摘要
目的探索活性维生素D2对维生素D受体（VDR）不同表达水平的糖尿病肾病大鼠模型的保护作用及可能机制。方法腹腔注射链脲佐菌素成功建立sD大鼠糖尿病肾病模型，将建模成功的36只大鼠随机分为3组（每组12只）：VDR过表达组（肾脏原位注射含有VDR的慢病毒）、正常表达组（注射单纯载体）和低表达组（注射干扰VDR表达的慢病毒）。在此基础上分别给予每组骨化三醇（即活性维生素D3，VD，0．03μg·kg-1·d-1）治疗或花生油对照处理后，分为6组：VDR过表达＋VD组、VDR过表达＋花生油组、VDR干扰＋VD组、VDR干扰＋花生油组、空载体＋VD组、空载体＋花生油组。37d后采集标本，以实时聚合酶链反应（RT．PCR）及免疫组化方法检测大鼠肾脏组织VDR表达，并进一步检测各组转化生长因子β1（TGF-β1）、人第10号染色体缺失的磷酸酶及张力蛋白同源基因（PTEN）和胶原蛋白I（COL—1）的表达情况。采用HE及Masson染色对大鼠肾脏形态结构及纤维化程度进行分析。采用单因素方差分析（ANOVA）及t检验法比较各组数据。结果RT．PCR及免疫组化结果显示，与VDR过表达＋花生油组相比，VDR过表达＋VD组VDR和PTENmRNA及蛋白表达均升高（t=-19．67、-5．18、-25．57、-10．06，均P〈O．01）；与VDR过表达＋花生油组相比，VDR过表达＋VD组TGF．131和COL-1mRNA及蛋白表达均降低（t=15．62、8．20、28．28、5．81，均P〈0．01）；与VDR干扰＋花生油组相比，VDR干扰＋VD组VDR和FFENmRNA及蛋白表达量均升高（t=53．67、-7．00、-19．81、-8．06，均P〈O．01）；与VDR干扰＋花生油组相比，VDR干扰＋VD组TGF．[31和COL-1mRNA及蛋白表达量均降低（t=12．0、8．50、21．22、5．42，均P〈0．01）；与空载体＋花生油组相比，空载体＋VD组VDR和PTENmRNA及蛋白表达均升高（t=-20．39、-5．46、-25．92、-6．38，均P〈0．01）；与空载体＋花生油组相比，空载体＋VD组TGF．131和COL-1mRNA及蛋白表达量均降低（t=17．80、10．61、47．36、8．23，均P〈0．01）；病理检查显示VDR表达越低组肾脏结构越紊乱、纤维化程度越严重。活性维生素D，干预后各组大鼠的肾脏结构及纤维化程度均可明显改善。结论活性维生素D，可通过VDR途径改善糖尿病肾病的肾脏纤维化。
Objective To explore the effects and mechanism of 1,25-dihydroxyvitamin D3 on the diabetic nephropathy with different levels of vitamin D receptor （VDR） expression in rats. Methods The models of diabetic nephropathy SD rats （n=36） were established by intraperitoneal injection of streptozotoein, then the rat models were divided into 3 groups： VDR over-expression group （kidney injection of VDR virus, n=12）, VDR normal expression group （kidney injection of viral vector, n=12） and VDR low expression group （kidney injection of VDR interfered virus, n=12）. Then the rats in the three groups were given calcitriol （0.03 μg. kg-1· d-1） or peanut oil for 37 days, six rats with one treatment in each group. So therats were divided into 6 groups： VDR over-expression＋VD group, VDR over-expression＋peanut oil group, VDR low expression＋VD group, VDR low expression＋peanut oil group, VDR normal expression＋VD group, VDR normal expression ＋ peanut oil group. The real time-polymerase chain reaction （RT-PCR） and immunohistochemistry staining were used to detect the mRNA and protein levels of VDR, transforming growth factor（TGF）-β1, phosphatase and tensin homolog deleted on chromosome ten （PTEN）, cologen-1 （COL- 1） in kidney. And the HE and Masson staining were used to detect changes of structure and the level of fibrosis in kidney. Data were compared by using t-test and one-way analysis of variance. Results The expressions of VDR and PTEN mRNA and protein in VDR over-expression ＋ VD group were significantly higher than those in VDR over-expression＋peanut oil group （t=- 19.67, -5.18, -25.57, -10.06, all P〈0.01）. The expressions of TGF-[31 and COL-1 mRNA and protein in VDR over-expression＋VD group significantly decreased than those in VDR over-expression＋peanut oil group（t=15.62, 8.20, 28.28, 5.81, all P〈0.01）. The expressions of VDR and PTEN mRNA and protein in VDR low-expression ＋ VD group were significantly higher than those in VDR low-expression ＋ peanut oil group （t=-53.67,- 7.00,- 19.81,-8.06, all P〈0.01）. The expressions of TGF-131 and COL-1 mRNA and protein in VDR low-expression＋VD group decreased significantly than those in VDR low-expression＋peanut oil group（t=12.00, 8.50, 21.22, 5.42, all P〈0.01）. The expressions of VDR and PTEN mRNA and protein in VDR normal expression ＋ VD group were significantly higher than those in VDR normal expression＋peanut oil group （t=-20.39, -5.46, -25.92, -6.38, all P〈0.01）. The expression of TGF-[31 and COL-1 mRNA and protein in VDR normal expression＋VD group significantly decreased than those in VDR normal expression＋peanut oil group （t=17.80, 10.61, 47.36, 8.23, all P〈0.01）. The pathological structure of the kidney in kidney disorders was associated with the low levels of VDR. These fibrosis changes could be improved by calcitriol interventions. Conclusions The 1,25- dihydroxyvitamin D3 has some anti-fibrosis effects on diabetic nephropathy by the pathway of VDR in kidney.
出处
《中华糖尿病杂志》
CAS
CSCD
2015年第11期676-683,共8页
CHINESE JOURNAL OF DIABETES MELLITUS
基金
国家自然科学基金（81160116）
作者简介
通信作者：朱筠，Email=Zhujun6677@163．com
