NADPH氧化酶4在高糖引起的血管内皮细胞损伤中的作用机理研究
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摘要
糖尿病(Diabetes mellitus, DM)是由绝对或者相对的胰岛素不足所造成的疾病,血糖升高(Hyperglycemia)是糖尿病的主要特点之一。糖尿病是心血管疾病重要的危险因素,其中血糖异常是糖尿病血管并发症发生的重要原因之一。高糖引起的血管内皮形态改变和功能异常,是糖尿病血管并发症的早期标志。研究已表明,高糖诱发的氧化应激是导致血管内皮损伤的主要原因。氧化应激(Oxidative stress)是指机体在遭受各种有害刺激时体内高活性分子如活性氧自由基(Reactive oxygen species, ROS)和活性氮自由基(Reactive nitrogen species, RNS)的产生过多,氧化程度超出抗氧化系统对氧化物的清除,氧化系统和抗氧化系统失衡,从而导致组织损伤。机体内有多种酶体参与了ROS的生成,如NADPH氧化酶(NADPH oxidase, NOX)、黄嘌呤氧化酶(Xanthine oxidase)、线粒体呼吸链酶复合体、内皮型一氧化氮合酶(Endothelial nitric oxide synthase, eNOS)及脂氧合酶(Lipoxygenase, LOX)等,研究认为NOX是血管内生成ROS的主要酶体。吞噬型NADPH氧化酶是由多个亚基组成的酶复合体,包括胞膜组分：细胞色素b558(gp91PHOX和P22PHOX)和胞浆组分：p47PHOX、p67PHOX及Rac(小的GTP结合蛋白)共五个亚组分。gp91PHOX(有NADPH、heme、FAD的潜在结合位点)和p22PHOX是NOX的酶促核心,它们受p47PHOX及Rac的调节。现已发现NOX的同源家族有NOX1、NOX2(gp91PHOX). NOX3、NOX4、NOX5、 Duox1(Dual oxidase1)和Duox2等成员,在血管内皮细胞主要表达NOX2和NOX4,且NOX4表达水平比NOX2高20倍。进一步的研究显示,血管内皮细胞产生的ROS主要来源于NOX4。NOX活性受其亚组分p47PHOX和Rac-1的调节。p47PHOX磷酸化之后,从胞浆转到胞膜上与细胞色素b558结合,在Rac-1的参与下激活NOX。
为了探讨高糖引起血管内皮细胞氧化应激的作用机制,我们用高浓度(20mmol/L)葡萄糖处理人脐静脉内皮细胞(Human umbilical vein endothelial cells,HUVECs),通过流式细胞术检测ROS水平的改变。结果显示,与正常培养条件相比,20mmol/L葡萄糖处理的HUVECs中ROS水平显著升高。分别用几个产生ROS途径的抑制剂DPI、Rotenone、Oxypurinol和L-NAME预处理后,NOX抑制剂DPI组ROS水平显著降低,而其他抑制剂组的ROS水平无明显变化。提示NOX是高葡萄糖诱导HUVECs产生ROS的主要来源。进一步应用RT-PCR、 Real-time PCR和Western blot分析NOX亚组分表达的变化。结果表明,]HUVECs表达NOX4及其亚组分p22PHOX、p47PHOX、p67PHOX和Rac-1。NOX4的表达在20mmol/L葡萄糖处理后显著升高,其它亚组分则无显著性改变。同时,我们应用SuperArray公司的第二代功能分类基因芯片技术对葡萄糖处理后HUVECs氧化应激与抗氧化相关基因的表达进行了检测。结果显示,20mmol/L葡萄糖引起氧化应激与抗氧化相关部分基因的表达升高或者降低,其中值得注意的是,与对照组相比,Dual oxidase2基因表达升高近13倍,而Superoxide dismutase3基因表达水平明显降低。观察葡萄糖对NOX4活性的调节机理的结果表明,在高浓度葡萄糖条件下,HUVECs的NOX4活性调节主要发生在两个水平上,一是通过PKC激活NOX4,二是通过p47PHOX和Rac-1的膜移位来调节NOX4的活性。为了研究NOX4表达水平的改变对ROS产生的影响及其对HUVECs损伤的作用,我们采取“失去功能”和“获得功能”两种策略,分别将NOX4-siRNA或NOX4基因转染入HUVECs,获得极低表达甚至无表达或高表达NOX4水平的HUVECs。分析比较这些内皮细胞和对照内皮细胞(未转入NOX4-siRNA和NOX4)中ROS水平与细胞凋亡。我们的实验结果表明,与未转染质粒的对照组和转染GFP质粒表达质粒组相比,转染NOX4质粒组的HUVECs中ROS明显升高(p<0.05)；流式细胞术、Hoechst染色和TUNEL染色结果均显示,转染NOX4质粒组的HUVECs发生明显凋亡(p<0.05)。高糖处理HUVECs之后,其NOX4mRNA水平升高,ROS产量增加,细胞凋亡的比例也增多,而经NOX4-siRNA转染抑制NOX4表达后,高糖所引起的ROS升高和凋亡均受到抑制。说明NOX4参与了高糖引起内皮细胞内ROS的生成和细胞凋亡。在此基础上,我们对NF-κB (nuclear factor-κ B)和p38MAPK信号通路在高糖诱导内皮细胞凋亡中的作用进行探讨。结果显示,20mmol/L葡萄糖处理HUVECs后,伴随ROS的升高,磷酸化的IκB和NF-κB均增多,IκB失去对NF-κB的抑制作用,NF-κB移位至细胞核内。另外,我们还发现,与对照组相比,葡萄糖处理组炎症相关因子ICAM-1、VCAM-1的表达均明显增强(p<0.05)。这些结果提示,高糖可以诱导ROS的升高,引起NF-κB的活化,使得炎症相关因子的表达增加。p38MAPK是细胞内重要的信号转导通路,参与内皮细胞凋亡的调节。研究已表明,高糖可导致p38MAPK的磷酸化从而进入细胞核启动下游信号通路的转录。我们用20mmol/L葡萄糖及p38MAPK特异性抑制剂SB203580处理HUVECs,用Western blot检测p38MAPK及磷酸化的p38MAPK (P-p38MAPK)的水平。结果显示,与正常对照组相比,葡萄糖处理组p38MAPK的表达无明显变化,而P-p38MAPK的水平明显升高(p<0.05)；与葡萄糖处理组相比,SB203580能明显抑制高糖所致P-p38MAPK的升高(p<0.05),而对p38MAPK的表达并无影响。Hoechst/PI染色、TUNEL和PI/Annexin V-FITC双染法的结果表明,SB203580预处理明显抑制葡萄糖引起的细胞凋亡。这些结果提示,p38MAPK在高糖诱导HUVECs凋亡的过程中起着重要的调节作用。近年研究提示,他汀类药物除了有降胆固醇的作用之外,还具有抗炎、抗氧化和稳定斑块的作用。我们在前期的研究中发现,辛伐他汀处理HUVECs能下调NOX4mRNA水平,但不影响NOX2的表达。高浓度(0.8g/L) LDL刺激HUVECs时,NOX4表达上调,细胞内ROS生成增加。用辛伐他汀预处理再加高浓度LDL刺激时,NOX4表达下降,ROS生成量减少。提示辛伐他汀可下调HUVECs NOX4表达,降低ROS水平。为了进一步在体内证明辛伐他汀的抗氧化作用,我们建立了Apo E-/-动脉粥样硬化小鼠模型,并用辛伐他汀进行干预。结果显示,与对照组小鼠相比,辛伐他汀能够明显降低Apo E-/-小鼠血液中TG和MDA,也降低动脉粥样硬化斑块的ROS水平和NOX亚组分的表达。此外,辛伐他汀干预后,动脉粥样硬化斑块变小,纤维帽的厚度和完整性得到明显改善。提示辛伐他汀可能起到稳定斑块的作用。
