Hydrogen sulfide decreases high glucose/palmitate-induced autophagy in endothelial cells by the Nrf2-ROS-AMPK signaling pathway.
Hydrogen sulfide decreases high glucose/palmitate-induced autophagy in endothelial cells by the Nrf2-ROS-AMPK signaling pathway.
复制标题
硫化氢通过 Nrf2-ROS-AMPK 信号通路降低高葡萄糖/棕榈酸酯诱导的内皮细胞自噬
DOI：
10.1186/s13578-016-0099-1
复制
发表时间：
2016
影响因子：
7.5
通讯作者：
Zhang W
中科院分区：
文献类型：
--
作者：
Liu J;Wu J;Sun A;Sun Y;Yu X;Liu N;Dong S;Yang F;Zhang L;Zhong X;Xu C;Lu F;Zhang W
背景过度氧化应激诱导的过度自噬是糖尿病血管内皮细胞功能障碍的主要原因。硫化氢具有抗氧化作用，但其对血管内皮细胞过度自噬的调节尚不清楚。方法采用40 mM葡萄糖和500μM棕榈酸酯处理的db/db小鼠(28周龄)和大鼠主动脉内皮细胞(RAECs)分别作为II型糖尿病动物模型和细胞模型，以100μMNaHS作为外源性H 2S供体。用末端脱氧核苷酸转移酶介导的dUTP缺口末端标记法和Hoechst 33342/PI染色检测细胞的凋亡率。测定超氧化物歧化酶(SOD)、过氧化氢酶(CAT)和呼吸复合体活性。实时荧光定量聚合酶链式反应检测超氧化物歧化酶和过氧化氢酶的基因表达水平。用AMPK-siRNA检测AMPK对自噬的影响。结果体内外H_2S生成均明显减少(P<0.05，P<0.01)，NaHS治疗可明显改善这种损伤(P<0.05，P<0.01)。体内外黏附蛋白表达均增加(P<0.05，P<0.01)，NaHS表达减弱(P<0.05，P<0.01)。NaHS可保护内皮细胞免受II型糖尿病诱导的细胞凋亡(P<0.05，P<0.01)。此外，糖尿病II组血管内皮细胞中超氧化物歧化酶和过氧化氢酶的表达和活性受损(P<0.05，P<0.01)；NaHS治疗可减轻这种损伤(P<0.05)。NaHS还增加了ATP的产生(p<0.05)和呼吸复合体的活性(p<0.05)，p-AMPK/AMPK的比值也被NaHS降低(p<0.01)。NaHS处理和AMPK-siRNA处理也降低了内皮细胞的自噬水平(P<0.05，P<0.01)。结论外源性硫化氢可能通过抑制Nrf2-ROS-AMPK信号通路，抑制氧化应激诱导的过度自噬，从而对动脉内皮细胞产生保护作用。
BackgroundExcessive autophagy induced by extravagant oxidative stress is the main reason for diabetes-induced vascular endothelial cells dysfunction. Hydrogen sulfide (H2S) has anti-oxidative effects but its regulation on excessive autophagy of vascular endothelial cells is unclear.MethodsIn this study, aorta of db/db mice (28 weeks old) and rat aortic endothelial cells (RAECs) treated with 40 mM glucose and 500 μM palmitate acted as type II diabetic animal and cellular models, respectively, and 100 μMNaHS was used as an exogenous H2S donor. The apoptosis level was measured by terminal deoxynucleotidyl transferase mediated dUTP nick-end labeling (TUNEL) staining and Hoechst 33342/PI staining. The activities of SOD, CAT and respiratory complexes were also measured. The mRNA levels of SOD and CAT were detected by real-time PCR. AMPK-siRNA was used to detect the effect of AMPK on autophagy. Western blotting was used to detected the protein level.ResultsH2S production was decreased (p < 0.05, p < 0.01) both in vitro and in vivo; NaHS treatment rescued this impairment (p < 0.05, p < 0.01). The expression of adhesive proteins was increased (p < 0.05, p < 0.01) both in vitro and in vivo; NaHS attenuated (p < 0.05, p < 0.01) these alterations. NaHS could protect endothelial cells against apoptosis induced by type II diabetes (p < 0.05, p < 0.01). Furthermore, the expressions and activities of SOD and CAT were impaired (p < 0.05, p < 0.01) in endothelial cells of diabetes II; NaHS treatment attenuated (p < 0.05) this impairment. NaHS also increased ATP production (p < 0.05) and activities of respiratory complexes (p < 0.05), and the ratio of p-AMPK to AMPK was also decreased by NaHS (p < 0.01). The level of autophagy in endothelial cells was also decreased (p < 0.05, p < 0.01) by NaHS treatment and AMPK-siRNA treatment. The expression of Nrf2 in the nuclei was increased (p < 0.05) by NaHS treatment.ConclusionExogenous H2S might protect arterial endothelial cells by suppressing excessive autophagy induced by oxidative stress through the Nrf2-ROS-AMPK signaling pathway.