白藜芦醇对糖尿病大鼠肾脏叉头转录因子O1的影响及机制研究
|
摘要
背景与目的
糖尿病肾病(diabetic nephropathy, DN)是糖尿病微血管并发症之一,是引起终末期肾功能衰竭的主要因素。目前其发生机制尚不完全清楚。近年来大量研究表明,活性氧(reactive oxygen species, ROS)是糖尿病血管并发症及糖尿病本身发生发展的一个主要因素,其与DN的关系尤为密切。叉头状转录因子O1 (Forkhead transcription factor O1, FoxO1)是Fox蛋白家族中的一员,其可通过上调抗氧化靶基因的水平,如,过氧化氢酶(catalase, CAT),起到抗氧化应激的作用。
沉默信息调节因子2(Sir2,哺乳动物为Sirt1)是细胞应激时FoxO乙酰化和转录功能的重要调节子。Sirt1/FoxO1通路对糖尿病肾病发生发展的影响国内外尚无报道,此通路对糖尿病肾脏是否起保护作用有待探讨。白藜芦醇(resveratrol, Res)是目前发现的最强的Sirt1激活剂,其可使Sirt1的表达增加。本实验应用白藜芦醇治疗实验性糖尿病大鼠,探讨白藜芦醇/Sirt1/FoxO1/CAT通路对糖尿病大鼠肾脏的保护作用,进一步研究DN发生发展的机制。
材料与方法
健康、雄性清洁级SD大鼠42只,体重(220±20)g,随机选取32只大鼠建立糖尿病(DM)模型,其余10只大鼠作为正常对照组(A组)。DM大鼠造模方法如下：禁食12h后,按60mg/kg次性大剂量腹腔注射1%STZ溶液,A组注射相当体积的柠檬酸钠-柠檬酸缓冲液。72h后测定血糖(BG)>16.7mmol/L,为DM成模标准(4只失败)。将糖尿病鼠再随机分为糖尿病未治疗组(B组)和糖尿病白藜芦醇治疗组(C组)。自成模之日起,C组每日固定时间以白藜芦醇混悬液30mg/kg,1次/日,灌胃12周。A、B组给予相同剂量的生理盐水。各组大鼠自由进食饮水,实验中不予任何降血糖药物。实验第12周末,代谢笼收集24小时尿,离心,保存于4℃冰箱,测定24小时尿蛋白(UPro/24h)及尿白蛋白定量(UAIb)。麻醉后腹腔静脉取血,分离血清检测血肌酐(Scr)、尿素氮(BUN)。剥离双肾,右肾称重后取肾皮质置于4%多聚甲醛中固定,光镜观察肾小球病理学变化,免疫组化检测大鼠肾小球胶原Ⅳ、纤连蛋白表达水平；取左肾上极1mm3大小的组织块,迅速用4%冷伐二醛溶液固定,透射电子显微镜观察肾脏超微结构变化,剩余肾皮质迅速置于-70。C冰箱里,分光光度计检测肾皮质丙二醛(MDA)含量和总超氧化物歧化酶(SOD)活性,RT-PCR检测肾皮质沉默信息调节因子1 (Sirtl)、FoxO1、过氧化氢酶(CAT) mRNA表达水平,Western blot检测FoxO1蛋白及其磷酸化的表达。
结果
1.实验12周末,与正常对照组比较,糖尿病未治疗组BG(28.2±2.7)mmol/L、KI (8.1±0.8)×10-3、UPro/24h (26.8±4.1) mg/24h、UAIb (0.89±0.13) mg/24h、Scr (73.5±10.9))μmol/L和BUN (15.3±1.4) mmol/L水平显著升高(均P<0.05), BW (250±27) g水平明显降低(P<0.05)；与糖尿病未治疗组比较,白藜芦醇治疗组KI(5.2±0.5)×10-3、Upro/24h(16.9±2.6)mg/24h、UAIb (0.65±0.11) mg/24h、Scr (56.7±5.9)μmol/L和BUN (11.3±1.1) mmol/L水平明显降低(均P<0.05),但仍高于正常对照组(P<0.05), BG (27.8±3.3) mmol/L水平稍有下降、BW(271±31)g水平稍有上升,但差异无统计学意义(P>0.05)。
2.糖尿病未治疗组肾皮质MDA含量(3.47±0.24) nmol/mg prot比正常对照组(1.35±0.29)nmol/mg prot明显升高(P<0.05),肾皮质总SOD活性(23.51±7.79)U/mg prot较正常对照组(65.74±8.71) U/mg prot显著降低(P<0.05)；与糖尿病未治疗组相比,白藜芦醇治疗组肾皮质MDA含量(2.48±0.27) nmol/mg prot显著减少(P<0.05),总SOD活性(43.31±5.57) U/mg prot则明显升高(P<0.05)。
3.免疫组化结果显示,与正常对照组比较(分别为1.09±0.11,1.81±0.44),糖尿病未治疗组大鼠肾小球纤连蛋白(10.65±1.27)、胶原Ⅳ(20.11±1.28))表达水平升高(均P<0.05)；白藜芦醇治疗组大鼠肾小球中两者蛋白的表达水平(分别为6.26±1.04,10.08±1.01)较糖尿病未治疗组均明显降低(均P<0.05),但仍高于正常对照组(P<0.05)。
4.糖尿病未治疗组大鼠肾皮质Sirt1 (0.63±0.04)、FoxO1 (0.37±0.03)、CAT mRNA (0.22±0.03)表达水平较正常对照组明显降低(分别为0.96±0.08,0.69±0.05,0.48±0.04；均P<0.05)；与糖尿病未治疗组比较,白藜芦醇治疗组糖尿病大鼠肾皮质三者的mRNA的表达水平显著增加(分别为0.83±0.05,0.53±0.04,0.40±0.03；均P<0.05),但仍高于正常对照组(P<0.05)。
5.糖尿病未治疗组FoxO1磷酸化水平(2.15±0.08)较对照组(1.14±0.08)明显升高,白藜芦醇治疗组FoxO1磷酸化水平(1.72±0.07)较糖尿病未治疗组显著降低(P<0.05),但仍高于对照组(P<0.05); FoxO1蛋白表达各组间比较无显著性差异(分别为0.45±0.09,0.48±0.08,0.47±0.09；均P>0.05)。
6.肾脏病理检测显示,光镜HE染色显示正常对照组大鼠肾小球结构未见异常,糖尿病未治疗组大鼠肾小球体积增大,系膜细胞增生,肾小球系膜基质增多,基底膜增厚。电镜超微结构显示正常对照组大鼠肾小球结构清晰,基底膜均匀无增厚,上皮细胞足突均匀分布,糖尿病未治疗组大鼠肾小球基底膜显著增厚,厚薄不均,上皮细胞足突融合。白藜芦醇治疗组大鼠肾小球病理变化明显改善,表现为系膜细胞增生程度降低,肾小球基底膜增厚程度减轻,足突融合减轻。
结论
1.糖尿病大鼠肾脏中FoxO1 mRNA水平降低,FoxO1蛋白磷酸化水平升高。糖尿病大鼠肾脏中FoxO1的抗氧化应激作用降低。
2.白藜芦醇增加糖尿病大鼠肾脏FoxO1 mRNA表达,降低FoxO1蛋白磷酸化水平。
3.白藜芦醇可能通过增加Sirt1的表达使FoxO1的表达及活性增加,继而FoxO1上调其抗氧化靶基因CAT的表达,降低肾脏氧化应激反应,减缓或抑制DN的发生。
4.对Sirtl/FoxO1通路的调节可能成为预防和治疗DN的潜在治疗靶点。
Background & Objective
Diabetes is increasing in an epidemic proportion throughout the world and diabetic nephropathy, one of microvascular complications of diabetes, is the leading cause of end-stage renal disease (ESRD). Numerous factors contribute to the pathogenesis and progression of DN. Experimental and clinical evidence indicates that excessive oxidative stress may contribute to the initiation and development of diabetic nephropathy. Transcription factor FoxO family members and FoxO1 in particular play important roles in oxidative stress resistance, which increases its antioxidant target genes (CAT) to reduce oxidative stress.
