂原蛋白激酶(extracellular signal-regulated kinase/mitogen-activated protein kinase，ERK/MAPK)相关蛋白表达情况。[结果] 分离的大鼠卵巢颗粒细胞鉴定正常。MTT结果显示，槲皮素在0～1 μg·mL-1时对正常颗粒细胞活力无影响，对于PCOS组细胞，槲皮素作用后各组细胞活力显著升高(P＜0.05，P＜0.01)。与PCOS组比较，PCOS+FZ组、PCOS+H-Q组、PCOS+H-Q+FZ组和PCOS+MH组细胞中E2、LH、T含量以及ROS活性均显著降低(P＜0.05，P＜0.01)，PCOS+H-Q组、PCOS+H-Q+FZ组、PCOS+MH组中RAGE、HMGB1、Bax的mRNA，AGEs和RAGE的蛋白表达水平均显著降低(P＜0.05，P＜0.01)，Bcl-2的mRNA及p-ERK、p-p38 MAPK蛋白表达水平均显著升高(P＜0.05，P＜0.01)。 [结论] 槲皮素能够通过降低AGEs/RAGE通路活性，促进PCOS大鼠卵巢颗粒的增殖，抑制细胞凋亡来改善PCOS。 |
| 英文摘要: |
| [Objective] To investigate the effects of quercetin(Q) on proliferation and apoptosis of ovarian granulosa cells in rats with polycystic ovarian syndrome(PCOS) and its mechanism. [Methods] After PCOS rat model was established， ovarian granulosa cells were extracted and identified. Then the ovarian granulosa cells were divided into PCOS group， PCOS+N-benzyl-4-chloro-N-cyclohexylbenzamide(FZ) group， PCOS+ L-Q(quercetin 0.5 μg·mL-1) group， PCOS+H-Q(quercetin 1 μg·mL-1) group， PCOS+H-Q+FZ group， and PCOS+metformin hydrochloride(MH) group， and normal granulosa cells as control group. Cell viability was detected by 3-(4，5)-dimethylthiahiazo(-z-y1)-3，5-di-phenytetrazoliumromide(MTT)， the contents of estradiol(E2)， progesterone(P)， follicle-stimulating hormone(FSH)， luteinizing hormone(LH) and testosterone(T) in cell supernatant were determined by enzyme linked immunosorbent assay(ELISA)， and cell apoptosis rate was determined by flow cytometry， and reactive oxygen species(ROS) activity was detected. The mRNA expressions of receptor of advanced glycation end products(RAGE)， Bax， Bcl-2 and inflammatory medium high mobility group protein B1(HMGB1) were detected by Real-time quantitative polymerase chain reaction(Real-time qPCR)， and the protein expressions of advanced glycation end products/receptor of advanced glycation end products(AGEs/RAGE) and extracellular signal-regulated kinase/mitogen-activated protein kinase(ERK/MAPK) related proteins were detected by Western blot. [Results] The isolated rat ovarian granulosa cells were successfully identified. MTT results showed that quercetin had no effect on viability of normal granulosa cells at 0～1 μg·mL-1， while for PCOS group， the viability of each group was significantly increased(P＜0.05， P＜0.01). Compared with PCOS group， E2， LH and T contents and ROS activity in PCOS+ FZ group， PCOS+H-Q group， PCOS+H-Q+FZ group and PCOS+MH group were significantly decreased(P＜0.05， P＜0.01). In PCOS+H-Q group， PCOS+H-Q+FZ group and PCOS+MH group， the mRNA expression levels of RAGE， HMGB1， Bax and protein expression levels of AGEs and RAGE were significantly decreased(P＜0.05， P＜0.01)， while the mRNA expression levels of Bcl-2 and p-ERK and p-p38 MAPK protein levels were significantly increased(P＜0.05， P＜0.01). [Conclusion] Quercetin can relieve PCOS by decreasing the activity of AGEs/RAGE pathway， promoting the proliferation of granulosa cells and inhibiting the apoptosis of PCOS rats. |
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