综上所述,本研究得出以下结论：1、高浓度(20mmol/L)葡萄糖主要通过上调NOX4的表达和活性显著提高]3UVECs的ROS水平；2、20mmol/L葡萄糖处理HUVECs引起氧化应激相关基因Dual Oxidase2表达升高近13倍,而Superoxide dismutase3基因表达水平明显降低；3、葡萄糖通过激活PKC和引起p47PHOX和Rac-1膜移位促进NOX4的活化；4、高浓度葡萄糖和NOX4过表达均可引起HUVECs凋亡,而干扰NOX4的表达可以抑制高糖引起的HUVECs凋亡;5、p38MAPK的磷酸化在高糖引起的HUVECs凋亡过程中发挥重要作用；6、葡萄糖能促进NF-κB的活化及其下游靶基因ICAM-1和VCAM-1的表达；7、辛伐他汀可能通过减少NOX亚组分的表达降低动脉硬化小鼠模型血管的氧化应激水平。本研究为探寻保护血管内皮细胞免受高糖状态下的氧化损伤提供新的线索。特异性抑制NOX活性可能成为防治糖尿病血管并发症的新靶点。
Diabetes mellitus is caused by absolute or relative insufficient insulin and hyperglycemia is one of the main characteristics of diabetes mellitus. Diabetes is an important risk factor of cardiovascular diseases and dysglycemia is one of major causes resulted in vascular complications of diabetes. High glucose-induced morphological changes and dysfunction of vascular endothelium are the early symbols of vascular complications of diabetes. Hyperglycemia-induced oxidative stress plays a key role in atherosclerosis accompanied with diabetes. Oxidative stress is caused by an imbalance between the production of reactive oxygen species (ROS) or reactive nitrogen species (RNS) and a biological system's ability to readily detoxify the reactive intermediates or easily repair the resulting damage. The potential sources of ROS in vascular system include nicotinamide vadenine dinucleotede phosphate oxidase (NADPH oxidase, NOX), xanthine oxidase, cytochrome p-450, uncoupled endothelial nitric oxide synthase and lipoxygenase. NOX is considered as the main source of ROS in vascular endothelium. NOX2, the complex enzyme NOX composed of five subunits including cytomembrane components (p22PHOX and NOX2) and cytosolic components (p47PHOX, p67PHOX and Rac), was firstly found in phagocyte cells. NOX2, which has potential binding sites of NADPH, heme and FAD, and p22PHOX are regulated by p47PHOX and Rac. Seven members of NOX family including NOX1, NOX2(formerly named gp91PHOX), NOX3, NOX4, NOX5, Duox1(Dual oxidase1) and Duox2have been identified in different cells. It is reported the both NOX2and NOX4are expressed in endothelial cells, and the level of NOX4mRNA in endothelial cells is20fold higher than NOX2. Further studies indicated that NOX4is the most important enzyme in the ROS generation in vascular endothelial cells. The enzyme activity of NOX is regulated through phosphorylation and translocation in membrane of p47PHOX and Rac-1.
In order to investigate the mechanisms of high glucose-induced oxidative stress in vascular endothelial cells, human umbilical vein endothelial cells (HUVECs) were treated with20mmol/L glucose, and ROS level was detected by flow cytometry. Comparing with the control group, treatment of20mmol/L glucose could lead to excessive production of ROS in HUVECs. However, preincubation of HUVECs with DPI, the NOX inhibitor, but not Rotenone, Oxypurinol or L-NAME could inhibit the generation of ROS, indicating that NOX is the main source of high glucose-induced ROS generation. Further, the expression of NOX subunits was investigated by RT-PCR, Real-time PCR and Western blot. The results showed that NOX4and sub-units p22PHOX, p47PHOX, p67PHOX, Rac-1were expressed in HUVECs, but only the expression of NOX4was