NE激活Nrf2/HO-1信号通路调节人子宫内膜上皮细胞的氧化应激  

NE activates Nrf2/HO-1 signaling pathway to regulate oxidative stress in human endometrial epithelial cells

作　　者：杨雪 马瑞欣 李佳鑫 孔雪瑞 李军平[1] 罗彦[1,2] Yang Xue;Ma Ruixin;Li Jiaxin;Kong Xuerui;Li Junping;Luo Yan(Basic Medical College,Ningxia Medical University,Yinchuan 750000;Basic Medical College,Zhaoqing Medical College,Zhaoqing 526000;Dept of Ophthalmology,People′s Hospital of Wuzhong Attached to The Ningxia Medical University,Wuzhong 751100;Dept of Gerontology,Xiangya Hospital,Central South University,Changsha 410000)

机构地区：[1]宁夏医科大学基础医学院,银川750000 [2]肇庆医学高等专科学校基础医学院,肇庆526000 [3]宁夏医科大学附属吴忠市人民医院眼科,吴忠751100 [4]中南大学湘雅医院老年医学科,长沙410000

出　　处：《安徽医科大学学报》2024年第5期767-773,共7页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81760265);宁夏自然科学基金(编号:2023AAC03218)。

摘　　要：目的明确去甲肾上腺素(NE)是否能通过核因子E2相关因子2(Nrf2)和血红素加氧酶-1(HO-1)调控培养的人子宫内膜上皮细胞(hEECs)的氧化应激状态。方法培养hEECs,通过逆转录多聚酶链式反应(RT-PCR)检测α和β肾上腺素能受体在hEECs的表达;细胞计数试剂盒-8(CCK-8)实验确定NE处理对细胞活性的影响,根据结果将细胞分为处理组和对照组,选择合适浓度进行处理;通过Western blot实验,检测闭锁蛋白(Occludin),闭锁小带蛋白-1(ZO-1),凋亡相关蛋白B细胞淋巴瘤-2蛋白(Bcl-2),Bcl-2相关X蛋白(Bax),抗氧化应激蛋白Nrf2、HO-1的表达,流式细胞术检测NE处理后细胞凋亡的情况;酶联免疫吸附试验试剂盒(ELISA)检测丙二醛(MDA)、超氧化物歧化酶(SOD)的水平。结果在hEECs上检测到α1 a、α1 b、α2 a、α2 b、α2 c、β1、β3肾上腺素能受体的表达。NE处理后,在低浓度(5、10μmol/L)不明显影响细胞活力,而15、20μmol/L NE处理细胞6 h或24 h均明显增加细胞活性。紧密连接蛋白ZO-1和Occludin在15μmol/L 24 h处理组显著升高。ZO-1在6 h处理组下降,15μmol/L处理显著下降,同时Occludin在6 h处理组升高。NE处理后与对照相比,细胞凋亡率升高,15μmol/L NE处理细胞6 h凋亡率显著升高,处理24 h后细胞凋亡率有所下降,凋亡相关蛋白Bcl-2/Bax>1。NE处理后,上调Nrf2及HO-1,细胞培养基上清液中的MDA水平没有明显升高,同时SOD水平明显上调。结论NE处理细胞后可通过激活Nrf2/HO-1信号,上调SOD,发挥抗氧化应激、抗凋亡的作用。Objective To investigate whether norepinephrine(NE)regulates the oxidative stress in human endometrial epithelial cells(hEECs)by activating nuclear factor E2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)signal pathway.Methods Cultured hEECs were used.The expression ofαandβadrenergic receptors was detected by reverse transcription-polymerase chain reaction(RT-PCR).Cell counting kit-8(CCK-8)assay was applied to test the effect of NE on cell viability,then the cells were divided into Control group and NE treatment group,and the appropriate concentrations were chosen.The expression of tight junction proteins Occludin and zona occludens-1(ZO-1),apoptosis-related proteins apoptosis-related protein B-cell lymphoma-2 protein(Bcl-2)and Bcl-2 associated X protein(Bax),antioxidant proteins Nrf2 and HO-1 were examined by Western blot.The apoptosis was detected by flow cytometry.The malonaldehyde(MDA)and superoxide dismutase(SOD)in the cell culture medium were detected by enzyme-linked immunosorbent assays kit(ELISA).Results The mRNA expression ofα1 a,α1 b,α2 a,α2 b,α2 c,β1,β3 was detected in the hEECs.After the NE treatment,no significant change in cell viability was observed in low concentration(5μmol/L and 10μmol/L)groups,while 15μmol/L and 20μmol/L NE treatments for 6 h or 24 h promoted the cell viability significantly.The expression of ZO-1 and Occludin increased significantly in 15μmol/L group after 24 h treatment,the expression of ZO-1 decreased in 6 h treatment group,significant down regulation was observed after 15μmol/L NE application,the expression of Occludin increased in 6 h group.The cell apoptosis increased compared with the control group after NE stimulation,especially a significantly increase in 6 h 15μmol/L group was detected,while the fall in increased apoptosis was observed after 24 h treatment.The ration of Bcl-2/Bax>1.The expression of Nrf2 and HO-1 was elevated by NE.There was no obvious change in MDA level while significant elevation in SOD was detected in cell culture medium.Conclusion

关 键 词：去甲肾上腺素 NRF2 HO-1 人子宫内膜上皮细胞 ZO-1 OCCLUDIN Bax Bcl-2 

分 类 号：R339.2[医药卫生—人体生理学]