Wu Xi-li, An Peng, Ye Bing-yu, Shi Xing-min, Sun Wan-sen, Fu Rong-guo, Wang Zhu, Qiao Cheng-lin
Department of Integrated Traditional Chinese and Western Medicine, Second Affiliated Hospital of Medical School of Xi'an Jiaotong University, Xi'an, 710004, China,
Chin J Integr Med. 2013 Dec;19(12):927-34. doi: 10.1007/s11655-013-1655-8. Epub 2013 Dec 5.
To study the effects and possible underlying mechanism of Qufeng Tongluo Prescription (, QFTL) on the regulation of mesangial cells (MCs) proliferation and apoptosis.
The MCs used in this experiment have undergone five passages induced by lipopolysaccharide (LPS). Changes in the proliferation, apoptosis, cell cycle regulatory proteins and mRNA expression levels of the MCs after administration of Benazepril or QFTL were measured by methyl thiazolyl tetrazolium (MTT) reduction assay, flow cytometry, Western blot and quantitative real-time polymerase chain reaction (qRT-PCR), respectively.
The addition of Benazepril or QFTL serum inhibited LPS-induced MC proliferation after treatment for 24, 48 and 72 h, respectively (P<0.05 or P<0.01). Moreover, the inhibitory effect is more significant in the QFTL group at 48 h (P<0.05). Compared with the control group, LPS-induced cell proliferation decreased the number of cells in G1 phase versus cells in S and G2/M phases, while the addition of QFTL and Benazepril serum increased the ratio of cells at G1 phase (P<0.05 or P<0.01) to cells at S phase (P<0.01), implicating the cell cycle inhibition effect exerted by QFTL. LPS decreased the level of MC apoptosis, compared with the control group (P<0.05), while QFTL and Benazepril serum increased the level of MC apoptosis (P<0.01). Moreover, the difference between the QFTL group and the Benazepril group was statistically significant (P<0.01). Compared with the control group, the protein and mRNA expression levels of cylinD1, cyclin dependent kinase 2 (CDK2) and p21 were significantly increased (P<0.05 or P<0.01), p27 was decreased but with no statistical significance (P>0.05); After being treated with QFTL and Benazepril serum, the protein and mRNA expression levels of cylinD1, CDK2, p21 were decreased and p27 increased significantly (P<0.05 or P<0.01); Compared with the Benazepril group, QFTL show better effects on protein and mRNA expression levels of cylinD1, CDK2 (P<0.05 or P<0.01) and p21 protein expression (P<0.05).
QFTL inhibits MCs proliferation, promotes MCs apoptosis through an underlying mechanism of down-regulating the protein and mRNA expression levels of cylinD1, CDK2, p21 and up-regulation of the expression level of p27.
研究祛风通络方（QFTL）对系膜细胞（MCs）增殖与凋亡调控的影响及其可能的潜在机制。
本实验所用MCs经脂多糖（LPS）诱导传代5次。分别采用甲基噻唑基四氮唑（MTT）比色法、流式细胞术、蛋白质免疫印迹法和实时定量聚合酶链反应（qRT-PCR）检测贝那普利或QFTL给药后MCs的增殖、凋亡、细胞周期调节蛋白及mRNA表达水平的变化。
分别给予贝那普利或QFTL含药血清处理24、48和72 h后，均可抑制LPS诱导的MCs增殖（P<0.05或P<0.01）。而且，48 h时QFTL组的抑制作用更显著（P<0.05）。与对照组相比，LPS诱导的细胞增殖使G1期细胞数量减少，S期和G2/M期细胞数量增加，而加入QFTL和贝那普利含药血清后，G1期细胞比例增加（P<0.05或P<0.01），S期细胞比例降低（P<0.01），提示QFTL具有细胞周期抑制作用。与对照组相比，LPS降低了MCs凋亡水平（P<0.05），而QFTL和贝那普利含药血清提高了MCs凋亡水平（P<0.01）。而且，QFTL组与贝那普利组之间差异具有统计学意义（P<0.01）。与对照组相比，细胞周期蛋白D1（cylinD1）、细胞周期蛋白依赖性激酶2（CDK2）和p21的蛋白及mRNA表达水平显著升高（P<0.05或P<0.01），p27降低但无统计学意义（P>0.05）；经QFTL和贝那普利含药血清处理后，cylinD1、CDK2、p21的蛋白及mRNA表达水平降低，p27显著升高（P<0.05或P<0.01）；与贝那普利组相比，QFTL对cylinD1、CDK2的蛋白及mRNA表达水平（P<0.05或P<0.01）和p21蛋白表达（P<0.05）的影响更好。
QFTL通过下调cylinD1、CDK2、p21的蛋白及mRNA表达水平，上调p27表达水平的潜在机制，抑制MCs增殖，促进MCs凋亡。
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