significantly upregulated after20mmol/L glucose treatment in HUVECs. The gene profile screened through SuperArray's PCR array indicated that the level of Dual Oxidase2mRNA could be increased by nearly13fold, while the level of Superoxide dismutase3mRNA could be significantly decreased in HUVECs treated with20mmol/L glucose. p47PHOx and Rac-1were translocated to cell membrane from cytoplasm to mediate the activation of NOX4and the generation of ROS, and PKC was also participated in regulating NOX4-mediated ROS generation. It was found that high concentration of glucose could stimulate the production of ROS and induce apoptosis through upregulating the expression of NOX4in HUVECs. Moreover, two strategies, lost of function and gain of function, were carried out to observe effects of NOX4on ROS generation and injury of HUVECs. The NOX4was knocked down by transfection with NOX4-siRNA, and overexpressed by transfection with NOX4-expressing vector in HUVECs. The results showed that overexpression of NOX4could induce the excessive production of ROS and apoptosis of HUVECs. Contrast, down-regulation of NOX4significantly reduced the level of ROS and prevented HUVECs from apoptosis. Furthermore, to study the mechanisms of apoptosis induced by glucose, nuclear factor-κ B (NF-κB) and p38MAPK pathways were analyzed by Western blot and immunofluorescence staining. After glucose treatment, phosphorylated IkB level was elevated and IκB was degraded, and NF-κB was also phosphorylated and translocated into nucleus in where NF-κB upregulated the expression of inflammatory factors such as ICAM-1and VCAM-1. p38MAPK pathway is considered as an important signaling pathway participated in regulation of endothelial apoptosis. The high glucose resulted in phosphorylation of p38MAPK and the translocation of p38MAPK into nucleus. SB203580, a specific inhibitor of p38MAPK, could inhibit high glucose-induced upregulation of P-p38MAPK and suppress high glucose-induced apoptosis, as measured by Hoechst/PI, TUNEL and PI/Annexin V-FITC staining. These pointed out that p38MAPK played a key role in the glucose-induced apoptosis. Recent results have revealed that beyond lipid-lowering activity, statins exhibit functions such as modulating inflammatory responses, improving antioxidant effects and maintaining plaque stability. Our previous results provided that Simvastatin could downregulate the expression of NOX4but not NOX2and impair the excessive production of ROS induced by treatment of0.8g/L LDL. These results indicated that Simvastatin could lower ROS level by suppression of NOX4expression. To further investgate Simvastin's antioxidant effects in vivo, oxidative stress was analyzed in atherosclerotic Apo E-/-mice treated with Simvastatin. The results showed that Simvastatin could reduce the levels of TG and MDA in serum and the expression of NOX in vascular system, in term impaired the oxidative stress of atherosclerotic mice. In addition, the average area of atherosclerotic lesion was decreased, and the thickness and integrity of fibrous cap were improved in Simvastatin-treated mice, suggesting that Simvastatin has the ability to stabilize the atherosclerotic plaque.
