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目的监测在缺氧条件下进行枸杞多糖处理后肺血管平滑肌细胞中沉默接合型信息调节因子2同源蛋白1(SIRT1)、基质金属蛋白酶9(MMP-9)及缺氧诱导因子1α(HIF-1α)的表达。方法血管平滑肌细胞分为常氧(200 m L/L O2)培养的正常细胞、2μmol/m L枸杞多糖处理的常氧细胞、DMSO处理的常氧细胞、DMSO处理的缺氧(100 m L/L O2)细胞、(0.5、1、2)μmol/m L枸杞多糖处理的缺氧细胞、SIRT1的特异性激动剂10μmol/L白芦藜醇处理的缺氧细胞、非特异性激动剂1μmol/L SRT-1720处理的缺氧细胞、SIRT1抑制剂0.5μmol/L EX-527处理的缺氧细胞,培养于6、12、24 h,实时定量PCR、Western blot法分别检测SIRT1、MMP-9及HIF-1α的mRNA和蛋白表达;在缺氧的血管平滑肌细胞中加入枸杞多糖处理12 h,实时定量PCR、Western blot法分别检测血管平滑肌细胞中SIRT1、MMP-9及HIF-1α的表达。结果缺氧条件下血管平滑肌细胞SIRT1mRNA和蛋白水平降低,MMP-9及HIF-1αmRNA和蛋白水平升高。缺氧条件下用枸杞多糖处理后,SIRT1的含量随枸杞多糖剂量的增加而增加,同时MMP-9、HIF-1α的含量降低。SIRT1的特异性激动剂白藜芦醇可抑制MMP-9的表达。结论枸杞多糖增加缺氧肺血管平滑肌细胞SIRT1水平、降低MMP-9及HIF-1α的水平。
Objective To detect the expression of SIRT1, MMP-9 and HIF-1α in pulmonary vascular smooth muscle cells treated with LBP under hypoxia conditions. 1α) expression. Methods Vascular smooth muscle cells were divided into normal cells cultured in normoxia (200 m L / L O2), normoxia cells treated with 2μmol / m L LBP, normoxic cells treated with DMSO, hypoxic (100 m L / L O2) cells, hypoxia cells treated with (0.5, 1, 2) μmol / mL LBP, hypoxia cells treated with 10μmol / L resveratrol specific agonist SIRT1, and non-specific agonist 1μmol / SRT-1720-treated hypoxia cells and SIRT1 inhibitor 0.5μmol / L EX-527-treated hypoxic cells were cultured at 6,12,24 h, real-time quantitative PCR, Western blot were detected SIRT1, MMP-9 and HIF 1α mRNA and protein expression was detected by flow cytometry. Lycium barbarum polysaccharide was added into hypoxic vascular smooth muscle cells for 12 h, and the expression of SIRT1, MMP-9 and HIF-1α in vascular smooth muscle cells were detected by real-time PCR and Western blot respectively. Results Under hypoxia, SIRT1mRNA and protein levels of vascular smooth muscle cells decreased, while MMP-9 and HIF-1αmRNA and protein levels increased. Under the condition of hypoxia, the content of SIRT1 increased with the increase of LBP, while the content of MMP-9 and HIF-1α decreased. Resveratrol, a specific agonist of SIRT1, inhibits the expression of MMP-9. Conclusion Lycium barbarum polysaccharides can increase the level of SIRT1 in hypoxic pulmonary vascular smooth muscle cells and decrease the levels of MMP-9 and HIF-1α.