摘要
利用半定量RT-PCR、免疫组化和Western blotting的方法,同时从mRNA水平和蛋白质水平对过敏毒素受体(C3aR)在不同病理阶段的2型糖尿病肾病模型小鼠——db/db小鼠肾脏中的表达情况进行了较为系统的分析.结果发现:a.在糖尿病前的db/db小鼠(4周龄的db/db小鼠),C3aR与作为正常对照的db/m小鼠相比没有明显差异.随着肥胖的加剧,高血糖、蛋白尿的发生和发展,C3aR在db/db小鼠肾脏中的表达显著升高.b.免疫组化分析显示,C3aR广泛地表达于db/m和db/db小鼠肾脏的皮质和髓质,分布于肾脏的上皮细胞中(包括肾小管上皮细胞、肾小球中的脏层上皮细胞(足细胞)和壁层上皮细胞).从部位来看,皮髓交界处的肾小管中C3aR表达量明显要比其他部位的多.在肾小球,C3aR特异地存在于足细胞部位.在db/m小鼠,不同周龄小鼠肾脏中C3aR的表达量并没有明显变化,但在db/db小鼠,从8周龄开始,分布在db/db小鼠肾小管上皮细胞和小球足细胞中的C3aR均随小鼠周龄的增加而增加,至少在时间上,与小鼠糖尿病肾病的发生发展相关,其中尤以足细胞中和皮髓交界处肾小管上皮细胞中的变化最为明显.c.在糖尿病肾病小鼠中高表达C3aR的肾小管上皮细胞常有空泡变性的情况.上述工作印证了先前对2型糖尿病肾病患者肾小球基因表达谱的分析结果,更加明确了C3aR与糖尿病肾病的相关性,同时揭示了C3aR在正常小鼠和糖尿病肾病小鼠肾脏中的表达、分布和变化规律,有利于进一步揭示C3aR的功能及其在糖尿病肾病发生、发展过程中的可能作用,探讨糖尿病肾病的分子机制。
C3aR is a receptor of anaphylatoxin C3a and have been reported associated with inflammation disease.Kidney samples were obtained from db/db mice and their db/m littermates at the age of 4,8,12,16 and 20 weeks.The expression of C3aR was assessed by RT-PCR,Western blotting and immunohistochemistry,and then correlated with biochemical and histological indices of kidney injury.No significant difference of C3aR expression was found in the kidney of db/db mice as compared with the control db/m mice at the age of 4 weeks,a time when all biochemical and histological indices indicated that the mice were in pre-diabetic condition.However,with the development of obesity,hyperglycemia and proteinuria,the renal expression of C3aR in the db/db mice increased as compared with the db/m mice of the same age.Immunohistochemistry analysis revealed that C3aR protein distributed specifically in renal epithelial cells,including tubular epithelial cells,glomerular podocytes and parietal epithelial cells both in db/m and db/db mice.Much more intensity of C3aR immunohistochemical staining was found in the epithelial cells of proximal tubule which just located at the intermedial of cortex and medulla.In the glomerulus,C3aR was found distributed specifically in the podocytes position and no C3aR staining was found in the mesangial and endothelial cells.In diabetic db/db mice,the increased expression of C3aR was found mainly in proximal tubule and podocytes.Interestingly,serious vacuolar degeneration was often observed in tubular cells with higher intenstity of C3aR immunohistochemical staining.All these suggested that C3aR might have something to do with the initiation or/and development of diabetic nephropathy,especially in the pathogenesis of tubular cells and podocytes.
出处
《生物化学与生物物理进展》
SCIE
CAS
CSCD
北大核心
2010年第8期847-854,共8页
Progress In Biochemistry and Biophysics
基金
南京军区南京总医院课题资助项目(2010M027)~~