p53蛋白表达对宫内生长受限大鼠肾脏发育的影响

The influence of p53 protein expression on kidney development in rats with intrauterine growth restriction

目的探讨p53蛋白表达对宫内生长受限(IUGR)大鼠肾脏发育的影响。方法孕鼠随机分为正常组、IUGR组和干预组。正常组孕鼠以普通饲料(含21%蛋白)喂养;IUGR组和干预组大鼠孕期均采用低蛋白饲料(含10%蛋白)喂养。仔鼠出生后,IUGR组母鼠普通饲料喂养;干预组母鼠在普通饲料喂养的基础上,每日于饮水中添加左旋精氨酸(L-Arg)200 mg/kg。仔鼠2月龄取肾脏组织,酸消化法进行肾小球数目计数,观察肾小球和肾小管细胞增生和凋亡情况,以及上皮细胞超微结构;采用Western Blot法,观察7天、21天、2月和3月龄仔鼠肾脏p53蛋白的表达。结果 2月龄时,IUGR组大鼠的肾小球数目低于正常组[(23 647±541)比(27 689±492)个/单侧,P<0.01];IUGR组大鼠肾脏细胞凋亡指数高于正常组[(21.9±2.0)比(16.7±2.5)%,P<0.05],两组增生指数差异无统计学意义(P>0.05);IUGR组和干预组肾小球系膜细胞均有增生,滤过膜足突减少及部分融合,但干预组系膜细胞增生少于IUGR组,正常组系膜细胞无明显增生,滤过膜足突形态正常。各组大鼠肾脏在7天和21天均未见p53蛋白的明显表达,2月龄和3月龄时IUGR组和干预组p53蛋白表达均高于正常组[2月:(0.28±0.03)、(0.21±0.01)比(0.10±0.02);3月:(0.39±0.04)、(0.26±0.02)比(0.17±0.03);P均<0.01],IUGR组高于干预组(P<0.05)。结论 IUGR大鼠肾小球数目减少,并存在肾脏细胞凋亡增加和p53蛋白表达增强,出现肾小球系膜细胞增生、滤过膜足突减少及部分融合。使用L-Arg后可以在一定程度上改善IUGR大鼠肾脏组织结构。

Objective To study the effect of renal p53 in rats with intrauterine growth restriction （IUGR）. Methods protein expression on kidney development Pregnant rats were randomly assigned into normal group, IUGR group and L-Arg treated group. Normal group were fed with normal diet （21% protein）. IUGR group and L-Arg treated group were fed with low-protein diet （10% protein）. During lactation, maternal rats in the three groups were all fed with normal diet. Maternal rats in L-Arg treated group were given additional special drinking water （L-Arg： 200 mg/kg）, while maternal rats in normal and IUGR group were given normal drinking water. Offspring weaned after 21 d and had free access to normal rodent diet and water. When the pups grew up to 2 m, the number of glomernli was counted using acid digestion method, the proliferation and apoptosis of glomerular and tubular cells were studied using TUNEL and Ki-67 immunohistochemistry, and the ultrastructure of epithelial cells was determined by electron microscopy. At 7 d, 21 d, 2 m and 3 m, p53 protein expression in renal tissue was measured by Western blot, respectively. Results At 2 m, the number of glomerulus （ right kidney） in IUGR group was significantly lower than normal group [ （23 647 ±541 ） vs. （27 689 ±492）, P 〈0. 01 ] ; the index of renal cell apoptosis in IUGR group was higher than normal group [ （21.9 ± 2. 0） vs. （16.7 ±2.5）, P 〈 0.05 ]; however, IUGR group and normal group had no statistically significant difference （ P 〉 0. 05 ）. The proliferation of mesangial cells was found in IUGR and L-Arg treated group, but not in normal group. And the extent of proliferation in L-Arg treated group was lesser than IUGR group. A reduction of foot process and partial fusion of foot process could be observed in IUGR group and L-Arg treated group while the foot process morphology in normal group was normal. At 2 m and 3 m, p53 protein level in IUGR group and L-Arg treated group were higher than normal group [ 2 m： （0. 28 ± 0.03） and （0.21±0.01） vs. （0.10±0.02）; 3 m： （0.39±0.04） and （0.26±0.02） vs. （0.17± 0. 03 ） ; P 〈 0.01] , while IUGR group higher than L-Arg treated group （P 〈 0. 05 ）. Conclusions The kidney of IUGR rats showed reduced glomerular number, increased renal cell apoptosis, enhanced p53 protein expression, increased proliferation of glomerular mesangial ceils, decreased foot process, and partial fusion of foot process. And L-Arg could to some extent improve the organizational structure of the kidney in IUGR rats.