CAJ | 학술논문

目的探讨去甲肾上腺素(NE)对体外培养的乳鼠心肌成纤维细胞(CFB)增殖和凋亡的影响及其机制.方法分离Sprague-Dawley (SD)大鼠乳鼠CFB,体外培养,设对照组(未给予任何干预)和NE不同浓度(0.1、1、10、50和100 μmol/L)处理组.水溶性四唑盐-1(WST-1)法测定CFB增殖情况,原位末端转移酶标记(TUNEL)法检测细胞凋亡情况,逆转录聚合酶链反应(RT-PCR)检测Ⅰ型胶原、Ⅲ型胶原和原癌基因c-myc mRNA的表达水平.蛋白印迹(Western blot)法检测磷酸化丝裂原活化蛋白激酶p38(p-p38MAPK)及半胱氨酸天冬氨酸蛋白酶3(caspase3)的表达水平.结果 NE 1 μmol/L处理组和NE 10μmol/L处理组CFB增殖数量明显高于对照组(1.05 ±0.05和1.09±0.02比1.00±0.03,P均＜0.05).NE 50 μmol/L处理组和NE 100 μmol/L处理组CFB凋亡率明显高于对照组[(22.69±2.18)％和(36.40±6.80)％比(4.50±1.08)％,P均＜ 0.05].在NE10 μmol/L处理组中Ⅰ型胶原的mRNA表达水平最高,Ⅲ型胶原和c-myc mRNA的表达水平随着NE浓度增加而增高,均明显高于对照组(P均＜ 0.05).NE 1μmol/L、10 μmol/L、50 μmol/L和100 μmol/L处理组p-p38MAPK及caspase3蛋白表达水平逐渐增高,均明显高于对照组(P均＜0.05).结论低浓度NE可诱导CFB增殖,高浓度NE可促进CFB凋亡.p38MAPK磷酸化可能介导了NE对CFB增殖和凋亡的调控.
목적 탐토거갑신상선소(NE)대체외배양적유서심기성섬유세포(CFB)증식화조망적영향급기궤제.방법 분리Sprague-Dawley (SD)대서유서CFB,체외배양,설대조조(미급여임하간예)화NE불동농도(0.1、1、10、50화100 μmol/L)처리조.수용성사서염-1(WST-1)법측정CFB증식정황,원위말단전이매표기(TUNEL)법검측세포조망정황,역전록취합매련반응(RT-PCR)검측Ⅰ형효원、Ⅲ형효원화원암기인c-myc mRNA적표체수평.단백인적(Western blot)법검측린산화사렬원활화단백격매p38(p-p38MAPK)급반광안산천동안산단백매3(caspase3)적표체수평.결과 NE 1 μmol/L처리조화NE 10μmol/L처리조CFB증식수량명현고우대조조(1.05 ±0.05화1.09±0.02비1.00±0.03,P균＜0.05).NE 50 μmol/L처리조화NE 100 μmol/L처리조CFB조망솔명현고우대조조[(22.69±2.18)％화(36.40±6.80)％비(4.50±1.08)％,P균＜ 0.05].재NE10 μmol/L처리조중Ⅰ형효원적mRNA표체수평최고,Ⅲ형효원화c-myc mRNA적표체수평수착NE농도증가이증고,균명현고우대조조(P균＜ 0.05).NE 1μmol/L、10 μmol/L、50 μmol/L화100 μmol/L처리조p-p38MAPK급caspase3단백표체수평축점증고,균명현고우대조조(P균＜0.05).결론 저농도NE가유도CFB증식,고농도NE가촉진CFB조망.p38MAPK린산화가능개도료NE대CFB증식화조망적조공.
Objective 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.Methods 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,prooncogene 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.Results 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 Pμmol/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).Conclusions 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.