In summary, our results provide the molecular evidence that NOX4-derived ROS could play an important role in oxidative stress of vascular endothelial cells induced by high concentration of glucose. Specifically inhibiting NOX activity may be a novel target preventing vascular complications of diabetes.
为了探讨高糖引起血管内皮细胞氧化应激的作用机制,我们用高浓度(20mmol/L)葡萄糖处理人脐静脉内皮细胞(Human umbilical vein endothelial cells,HUVECs),通过流式细胞术检测ROS水平的改变。结果显示,与正常培养条件相比,20mmol/L葡萄糖处理的HUVECs中ROS水平显著升高。分别用几个产生ROS途径的抑制剂DPI、Rotenone、Oxypurinol和L-NAME预处理后,NOX抑制剂DPI组ROS水平显著降低,而其他抑制剂组的ROS水平无明显变化。提示NOX是高葡萄糖诱导HUVECs产生ROS的主要来源。进一步应用RT-PCR、 Real-time PCR和Western blot分析NOX亚组分表达的变化。结果表明,]HUVECs表达NOX4及其亚组分p22PHOX、p47PHOX、p67PHOX和Rac-1。NOX4的表达在20mmol/L葡萄糖处理后显著升高,其它亚组分则无显著性改变。同时,我们应用SuperArray公司的第二代功能分类基因芯片技术对葡萄糖处理后HUVECs氧化应激与抗氧化相关基因的表达进行了检测。结果显示,20mmol/L葡萄糖引起氧化应激与抗氧化相关部分基因的表达升高或者降低,其中值得注意的是,与对照组相比,Dual oxidase2基因表达升高近13倍,而Superoxide dismutase3基因表达水平明显降低。观察葡萄糖对NOX4活性的调节机理的结果表明,在高浓度葡萄糖条件下,HUVECs的NOX4活性调节主要发生在两个水平上,一是通过PKC激活NOX4,二是通过p47PHOX和Rac-1的膜移位来调节NOX4的活性。为了研究NOX4表达水平的改变对ROS产生的影响及其对HUVECs损伤的作用,我们采取“失去功能”和“获得功能”两种策略,分别将NOX4-siRNA或NOX4基因转染入HUVECs,获得极低表达甚至无表达或高表达NOX4水平的HUVECs。分析比较这些内皮细胞和对照内皮细胞(未转入NOX4-siRNA和NOX4)中ROS水平与细胞凋亡。我们的实验结果表明,与未转染质粒的对照组和转染GFP质粒表达质粒组相比,转染NOX4质粒组的HUVECs中ROS明显升高(p<0.05)；流式细胞术、Hoechst染色和TUNEL染色结果均显示,转染NOX4质粒组的HUVECs发生明显凋亡(p<0.05)。高糖处理HUVECs之后,其NOX4mRNA水平升高,ROS产量增加,细胞凋亡的比例也增多,而经NOX4-siRNA转染抑制NOX4表达后,高糖所引起的ROS升高和凋亡均受到抑制。说明NOX4参与了高糖引起内皮细胞内ROS的生成和细胞凋亡。在此基础上,我们对NF-κB (nuclear factor-κ B)和p38MAPK信号通路在高糖诱导内皮细胞凋亡中的作用进行探讨。结果显示,20mmol/L葡萄糖处理HUVECs后,伴随ROS的升高,磷酸化的IκB和NF-κB均增多,IκB失去对NF-κB的抑制作用,NF-κB移位至细胞核内。另外,我们还发现,与对照组相比,葡萄糖处理组炎症相关因子ICAM-1、VCAM-1的表达均明显增强(p<0.05)。这些结果提示,高糖可以诱导ROS的升高,引起NF-κB的活化,使得炎症相关因子的表达增加。p38MAPK是细胞内重要的信号转导通路,参与内皮细胞凋亡的调节。研究已表明,高糖可导致p38MAPK的磷酸化从而进入细胞核启动下游信号通路的转录。我们用20mmol/L葡萄糖及p38MAPK特异性抑制剂SB203580处理HUVECs,用Western blot检测p38MAPK及磷酸化的p38MAPK (P-p38MAPK)的水平。结果显示,与正常对照组相比,葡萄糖处理组p38MAPK的表达无明显变化,而P-p38MAPK的水平明显升高(p<0.05)；与葡萄糖处理组相比,SB203580能明显抑制高糖所致P-p38MAPK的升高(p<0.05),而对p38MAPK的表达并无影响。Hoechst/PI染色、TUNEL和PI/Annexin V-FITC双染法的结果表明,SB203580预处理明显抑制葡萄糖引起的细胞凋亡。这些结果提示,p38MAPK在高糖诱导HUVECs凋亡的过程中起着重要的调节作用。近年研究提示,他汀类药物除了有降胆固醇的作用之外,还具有抗炎、抗氧化和稳定斑块的作用。我们在前期的研究中发现,辛伐他汀处理HUVECs能下调NOX4mRNA水平,但不影响NOX2的表达。高浓度(0.8g/L) LDL刺激HUVECs时,NOX4表达上调,细胞内ROS生成增加。用辛伐他汀预处理再加高浓度LDL刺激时,NOX4表达下降,ROS生成量减少。提示辛伐他汀可下调HUVECs NOX4表达,降低ROS水平。为了进一步在体内证明辛伐他汀的抗氧化作用,我们建立了Apo E-/-动脉粥样硬化小鼠模型,并用辛伐他汀进行干预。结果显示,与对照组小鼠相比,辛伐他汀能够明显降低Apo E-/-小鼠血液中TG和MDA,也降低动脉粥样硬化斑块的ROS水平和NOX亚组分的表达。此外,辛伐他汀干预后,动脉粥样硬化斑块变小,纤维帽的厚度和完整性得到明显改善。提示辛伐他汀可能起到稳定斑块的作用。
