摘要
目的观察川芎嗪(TMP)对人脐静脉内皮细胞(HUVECs)缺氧相关因子表达的影响。方法体外培养HUVECs,取2~5代细胞进行实验。将HUVECs分为对照组、氯化钻(CoCl2)缺氧组及50、100、200μmol/L TMP药物处理组。对照组不作任何处理。CoCl2缺氧组利用150μmol/L。的CoCl2干预HUVECs4h;50、100、200μmol/L TMP药物处理组在CoCl2干预HUVECs4h后,再加入不同浓度TMP继续培养8h。分别提取各组细胞RNA和总蛋白,采用实时荧光定量聚合酶链反应(RT—PCR)和蛋白免疫印迹法(Westernblot)检测脯氨酰羟化酶2(PHD2)、缺氧诱导因子-1a(HIF-1a)、血管内皮生长因子(VEGF)mRNA和蛋白的表达。结果实时RT—PCR检测结果显示,与对照组比较,CoCl2缺氧组PHD2mRNA表达下降(t=3.734),HIF-1a和VEGFmRNA表达升高(t=4.589、3.926),差异均有统计学意义(P〈O.05)。50、100、200ffmol/L。TMP药物处理组分别与CoCl。缺氧组比较,PHD2mRNA表达均升高(t=3.286、3.617、3.886),差异有统计学意义(P〈0.05);HIF-1amRNA表达均无明显变化(t=1.025、0.726、-1.386),差异无统计学意义(P〉0.05);VEGFmRNA表达均有所下降,但50μmol/L。TMP药物处理组与之差异无统计学意义(t=1.696,P〉0.05),100、200umol/L。TMP药物处理组与之差异有统计学意义(t=2.974、3.492,P%0.05)。Westernblot检测结果显示,与对照组比较,CoCl2缺氧组PHD2蛋白表达下降,差异有统计学意义(t=3.122,P〈0.05);HIF-1a及VEGF蛋白表达均有不同程度升高,差异有统计学意义(t=3.778、3.257,P〈0.05)。50、100、200umol/L。TMP药物处理组分别与CoCl。缺氧组比较,PHD2蛋白表达均升高(t=2.813、3.026、3.078),差异有统计学意义(P%0.05);HIF-1d、VEGF蛋白表达均有所下降,但50umol/L.TMP药物处理组与之差异无统计学意义(t=2.056、1.986,P〉0.05),100ffmol/L。TMP药物处理组(t=3.058、2.976)及200ffmol/L。TMP药物处理组(t=3.828、3.136)与之差异有统计学意义(P〈0.05)。结论TMP能上调缺氧HUVECs中PHD2mRNA和蛋白的表达,下调HIF-1“蛋白、VEGFmRNA和蛋白的表达。
Objective To observe the effect of tetramethypyrazine (TMP) on the expression of hypoxia-related factors in human umbilical vein endothelial cells (HUVECs). Methods The second to fifth passage cultured HUVECs were divided into five groups: control group, CoC12-induced hypoxic group and 50, 100, 200 μmol/L TMP treatment groups. HUVECs in control group were not treated. HUVECs in CoCl2 induced bypoxic group were treated with 150 μmol/L COC12 for four hours. HUVECs in 50, 100, 200 μmol/L TMP treated groups were pretreated with 150 /,mol/L CoOl2 for four hours, followed by treatment with 50, 100, 200μmol/L TMP for eight hours. Real-time reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA levels of prolyl hydroxylase 2 (PHD2), hypoxia-induced factor-lc^(HIF-lcO and vascular endothelial growth factor (VEGF). Protein levels of PHD2, HIF-I~, and VEGF were detected using Western blot. Results Compared with the control group, the COC12 induced hypoxic group showed decreased mRNA and protein levels of PHD2 (t=3. 734, 3. 122; P〈0.05), while
出处
《中华眼底病杂志》
CAS
CSCD
北大核心
2013年第3期284-289,共6页
Chinese Journal of Ocular Fundus Diseases
基金
国家自然科学基金(30973899)
高等学校博士学科点专项科研基金(20120171110086)
关键词
内皮细胞
细胞
培养的
药物作用
川芎嗪
治疗应用
血管内皮生长因子类
Endothelial cells
Cells, cultured/drug effects
Tetramethylpyrazine/therapeu*ic use
Vascular endothelial growth factors