Metformin Protects Type II Diabetic Kidneys by Regulating PARP-1 Activity
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摘要:
目的 研究二甲双胍通过调控多聚（ADP-核糖）聚合酶1（PARP-1）的活性，对2型糖尿病肾脏的保护作用及其机制。 方法 Wistar大鼠分为正常组（n = 12），DN组（n = 12），DN+DPQ组（n = 12），DN+二甲双胍组（n = 12）以及DN+二甲双胍+DPQ组（n = 12）。建模后检测各组大鼠空腹血糖含量，尿素氮含量，肌酐含量以及尿蛋白浓度等生化指标，HE染色和TUNEL染色观察肾脏病理情况，Western blot 检测PARP-1，iNOS，NF-κB及caspase-3的蛋白表达，ELISA检测炎性因子TNF-α及IL-1β的表达，免疫组化测定3-硝基酪氨酸（3-nitrotyrosine，3-NT）的表达。 结果 （1）3组治疗组大鼠的各项生化指标相较于DN组均有下降，其中DN + 二甲双胍 + DPQ组大鼠生化指标改变最为明显（P < 0. 01）；（2）3组治疗组大鼠PARP-1的表达较于DN组下降，其中DN + 二甲双胍 + DPQ组的表达下降最为明显（ P < 0.05）；（3）3组治疗组大鼠肾脏组织的病理变化及肾脏细胞的凋亡较于DN组减缓，其中DN + 二甲双胍 + DPQ组的改善最为明显；（4）3组治疗组大鼠炎性因子的表达以及NF-kB，iNOS及3-NT的表达较于DN组下降，其中DN + 二甲双胍 + DPQ组的表达下降最为明显（ P < 0.05）。 结论 二甲双胍通过调控PARP-1的表达，下调DN模型中NF-kB的表达，抑制NF-kB/iNOS/NO通路，抑制氧化损伤，减轻炎症反应，从而起到在糖尿病导致的高糖环境下，对肾脏的保护作用。 Abstract:Objective To investigate the protective effect of metformin on the kidney in type II diabetes mellitus （T2DM） by regulating the activity of poly （ADP-ribose） polymerase 1 （PARP-1） and investigate the mechanism of this effect. Methods Wistar rats were divided into normal group （n = 12）, DN group （n = 12）, DN + DPQ group （n = 12）, DN + metformin group （n = 12） and DN + metformin + DPQ group （n = 12）. After model establishment, biochemical parameters such as fasting glucose content, urea nitrogen content, creatinine content, and urine protein concentration were measured in each group of rats.HE staining and TUNEL staining were used to observe the renal pathology, Western blot was applied to detect the protein expression of PARP-1, iNOS, NF-κB and caspase-3.The expression of inflammatory factors TNF-α and IL-1β was detected by ELISA, and the expression of 3-nitrotyrosine （3-NT）was determined by immunohistochemistry. Results （1）The biochemical indexes of rats in the three treatment groups decreased compared with those in the DN group, with the most obvious changes in the DN + metformin + DPQ group （P < 0.01） ; （2） The expression of PARP-1 in rats in the three treatment groups decreased compared to the DN group, with the most significant decrease in the DN + metformin + DPQ group （ P < 0.05）; （3） The pathological changes of kidney tissue and apoptosis of kidney cells in the three treatment groups were alleviatedcompared with the DN group, with the most significant decrease in the expression of DN + metformin + DPQ group; （4） The expression of inflammatory factors as well as the expression of NF-kB, iNOS and 3-NT decreased in the three treatment groups compared with the DN group, with the most significant decrease in the DN + metformin + DPQ group （ P < 0.05）. Conclusion Metformin plays a role in protecting the kidney under the high glucose environment caused by diabetes through regulating the expression of PARP-1, down-regulating the expression of NF-kB in the DN model, inhibiting NF-kB/iNOS/NO pathway, inhibiting oxidative damage, and reducing inflammation. -
Key words:
- Diabetic nephropathy /
- Metformin /
- PARP-1 /
- Oxidative stress
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图 1 各组大鼠空腹血糖含量，尿素氮含量，肌酐含量以及尿蛋白浓度
A：空腹血糖含量；B：尿素氮含量；C：肌酐含量；D：尿蛋白浓度。两两组间比较，**P < 0.01，***P < 0.001，****P < 0.000 1。
Figure 1. Fasting glucose content，urea nitrogen content，creatinine content and urine protein concentration in each group of rats
图 2 各组大鼠PARP-1相对蛋白表达
A：各组大鼠PARP-1蛋白表达条带；B：各组大鼠PARP-1相对蛋白表达量。两两组间比较，*P < 0.05，***P < 0.001，****P < 0.0001。
Figure 2. Relative expression of PARP-1 in each group
图 4 二甲双胍通过降低PARP-1表达减轻大鼠肾脏细胞凋亡
A：肾脏细胞凋亡（40*）；B：Caspase-3 在各组中的蛋白表达条带；C：Caspase-3 在各组中的相对蛋白表达。两两组间比较，*P < 0.05，***P < 0.001，****P < 0.000 1。
Figure 4. Metformin attenuates renal cell apoptosis in rats by reducing PARP-1 expression
图 5 各组大鼠TNF-α及IL-1β浓度
A：各组大鼠TNF-α浓度；B：各组大鼠IL-1β浓度。两两组间比较，****P < 0.000 1。
Figure 5. Relative expression of TNF-α and IL-1β in each group
图 6 各组大鼠NF-κB相对蛋白表达
A：各组大鼠NF-κB p65 以及p-P65 蛋白表达条带；B：各组大鼠NF-κB p65 以及p-P65相对蛋白表达量。两两组间比较，**P < 0.01，***P < 0.001，****P < 0.000 1。
Figure 6. Relative expression of NF-κB in each group
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