【作者】 修春英；

【导师】 蔡辉；

【作者基本信息】 南京中医药大学 ， 中西医结合基础， 2006， 硕士

【摘要】 目的：观察溶血磷脂酸（LPA）对体外培养心脏成纤维细胞（CFs）的增殖、胶原合成及促丝裂原蛋白活化激酶（MAPK）的影响，进一步研究中药单体川芎嗪（TMP）的干预作用。 方法：用消化法培养新生SD大鼠的CFs，用四氮唑蓝（MTT）比色法测定细胞增殖，羟脯氨酸碱水解法测定胶原合成，并用Western blot的方法检测MAPK中细胞外信号调节激酶1／2（ERK1／2）的含量，分别观察不同浓度LPA对CFs增殖和胶原合成的影响及TMP的干预作用。 结果：①随着LPA浓度的升高，MTT比色法A₄₉₀值呈上升趋势。在药物作用48h后，只有10^-6μmol／L组A₄₉₀值较空白对照组有显著升高（P＜0.05）；作用至72h后，10^-7、10^-6μmol／L组A₄₉₀值较空白对照组有显著升高（P＜0.05）；作用至96h后，各组A₄₉₀值较空白对照组均有显著升高（P＜0.05或P＜0.01）；②CFs分泌的胶原随着LPA作用浓度和时间的增加呈递增趋势。药物作用48、72h，只有10^-6μmol／L组胶原浓度较空白对照组有显著升高（P＜0.01）；作用至96h，各浓度组胶原含量均较对照组有显著升高（P＜0.05或P＜0.01），其中10^-6与10^-7、10^-8μmol／L组之间差异有显著性（P＜0.05）；③在LPA作用48h后，10^-8、10^-7μmol／L组胶原含量／A₄₉₀比值高于空白对照组，但与空白对照组比较无统计学差异（P＞0.05），其中10^-6μmol／L组的比值显著高于空白对照组（P＜0.05）；药物作用至96h，中、高浓度的胶原含量／A₄₉₀比值均显著高于空白对照组（P＜0.05）；④在药物作用的48h，20mg／L浓度的TMP即表现明显抑制LPA(10^-6μmol／L)诱导的细胞增殖作用（P＜0.05），72h后抑制细胞增殖现象逐渐明显，其中10、20 mg／L浓度组A₄₉₀伽值显著低于LPA组（P＜0.001），5mg／L浓度组TMP也表现出明显抑制增殖作用（P＜0.05）：⑤在TMP作用48h后，低浓度组（5mg／L）能轻度抑制LPA诱导的胶原合成作用，但与LPA组比较无显著差异（P＞0.05），10、20mg／L浓度组产生明显抑制作用（P＜0.05或P＜0.001），TMP作用72h后，各浓度组均表现为显著抑制胶原合成作用，并随浓度的增加作用加强（P＜0.05和P＜0.001）；⑥在药物作用的各个时间段，LPA组的胶原含量／A₄₉₀比值均高于空白对照组，在TMP作用的48、72h，低浓度组（5mg／L）的胶原含量／A₄₉₀比值即低于LPA组，但与LPA组比较无显著差异（P＞0.05），而中、高浓度组更多还原

【Abstract】 Objective: To investigate the effects of lysophosphatidic acid （LPA） on the proliferation and collagen synthesis of cardiac fibroblasts（CFs） in vitro culture. And to investigate the interactions between the effects of tetramethylpyrazine（TMP） and LPA on cell proliferation, collagen synthesis and mitogen activated protein kinase（MAPK）. Methods: Isolated and cultured neonatal Sprague-Daeley（SD） rats were used as experimental model. CFs proliferation was measured by MTT assay. Collagen synthesis was measured by hydroxyproline alkaline hydrolysis assay. And extracellular signal-regulated kinase 1/2 （ERK1/2） in CFs was measured by western blot.Results: ①A₄₉₀ value of CFs by MTT assay increased with the elevation of LPA concentration. Forty-eight hours later, Only A₄₉₀ value of 10^-6μ mol/L group was significantly higher than that of control group （ P<0.05）. Seventy-two hours later, A₄₉₀ value of 10^-7,10^-6 μ- mol/L groups were significantly higher than that of control group （ P<0.05）. Ninety-six hours later, A₄₉₀ value of 10^-8, 10^-7,10^-6 μ mol/L groups were all significantly higher than that of control group（P<0.05 or P<0.01）. ②The secretion of collagen increased with the elevation of LPA concentration and action time. Seventy-two hours later, Only the collagen concentration of 10^-6 μ mol/L group was increased significantly compared to the control group（P<0.01）. Ninety-six hours later, the concentration of all three groups were increased significantly compared to the control group（P<0.05 or P<0.01）, in which that of 10^-6μ mol/L groups was increased significantly compared to that of the 10^-7, 10^-8 μ mol/L groups（P<0.05）. ③ Forty-eight hours later, the ratios of the collagen content and A₄₉₀ in 10^-8、 10^-7 μ mol/L group were higher than that of the control group, but there was significant更多还原