SIRT1 is a mediator of resveratrol action in FoxO1-mediated gene expression, and the dephosphorylated form of FoxO1, which is distributed in the nucleus, is deacetylated by SIRT1. Resveratrol is known to be an activator of sirtuinl (SIRT1). To investigate the effect of FoxO1 on oxidant stress in streptozotocin-induced diabetic rats, we sought to determine the effect of resveratrol on FoxO1. We determined the relationship of FoxO1 and Sirt1 activation to oxidative stress in kidney of DN.
Materials & Methods
Male Sprague-Dawley rats (220±20 g) were obtained from Experimental Animal Center of Henan Province. Diabetes was induced by a single intraperitoneal injection of 60 mg/kg STZ (dissolved in 0.05M citrate buffer, pH 4.5) after fasting for 12 h. Blood glucose was measured to confirm the development of diabetes 3 days after STZ injection. Animals in the diabetic group were excluded from the study if plasma glucose was less than 16.7mM. Rats that received an injection of diluent buffer alone served as control. After developed diabetes, rats were divided into two groups: diabetic rats without treatment and diabetic rats treated with resveratrol. At the end of 12-week study,24-hour urine from each rat was collected for 24 hours urinary protein (24 hUPro) and urine albumin (UAIb) examination,then the rats were anesthetized with pentobarbital sodium (50 mg/kg body wt) and blood was collected from abdominal vein for Blood glucose (BG), blood urea nitrogen (BUN) and serum creatinine (Scr) detection. Kidney weight/body weight ratio (KI) of rats were measured and calculated. The right kidney from each rat was fixed in 4% paraformaldehyde and embedded in paraffin for immunohistochemistry and HE stain. The renal cortex of the left kidney from each rat was cut into small pieces (1 mm3) for electron microscope examination and the left cortical tissues from rats kidneys were removed and stored at-70℃for further study. The content of malondiadehyde (MDA) and the activity of superoxide dismutas (SOD) was detected using spectrophotometer.. Expressions of collagen IV and fibronectin in glomeruli were detected by immunohistochemical staining. The expressions of silent information regulator 1 (Sirt1), Foxol and catalase (CAT) mRNA were respectively detected using RT-RCR. The expressions of FoxO1 protein and FoxO1 phosphorylation level were detected using Western blot.
Results
1. Compared with normal group, BG (28.2±2.7) mmol/L. KI (8.1±0.8)×10-3、UPro/24h (26.8±4.1) mg/24h、UAIb (0.89±0.13) mg/24h、Scr (73.5±10.9)μmol/ L and BUN (15.3±1.4) mmol/L levels were elevated significantly (all P< 0.05), and body weight (250±27) g was decreased in diabetic rats without treatment (P< 0.05). We also observed that KI (5.2±0.5)×10-3、Upro/24h (16.9±2.6) mg/24h, UAIb (0.65±0.11) mg/24h、Scr (56.7±5.9)μmol/Land BUN (11.3±1.1) mmol/L levels were decrease significantly in diabetic rats treated with resveratrol (RSV) (all P< 0.05). But there were no significant differences in BW and BG levels between diabetic rats treated with resveratrol [(271±31) g and (27.8±3.3) mmol/L, respectively] and without treatment (P>0.05).
2. Compared with normal group[(1.35±0.29) nmol/mg prot and (65.74±8.71) U/mg prot, respectively], level of MDA generation (3.47±0.24) nmol/mg prot was increased markedly and the activity of SOD (23.51±7.79) U/mg prot was decreased in group of diabetic rats without treatment (all P< 0.05). Similarly, after treated with resveratrol for 12 weeks, level of MDA generation (2.48±0.27) nmol/mg prot was significantly decreased and the activity of SOD (43.31±5.57) U/mg prot in diabetic rats treated with resveratrol (all P< 0.05).
3. Compared with normal group (1.09±0.11 and 1.81±0.44, respectively) the expressions of fibronectin (10.65±1.27) and collagen IV (20.11±1.28) in glomeruli were significantly increased in group of diabetic rats without treatment (all P< 0.05). Similarly, after treated with resveratrol for 12 weeks, the expressions of collagenⅣand fibronectin (6.26±1.04 and 10.08±1.01, respectively) in glomeruli were significantly decreased in diabetic rats treated with resveratrol (all P< 0.05).
4. The expressions of Sirtl (0.63±0.04), FoxO1 (0.37±0.03) and CAT (0.22±0.03) mRNA in kidneys of diabetic rats without treatment were decreased compared with normal group (0.96±0.08,0.69±0.05 and 0.48±0.04, respectively; all P<0.05).After treated with resveratrol for 12 weeks, the relative amounts of mRNA (0.83±0.05, 0.53±0.04 and 0.40±0.03, respectively; all P<0.05) were higher in diabetic rat treated with resveratrol compared to the diabetic rats without treatment (all P< 0.05).
5. Western blot showed that there was no difference FoxO1 protein expression among groups (0.45±0.09,0.48±0.08 and 0.47±0.09, respectively; all P>0.05), but FoxO1 phosphorylation level in group of diabetic rats without treatment (2.15±0.08) was highter than those of normal group (1.14±0.08, P< 0.05), and FoxO1 phosphorylation level in group of diabetic rats treated with resveratrol (1.72±0.07) was lower than those of diabetic rats without treatment (P< 0.05).
Conclusions
1. The expression of FoxO1 mRNA was decreased, and FoxO1 phosphorylation level was increased in kidney of diabetic rats.The role in defensing against oxidative stress of FoxO1 was decreased in kidney of diabetic rats.
2. Resveratrol significantly increased the FoxOl mRNA and decreased FoxO1 phosphorylation levels in kidneys of diabetic rats treated with resveratrol.
3. Sirtl activated by resveratrol may increase the expression of FoxO1, which increases its antioxidant target genes (CAT) to reduce ROS level in the kidneys.
