Ma Miaomiao, Wang Li, Ma Yitong, Yang Yining, Chen Bangdang, Zhu Xiaoli
Department of Cardiology, First Affiliated Hospital, School of Medicine, Shihezi University, Shihezi 832008, China.
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Zhonghua Xin Xue Guan Bing Za Zhi. 2015 Jun;43(6):542-7.
To investigate the effects of different concentrations of norepinephrine (NE) on proliferation and apoptosis of cultured cardiac fibroblasts (CFBs) from neonatal mice and to elucidate related mechanisms.
CFBs of Sprague-Dawley (SD) rats were isolated and cultured and divided into normal control group and different concentration of NE intervention groups (0.1, 1, 10, 50, and 100 µmol/L). Water soluble tetrazolium-1 (WST-1) assay was carried out to detect the viability of CFBs. Morphology of apoptosis cells was evaluated by fluorescence microscope with terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. The expressions of collagen I, collagen III, pro-oncogene c-myc in CFBs were detected by reverse transcription-polymerase chain reaction (RT-PCR). The phospho-mitogen activated protein kinase (p-p38MAPK) and caspase3 protein levels were examined by Western blot.
Proliferation was significantly increased in 1 µmol/L and 10 µmol/L groups compared with the normal control group (1.05 ± 0.05 and 1.09 ± 0.02 vs. 1.00 ± 0.03, all P < 0.05).CFBs apoptosis was significantly enhanced in 50 µmol/L and 100 µmol/L groups ((22.69 ± 2.18)% and (36.40 ± 6.80)% vs.(4.50 ± 1.08)%, all P < 0.05). Expression of Collagen I peaked in 10 µmol/L group, expression of collagen III and c-myc increased dose-dependently in proportion to increasing NE concentrations (all P < 0.05 vs. control group). The expression of p-p38MAPK and caspase3 was also significantly upregulated in a dose-dependent manner in NE groups (all P < 0.05 vs. control group).
Low concentration NE induces CFBs proliferation and high concentration NE promotes CFBs apoptosis. p38MAPK phosphorylation may be a major mediator of NE-induced effects on CFBs.
探讨不同浓度去甲肾上腺素（NE）对新生小鼠心脏成纤维细胞（CFBs）增殖和凋亡的影响，并阐明相关机制。
分离培养Sprague-Dawley（SD）大鼠的CFBs，分为正常对照组和不同浓度NE干预组（0.1、1、10、50和100 μmol/L）。采用水溶性四氮唑盐-1（WST-1）法检测CFBs的活力。用末端脱氧核苷酸转移酶dUTP缺口末端标记（TUNEL）染色，通过荧光显微镜评估凋亡细胞的形态。采用逆转录-聚合酶链反应（RT-PCR）检测CFBs中Ⅰ型胶原、Ⅲ型胶原、原癌基因c-myc的表达。通过蛋白质印迹法检测磷酸化丝裂原活化蛋白激酶（p-p38MAPK）和半胱天冬酶3蛋白水平。
与正常对照组相比，1 μmol/L和10 μmol/L组的增殖显著增加（1.05±0.05和1.09±0.02 vs. 1.00±0.03，均P<0.05）。50 μmol/L和100 μmol/L组的CFBs凋亡显著增强（分别为（22.69±2.18）%和（36.40±6.80）% vs.（4.50±1.08）%，均P<0.05）。Ⅰ型胶原表达在10 μmol/L组达到峰值，Ⅲ型胶原和c-myc的表达随NE浓度增加呈剂量依赖性增加（与对照组相比，均P<0.05）。NE组中p-p38MAPK和半胱天冬酶3的表达也呈剂量依赖性显著上调（与对照组相比，均P<0.05）。
低浓度NE诱导CFBs增殖，高浓度NE促进CFBs凋亡。p38MAPK磷酸化可能是NE对CFBs作用的主要介导因素。