综上所述,本研究得出以下结论：1、高浓度(20mmol/L)葡萄糖主要通过上调NOX4的表达和活性显著提高]3UVECs的ROS水平；2、20mmol/L葡萄糖处理HUVECs引起氧化应激相关基因Dual Oxidase2表达升高近13倍,而Superoxide dismutase3基因表达水平明显降低；3、葡萄糖通过激活PKC和引起p47PHOX和Rac-1膜移位促进NOX4的活化；4、高浓度葡萄糖和NOX4过表达均可引起HUVECs凋亡,而干扰NOX4的表达可以抑制高糖引起的HUVECs凋亡;5、p38MAPK的磷酸化在高糖引起的HUVECs凋亡过程中发挥重要作用；6、葡萄糖能促进NF-κB的活化及其下游靶基因ICAM-1和VCAM-1的表达；7、辛伐他汀可能通过减少NOX亚组分的表达降低动脉硬化小鼠模型血管的氧化应激水平。本研究为探寻保护血管内皮细胞免受高糖状态下的氧化损伤提供新的线索。特异性抑制NOX活性可能成为防治糖尿病血管并发症的新靶点。
Diabetes mellitus is caused by absolute or relative insufficient insulin and hyperglycemia is one of the main characteristics of diabetes mellitus. Diabetes is an important risk factor of cardiovascular diseases and dysglycemia is one of major causes resulted in vascular complications of diabetes. High glucose-induced morphological changes and dysfunction of vascular endothelium are the early symbols of vascular complications of diabetes. Hyperglycemia-induced oxidative stress plays a key role in atherosclerosis accompanied with diabetes. Oxidative stress is caused by an imbalance between the production of reactive oxygen species (ROS) or reactive nitrogen species (RNS) and a biological system's ability to readily detoxify the reactive intermediates or easily repair the resulting damage. The potential sources of ROS in vascular system include nicotinamide vadenine dinucleotede phosphate oxidase (NADPH oxidase, NOX), xanthine oxidase, cytochrome p-450, uncoupled endothelial nitric oxide synthase and lipoxygenase. NOX is considered as the main source of ROS in vascular endothelium. NOX2, the complex enzyme NOX composed of five subunits including cytomembrane components (p22PHOX and NOX2) and cytosolic components (p47PHOX, p67PHOX and Rac), was firstly found in phagocyte cells. NOX2, which has potential binding sites of NADPH, heme and FAD, and p22PHOX are regulated by p47PHOX and Rac. Seven members of NOX family including NOX1, NOX2(formerly named gp91PHOX), NOX3, NOX4, NOX5, Duox1(Dual oxidase1) and Duox2have been identified in different cells. It is reported the both NOX2and NOX4are expressed in endothelial cells, and the level of NOX4mRNA in endothelial cells is20fold higher than NOX2. Further studies indicated that NOX4is the most important enzyme in the ROS generation in vascular endothelial cells. The enzyme activity of NOX is regulated through phosphorylation and translocation in membrane of p47PHOX and Rac-1.
