黄柏碱对高糖诱导的晶状体上皮细胞自噬和凋亡影响  

Effect of phellodendrine on the autophagy and apoptosis of lens epithelial cells induced by high glucose

作　　者：李成茂 吴晓玉 李维娜 LI Chengmao;WU Xiaoyu;LI Weina(Ophthalmology Department,910 Hospital of the Joint Support Force of the People’s Liberation Army of China,Quanzhou 362000,China)

机构地区：[1]中国人民解放军联勤保障部队第九一O医院眼科,福建泉州362000

出　　处：《青岛大学学报（医学版）》2024年第5期673-679,共7页Journal of Qingdao University(Medical Sciences)

基　　金：泉州市科技局/卫健委科技计划项目(2021N169S)。

摘　　要：目的探讨黄柏碱对高糖(HG)诱导的晶状体上皮细胞(LEC)自噬和凋亡的影响及机制。方法将HLEB3细胞随机分为对照组(A组)、HG组(B组)、黄柏碱组(C组)、Compound C(AMPK抑制剂)组(D组)。应用CCK-8法检测细胞活力,流式细胞术测定细胞凋亡,活性氧荧光探针(DCFH-DA)法检测细胞内活性氧(ROS)含量,ELISA试剂盒检测细胞内丙二醛(MDA)和超氧化物歧化酶(SOD)水平,Western blot法检测Bcl-2、Bax、Beclin-1、LC3-Ⅱ、LC3-Ⅰ、p-AMPK、AMPK、p-mTOR、mTOR蛋白表达。结果5 mg/L为黄柏碱对HLEB3细胞最佳作用浓度。与A组比较,B组细胞活力、自噬体数量以及SOD、Beclin-1、LC3-Ⅱ/LC3-Ⅰ、Bcl-2、p-AMPK/AMPK水平显著降低(F=30.857~213.971,P<0.001),细胞凋亡率及ROS、MDA、Bax、p-mTOR/mTOR水平显著升高(F=19.111~156.780,P<0.001);与B组相比,C组细胞活力、自噬体数量以及SOD、Beclin-1、LC3-Ⅱ/LC3-Ⅰ、Bcl-2、p-AMPK/AMPK水平显著升高(P<0.05),细胞凋亡率及ROS、MDA、Bax、p-mTOR/mTOR水平显著降低(P<0.05);Compound C可逆转黄柏碱对HLEB3细胞自噬和凋亡的改善作用(P<0.05)。结论黄柏碱能减少HG诱导的LEC氧化应激和凋亡,促进细胞自噬,其作用机制可能与调节AMPK/mTOR信号通路有关。Objective To investigate the effect of phellodendrine on the autophagy and apoptosis of lens epithelial cells(LECs)induced by high glucose(HG)and its mechanism.Methods HLEB3 cells were randomly divided into control group(group A),HG group(group B),phellodendrine group(group C),and Compound C(AMPK inhibitor)group(group D).CCK-8 assay was used to measure cell viability;flow cytometry was used to measure cell apoptosis;the DCFH-DA probe was used to measure the content of reactive oxygen species(ROS)in cells;ELISA kits were used to measure the levels of malondialdehyde(MDA)and superoxide dismutase(SOD);Western blot was used to measure the protein expression levels of B-cell lymphoma-2(Bcl-2),Bcl-2 related X gene(Bax),Beclin-1,microtubule-associated protein 1 light chain 3-Ⅱ(LC3-Ⅱ),microtubule-associated protein 1 light chain 3-Ⅰ(LC3-Ⅰ),AMP-activated protein kinase(AMPK),phosphorylated AMPK(p-AMPK),mammalian target of rapamycin(mTOR),and phosphorylated mTOR(p-mTOR).Results The results showed that 5 mg/L phellodendrine was the best concentration for the culture of HLEB3 cells.Compared with group A,group B had significant reductions in cell viabi-lity,the number of autophagosomes,and the levels of SOD,Beclin-1,LC3-Ⅱ/LC3-Ⅰ,Bcl-2,and p-AMPK/AMPK(F=30.857-213.971,P<0.001)and significant increases in cell apoptosis rate and the levels of ROS,MDA,Bax,and p-mTOR/mTOR(F=19.111-156.780,P<0.001).Compared with group B,group C had significant increases in cell viability,the number of autophagosomes,and the levels of SOD,Beclin-1,LC3-Ⅱ/LC3-Ⅰ,Bcl-2,and p-AMPK/AMPK(P<0.05)and significant reductions in cell apoptosis rate and the levels of ROS,MDA,Bax,and p-mTOR/mTOR(P<0.05).The AMPK inhibitor Compound C could reverse the effect of phellodendrine in improving the autophagy and apoptosis of HLEB3 cells(P<0.05).Conclusion Phellodendrine can reduce the oxidative stress and apoptosis of LEC induced by HG and promote autophagy,possibly by re-gulating the AMPK/mTOR signaling pathway.

关 键 词：黄柏 晶体 上皮细胞 自噬 细胞凋亡 AMP活化蛋白激酶激酶类 TOR丝氨酸-苏氨酸激酶 

分 类 号：R285.5[医药卫生—中药学] R776.1[医药卫生—中医学]