血府逐瘀汤延缓大鼠骨髓源内皮祖细胞衰老的机制研究
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摘要
目的观察血府逐瘀汤对血管紧张素Ⅱ(AngⅡ)诱导大鼠骨髓源内皮祖细胞(EPCs)衰老及肿瘤抑制基因(p53)、沉默信息调节因子1(SIRT1)表达的影响。方法体外培养EPCs,加入1×10~(-7)mol·L~(-1)AngⅡ诱导细胞衰老,建立内皮祖细胞衰老模型。将EPCs随机分为6组:正常组(不做处理)、模型组(空白血清),对照组(miR-34a抑制剂)和低、中、高3个剂量实验组(5%,10%,15%含药血清)。分别处理24,48,72h后,收集细胞。以β-半乳糖苷酶染色法检测内皮祖细胞衰老状态;以实时荧光定量法检测p53 mRNA、SIRT1 mRNA的表达。结果正常组、模型组、对照组和低、中、高3个剂量实验组的衰老细胞百分数分别为(10. 67±1. 52)%,(66. 33±2. 08)%,(23. 66±1. 52)%,(51. 33±1. 52)%,(43. 66±2. 08)%和(32. 00±2. 00)%;这6组p53 mRNA的表达量(相对值)分别为1. 00±0. 00,2. 99±0. 15,1. 47±0. 10,2. 96±0. 18,2. 02±0. 08和2. 00±0. 13;这6组SIRT1mRNA的表达量(相对值)分别为1. 00±0. 00,0. 49±0. 03,0. 90±0. 06,0. 50±0. 02,0. 60±0. 03和0. 69±0. 03。模型组的上述指标与正常组比较,差异均有统计学意义(均P <0. 01);对照组和3个剂量实验组与模型组比较,差异均有统计学意义(均P <0. 01)。结论血府逐瘀汤能够延缓EPCs的衰老,可以减少衰老EPCs中p53 mRNA的表达而增加SIRT1 mRNA的表达,其中以高剂量组效果最佳。
Objective To observe the effect of Xuefuzhuyu decoction on the senescence of bone marrow-derived endothelial progenitor cells( EPCs) induced by angiotensin Ⅱ( AngⅡ) and the expression of tumor-suppressing gene( p53) mRNA and silent information regulator 1( SIR1) mRNA in cell senescence,in order to explore its anti-aging effect and mechanism. Methods EPCs were cultured in vitro,and 1 ×10~(-7) mol·L~(-1) AngⅡwas used to induce cell senescence. The model of EPCs senescence was established. The EPCs were divided into 6 groups:normal group( no treatment),model group( blank serum),control group( miR-34 a inhibitor),experimental-L,-M,-H groups( 5%,10%,15% XuefuZhuyu Decoction serum),cells were collected after treated for24,48,72 h. β-galactosidase staining was used to detect the senescence of EPCs. The expressions of p53 mRNA and SIRT1 mRNA in EPCs weredetected by real time quantitative polymerase chain reaction. Results After treatment,No.( %) of aging EPCs in normal group,model group,control group,experimental-L,-M,-H groups were( 10. 67 ± 1. 52) %,( 66. 33 ± 2. 08) %,( 23. 66 ± 1. 52) %,( 51. 33 ± 1. 52) %,( 43. 66 ± 2. 08) %,( 32. 00 ± 2. 00) %. The expression levels( relative values)of p53 mRNA in the 6 groups were 1. 00 ± 0. 00,2. 99 ± 0. 15,1. 47 ± 0. 10,2. 96 ± 0. 18,2. 02 ± 0. 08,2. 00 ± 0. 13;The expression levels( relative values) of SIRT1 mRNA in the 6 groups were 1. 00 ± 0. 00,0. 49 ± 0. 03,0. 90 ± 0. 06,0. 50 ± 0. 02,0. 60 ± 0. 03,0. 69 ± 0. 03. By comparing between model group with normal group,the differences of above indicators were statistically significant( P < 0. 01); comparing between 4 drugs groups with model group,the differences of above indicators were statistically significant( all P < 0. 01). Conclusion Xuefuzhuyu decoction can delay the senescence of EPCs,and its mechanism may be related to the down-regulation of p53 mRNA and up-regulation of SIRT1 mRNA. Experimental-H group had the best effect.
