嗜铁蛋白对血管内皮细胞的损伤作用及葛根素的保护机制

Protective effects and mechanisms of puerarin on alleviating siderocalin-induced vascular endothelial cell injury

目的探讨葛根素对嗜铁蛋白诱导EA.hy926型血管内皮细胞损伤的保护作用及机制。方法嗜铁蛋白以不同作用时间(0、24、48、72 h)及浓度(0、5、10、20μmol·L-1)刺激EA.hy926细胞,MTT法及流式细胞术分别测细胞增殖及凋亡,比色法测乳酸脱氢酶(LDH)活性、ELISA测高敏C反应蛋白(hs-CRP)及单核细胞趋化蛋白-1(MCP-1)水平,Western blot测p-p38 MAPK、t-p38 MAPK、Bax、Bcl-2蛋白表达,加入葛根素(10、50、100μmol·L-1)和p38 MAPK抑制剂SB203580(20μmol·L-1)检测葛根素的干预作用及机制。结果与空白组比较,10μmol·L-1嗜铁蛋白作用48 h可显著抑制EA.hy926细胞增殖,上调p-p38 MAPK及Bax蛋白表达促进细胞凋亡,诱导分泌hs-CRP及MCP-1,并增加LDH活性(P<0.05);50、100μmol·L-1葛根素均可显著减轻嗜铁蛋白诱导的EA.hy926细胞增殖抑制、下调p-p38 MAPK及Bax蛋白表达以抑制细胞凋亡、减少嗜铁蛋白诱导EA.hy926细胞分泌hs-CRP及MCP-1并降低LDH活性(均P<0.01),且较SB203580更为显著(均P<0.05)。结论嗜铁蛋白可诱导血管内皮细胞损伤,葛根素可抑制p38 MAPK通路减轻嗜铁蛋白诱导的血管内皮细胞损伤。

AIM To induced vascular endothelial investigate the effects and the mechanism of puerarin in alleviating siderocalin- cells （EA.hy926） injury. METHODS Siderocalin at different concentration （0, 5, 10, 20 μmol.L-1） or different time （0, 24, 48, and apoptosis were measured by MTF assay and flow determined by colorimetry, contents of high-sensitivity 72 h） were added in EA.hy 926 cells, the proliferation cytometry, lactic dehydrogenase （LDH） activities were C- reactive protein （hs- CRP） and monocyte chemotacticprotein- 1 （MCP- 1） were quantified by ELISA and the expressions of p- p38 MAPK, t- p38 MAPK, Bax and Bcl-2 in EA.hy 926 ceils were subjected by Western blot analysis. Then, puerarin （10, 50, 100 μmol.L-1） or SB203580 （20 μmol .L-1） was added to observe the effects and mechanism of puerarin on siderocalin-induced EA.hy926 cells injury. RESULTS Compared with the untreated EA.hy926 cells, 10 μ mol.L-1 siderocalin significantly increased the apoptosis, LDH activities, hs-CRP and MCP-l contents, p-p38 MAPK and Bax expressions but restrained the proliferation in EA.hy926 cells （P 〈 0.05）. Compared with the siderocalin- induced EA.hy926 cells, 50 and 100 μmol.L-1 puerarin significantly restrained the apoptosis, LDH activities, hs-CRP and MCP-1 contents, p-p38 MAPK and Bax expressions but increased the proliferation in EA.hy926 cells （P 〈 0.01） , and had significant difference with the SB203580 treatment （P 〈 0.05）. CONCLUSION Siderocalin induces vascular endothelial cells injury. Puerarin could alleviate siderocalin- induced vascular endothelial cells injury via the inhibition of p38 MAPK pathway.