摘要
目的探讨达格列净(DAPA)对糖尿病(DM)小鼠肾的保护作用及机制。方法选取10只8周龄SPF级db/db雄性DM小鼠为研究对象。将小鼠随机分为DM组与DAPA组,每组各5只。DM组连续口服喂养水12周;DAPA组连续口服喂养DAPA 12周。定期监测体质量和血糖。12周后,处死小鼠,留取小鼠血清及肾组织,检测小鼠血清中的尿素氮水平。对小鼠肾组织进行HE染色、Masson染色及PAS染色,评价小鼠肾的功能。小鼠肾组织行蛋白免疫印迹法和免疫组化染色检测炎症因子的表达水平。体外应用过氧化氢(H_(2)O_(2))刺激人胚肾细胞(AD293)模拟DM小鼠肾组织炎症,同时,给予达格列净0.4μmol/L、0.8μmol/L的剂量治疗,蛋白免疫印迹法检测细胞内炎症因子及相关基因的表达情况。结果12周后,DAPA组小鼠血糖低于DM组,差异有统计学意义(P<0.05)。与DM组小鼠相比,DAPA组小鼠的肾结构比较完整,肾小球内细胞数目有所增加,但未减小肾小球体积大小;DAPA组的肾小球硬化程度以及肾小球纤维化的现象有所缓解。与DM组比较,DAPA组小鼠血清尿素氮水平降低,肾质量及肾质量/体质量增加,差异有统计学意义(P<0.05)。与DM组比较,DAPA组小鼠肾组织中SGLT2表达量降低,但差异无统计学意义(P>0.05)。DAPA组小鼠肾组织中P-P65(Ser-468)蛋白表达低于DM组,差异有统计学意义(P<0.05)。与H_(2)O_(2)组比较,H_(2)O_(2)+DAPA 0.4μmol/L组、H_(2)O_(2)+DAPA 0.8μmol/L组AD293细胞中SGLT2、P-P65(Ser-468)、P-P65(Ser-536)的蛋白表达明显降低,且H_(2)O_(2)+DAPA 0.8μmol/L组低于H_(2)O_(2)+DAPA 0.4μmol/L组。结论达格列净可能通过影响P65的磷酸化水平调节下游靶基因的活性,减轻炎症,缓解肾组织损伤。
Objective To explore the protective effect and mechanism of Dapagliflozin(DAPA)on the kidneys of diabetes mellitus(DM)mice.Methods A total of 10 SPF db/db male diabetic mice aged 8 weeks were selected as the research object.Mice were randomly divided into DM group and DAPA group,with 5 mice in each group.DM group received oral feeding water for 12 weeks.DAPA group received DAPA orally for 12 weeks.Monitored body mass and blood glucose regularly.After 12 weeks,the mice were sacrificed,and the mouse serum and kidney tissues were collected.At the same time,the level of urea nitrogen in the mouse serum was measured.HE staining,Masson staining and PAS staining were performed to evaluate the renal function of mice.Western blot and immunohistochemical staining were performed to detect the expression levels of inflammatory factors.Human embryonic kidney cells(AD293)were stimulated by hydrogen peroxide(H_(2)O_(2))in vitro to simulate renal tissue inflammation in diabetic mice.Meanwhile,dapagliflozin was given 0.4μmol/L and 0.8μmol/L,and the expression of intracellular inflammatory factors and related genes was detected by western blot.Results After 12 weeks,blood glucose in DAPA group was lower than that in DM group,the difference was statistically significant(P<0.05).Compared with DM group,the renal structure of DAPA group was intact,and the number of cells in the glomerular increased,but the glomerular volume did not decrease.The DAPA group showed remission of glomerular sclerosis and glomerular fibrosis.Compared with DM group,serum urea nitrogen level of DAPA group was decreased,kidney weight and kidney weight/body weight were increased,the differences were statistically significant(P<0.05).Compared with DM group,the expression of SGLT2 in renal tissues of DAPA group was decreased,but the difference was not statistically significant(P>0.05).The expression of P-P65(Ser-468)protein in renal tissues of DAPA group was lower than that of DM group,and the difference was statistically significant(P<0.05).Compared with H_(2)O_(2) group,the protein expressions of SGLT2,P-P65(Ser-468)and P-P65(Ser-536)in AD293 cells in H_(2)O_(2)+DAPA 0.4μmol/L group and H_(2)O_(2)+DAPA 0.8μmol/L group were significantly decreased,and H_(2)O_(2)+DAPA 0.8μmol/L group was lower than H_(2)O_(2)+DAPA 0.4μmol/L group.Conclusion DAPA may affect the activity of downstream target genes by affecting the phosphorylation level of P65,thereby reducing inflammation and alleviating kidney tissue damage.
作者
杨秋旻
李佳荫
张艳
闫承慧
仲崇斌
YANG Qiu-min;LI Jia-yin;ZHANG Yan;YAN Cheng-hui;ZHONG Chong-bin(School of Life Sciences and Health,Northeastern University,Shenyang 110167,China;Department of Cardiology,General Hospital of Northern Theater Command,Shenyang 110016,China)
出处
《临床军医杂志》
CAS
2021年第10期1127-1132,共6页
Clinical Journal of Medical Officers
基金
国家自然科学基金面上项目(82070300)。
关键词
达格列净
糖尿病
炎症
磷酸化
P65
Dapagliflozin
Diabetes
Inflammation
Phosphorylated
P65
