观察阿霉素(ADR)诱导的局灶节段性肾小球硬化(FSGS)模型小鼠肾脏转化生长因子-β1 (TGF-β1)/丝裂原活化蛋白激酶(MAPK)信号通路的表达。

将8周龄雄性BALB/c小鼠(18～23 g)分为实验组和对照组，用ADR尾静脉注射进实验组小鼠，生理盐水注射入对照组，分别收集各组小鼠血、尿及肾脏组织。利用ELISA方法检测尿蛋白及血清肌酐、尿素氮，观察肾脏病理改变，实时荧光定量PCR技术检测转化生长因子-β1(TGF-β1)、Smad7和IV型胶原(Col IV)mRNA水平，Western印迹方法检测磷酸化细胞外信号调节激酶(p-ERK1/2)和磷酸化p38丝裂原活化蛋白激酶(p-p38MAPK)的蛋白水平。采用Graphpad Prism 5软件对结果进行单因素方差分析，P<0.05为差异有统计学意义。

实验组显示在第3天出现24 h尿蛋白增多，第22天升高更明显，与对照组相比差异有统计学意义(t＝ 8.718，P<0.001)；实验组中血清肌酐及尿素氮与对照组相比，第22天升高(t＝5.12，t＝11.7；P<0.001)，第32天达高峰(t＝25.7，t＝16.21；P<0.001)；肾脏病理：实验组第22天可见部分肾小球系膜基质增多和系膜细胞增生，第32天出现系膜基质和系膜细胞轻至中度增生，节段加重，伴足细胞损伤，且可发现血管袢和肾小球囊粘连，符合FSGS的病理特点；荧光实时定量PCR结果显示：实验组第22天时肾脏TGF-β1 mRNA水平明显高于对照组(q＝5.091，P<0.05)，Col IV mRNA相对表达量随着诱导天数而增加，在第32天时达到最高，与对照组相比差异有统计学意义(q＝5.222，P<0.05)；Smad7的表达量在第32天时显著减少，与对照组比较差异有统计学意义(q＝10.64，P<0.01)，与第22天比较差异亦有统计学意义(q＝9.643，P<0.01)；Western印迹结果显示，第22天实验组p-ERK1/2蛋白水平较对照组降低(q＝5.570，P<0.05)，第32天较对照组降低更明显(q＝8.881，P<0.01)；第22天实验组p-p38MAPK表达较对照组增加(q＝30.65，P<0.001)，第32天p-p38MAPK较对照组比较差异仍有统计学意义(q＝31.76，P<0.001)。

FSGS模型小鼠TGF-β/MAPK信号通路的表达量较对照组相比明显增加，提示TGF-β/MAPK通路有可能被激活从而参与FSGS的发病过程。

To observe adriamycin (ADR)-induced expression of transforming growth factor-β (TGF-β) / mitogen activated protein kinase (MAPK) signaling pathway in kidneys of focal segmental glomerulosclerosis (FSGS) model mice.

8-week-old male BALB/c mice (18-23 g) were randomly divided into experimental group and control group. The ADR was injected into the tail vein of mice of experimental group; normal saline was injected into the tail vein of mice of control group. The blood, urine, and renal tissue samples of each group were collected. ELISA method was used for detection of urinary protein, serum creatinine, and blood urea nitrogen; renal pathological changes were observed; real-time fluorescence quantitative PCR was used or detection of TGF-β1, Smad 7, and type IV collagen mRNA; and Western blot was used to detect protein levels of phosphorylaed-extracellular signal regulated kinase (p-ERK1/2) and phosphorylated mitogen-activated protein kinases (p-p38MAPK). Graphpad Prism 5 software was used for single factor analysis of variance. The differences were considered statistically significant when P<0.05.

The experimental group showed that 24-hour urinary protein level increased from the third day, and increased more on the twenty-second day, which was significantly different from that of the control group (t=8.718, P<0.001). Compared with the control group, the experimental group showed higher levels of both serum creatinine and blood urea nitrogen on the 22nd day (t=5.12, t=11.7; P<0.001), as well as on the 32nd day when they peaked (t=25.7, t=16.21; P<0.001). Pathology examination, in the experimental group, showed some glomerular mesangial matrix accumulation and mesangial cell hyperplasia on the 22nd day, and mild to moderate glomerular mesangial matrix accumulation and mesangial cell hyperplasia on the 32nd day, with aggravated segmentation and mild podocyte hyperplasia; adhesion between vascular loops and bowman′s capsule could also be found; and all these changes accorded with the pathological features of FSGS. The results of real-time fluorescence quantitative PCR showed that the level of TGF-β1 mRNA was significantly higher in the experimental group than in the control group (q=5.091, P<0.05) on the 22nd day; the relative expression of collagen IV mRNA increased with the days of induction and peaked on the 32nd day, which was higher in the experimental group than in the control group (q=5.222, P<0.05); and Smad7 expression was significantly lower on the 32nd day in the experimental group than in the control group (q=10.64, P<0.01), and significantly lower than on the 22nd day (q=9.643, P<0.01). The results of Western blot showed that, in the experimental group, the level of p-ERK1/2 protein was lower on the 22nd day than that in the control group (q=5.570, P<0.05), and was more lower on the 32nd day than in the control group (q=8.881, P<0.01). Compared with the control group, the expression of p-p38MAPK in the experimental group was higher (q=30.65, P<0.001) on the 22nd day, and was still higher on the 32nd day (q=31.76, P<0.001).

Compared with the control group, the expression of TGF-β/MAPK signaling pathway in FSGS model mice was significantly increased, indicating that the TGF-β/MAPK pathway might be activated to participate in the pathogenesis of FSGS.