4. Alterations of the Sirtl/FoxO1 pathway may be considered a potential therapeutic target for DN.
糖尿病肾病(diabetic nephropathy, DN)是糖尿病微血管并发症之一,是引起终末期肾功能衰竭的主要因素。目前其发生机制尚不完全清楚。近年来大量研究表明,活性氧(reactive oxygen species, ROS)是糖尿病血管并发症及糖尿病本身发生发展的一个主要因素,其与DN的关系尤为密切。叉头状转录因子O1 (Forkhead transcription factor O1, FoxO1)是Fox蛋白家族中的一员,其可通过上调抗氧化靶基因的水平,如,过氧化氢酶(catalase, CAT),起到抗氧化应激的作用。
沉默信息调节因子2(Sir2,哺乳动物为Sirt1)是细胞应激时FoxO乙酰化和转录功能的重要调节子。Sirt1/FoxO1通路对糖尿病肾病发生发展的影响国内外尚无报道,此通路对糖尿病肾脏是否起保护作用有待探讨。白藜芦醇(resveratrol, Res)是目前发现的最强的Sirt1激活剂,其可使Sirt1的表达增加。本实验应用白藜芦醇治疗实验性糖尿病大鼠,探讨白藜芦醇/Sirt1/FoxO1/CAT通路对糖尿病大鼠肾脏的保护作用,进一步研究DN发生发展的机制。
材料与方法
健康、雄性清洁级SD大鼠42只,体重(220±20)g,随机选取32只大鼠建立糖尿病(DM)模型,其余10只大鼠作为正常对照组(A组)。DM大鼠造模方法如下：禁食12h后,按60mg/kg次性大剂量腹腔注射1%STZ溶液,A组注射相当体积的柠檬酸钠-柠檬酸缓冲液。72h后测定血糖(BG)>16.7mmol/L,为DM成模标准(4只失败)。将糖尿病鼠再随机分为糖尿病未治疗组(B组)和糖尿病白藜芦醇治疗组(C组)。自成模之日起,C组每日固定时间以白藜芦醇混悬液30mg/kg,1次/日,灌胃12周。A、B组给予相同剂量的生理盐水。各组大鼠自由进食饮水,实验中不予任何降血糖药物。实验第12周末,代谢笼收集24小时尿,离心,保存于4℃冰箱,测定24小时尿蛋白(UPro/24h)及尿白蛋白定量(UAIb)。麻醉后腹腔静脉取血,分离血清检测血肌酐(Scr)、尿素氮(BUN)。剥离双肾,右肾称重后取肾皮质置于4%多聚甲醛中固定,光镜观察肾小球病理学变化,免疫组化检测大鼠肾小球胶原Ⅳ、纤连蛋白表达水平；取左肾上极1mm3大小的组织块,迅速用4%冷伐二醛溶液固定,透射电子显微镜观察肾脏超微结构变化,剩余肾皮质迅速置于-70。C冰箱里,分光光度计检测肾皮质丙二醛(MDA)含量和总超氧化物歧化酶(SOD)活性,RT-PCR检测肾皮质沉默信息调节因子1 (Sirtl)、FoxO1、过氧化氢酶(CAT) mRNA表达水平,Western blot检测FoxO1蛋白及其磷酸化的表达。
结果
1.实验12周末,与正常对照组比较,糖尿病未治疗组BG(28.2±2.7)mmol/L、KI (8.1±0.8)×10-3、UPro/24h (26.8±4.1) mg/24h、UAIb (0.89±0.13) mg/24h、Scr (73.5±10.9))μmol/L和BUN (15.3±1.4) mmol/L水平显著升高(均P<0.05), BW (250±27) g水平明显降低(P<0.05)；与糖尿病未治疗组比较,白藜芦醇治疗组KI(5.2±0.5)×10-3、Upro/24h(16.9±2.6)mg/24h、UAIb (0.65±0.11) mg/24h、Scr (56.7±5.9)μmol/L和BUN (11.3±1.1) mmol/L水平明显降低(均P<0.05),但仍高于正常对照组(P<0.05), BG (27.8±3.3) mmol/L水平稍有下降、BW(271±31)g水平稍有上升,但差异无统计学意义(P>0.05)。
2.糖尿病未治疗组肾皮质MDA含量(3.47±0.24) nmol/mg prot比正常对照组(1.35±0.29)nmol/mg prot明显升高(P<0.05),肾皮质总SOD活性(23.51±7.79)U/mg prot较正常对照组(65.74±8.71) U/mg prot显著降低(P<0.05)；与糖尿病未治疗组相比,白藜芦醇治疗组肾皮质MDA含量(2.48±0.27) nmol/mg prot显著减少(P<0.05),总SOD活性(43.31±5.57) U/mg prot则明显升高(P<0.05)。
3.免疫组化结果显示,与正常对照组比较(分别为1.09±0.11,1.81±0.44),糖尿病未治疗组大鼠肾小球纤连蛋白(10.65±1.27)、胶原Ⅳ(20.11±1.28))表达水平升高(均P<0.05)；白藜芦醇治疗组大鼠肾小球中两者蛋白的表达水平(分别为6.26±1.04,10.08±1.01)较糖尿病未治疗组均明显降低(均P<0.05),但仍高于正常对照组(P<0.05)。
4.糖尿病未治疗组大鼠肾皮质Sirt1 (0.63±0.04)、FoxO1 (0.37±0.03)、CAT mRNA (0.22±0.03)表达水平较正常对照组明显降低(分别为0.96±0.08,0.69±0.05,0.48±0.04；均P<0.05)；与糖尿病未治疗组比较,白藜芦醇治疗组糖尿病大鼠肾皮质三者的mRNA的表达水平显著增加(分别为0.83±0.05,0.53±0.04,0.40±0.03；均P<0.05),但仍高于正常对照组(P<0.05)。
5.糖尿病未治疗组FoxO1磷酸化水平(2.15±0.08)较对照组(1.14±0.08)明显升高,白藜芦醇治疗组FoxO1磷酸化水平(1.72±0.07)较糖尿病未治疗组显著降低(P<0.05),但仍高于对照组(P<0.05); FoxO1蛋白表达各组间比较无显著性差异(分别为0.45±0.09,0.48±0.08,0.47±0.09；均P>0.05)。
6.肾脏病理检测显示,光镜HE染色显示正常对照组大鼠肾小球结构未见异常,糖尿病未治疗组大鼠肾小球体积增大,系膜细胞增生,肾小球系膜基质增多,基底膜增厚。电镜超微结构显示正常对照组大鼠肾小球结构清晰,基底膜均匀无增厚,上皮细胞足突均匀分布,糖尿病未治疗组大鼠肾小球基底膜显著增厚,厚薄不均,上皮细胞足突融合。白藜芦醇治疗组大鼠肾小球病理变化明显改善,表现为系膜细胞增生程度降低,肾小球基底膜增厚程度减轻,足突融合减轻。
结论
1.糖尿病大鼠肾脏中FoxO1 mRNA水平降低,FoxO1蛋白磷酸化水平升高。糖尿病大鼠肾脏中FoxO1的抗氧化应激作用降低。
2.白藜芦醇增加糖尿病大鼠肾脏FoxO1 mRNA表达,降低FoxO1蛋白磷酸化水平。
3.白藜芦醇可能通过增加Sirt1的表达使FoxO1的表达及活性增加,继而FoxO1上调其抗氧化靶基因CAT的表达,降低肾脏氧化应激反应,减缓或抑制DN的发生。
4.对Sirtl/FoxO1通路的调节可能成为预防和治疗DN的潜在治疗靶点。
Background & Objective
Diabetes is increasing in an epidemic proportion throughout the world and diabetic nephropathy, one of microvascular complications of diabetes, is the leading cause of end-stage renal disease (ESRD). Numerous factors contribute to the pathogenesis and progression of DN. Experimental and clinical evidence indicates that excessive oxidative stress may contribute to the initiation and development of diabetic nephropathy. Transcription factor FoxO family members and FoxO1 in particular play important roles in oxidative stress resistance, which increases its antioxidant target genes (CAT) to reduce oxidative stress.
SIRT1 is a mediator of resveratrol action in FoxO1-mediated gene expression, and the dephosphorylated form of FoxO1, which is distributed in the nucleus, is deacetylated by SIRT1. Resveratrol is known to be an activator of sirtuinl (SIRT1). To investigate the effect of FoxO1 on oxidant stress in streptozotocin-induced diabetic rats, we sought to determine the effect of resveratrol on FoxO1. We determined the relationship of FoxO1 and Sirt1 activation to oxidative stress in kidney of DN.
Materials & Methods
Male Sprague-Dawley rats (220±20 g) were obtained from Experimental Animal Center of Henan Province. Diabetes was induced by a single intraperitoneal injection of 60 mg/kg STZ (dissolved in 0.05M citrate buffer, pH 4.5) after fasting for 12 h. Blood glucose was measured to confirm the development of diabetes 3 days after STZ injection. Animals in the diabetic group were excluded from the study if plasma glucose was less than 16.7mM. Rats that received an injection of diluent buffer alone served as control. After developed diabetes, rats were divided into two groups: diabetic rats without treatment and diabetic rats treated with resveratrol. At the end of 12-week study,24-hour urine from each rat was collected for 24 hours urinary protein (24 hUPro) and urine albumin (UAIb) examination,then the rats were anesthetized with pentobarbital sodium (50 mg/kg body wt) and blood was collected from abdominal vein for Blood glucose (BG), blood urea nitrogen (BUN) and serum creatinine (Scr) detection. Kidney weight/body weight ratio (KI) of rats were measured and calculated. The right kidney from each rat was fixed in 4% paraformaldehyde and embedded in paraffin for immunohistochemistry and HE stain. The renal cortex of the left kidney from each rat was cut into small pieces (1 mm3) for electron microscope examination and the left cortical tissues from rats kidneys were removed and stored at-70℃for further study. The content of malondiadehyde (MDA) and the activity of superoxide dismutas (SOD) was detected using spectrophotometer.. Expressions of collagen IV and fibronectin in glomeruli were detected by immunohistochemical staining. The expressions of silent information regulator 1 (Sirt1), Foxol and catalase (CAT) mRNA were respectively detected using RT-RCR. The expressions of FoxO1 protein and FoxO1 phosphorylation level were detected using Western blot.
