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目的探讨内皮素-1(ET-1)对大鼠骨髓内皮前体细胞(EPC)增殖、细胞周期及一氧化氮(NO)分泌功能的影响。方法采用密度梯度离心法获取大鼠骨髓单核细胞,在M199培养液中培养扩增EPC并进行鉴定。不同浓度ET-1(A组:0mol/L、B组:10-9mol/L、C组:10-8mol/L、D组:10-7mol/L、E组:10-6mol/L)作用于EPC,MTT法检测ET-1对EPC增殖能力的影响,流式细胞仪检测ET-1对EPC细胞周期的影响,硝酸还原酶法检测细胞培养液中NO含量的变化,观察ET-1对EPC的NO分泌功能的影响。结果与对照组(A组,0.405±0.017)相比,ET-1浓度C组(0.434±0.016)、D组(0.463±0.016)、E组(0.473±0.015)EPC增殖能力显著增高(n=8,P<0.01)。EPC的G0/G1期细胞百分数C组(57.28±3.65)%、D组(44.99±3.19)%、E组(40.29±3.74)%较对照组(70.55±1.37)%明显降低(n=5,P<0.01),EPC的S期细胞百分数C组(26.75±2.87)%、D组(32.79±2.41)%、E组(35.74±2.94)%较对照组(20.04±1.64)%明显升高(n=5,P<0.01),EPC的G2/M期细胞百分数C组(15.96±1.71)%、D组(22.22±2.22)%、E组(23.64±2.86)%较对照组(9.41±0.81)%明显升高(n=5,P<0.01)。EPC培养液中NO含量(μmol/L)ET-1浓度C组(11.70±1.80)、D组(15.69±1.86)、E组(16.89±1.55)较对照组(7.45±2.41)明显增加(n=8,P<0.01)。B组检测结果与对照组相比,各项指标均无统计学差异。结论ET-1能够促进EPC的增殖,促进细胞向S期和G2期的转化,并提高细胞NO分泌功能,可能参与缺血性疾病的血管再生过程。
Objective To investigate the effects of endothelin-1 (ET-1) on the proliferation, cell cycle and nitric oxide (NO) secretion of rat bone marrow endothelial progenitor cells (EPCs). Methods Rat bone marrow mononuclear cells were obtained by density gradient centrifugation. EPCs were cultured in M199 medium and identified. The effects of different concentration of ET-1 (A group: 0mol / L, B group: 10-9mol / L, C group: 10-8mol / L, D group: 10-7mol / L, E group: 10-6mol / L) The effect of ET-1 on the proliferation of EPCs was examined by EPC and MTT assay. The effect of ET-1 on the cell cycle of EPC was detected by flow cytometry. The content of NO in cell culture medium was detected by nitrate reductase method. The effect of EPC on NO secretion. Results Compared with the control group (group A, 0.405 ± 0.017), the proliferative capacity of ET-1 in C group (0.434 ± 0.016), D group (0.463 ± 0.016) and E group (0.473 ± 0.015) 8, P <0.01). The percentage of cells in G0 / G1 phase in EPC group was significantly lower than that in control group (57.28 ± 3.65)%, 44.99 ± 3.19% in group D and 40.29 ± 3.74% in group E (70.55 ± 1.37% (P <0.01). The percentage of cells in S phase in EPC group was significantly higher than that in control group (26.75 ± 2.87)%, D group (32.79 ± 2.41)% and E group (35.74 ± 2.94)% The percentage of cells in G2 / M phase of EPC was (15.96 ± 1.71)% in group C and (22.22 ± 2.22)% in group D compared with 9.41 ± 0.81 (23.64 ± 2.86)% in group E )% Was significantly higher (n = 5, P <0.01). The concentration of ET-1 in group C (11.70 ± 1.80), group D (15.69 ± 1.86), group E (16.89 ± 1.55) was significantly higher than that in control group (7.45 ± 2.41) = 8, P <0.01). B group test results compared with the control group, the indicators were not statistically different. Conclusion ET-1 can promote the proliferation of EPCs, promote the transformation of cells to S phase and G2 phase, increase the secretion of NO, and may be involved in the process of angiogenesis in ischemic diseases.