2型糖尿病肾病大鼠白细胞介素-6、白细胞介素-10、转化生长因子-β_(1)、组蛋白去乙酰化酶4表达及其与血糖、尿蛋白-肌酐比值的相关性

Changes of interleukin-6,interleukin-10,transforming growth factor-β_(1)and histone deacetylases 4 expression and their correlations with blood glucose and urinary protein-creatinine ratio in rats with type 2 diabetic nephropathy

目的探讨2型糖尿病肾病(DN)大鼠白细胞介素-6(IL-6)、白细胞介素-10(IL-10)、转化生长因子-β_(1)(TGF-β_(1))、组蛋白去乙酰化酶4(HDAC4)表达及其与血糖、尿蛋白-肌酐比值(UACR)的相关性。方法采用随机数字表法将64只健康雄性Sprague Dawley大鼠分为对照组(n=32)和DN组(n=32)。DN组大鼠给予高脂高糖饲料及腹腔注射链脲佐菌素40 mg·kg^(-1)制备DN模型,DN模型大鼠造模成功后正常饮食饮水。对照组大鼠正常饮食饮水,经腹腔注射等剂量柠檬酸-柠檬酸钠缓冲液。于造模后第0、4、8、12周分批处死2组大鼠,于每次处死前称大鼠体质量;使用血糖仪及配套试纸检测大鼠血糖;收集尿液,使用全自动生物化学分析仪检测大鼠尿微量白蛋白、尿肌酐水平,并计算UACR;于大鼠处死后立即取出双侧肾脏,其中一侧肾脏去除包膜后称肾质量,并计算肾脏指数(KI)。采用Western blot法检测2组大鼠肾组织中IL-6、IL-10、TGF-β_(1)、HDAC4蛋白表达,采用酶联免疫吸附试验(ELISA)法检测2组大鼠血清中IL-6、IL-10、TGF-β_(1)和HDAC4水平,采用Pearson相关分析DN大鼠血清IL-6、IL-10、TGF-β_(1)、HDAC4水平与血糖、UACR的相关性。结果造模后第0、4、8、12周,DN组大鼠血糖水平及UACR均显著高于对照组(P<0.05)。造模后第0周,2组大鼠的体质量比较差异无统计学意义(P>0.05);造模后第4、8、12周,DN组大鼠的体质量均显著低于对照组(P<0.05)。造模后第0、4周,2组大鼠的KI比较差异无统计学意义(P>0.05);造模后第8、12周,DN组大鼠的KI显著高于对照组(P<0.05)。造模后第0、4、8、12周,DN组大鼠血清中IL-6、HDAC4、TGF-β_(1)水平均显著高于对照组,IL-10水平显著低于对照组(P<0.05)。造模后第0周,DN组大鼠肾组织中IL-10相对表达量显著高于对照组(P<0.05);造模后第8周,DN组大鼠肾组织中IL-10相对表达量显著低于对照组(P<0.05);造模后第4、12周,2组大鼠肾组织中IL-10相对表达量比较差异无统计学意义(P>0.05)。造模后第0、4、8、12周,DN组大鼠肾组织中IL-6、HDAC4相对表达量均显著高于对照组(P<0.05)。造模后第0周,2组大鼠肾组织中TGF-β_(1)相对表达量比较差异无统计学意义(P>0.05);造模后第4、8、12周,DN组大鼠肾组织中TGF-β_(1)相对表达量显著高于对照组(P<0.05)。Pearson相关性分析显示,DN组大鼠血清IL-6、TGF-β_(1)水平与血糖呈正相关(r=0.614、0.514,P<0.05),血清IL-10水平与血糖呈负相关(r=-0.448,P<0.05),血清HDAC4水平与血糖无相关性(r=-0.177,P>0.05)。DN组大鼠血清IL-6、TGF-β_(1)水平与UACR呈正相关(r=0.405、0.470,P<0.05),血清IL-10、HDAC4水平与UACR无相关性(r=-0.134、-0.124,P>0.05)。结论DN大鼠IL-6、TGF-β_(1)、HDAC4表达上调,IL-10表达下调;DN大鼠血清IL-6、TGF-β_(1)水平与血糖、UACR呈显著正相关,血清IL-10水平与血糖呈显著负相关。