Results
1. Compared with normal group, BG (28.2±2.7) mmol/L. KI (8.1±0.8)×10-3、UPro/24h (26.8±4.1) mg/24h、UAIb (0.89±0.13) mg/24h、Scr (73.5±10.9)μmol/ L and BUN (15.3±1.4) mmol/L levels were elevated significantly (all P< 0.05), and body weight (250±27) g was decreased in diabetic rats without treatment (P< 0.05). We also observed that KI (5.2±0.5)×10-3、Upro/24h (16.9±2.6) mg/24h, UAIb (0.65±0.11) mg/24h、Scr (56.7±5.9)μmol/Land BUN (11.3±1.1) mmol/L levels were decrease significantly in diabetic rats treated with resveratrol (RSV) (all P< 0.05). But there were no significant differences in BW and BG levels between diabetic rats treated with resveratrol [(271±31) g and (27.8±3.3) mmol/L, respectively] and without treatment (P>0.05).
2. Compared with normal group[(1.35±0.29) nmol/mg prot and (65.74±8.71) U/mg prot, respectively], level of MDA generation (3.47±0.24) nmol/mg prot was increased markedly and the activity of SOD (23.51±7.79) U/mg prot was decreased in group of diabetic rats without treatment (all P< 0.05). Similarly, after treated with resveratrol for 12 weeks, level of MDA generation (2.48±0.27) nmol/mg prot was significantly decreased and the activity of SOD (43.31±5.57) U/mg prot in diabetic rats treated with resveratrol (all P< 0.05).
3. Compared with normal group (1.09±0.11 and 1.81±0.44, respectively) the expressions of fibronectin (10.65±1.27) and collagen IV (20.11±1.28) in glomeruli were significantly increased in group of diabetic rats without treatment (all P< 0.05). Similarly, after treated with resveratrol for 12 weeks, the expressions of collagenⅣand fibronectin (6.26±1.04 and 10.08±1.01, respectively) in glomeruli were significantly decreased in diabetic rats treated with resveratrol (all P< 0.05).
4. The expressions of Sirtl (0.63±0.04), FoxO1 (0.37±0.03) and CAT (0.22±0.03) mRNA in kidneys of diabetic rats without treatment were decreased compared with normal group (0.96±0.08,0.69±0.05 and 0.48±0.04, respectively; all P<0.05).After treated with resveratrol for 12 weeks, the relative amounts of mRNA (0.83±0.05, 0.53±0.04 and 0.40±0.03, respectively; all P<0.05) were higher in diabetic rat treated with resveratrol compared to the diabetic rats without treatment (all P< 0.05).
5. Western blot showed that there was no difference FoxO1 protein expression among groups (0.45±0.09,0.48±0.08 and 0.47±0.09, respectively; all P>0.05), but FoxO1 phosphorylation level in group of diabetic rats without treatment (2.15±0.08) was highter than those of normal group (1.14±0.08, P< 0.05), and FoxO1 phosphorylation level in group of diabetic rats treated with resveratrol (1.72±0.07) was lower than those of diabetic rats without treatment (P< 0.05).
Conclusions
1. The expression of FoxO1 mRNA was decreased, and FoxO1 phosphorylation level was increased in kidney of diabetic rats.The role in defensing against oxidative stress of FoxO1 was decreased in kidney of diabetic rats.
2. Resveratrol significantly increased the FoxOl mRNA and decreased FoxO1 phosphorylation levels in kidneys of diabetic rats treated with resveratrol.
3. Sirtl activated by resveratrol may increase the expression of FoxO1, which increases its antioxidant target genes (CAT) to reduce ROS level in the kidneys.
4. Alterations of the Sirtl/FoxO1 pathway may be considered a potential therapeutic target for DN.
引文
[1]Minoru Satoh, Sohachi Fujimoto, et al. NAD(P)H oxidase and uncoupled nitric oxide synthase are major sources of glomerular superoxide in rats with experimental diabetic nephropathy. Am J Physiol Renal Physiol,2005,288:F1144-F1152.
[2]L.Xia, H.Wang. Mesangial cell NADPH oxidase upregulation in high glucose is PKC dependent and required for collagen Ⅳ expression. Am J Physiol Renal Physiol,2006,290: F345-F356.
[3]Yves Gorin, Laren Block. NOX4 NADPH mediates hypertrophy and fibronectin expression in the diabetic kidney. The Journal of Biological chemistry,2005,280:39616-39626.
[4]Susumu Ogawa, Hiroyuki Kobori. Angiotensin Ⅱ type 1 receptor blockers reduce urinary angiotensinogen excretion and the levels of urinary markers of oxidative stress and inflammation in patients with type 2 diabetic nephropathy. Biomarker Insights,2009,4: 97-102.
[5]曹冬梅,卢建.叉头框(Fox)转录因子家族的结构与功能.Chinese Bulletin of Life Sciences,2006,18(5):491-496.
[6]Marieke AG Essers, Sanne Weijzen, Alida MM de Vries-Smits. FoxO transcription factor activation by oxidative stress mediated by the small GTPase Ral and JNK. The EMBO Journal,2004,23:4802-4812.
[7]Zuzana Tothova, Ramya Kollipara, Brian J. Huntly, et al. FoxOs Are Critical Mediators of Hematopoietic Stem Cell Resistance to Physiologic Oxidative Stress. Cell,2007,128: 325-339.
[8]Angela R. Subauste, Charles F. Burant. Role of FoxO1 in FFA-induced oxidative stress in adipocytes. Am J Physiol Endocrinol Metab,2007,293:E159-E164.
[9]Matsumoto M, Han S, K itamura T, et al. Dual role of transcription factor FoxO1 in controlling hepatic insulin sensitivity and lipid metabolism. J Clin Invest,2006, 116:2464-2472.
[10]Brunet. A., Sweeney. L.B., Sturgill. J.F., et al. Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. Science,2004,303:2011-2015.
[11]Alcendor.R.R., Gao,S., Zhai,P., et al. Sirtl regulates aging and resistance to oxidative stress in the heart. Circ Res,2007,100:1512-1521.
[12]Borra MT, Smith BC, Denu JM. Mechanism of human SIRT1 activation by resveratrol. Biol Chem,2005,280:17187-17195.
[13]Hunjoo Ha, In-A Hwang, et al. Role of reactive oxygen species in the pathogenesis of diabetic nephropathy. Diabetes Res Clin Pr,2008,82:S42-S45.
[14]Josephine M. Forbes, Melinda T. coughlan. Oxidative stress as a major culprit in kidney disease in diabetes. Diabetes,2008,57:1446-1454.
[15]Dae Hyun Kim, Ji Young Kim. The activation of NF-κB through AKT-induced FoxO1 phosphorylation during aging and its modulation by calorie restriction. Biogerontology, 2008,9:33-47.
[16]Mitsuo Kato, Hang Yuan, Zhong-Gao Xu, et al. Role of the Akt/FoxO3a pathway in TGF-β1-mediated mesangial cell dysfunction:a novel mechanism related to diabetic kidney disease. J Am Soc Nephrol,2006,17:3325-3335.
[17]Balachandar Venkatesan, Lenin Mahimainathan, Falguni Das, et al. Downregulation of catalase by reactive oxygen species via PI3 Kinase/Akt signaling in mesangial cells. J. Cell. Physiol,2007,211:457-467.
[18]丁洪成,程长明,廖勇敢.白藜芦醇对糖尿病大鼠肾脏内氧化应激的影响.成都中医药大学学报,2009,32：62-65.
[19]Frescas D, Valenti L, Accili D. Nuclear trapping of the forkhead transcription factor FoxO1 via Sirt-dependent deacetylation promotes expression of glucogenetic genes. J Biol Chem, 2005,280:20589-20595.
[20]Qinghe Chen, Suthakar Ganapathy, et al. Resveratrol Induces Growth Arrest and Apoptosis through Activation of FOXO Transcription Factors in Prostate Cancer Cells. PLoS One. 2010,5:e15288.