In order to investigate the mechanisms of high glucose-induced oxidative stress in vascular endothelial cells, human umbilical vein endothelial cells (HUVECs) were treated with20mmol/L glucose, and ROS level was detected by flow cytometry. Comparing with the control group, treatment of20mmol/L glucose could lead to excessive production of ROS in HUVECs. However, preincubation of HUVECs with DPI, the NOX inhibitor, but not Rotenone, Oxypurinol or L-NAME could inhibit the generation of ROS, indicating that NOX is the main source of high glucose-induced ROS generation. Further, the expression of NOX subunits was investigated by RT-PCR, Real-time PCR and Western blot. The results showed that NOX4and sub-units p22PHOX, p47PHOX, p67PHOX, Rac-1were expressed in HUVECs, but only the expression of NOX4was significantly upregulated after20mmol/L glucose treatment in HUVECs. The gene profile screened through SuperArray's PCR array indicated that the level of Dual Oxidase2mRNA could be increased by nearly13fold, while the level of Superoxide dismutase3mRNA could be significantly decreased in HUVECs treated with20mmol/L glucose. p47PHOx and Rac-1were translocated to cell membrane from cytoplasm to mediate the activation of NOX4and the generation of ROS, and PKC was also participated in regulating NOX4-mediated ROS generation. It was found that high concentration of glucose could stimulate the production of ROS and induce apoptosis through upregulating the expression of NOX4in HUVECs. Moreover, two strategies, lost of function and gain of function, were carried out to observe effects of NOX4on ROS generation and injury of HUVECs. The NOX4was knocked down by transfection with NOX4-siRNA, and overexpressed by transfection with NOX4-expressing vector in HUVECs. The results showed that overexpression of NOX4could induce the excessive production of ROS and apoptosis of HUVECs. Contrast, down-regulation of NOX4significantly reduced the level of ROS and prevented HUVECs from apoptosis. Furthermore, to study the mechanisms of apoptosis induced by glucose, nuclear factor-κ B (NF-κB) and p38MAPK pathways were analyzed by Western blot and immunofluorescence staining. After glucose treatment, phosphorylated IkB level was elevated and IκB was degraded, and NF-κB was also phosphorylated and translocated into nucleus in where NF-κB upregulated the expression of inflammatory factors such as ICAM-1and VCAM-1. p38MAPK pathway is considered as an important signaling pathway participated in regulation of endothelial apoptosis. The high glucose resulted in phosphorylation of p38MAPK and the translocation of p38MAPK into nucleus. SB203580, a specific inhibitor of p38MAPK, could inhibit high glucose-induced upregulation of P-p38MAPK and suppress high glucose-induced apoptosis, as measured by Hoechst/PI, TUNEL and PI/Annexin V-FITC staining. These pointed out that p38MAPK played a key role in the glucose-induced apoptosis. Recent results have revealed that beyond lipid-lowering activity, statins exhibit functions such as modulating inflammatory responses, improving antioxidant effects and maintaining plaque stability. Our previous results provided that Simvastatin could downregulate the expression of NOX4but not NOX2and impair the excessive production of ROS induced by treatment of0.8g/L LDL. These results indicated that Simvastatin could lower ROS level by suppression of NOX4expression. To further investgate Simvastin's antioxidant effects in vivo, oxidative stress was analyzed in atherosclerotic Apo E-/-mice treated with Simvastatin. The results showed that Simvastatin could reduce the levels of TG and MDA in serum and the expression of NOX in vascular system, in term impaired the oxidative stress of atherosclerotic mice. In addition, the average area of atherosclerotic lesion was decreased, and the thickness and integrity of fibrous cap were improved in Simvastatin-treated mice, suggesting that Simvastatin has the ability to stabilize the atherosclerotic plaque.
In summary, our results provide the molecular evidence that NOX4-derived ROS could play an important role in oxidative stress of vascular endothelial cells induced by high concentration of glucose. Specifically inhibiting NOX activity may be a novel target preventing vascular complications of diabetes.
引文
1. Sorescu, D., et al., Superoxide Production and Expression of Nox Family Proteins in Human Atherosclerosis. Circulation,2002.105(12):p.1429-1435.
2. Ago, T., et al., Nox4 as the Major Catalytic Component of an Endothelial NAD(P)H Oxidase. Circulation,2004.109(2):p.227-233.