Objective To observe the effect of Xuefuzhuyu decoction on the senescence of bone marrow-derived endothelial progenitor cells( EPCs) induced by angiotensin Ⅱ( AngⅡ) and the expression of tumor-suppressing gene( p53) mRNA and silent information regulator 1( SIR1) mRNA in cell senescence,in order to explore its anti-aging effect and mechanism. Methods EPCs were cultured in vitro,and 1 ×10~(-7) mol·L~(-1) AngⅡwas used to induce cell senescence. The model of EPCs senescence was established. The EPCs were divided into 6 groups:normal group( no treatment),model group( blank serum),control group( miR-34 a inhibitor),experimental-L,-M,-H groups( 5%,10%,15% XuefuZhuyu Decoction serum),cells were collected after treated for24,48,72 h. β-galactosidase staining was used to detect the senescence of EPCs. The expressions of p53 mRNA and SIRT1 mRNA in EPCs weredetected by real time quantitative polymerase chain reaction. Results After treatment,No.( %) of aging EPCs in normal group,model group,control group,experimental-L,-M,-H groups were( 10. 67 ± 1. 52) %,( 66. 33 ± 2. 08) %,( 23. 66 ± 1. 52) %,( 51. 33 ± 1. 52) %,( 43. 66 ± 2. 08) %,( 32. 00 ± 2. 00) %. The expression levels( relative values)of p53 mRNA in the 6 groups were 1. 00 ± 0. 00,2. 99 ± 0. 15,1. 47 ± 0. 10,2. 96 ± 0. 18,2. 02 ± 0. 08,2. 00 ± 0. 13;The expression levels( relative values) of SIRT1 mRNA in the 6 groups were 1. 00 ± 0. 00,0. 49 ± 0. 03,0. 90 ± 0. 06,0. 50 ± 0. 02,0. 60 ± 0. 03,0. 69 ± 0. 03. By comparing between model group with normal group,the differences of above indicators were statistically significant( P < 0. 01); comparing between 4 drugs groups with model group,the differences of above indicators were statistically significant( all P < 0. 01). Conclusion Xuefuzhuyu decoction can delay the senescence of EPCs,and its mechanism may be related to the down-regulation of p53 mRNA and up-regulation of SIRT1 mRNA. Experimental-H group had the best effect.
引文
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[12]王铭,雷燕,陈连凤.人参三七川芎提取物抑制p53基因表达促进衰老内皮细胞增殖的研究[J].中华中医药杂志,2015,30(2):519-523.
[2] SEILER C. Assessment and impact of the human coronary collateral circulation on myocardial ischemia and outcome[J]. Circ Cardiovasc Interv,2013,6(6):719-728.
[3] HUR J,YANG H M,YOON C H,et al. Identification of a novel role of T cells in postnatal vasculogenesis:characterization of endothelial progenitor cell colonies[J]. Circulation,2007,116(15):1671-1682.
[4] ZHU S,DENG S,MA Q,et al. MicroRNA-10A*and microRNA-21 modulate endothelial progenitor cell senescence via suppressing high-mobility group A2[J]. Circ Res,2013,112(1):152-164.
[5]王丽,张会峰,袁慧娟,等.大鼠骨髓内皮祖细胞的分离培养与鉴定[J].中国组织工程研究,2012,25(10):1733-1736.
[6]高冬,焦雨欢,武一曼,等.血府逐瘀汤诱导内皮祖细胞参与缺血区血管新生的实验研究[J].中国中西医结合杂志,2012,32(2):224-228.
[7] IMANISHI T,HANO T,NISHIO I. AngiotensinⅡaccelerates endothelial progenitor cell senescence through induction of oxidative stress[J]. JHyperten,2005,23(1):97-104.
[8]曹建明,郝东杰,陈华乐,等. D-半乳糖诱导细胞衰老模型的线粒体能量代谢功能研究[J].中华老年医学杂志,2017,45(2):199-203.
[9]李晶,欧芹,江旭东,等.山茱萸多糖对衰老模型小鼠肝组织SIRT1和p53基因表达的影响[J].中国老年学杂志,2015,43(7):1885-1886.
[10] BLACKBURN E H. Telomere states and cell fates[J]. Nature,2000,408(1):53-56
[11]贾春平.抑癌基因p53与肿瘤研究的最新进展[J].生命科学杂志,2008,20(3):450-453
[12]王铭,雷燕,陈连凤.人参三七川芎提取物抑制p53基因表达促进衰老内皮细胞增殖的研究[J].中华中医药杂志,2015,30(2):519-523.