[21]Joo-Man Park, Tae-Hyun Kima, Jin-Sik Bae, et al. Role of resveratrol in FOXO 1-mediated gluconeogenic gene expression in the liver. Biochem Biophys Res Commun,2010, 403:329-334.
[22]Shiqin Xiong, Gloria Salazar, Nikolay Patrushev, et al. FoxO1 mediates an auto-feedback loop regulating SIRT1 expression. J Biol Chem,2010,10:1-20.
[23]Nakae J, Cao Y, Daitoku H, et al. The LXXLL motif of murine forkhead transcription factor FoxO1 mediates Sirtl-dependent transcriptional activity. J Clin Invest.2006, 116:2473-2483.
[1]Josephine M. Forbes, Melinda T. coughlan. Oxidative stress as a major culprit in kidney disease in diabetes. Diabetes,2008,57:1446-1454.
[2]Hunjoo Ha, In-A Hwang, et al. Role of reactive oxygen species in the pathogenesis of diabetic nephropathy. Diabetes Res Clin Pr,2008,82:S42-S45.
[3]Fioretto P, Stehouwer CDA, Mauer Metal. Heterogeneous nature of microalbuminuria in NIDDM:studies of endothelial function and renal structure. Diabetologia,1998,41:233-236.
[4]Osterby R, Gundersen HJG, Horlyck A, et al. Diabetic glomerulopathy-structural characteristics of the early and advanced stages. Diabetes,1983.32:79-82.
[5]Dalla VM, Saller A,Mauer M, Fioretto P. Role of mesangial expansion in the pathogenesis of iabetic nephropathy. J Nephro,2001,114:S51-S57.
[6]vanden Hoven MJ, Rops AL, Bakker Maetal. Increased expression of heparanase in overt diabetic nephropathy. Kidney Int,2006,70:2100-2108.
[7]vanden Hoven MJ, Rops AL, Vlodavsky I, et al. Heparanase in glomerular diseases. Kidney Int,2007,72:543-548.
[8]vanden Hoven MJ, Wijnhoven TJ, Li J Petal. Reduction of anionic sites in the glomerular basement membrane by heparanase does not lead to proteinuria. Kidney Int,2008, 73:278-287.
[9]Wijnhoven TJM, Lensen JFM, Rops ALW Metal. Aberrant heparan sulfate profile in the human diabetic kidney offers new clues for the rapeutic glycomimetics. Am J Kidney Dis,2006,48:250-261.
[10]Wijnhoven TJM, vanden Hoven MJW, Ding Hetal. Heparanase induces a differential loss of heparan sulphate domains in overt diabetic nephropathy. Diabetologia,2008,51:372-382.
[11]Singh DK, Winocour P, Farrington K. Mechanisms of disease:the hypoxic tubular hypothesis of diabetic nephropathy. Nat Clin Pract Nephrol,2008,4:216-226.
[12]PITKANEN, S.&B. H.ROBINSON. Mitochondrial complex I deficiency leads to increased production of superoxide radicals and induction of superoxide dismutase. J Clin Invest, 1996,98:345-351.
[13]ROSCA, M.G. et al. Glycation of mitochondrial proteins from diabetic rat kidney is associated with excess superoxide formation. Am J Physiol Renal Physiol,2005,289: F420-F430.
[14]JAMES, A.M.&M.P.MURPHY. How mitochondrial damage affects cell function. J Biomed Sci,2002,9:475-487.
[15]GRIENDLING, K.K., D.SORESCU, M.USHIO-FUKAI. NAD(P)H oxidase:role in cardiovascular biology and disease. Circ Res,2000,86:494-501.
[16]JONES, S.A. et al. The expression of NADPH oxidase components in human glomerular mesangial cells:detection of protein and mRNA for p47phox, p67phox, and p22phox. J Am Soc Nephrol,1995,5:1483-1491.
[17]RADEKE, H.H. et al. Functional expression of NADPH Oxidase components (alpha-and beta-subunits of cytochrome b558 and 45-kDa flavoprotein) by intrinsic human glomerular mesangial cells.J Biol Chem,1991,266:21025-21029.
[18]SHIOSE, A. et al. A novel superoxide-producing NAD(P)Hoxidase in kidney. J Biol Chem, 2001.276:1417-1423.
[19]ASABA, K. et al. Effects of NADPH oxidase inhibitor in diabetic nephropathy. Kidney Int,2005,67:1890-1898.
[20]L.Xia, H.Wang. Mesangial cell NADPH oxidase upregulation in high glucose is PK.C dependent and required for collagen Ⅳ expression. Am J Physiol Renal Physiol,2006, 290:F345-F356.
[21]Yves Gorin, Laren Block. NOX4 NADPH Oxidase mediates hypertrophy and fibronectin expression in the diabetic kidney. J Biol Chem,2005,280:39616-39626.
[22]R.K. Studer, P.A. Craven, F.R. De Rubertis. Antioxidant inhibition of protein kinase C-signaled increases in transfonning growth factor-beta in mesangial cells. Metabolism, 1997,46:918-925.
[23]H. Ha, M.R. Yu, Y.J. Choi, H.B. Lee. Activation of protein kinase C-δ and-ε by oxidative stress in early diabetic kidney. Am J Kidney Dis.2001,38:S204-S207.
[24]M. Haneda, S. Araki, M. Togawa, et al. Mitogen-activated protein kinase cascade is activated in glomeruli of diabetic rats and glomerular mesangial cells cultured under high glucose conditions. Diabetes,1997,46:847-853.
[25]H. Ha, M.R. Yu, Y.J. Choi, et al. Role of high glucose-induced nuclear factor-kappa B activation in monocyte chemoattractant protein-1 expression by mesangial cells. J Am Soc Nephrol,2002,13:894-902.
[26]E.A. Lee, J.Y. Seo, Z. Jiang, et al. Reactive oxygen species mediate high glucose-induced plasminogen activator inhibitor-1 upregulation in mesangial cells and in diabetic kidney. Kidney Int,2005,67:1762-1771.
[27]Z. Jiang, J.Y. Seo, H. Ha, et al. Reactive oxygen species mediated TGF-β1-induced plasminogen activator inhibitor-1 upregulation in mesangial cells. Biochem Biophys Res, 2003,309:961-966.
[28]D.Y. Rhyu, Y. Yang, H. Ha, et al. Role of reactive oxygen species in TGF-betal-induced mitogen-activated protein kinase activation and epithelial-mesenchymal transition in renal tubular epithelial cells. J Am Soc Nephrol,2006,16:667-675.
[29]H. Ha, M.R. Yu, K.H. Kim. Melatonin and taurine reduce early glomerulopathy in diabetic rats. Free Radic Biol Med,1999,26:944-950.
[30]M.C. Iglesias-De La Cruz, P. Ruiz-Torres, et al. Hydrogen peroxide increases extracellula rmatrix mRNA through TGF-β in human mesangial cells. Kidney Int,2001:87-95.
[31]H.B. Lee, M.R. Yu, Y. Yang, et al. Reactive oxygen species-regulated signaling pathways in diabetic nephropathy. J Am Soc Nephrol,2003,14:S241-S245.
[32]T.J. Hsieh, P. Fustier, C.C. Wei, et al. Reactive oxygen species blockade and action of insulin on expression of angiotensinogen gene in proximal tubular cells. J Endocrinol,2004, 183:535-550.
[33]M. Ushio-Fukai, A.M. Zafari, T. Fukui, et al. Grindling. p22Phox is a critical component of the superoxide-generating NADH/NADPH oxidase system and regulates angiotensin Ⅱ-induced hypertrophy in vascular smooth musclec ells. J Biol Chem,1996,271:23317-23321.
[34]D. Koya, K. Hayashi, M. Kitada, et al. Effect of antioxidants in diabetes-induced oxidative stress in the glomeruli of diabetic rats. J Am Soc Nephrol,2003,14:S250-S253.
