Leprdb/db小鼠肾损伤机制探讨

  目的  探讨瘦素受体完全缺陷的Leprdb/db小鼠肾损伤机制。  方法  选取28周龄瘦素受体杂合缺陷的Leprdb/+（作为对照）与Leprdb/db雄性小鼠各10只，禁食8 h后，分别测量两组小鼠体质量、空腹血糖（FBG）和糖化血红蛋白（HbA1c）水平。小鼠经股动脉采血后处死。采用试剂盒检测血清中肌酐（CRE）、尿素氮（BUN）、超氧化物歧化酶（SOD）、还原型谷胱甘肽（GSH） 、丙二醛（MDA）的水平。酶联免疫吸附法（ELISA）检测血清中炎性细胞因子白细胞介素-1β（IL-1β）、单核细胞趋化因子-1（MCP-1)、肿瘤坏死因子-α（TNF-α）的表达水平。取肾进行病理观察。Western blot检测肾组织中核因子E2相关因子2（Nrf2）及核因子-κB（NF-κB）蛋白表达。提取肾组织细胞线粒体，Western blot检测线粒体中硫辛酸合成酶(lipoic acid synthase, LIAS)蛋白的表达水平。  结果  与Leprdb/+小鼠相比，Leprdb/db小鼠体质量、FPG、HbA1c、CRE、BUN水平均升高，差异有统计学意义（P<0.05）。病理观察发现，Leprdb/+小鼠肾细胞结构完整，而Leprdb/db小鼠肾小球体积增大，基底膜及毛细血管壁增厚，系膜细胞和系膜基质增多。与Leprdb/+小鼠相比，Leprdb/db小鼠血清中GSH水平降低，MDA和炎性因子MCP-1、IL-1β、TNF-α水平均升高，差异均有统计学意义（P<0.05）；Leprdb/db组小鼠肾脏线粒体内LIAS和肾组织中Nrf2蛋白表达量均降低，而NF-κB蛋白水平升高，差异均有统计学意义（P<0.05）。  结论  28周龄Leprdb/db小鼠存在氧化应激和炎症反应，肾损伤机制可能与LIAS调控Nrf2和NF-κB有关。

Renal Injury and Its Mechanisms in Leprdb/db Mice

  Objective   To study the mechanism of renal injury in Leprdb/db mice with the leptin receptor homozygous deficiency.  Methods  Ten male of 28-week-old Leprdb/+ mice with leptin receptor heterozygous deficiency were selected as control group and ten male Leprdb/db mice with leptin receptor homozygous deficiency were used in this study. After fasting for 8 hours, the body mass, fasting blood glucose (FBG) and glycosylated hemoglobulin (HbA1c) of the mice were measured. Blood of the mice was obtained from femoral artery before euthanasia. Serum creatinine (CRE), blood urea nitrogen (BUN), superoxide dismutase (SOD), glutathione (GSH) and malonaldehyde (MDA) were detected by corresponding kits, and serum interleukin-1β (IL-1β), monocyte chemotactic protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α) were measured using enzyme-linked immunosorbent assay (ELISA) method. The kidney was taken for pathological observation. The expression levels of nuclear factor E2-related factor 2 (Nrf2) and nuclear factor kappa B (NF-κB) in renal were analyzed by Western blotting. The mitochondria of renal was isolated by the corresponding kit. Meanwhile, the expression level of lipoic acid synthase (LIAS) in renal mitochondria was measured by Western blotting.  Results  The body mass, FPG, HbA1c, CRE and BUN levels of the Leprdb/db mice were significantly increased in comparison with the Leprdb/+ mice (P<0.05). Compared with the Leprdb/+ mice, the Leprdb/db mice renal exhibited glomerular hypertrophy, thickened basement membrane and capillary wall, the mesangial matrix expansion and mesangial cell hyperplasia. Compared with the Leprdb/+ mice, the serum level of GSH in the Leprdb/db mice was decreased significantly (P<0.05). The levels of MDA and concentrations of MCP-1, IL-1β and TNF-α in serum of the Leprdb/db mice were higher than those of the Leprdb/+ mice (P<0.05). Compared with the Leprdb/+ mice, the expression of LIAS and Nrf2 protein in the Leprdb/db mice renal were decreased (P<0.05), while the expression of NF-κB protein was increased (P<0.05).  Conclusion  LIAS, Nrf2 and NF-κB might play significant roles through regulation of oxidative stress and inflammation in the renal injury of Leprdb/db mice.