Objective To explore the expressions of interleukin-6(IL-6),interleukin-10(IL-10),transforming growth factor-β_(1)(TGF-β_(1))and histone deacetylases 4(HDAC4)and their correlations with blood glucose and urinary protein-creatinine ratio(UACR)in rats with type 2 diabetic nephropathy(DN).Methods A total of 64 healthy male Sprague Dawley rats were randomly divided into the control group(n=32)and the DN group(n=32)using a random number table method.The rats in the DN group were given high fat and high sugar feed and intraperitoneal injection of 40 mg·kg^(-1)of streptozotocin to prepare DN model.After successful modeling,DN model rats ate and drank water normally.The rats in the control group were given normal diet and drinking water,and an equal dose of citrate sodium citrate buffer was injected intraperitoneally.The rats in the two groups were euthanized in batches at 0,4,8,and 12 weeks after modeling,and their body weight was measured before each execution;the blood glucose level of rats was detected by blood glucose meter and matching test strips;the levels of microalbumin and creatinine in urine of rats were detected by automatic biochemical analyzer,and UACR was calculated;immediately after the rat was euthanized,both kidneys were removed.After removing the capsule from one side of the kidney,the kidney mass was weighted and the kidney index(KI)was calculated.The expressions of IL-6,IL-10,TGF-β_(1)and HDAC4 protein in renal tissue of rats in the two groups were detected by Western blot method.The levels of IL-6,IL-10,TGF-β_(1)and HDAC4 in serum of rats in the two groups were detected by enzyme-linked immunosorbent assay.The correlation between serum IL-6,IL-10,TGF-β_(1)and HDAC4 levels and blood glucose,UACR in DN rats was analyzed by Pearson correlation analysis.Results At 0,4,8 and 12 weeks after modeling,the blood glucose levels and UACR of rats in the DN group were significantly higher than those in the control group(P<0.05).At 0 week after modeling,there was no significant difference in body mass of rats between the two groups(P>0.05);at 4,8 and 12 weeks after modeling,the body mass of rats in the DN group was significantly lower than that in the control group(P<0.05).At 0 and 4 weeks after modeling,there was no significant difference in KI of rats between the two groups(P>0.05);at 8 and 12 weeks after modeling,the KI of rats in the DN group was significantly higher than that in the control group(P<0.05).At 0,4,8 and 12 weeks after modeling,the levels of IL-6,HDAC4 and TGF-β_(1)in the serum of rats in the DN group were significantly higher than those in the control group,while the IL-10 level was significantly lower than that in the control group(P<0.05).At 0 week after modeling,the relative expression level of IL-10 in the kidney tissue of rats in the DN group was significantly higher than that in the control group(P<0.05);at 8 weeks after modeling,the relative expression level of IL-10 in the kidney tissue of rats in the DN group was significantly lower than that in the control group(P<0.05);at 4 and 12 weeks after modeling,there was no significant difference in the relative expression level of IL-10 in the renal tissues of rats between the two groups(P>0.05).At 0,4,8 and 12 weeks after modeling,the relative expression levels of IL-6 and HDAC4 in the renal tissue of rats in the DN group were significantly higher than those in the control group(P<0.05).At 0 week after modeling,there was no significant difference in the relative expression level of TGF-β_(1)in renal tissue of rats between the two groups(P>0.05);at 4,8 and 12 weeks after modeling,the relative expression level of TGF-β_(1)in renal tissue of rats in the DN group was significantly higher than that in the control group(P<0.05).Pearson correlation analysis showed that the serum IL-6 and TGF-β_(1)levels of rats in the DN group was positively correlated with blood glucose(r=0.614,0.514;P<0.05),serum IL-10 level was negatively correlated with blood glucose(r=-0.448,P<0.05),and there was no significant correlation between the serum HDAC4 level and blood glucose(r=-0.177,P>0.05).The serum IL-6 and TGF-β_(1)levels of rats in the DN group was positively correlated with UACR(r=0.405,0.470;P<0.05),and there was no significant correlation between the serum IL-10,HDAC4 levels and UACR(r=-0.134,-0.124;P>0.05).Conclusion The expressions of IL-6,TGF-β_(1)and HDAC4 is upregulated in DN rats,while the expression of IL-10 is downregulated.The serum IL-6 and TGF-β_(1)levels in DN rats is significantly positively correlated with blood glucose and UACR,while the serum IL-10 level is significantly negatively correlated with blood glucose.