[35]H. Ha, C. Kim, Y. Son, et al. DNA damage in the kidneys of diabetic rats exhibiting microalbuminuria. Free Radic Biol Med,1994,16:271-274.
[36]P. Dandona, K. Thusu, S. Cook, et al. Oxidative damage to DNA in diabetes mellitus. Lancet,1996,347:444-445.
[37]Minoru Satoh, Sohachi Fujimoto, et al. NAD(P)H oxidase and uncoupled nitric oxide synthase are major sources of glomerular superoxide in rats with experimental diabetic nephropathy. Am J Physiol Renal Physiol,2005,288:F1144-F1152.
[38]Susumu Ogawa, Hiroyuki Kobori. Angiotensin Ⅱ type Ⅰ receptor blockers reduce urinary angiotensinogen excretion and the levels of urinary markers of oxidative stress and inflammation in patients with type 2 diabetic nephropathy. Biomarker Insights,2009,4: 97-102.
[39]Brownlee M. Biochemistry and molecular cell biology of diabetic complications. Nature, 2001,414:813-820.
[40]Nishikawa T, Edelstein D, Du Xletal. Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature,2000,404:787-790.
[41]Ha H, Hwang IA, Park JH, et al. Role of reactive oxygen species in the pathogenesis of diabetic nephropathy. Diabetes Res Clin Pract,2008,82:S42-S45.
[42]Kang BPS, Urbonas A, Baddoo A, et al. IGF-1 inhibits the mitochondrial apoptosis program in mesangial cells exposed to high glucose. Am J Physiol Renal Physio,2003,1285: F1013-F1024.
[43]Verzola D, Bertolotto MB, Villaggio Betal. Taurine prevents apoptosis induced by high ambient glucose in human tubulerenal cells. Jinvestig Med,2002,50:443-451.
[44]Isermann B, Vinnikov IA, Madhusudhan T, et al. Activated protein C protects against diabetic nephropathy by inhibiting endothelial and podocyte apoptosis. Nat Med,2007, 13:1349-1358.
[45]Brownlee M. Preventing kidney cell suicide. Nat Med,2007,13:1284-1285.
[46]Heilig CW, Concepcion LA, Riser BL, et al. Overexpression of glucose transporters in rat mesangial cells cultured in a normal glucose milieu mimics the diabetic phenotype. J Clin Invest,1995,96:1802-1814.
[47]Coward RJ, Welsh GI, Yang J, et al. The human glomerular podocyte is a novel target for insulin action. Diabetes,2005,54:3095-3102.
[48]UK Prospective Diabetes Study (UKPDS) Group. Intensive blood-glucose control with sulphonylureas or insulin compared with conventional treatment and risk of complications in patients with type 2 diabetes (UKPDS33). Lancet,1998 352:837-853.
[49]Asada T, Ogawa T, Iwai M, et al. Recombinant insulin-like growth factor I normalizes expression of renal glucose transporters in diabetic rats. Am J Physiol,1997,273:F27-F37.
[50]E.A. Lee, J.Y. Seo, Z. Jiang, et al. Kwon, H. Ha, et al. Reactive oxygen species mediate high glucose-induced plasminogen activator inhibitor-1 upregulation in mesangial cells and in diabetic kidney. Kidney Int,2005,67:1762-1771.
[51]P.A. Craven, M.F. Melhem, S.L. Phillips, et al. Overexpression of Cu2-/Zn2- superoxide dismutase protects against early diabetic glomerular injury in transgenic mice. Diabetes,2001,50:2114-2125.
[52]Y. Hamada, S. Miyata, T. Nii-Kono, et al. Overexpression of thioredoxinl in transgenic mice suppresses development of diabetic nephropathy. Nephrol Dial Transplant, 2007,22:1547-1557.
[53]M.L. Brezniceanu, F. Liu, C.C. Wei, et al. Catalase overexpression attenuates angiotensinogen expression and apoptosis in diabetic mice. Kidney Int,2007,71:912-923.
[54]Nishikawa T, Edelstein D, Du XL, et al. Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature,2000,404:787-790.
[55]曹冬梅,卢建. 义头框(Fox)转录因子家族的结构与功能.Chinese Bulletin of Life Sciences,2006,18(5):491-496.
[56]Marieke AG Essers, Sanne Weijzen, Alida MM de Vries-Smits. FoxO transcription factor activation by oxidative stress mediated by the small GTPase Ral and JNK. The EMBO Journal,2004,23:4802-4812.
[57]Zuzana Tothova, Ramya Kollipara, Brian J. Huntly, et al. FoxOs Are Critical Mediators of Hematopoietic Stem Cell Resistance to Physiologic Oxidative Stress. Cell,2007,128: 325-339.
[58]Angela R. Subauste, Charles F. Burant. Role of FoxO 1 in FEA-induced oxidative stress in adipocytes. Am J Physiol Endocrinol Metab,2007,293:E159-E164.
[59]Dae Hyun Kim, Ji Young Kim. The activation of NF-κB through AKT-induced FoxO1 phosphorylation during aging and its modulation by calorie restriction. Biogerontology, 2008,9:33-47.
[60]Mitsuo Kato, Hang Yuan, Zhong-Gao Xu, et al. Role of the Akt/FoxO3a pathway in TGF-β1-mediated mesangial cell dysfunction:a novel mechanism related to diabetic kidney disease. J Am Soc Nephrol,2006,17:3325-3335.
[61]Balachandar Venkatesan, Lenin Mahimainathan, Falguni Das, et al. Downregulation of catalase by reactive oxygen species via PI3 Kinase/Akt signaling in mesangial cells. J Cell Physiol,2007,211:457-467.
[62]Brunet. A., Sweeney. L.B., Sturgill. J.F., et al. Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. Science,2004,303:2011-2015.
[63]Alcendor.R.R., Gao,S., Zhai,P., et al. Sirtl regulates aging and resistance to oxidative stress in the heart. Circ Res,2007,100:1512-1521.
[64]Yugal Behl, Padmaja Krothapalli, Tesfahun Desta, et al. FoxO1 plays an important role in enhanced microvascular cell apoptosis and microvascular cell loss in type 1 and type 2 diabetic rats. Diabetes,2009,58:917-925.
[2]L.Xia, H.Wang. Mesangial cell NADPH oxidase upregulation in high glucose is PKC dependent and required for collagen Ⅳ expression. Am J Physiol Renal Physiol,2006,290: F345-F356.
[3]Yves Gorin, Laren Block. NOX4 NADPH mediates hypertrophy and fibronectin expression in the diabetic kidney. The Journal of Biological chemistry,2005,280:39616-39626.
[4]Susumu Ogawa, Hiroyuki Kobori. Angiotensin Ⅱ type 1 receptor blockers reduce urinary angiotensinogen excretion and the levels of urinary markers of oxidative stress and inflammation in patients with type 2 diabetic nephropathy. Biomarker Insights,2009,4: 97-102.
[5]曹冬梅,卢建.叉头框(Fox)转录因子家族的结构与功能.Chinese Bulletin of Life Sciences,2006,18(5):491-496.
[6]Marieke AG Essers, Sanne Weijzen, Alida MM de Vries-Smits. FoxO transcription factor activation by oxidative stress mediated by the small GTPase Ral and JNK. The EMBO Journal,2004,23:4802-4812.
[7]Zuzana Tothova, Ramya Kollipara, Brian J. Huntly, et al. FoxOs Are Critical Mediators of Hematopoietic Stem Cell Resistance to Physiologic Oxidative Stress. Cell,2007,128: 325-339.
[8]Angela R. Subauste, Charles F. Burant. Role of FoxO1 in FFA-induced oxidative stress in adipocytes. Am J Physiol Endocrinol Metab,2007,293:E159-E164.
[9]Matsumoto M, Han S, K itamura T, et al. Dual role of transcription factor FoxO1 in controlling hepatic insulin sensitivity and lipid metabolism. J Clin Invest,2006, 116:2464-2472.
[10]Brunet. A., Sweeney. L.B., Sturgill. J.F., et al. Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. Science,2004,303:2011-2015.
