高盐促大鼠血管平滑肌细胞增殖的时效量效关系及相关机制
发布时间：2019-05-10 23:39
【摘要】：目的比较不同Na~+浓度及其作用时间促大鼠血管平滑肌细胞(VSMC)增殖的差异,探讨高盐诱导VSMC增殖的可能机制。方法组织贴壁法培养原代VSMC,传至第四代去血清同步化1d,分别用139、141、144、147、150、153、156、159、162和165mmol/L的Na~+干预1、2、3和4d后,MTT比色法和细胞增殖检测试剂盒(CCK-8)检测VSMC增殖情况。选取促增殖最显著的盐浓度和作用时间,以Na~+139mmol/L为对照进行后续实验。CCK-8法检测渗透压对VSMC增殖的影响;流式细胞仪分析VSMC增殖周期;免疫荧光染色和Western blot检测VSMC增殖细胞核抗原(PCNA)、血管紧张素Ⅱ(AngⅡ)、血管紧张素Ⅱ1型受体(AT_1R)的表达情况;实时荧光定量PCR检测VSMC中血管紧张素原(AGT)、AT_1R mRNA的表达。结果MTT和CCK-8结果显示,Na~+153~165mmol/L作用2、3d可促进VSMC增殖(Na~+159mmol/L作用3d最明显);CCK-8检测渗透压对VSMC增殖结果显示,与Na~+139mmol/L组比较,Na~+159mmol/L细胞明显增殖[(1.21±0.16)%比(1.00±0.25)%,P0.05],而甘露醇组与Na~+139 mmol/L组比较差异无统计学意义;流式细胞仪分析VSMC增殖周期结果显示,Na~+159mmol/L组G_0/G_1期细胞比例降低,S期细胞比例增多(均P0.05);PCNA免疫荧光染色可见Na~+159mmol/L组VSMC中PCNA阳性细胞核增加,Western blot结果显示Na~+159mmol/L组PCNA蛋白表达明显增加(均P0.05);实时荧光定量PCR显示,与Na~+139mmol/L组相比,Na~+159mmol/L组中AGT、AT_1R mRNA表达增加(均P0.05);Western blot显示,与Na~+139 mmol/L组相比,Na~+159 mmol/L组中AngⅡ、AT_1R蛋白表达增多(均P0.05)。结论高Na~+(153~165 mmol/L)作用2~3d能诱导VSMC增殖,且Na~+159mmol/L作用3d增殖效应最为显著;作用机制可能与高Na~+促进AngⅡ和AT_1R表达有关。
[Abstract]:Objective to compare the effects of different concentrations of Na~ and their time on (VSMC) proliferation of rat vascular smooth muscle cells (VSMCs), and to explore the possible mechanism of VSMC proliferation induced by high salt. Methods Primary VSMC, cultured by tissue adherent method was transferred to the fourth generation of serum for 1 day. The proliferation of VSMC was detected by MTT colorimetric assay and cell proliferation kit (CCK-8) after intervention with 139141144147150153156159162 and 165mmol/L Na~ for 2, 3 and 4 days, respectively. The most significant salt concentration and action time were selected to carry out the follow-up experiment with Na~ 139mmol/L as control. CCK-8 method was used to detect the effect of osmotic pressure on the proliferation of VSMC, flow cytometry was used to analyze the proliferation cycle of VSMC, and CCK-8 method was used to detect the effect of osmotic pressure on proliferation of VSMC. The expression of proliferating cell nuclear antigen (PCNA), angiotensin 鈪，
本文编号：2474089
[Abstract]:Objective to compare the effects of different concentrations of Na~ and their time on (VSMC) proliferation of rat vascular smooth muscle cells (VSMCs), and to explore the possible mechanism of VSMC proliferation induced by high salt. Methods Primary VSMC, cultured by tissue adherent method was transferred to the fourth generation of serum for 1 day. The proliferation of VSMC was detected by MTT colorimetric assay and cell proliferation kit (CCK-8) after intervention with 139141144147150153156159162 and 165mmol/L Na~ for 2, 3 and 4 days, respectively. The most significant salt concentration and action time were selected to carry out the follow-up experiment with Na~ 139mmol/L as control. CCK-8 method was used to detect the effect of osmotic pressure on the proliferation of VSMC, flow cytometry was used to analyze the proliferation cycle of VSMC, and CCK-8 method was used to detect the effect of osmotic pressure on proliferation of VSMC. The expression of proliferating cell nuclear antigen (PCNA), angiotensin 鈪，
本文编号：2474089
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