3. Kwan, J., et al., In high glucose protein kinase C-zeta activation is required for mesangial cell generation of reactive oxygen species. Kidney Int,2005.68(6): p.2526-41.
4. Pricci, F., et al., Oxidative stress in diabetes-induced endothelial dysfunction involvement of nitric oxide and protein kinase C. Free Radic Biol Med,2003. 35(6):p.683-94.
5. Lee, H.B., et al., Reactive oxygen species amplify protein kinase C signaling in high glucose-induced fibronectin expression by human peritoneal mesothelial cells. Kidney Int,2004.65(4):p.1170-1179.
6. Jiang, C., et al., Methyl selenium-induced vascular endothelial apoptosis is executed by caspases and principally mediated by p38 MAPK pathway. Nutr Cancer,2004.49(2):p.174-83.
7. Osone, S., et al., Fenretinide induces sustained-activation of JNK/p38 MAPK and apoptosis in α reactive oxygen species-dependent manner in neuroblastoma cells. Int J Cancer,2004.112(2):p.219-24.
8. Pearl-Yafe, M., et al., The p38 pathway partially mediates caspase-3 activation induced by reactive oxygen species in Fanconi anemia C cells. Biochem Pharmacol,2004.67(3):p.539-46.
9. Ho, F.M., et al., High Glucose-Induced Apoptosis in Human Endothelial Cells Is Mediated by Sequential Activations of c-Jun NH2-Terminal Kinase and Caspase-3. Circulation,2000.101(22):p.2618-2624.
10. Nakagami, H., et al., Phosphorylation of p38 Mitogen-Activated Protein Kinase Downstream of Bax-Caspase-3 Pathway Leads to Cell Death Induced by High D-Glucose in Human Endothelial Cells. Diabetes,2001.50(6):p. 1472-1481.
11. Jiang, D.J., et al., Asymmetric dimethylarginine induces apoptosis via p38 MAPK/caspase-3-dependent signaling pathway in endothelial cells. J Mol Cell Cardiol,2006.40(4):p.529-39.
12. Quagliaro, L., et al., Intermittent High Glucose Enhances Apoptosis Related to Oxidative Stress in Human Umbilical Vein Endothelial Cells:The Role of Protein Kinase C and NAD(P)H-Oxidase Activation. Diabetes,2003.52(11):p. 2795-2804.
13. Bedard, K. and K.-H. Krause, The NOX Family of ROS-Generating NADPH Oxidases:Physiology and Pathophysiology. Physiol. Rev.,2007.87(1):p. 245-313.
14. Riganti, C., et al., Diphenyleneiodonium Inhibits the Cell Redox Metabolism and Induces Oxidative Stress. J. Biol. Chem.,2004.279(46):p.47726-47731.
15. Balcerczyk, A., et al., Induction of apoptosis and modulation of production of reactive oxygen species in human endothelial cells by diphenyleneiodonium. Biochem Pharmacol,2005.69(8):p.1263-73.
16. Guzik, T.J., et al., Mechanisms of Increased Vascular Superoxide Production in Human Diabetes Mellitus:Role of NAD(P)H Oxidase and Endothelial Nitric Oxide Synthase. Circulation,2002.105(14):p.1656-1662.
17. Zhang, Z., et al., High Glucose Inhibits Glucose-6-phosphate Dehydrogenase via cAMP in Aortic Endothelial Cells. J. Biol. Chem.,2000.275(51):p. 40042-40047.
18. Inoguchi, T., et al., Protein Kinase C-Dependent Increase in Reactive Oxygen Species (ROS) Production in Vascular Tissues of Diabetes:Role of Vascular NAD(P)H Oxidase. J Am Soc Nephrol,2003.14(90003):p. S227-232.
19. Babior, B.M., NADPH Oxidase:An Update. Blood,1999.93(5):p. 1464-1476.
20. DeLeo, F.R., et al., NADPH Oxidase Activation and Assembly During Phagocytosis. J Immunol,1999.163(12):p.6732-6740.
21. Bissonnette, S.A., et al., Phosphatidylinositol 3-Phosphate-dependent and-independent Functions of p40phox in Activation of the Neutrophil NADPH Oxidase. J. Biol. Chem.,2008.283(4):p.2108-2119.
22. Li, J.-M., et al., Essential Role of the NADPH Oxidase Subunit p47phox in Endothelial Cell Superoxide Production in Response to Phorbol Ester and Tumor Necrosis Factor-(alpha). Circ Res,2002.90(2):p.143-150.
23. Xia, L., et al., Mesangial cell NADPH oxidase upregulation in high glucose is protein kinase C dependent and required for collagen TV expression. Am J Physiol Renal Physiol,2006.290(2):p. F345-356.