[11]Alcendor.R.R., Gao,S., Zhai,P., et al. Sirtl regulates aging and resistance to oxidative stress in the heart. Circ Res,2007,100:1512-1521.
[12]Borra MT, Smith BC, Denu JM. Mechanism of human SIRT1 activation by resveratrol. Biol Chem,2005,280:17187-17195.
[13]Hunjoo Ha, In-A Hwang, et al. Role of reactive oxygen species in the pathogenesis of diabetic nephropathy. Diabetes Res Clin Pr,2008,82:S42-S45.
[14]Josephine M. Forbes, Melinda T. coughlan. Oxidative stress as a major culprit in kidney disease in diabetes. Diabetes,2008,57:1446-1454.
[15]Dae Hyun Kim, Ji Young Kim. The activation of NF-κB through AKT-induced FoxO1 phosphorylation during aging and its modulation by calorie restriction. Biogerontology, 2008,9:33-47.
[16]Mitsuo Kato, Hang Yuan, Zhong-Gao Xu, et al. Role of the Akt/FoxO3a pathway in TGF-β1-mediated mesangial cell dysfunction:a novel mechanism related to diabetic kidney disease. J Am Soc Nephrol,2006,17:3325-3335.
[17]Balachandar Venkatesan, Lenin Mahimainathan, Falguni Das, et al. Downregulation of catalase by reactive oxygen species via PI3 Kinase/Akt signaling in mesangial cells. J. Cell. Physiol,2007,211:457-467.
[18]丁洪成,程长明,廖勇敢.白藜芦醇对糖尿病大鼠肾脏内氧化应激的影响.成都中医药大学学报,2009,32：62-65.
[19]Frescas D, Valenti L, Accili D. Nuclear trapping of the forkhead transcription factor FoxO1 via Sirt-dependent deacetylation promotes expression of glucogenetic genes. J Biol Chem, 2005,280:20589-20595.
[20]Qinghe Chen, Suthakar Ganapathy, et al. Resveratrol Induces Growth Arrest and Apoptosis through Activation of FOXO Transcription Factors in Prostate Cancer Cells. PLoS One. 2010,5:e15288.
[21]Joo-Man Park, Tae-Hyun Kima, Jin-Sik Bae, et al. Role of resveratrol in FOXO 1-mediated gluconeogenic gene expression in the liver. Biochem Biophys Res Commun,2010, 403:329-334.
[22]Shiqin Xiong, Gloria Salazar, Nikolay Patrushev, et al. FoxO1 mediates an auto-feedback loop regulating SIRT1 expression. J Biol Chem,2010,10:1-20.
[23]Nakae J, Cao Y, Daitoku H, et al. The LXXLL motif of murine forkhead transcription factor FoxO1 mediates Sirtl-dependent transcriptional activity. J Clin Invest.2006, 116:2473-2483.
[1]Josephine M. Forbes, Melinda T. coughlan. Oxidative stress as a major culprit in kidney disease in diabetes. Diabetes,2008,57:1446-1454.
[2]Hunjoo Ha, In-A Hwang, et al. Role of reactive oxygen species in the pathogenesis of diabetic nephropathy. Diabetes Res Clin Pr,2008,82:S42-S45.
[3]Fioretto P, Stehouwer CDA, Mauer Metal. Heterogeneous nature of microalbuminuria in NIDDM:studies of endothelial function and renal structure. Diabetologia,1998,41:233-236.
[4]Osterby R, Gundersen HJG, Horlyck A, et al. Diabetic glomerulopathy-structural characteristics of the early and advanced stages. Diabetes,1983.32:79-82.
[5]Dalla VM, Saller A,Mauer M, Fioretto P. Role of mesangial expansion in the pathogenesis of iabetic nephropathy. J Nephro,2001,114:S51-S57.
[6]vanden Hoven MJ, Rops AL, Bakker Maetal. Increased expression of heparanase in overt diabetic nephropathy. Kidney Int,2006,70:2100-2108.
[7]vanden Hoven MJ, Rops AL, Vlodavsky I, et al. Heparanase in glomerular diseases. Kidney Int,2007,72:543-548.
[8]vanden Hoven MJ, Wijnhoven TJ, Li J Petal. Reduction of anionic sites in the glomerular basement membrane by heparanase does not lead to proteinuria. Kidney Int,2008, 73:278-287.
[9]Wijnhoven TJM, Lensen JFM, Rops ALW Metal. Aberrant heparan sulfate profile in the human diabetic kidney offers new clues for the rapeutic glycomimetics. Am J Kidney Dis,2006,48:250-261.
[10]Wijnhoven TJM, vanden Hoven MJW, Ding Hetal. Heparanase induces a differential loss of heparan sulphate domains in overt diabetic nephropathy. Diabetologia,2008,51:372-382.
[11]Singh DK, Winocour P, Farrington K. Mechanisms of disease:the hypoxic tubular hypothesis of diabetic nephropathy. Nat Clin Pract Nephrol,2008,4:216-226.
[12]PITKANEN, S.&B. H.ROBINSON. Mitochondrial complex I deficiency leads to increased production of superoxide radicals and induction of superoxide dismutase. J Clin Invest, 1996,98:345-351.
[13]ROSCA, M.G. et al. Glycation of mitochondrial proteins from diabetic rat kidney is associated with excess superoxide formation. Am J Physiol Renal Physiol,2005,289: F420-F430.
[14]JAMES, A.M.&M.P.MURPHY. How mitochondrial damage affects cell function. J Biomed Sci,2002,9:475-487.
[15]GRIENDLING, K.K., D.SORESCU, M.USHIO-FUKAI. NAD(P)H oxidase:role in cardiovascular biology and disease. Circ Res,2000,86:494-501.
[16]JONES, S.A. et al. The expression of NADPH oxidase components in human glomerular mesangial cells:detection of protein and mRNA for p47phox, p67phox, and p22phox. J Am Soc Nephrol,1995,5:1483-1491.
[17]RADEKE, H.H. et al. Functional expression of NADPH Oxidase components (alpha-and beta-subunits of cytochrome b558 and 45-kDa flavoprotein) by intrinsic human glomerular mesangial cells.J Biol Chem,1991,266:21025-21029.
[18]SHIOSE, A. et al. A novel superoxide-producing NAD(P)Hoxidase in kidney. J Biol Chem, 2001.276:1417-1423.
[19]ASABA, K. et al. Effects of NADPH oxidase inhibitor in diabetic nephropathy. Kidney Int,2005,67:1890-1898.
[20]L.Xia, H.Wang. Mesangial cell NADPH oxidase upregulation in high glucose is PK.C dependent and required for collagen Ⅳ expression. Am J Physiol Renal Physiol,2006, 290:F345-F356.
[21]Yves Gorin, Laren Block. NOX4 NADPH Oxidase mediates hypertrophy and fibronectin expression in the diabetic kidney. J Biol Chem,2005,280:39616-39626.
[22]R.K. Studer, P.A. Craven, F.R. De Rubertis. Antioxidant inhibition of protein kinase C-signaled increases in transfonning growth factor-beta in mesangial cells. Metabolism, 1997,46:918-925.
[23]H. Ha, M.R. Yu, Y.J. Choi, H.B. Lee. Activation of protein kinase C-δ and-ε by oxidative stress in early diabetic kidney. Am J Kidney Dis.2001,38:S204-S207.
[24]M. Haneda, S. Araki, M. Togawa, et al. Mitogen-activated protein kinase cascade is activated in glomeruli of diabetic rats and glomerular mesangial cells cultured under high glucose conditions. Diabetes,1997,46:847-853.
[25]H. Ha, M.R. Yu, Y.J. Choi, et al. Role of high glucose-induced nuclear factor-kappa B activation in monocyte chemoattractant protein-1 expression by mesangial cells. J Am Soc Nephrol,2002,13:894-902.
[26]E.A. Lee, J.Y. Seo, Z. Jiang, et al. Reactive oxygen species mediate high glucose-induced plasminogen activator inhibitor-1 upregulation in mesangial cells and in diabetic kidney. Kidney Int,2005,67:1762-1771.
[27]Z. Jiang, J.Y. Seo, H. Ha, et al. Reactive oxygen species mediated TGF-β1-induced plasminogen activator inhibitor-1 upregulation in mesangial cells. Biochem Biophys Res, 2003,309:961-966.
[28]D.Y. Rhyu, Y. Yang, H. Ha, et al. Role of reactive oxygen species in TGF-betal-induced mitogen-activated protein kinase activation and epithelial-mesenchymal transition in renal tubular epithelial cells. J Am Soc Nephrol,2006,16:667-675.
[29]H. Ha, M.R. Yu, K.H. Kim. Melatonin and taurine reduce early glomerulopathy in diabetic rats. Free Radic Biol Med,1999,26:944-950.
[30]M.C. Iglesias-De La Cruz, P. Ruiz-Torres, et al. Hydrogen peroxide increases extracellula rmatrix mRNA through TGF-β in human mesangial cells. Kidney Int,2001:87-95.
[31]H.B. Lee, M.R. Yu, Y. Yang, et al. Reactive oxygen species-regulated signaling pathways in diabetic nephropathy. J Am Soc Nephrol,2003,14:S241-S245.
[32]T.J. Hsieh, P. Fustier, C.C. Wei, et al. Reactive oxygen species blockade and action of insulin on expression of angiotensinogen gene in proximal tubular cells. J Endocrinol,2004, 183:535-550.
[33]M. Ushio-Fukai, A.M. Zafari, T. Fukui, et al. Grindling. p22Phox is a critical component of the superoxide-generating NADH/NADPH oxidase system and regulates angiotensin Ⅱ-induced hypertrophy in vascular smooth musclec ells. J Biol Chem,1996,271:23317-23321.
[34]D. Koya, K. Hayashi, M. Kitada, et al. Effect of antioxidants in diabetes-induced oxidative stress in the glomeruli of diabetic rats. J Am Soc Nephrol,2003,14:S250-S253.
[35]H. Ha, C. Kim, Y. Son, et al. DNA damage in the kidneys of diabetic rats exhibiting microalbuminuria. Free Radic Biol Med,1994,16:271-274.
[36]P. Dandona, K. Thusu, S. Cook, et al. Oxidative damage to DNA in diabetes mellitus. Lancet,1996,347:444-445.
[37]Minoru Satoh, Sohachi Fujimoto, et al. NAD(P)H oxidase and uncoupled nitric oxide synthase are major sources of glomerular superoxide in rats with experimental diabetic nephropathy. Am J Physiol Renal Physiol,2005,288:F1144-F1152.
[38]Susumu Ogawa, Hiroyuki Kobori. Angiotensin Ⅱ type Ⅰ receptor blockers reduce urinary angiotensinogen excretion and the levels of urinary markers of oxidative stress and inflammation in patients with type 2 diabetic nephropathy. Biomarker Insights,2009,4: 97-102.
[39]Brownlee M. Biochemistry and molecular cell biology of diabetic complications. Nature, 2001,414:813-820.
[40]Nishikawa T, Edelstein D, Du Xletal. Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature,2000,404:787-790.
[41]Ha H, Hwang IA, Park JH, et al. Role of reactive oxygen species in the pathogenesis of diabetic nephropathy. Diabetes Res Clin Pract,2008,82:S42-S45.
[42]Kang BPS, Urbonas A, Baddoo A, et al. IGF-1 inhibits the mitochondrial apoptosis program in mesangial cells exposed to high glucose. Am J Physiol Renal Physio,2003,1285: F1013-F1024.
[43]Verzola D, Bertolotto MB, Villaggio Betal. Taurine prevents apoptosis induced by high ambient glucose in human tubulerenal cells. Jinvestig Med,2002,50:443-451.
[44]Isermann B, Vinnikov IA, Madhusudhan T, et al. Activated protein C protects against diabetic nephropathy by inhibiting endothelial and podocyte apoptosis. Nat Med,2007, 13:1349-1358.
[45]Brownlee M. Preventing kidney cell suicide. Nat Med,2007,13:1284-1285.
[46]Heilig CW, Concepcion LA, Riser BL, et al. Overexpression of glucose transporters in rat mesangial cells cultured in a normal glucose milieu mimics the diabetic phenotype. J Clin Invest,1995,96:1802-1814.
[47]Coward RJ, Welsh GI, Yang J, et al. The human glomerular podocyte is a novel target for insulin action. Diabetes,2005,54:3095-3102.
[48]UK Prospective Diabetes Study (UKPDS) Group. Intensive blood-glucose control with sulphonylureas or insulin compared with conventional treatment and risk of complications in patients with type 2 diabetes (UKPDS33). Lancet,1998 352:837-853.
[49]Asada T, Ogawa T, Iwai M, et al. Recombinant insulin-like growth factor I normalizes expression of renal glucose transporters in diabetic rats. Am J Physiol,1997,273:F27-F37.
[50]E.A. Lee, J.Y. Seo, Z. Jiang, et al. Kwon, H. Ha, et al. Reactive oxygen species mediate high glucose-induced plasminogen activator inhibitor-1 upregulation in mesangial cells and in diabetic kidney. Kidney Int,2005,67:1762-1771.
[51]P.A. Craven, M.F. Melhem, S.L. Phillips, et al. Overexpression of Cu2-/Zn2- superoxide dismutase protects against early diabetic glomerular injury in transgenic mice. Diabetes,2001,50:2114-2125.
[52]Y. Hamada, S. Miyata, T. Nii-Kono, et al. Overexpression of thioredoxinl in transgenic mice suppresses development of diabetic nephropathy. Nephrol Dial Transplant, 2007,22:1547-1557.
[53]M.L. Brezniceanu, F. Liu, C.C. Wei, et al. Catalase overexpression attenuates angiotensinogen expression and apoptosis in diabetic mice. Kidney Int,2007,71:912-923.
[54]Nishikawa T, Edelstein D, Du XL, et al. Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature,2000,404:787-790.
[55]曹冬梅,卢建. 义头框(Fox)转录因子家族的结构与功能.Chinese Bulletin of Life Sciences,2006,18(5):491-496.
[56]Marieke AG Essers, Sanne Weijzen, Alida MM de Vries-Smits. FoxO transcription factor activation by oxidative stress mediated by the small GTPase Ral and JNK. The EMBO Journal,2004,23:4802-4812.
[57]Zuzana Tothova, Ramya Kollipara, Brian J. Huntly, et al. FoxOs Are Critical Mediators of Hematopoietic Stem Cell Resistance to Physiologic Oxidative Stress. Cell,2007,128: 325-339.
[58]Angela R. Subauste, Charles F. Burant. Role of FoxO 1 in FEA-induced oxidative stress in adipocytes. Am J Physiol Endocrinol Metab,2007,293:E159-E164.
[59]Dae Hyun Kim, Ji Young Kim. The activation of NF-κB through AKT-induced FoxO1 phosphorylation during aging and its modulation by calorie restriction. Biogerontology, 2008,9:33-47.
[60]Mitsuo Kato, Hang Yuan, Zhong-Gao Xu, et al. Role of the Akt/FoxO3a pathway in TGF-β1-mediated mesangial cell dysfunction:a novel mechanism related to diabetic kidney disease. J Am Soc Nephrol,2006,17:3325-3335.
[61]Balachandar Venkatesan, Lenin Mahimainathan, Falguni Das, et al. Downregulation of catalase by reactive oxygen species via PI3 Kinase/Akt signaling in mesangial cells. J Cell Physiol,2007,211:457-467.
[62]Brunet. A., Sweeney. L.B., Sturgill. J.F., et al. Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase. Science,2004,303:2011-2015.
[63]Alcendor.R.R., Gao,S., Zhai,P., et al. Sirtl regulates aging and resistance to oxidative stress in the heart. Circ Res,2007,100:1512-1521.
[64]Yugal Behl, Padmaja Krothapalli, Tesfahun Desta, et al. FoxO1 plays an important role in enhanced microvascular cell apoptosis and microvascular cell loss in type 1 and type 2 diabetic rats. Diabetes,2009,58:917-925.