24. Silver, A.E., et al., Overweight and Obese Humans Demonstrate Increased Vascular Endothelial NAD(P)H Oxidase-p47phox Expression and Evidence of Endothelial Oxidative Stress. Circulation,2007.115(5):p.627-637.
25. Baumer, A.T., et al., Phosphatidylinositol 3-Kinase-dependent Membrane Recruitment of Rac-1 and p47phox Is Critical for{alpha}-Platelet-derived Growth Factor Receptor-induced Production of Reactive Oxygen Species. J. Biol. Chem.,2008.283(12):p.7864-7876.
26. Usatyuk, P.V., et al., Regulation of Hyperoxia-induced NADPH Oxidase Activation in Human Lung Endothelial Cells by the Actin Cytoskeleton and Cortactin. J. Biol. Chem.,2007.282(32):p.23284-23295.
27. Kitada, M., et al., Translocation of Glomerular p47phox and p67phox by Protein Kinase C-{beta} Activation Is Required for Oxidative Stress in Diabetic Nephropathy. Diabetes,2003.52(10):p.2603-2614.
28. Xia, L., et al., Reactive oxygen species, PKC-betal, and PKC-{zeta} mediate high-glucose-induced vascular endothelial growth factor expression in mesangial cells. Am J Physiol Endocrinol Metab,2007.293(5):p. E1280-1288.
29. Lassegue, B. and R.E. Clempus, Vascular NAD(P)H oxidases:specific features, expression, and regulation. Am J Physiol Regul Integr Comp Physiol, 2003.285(2):p. R277-297.
30. Iwashima, F., et al., Aldosterone Induces Superoxide Generation via Racl Activation in Endothelial Cells. Endocrinology,2008.149(3):p.1009-1014.
31. Krotz, F., et al., Mycophenolate Acid Inhibits Endothelial NAD(P)H Oxidase Activity and Superoxide Formation by a Racl-Dependent Mechanism. Hypertension,2007.49(1):p.201-208.
32. Cheng, G., et al., Noxl-dependent Reactive Oxygen Generation Is Regulated by Racl. J. Biol. Chem.,2006.281(26):p.17718-17726.
33. Ueyama, T., M. Geiszt, and T.L. Leto, Involvement of Racl in Activation of Multicomponent Noxl-and Nox3-Based NADPH Oxidases. Mol. Cell. Biol., 2006.26(6):p.2160-2174.
34. Furst, R., et al., Atrial Natriuretic Peptide Induces Mitogen-Activated Protein Kinase Phosphatase-1 in Human Endothelial Cells via Racl and NAD(P)H Oxidase/Nox2-Activation. Circ Res,2005.96(1):p.43-53.
35. Vecchione, C., et al., Selective Rac-1 Inhibition Protects From Diabetes-Induced Vascular Injury. Circ Res,2006.98(2):p.218-225.
36. Pagano, P.J., et al., Localization of a constitutively active, phagocyte-like NADPH oxidase in rabbit aortic adventitia:Enhancement by angiotensin 营Ⅰ. Proceedings of the National Academy of Sciences,1997.94(26):p. 14483-14488.
37. Li, X.J., et al., Leu505 of Nox2 is crucial for optimal p67phox-dependent activation of the flavocytochrome b558 during phagocytic NADPH oxidase assembly. J Leukoc Biol,2007.81(1):p.238-249.
38. Weaver, M., et al., Adventitial delivery of dominant-negative p67phox attenuates neointimal hyperplasia of the rat carotid artery. Am J Physiol Heart Circ Physiol,2006.290(5):p. H1933-1941.
39. Yamakawa, T., et al., Lysophosphatidylcholine Activates Extracellular Signal-Regulated Kinases 1/2 Through Reactive Oxygen Species in Rat Vascular Smooth Muscle Cells. Arterioscler Thromb Vase Biol,2002.22(5):p. 752-758.
40. Frey, R.S., et al., PKC{zeta} Regulates TNF-{alpha}-Induced Activation of NADPH Oxidase in Endothelial Cells. Circ Res,2002.90(9):p.1012-1019.
41. Banfi, B., et al., A Ca2+ -activated NADPH Oxidase in Testis, Spleen, and Lymph Nodes. J. Biol. Chem.,2001.276(40):p.37594-37601.
42. Thallas-Bonke, V., et al., Inhibition of NADPH Oxidase Prevents Advanced Glycation End Product-Mediated Damage in Diabetic Nephropathy Through a Protein Kinase C-(alpha)-Dependent Pathway. Diabetes,2008.57(2):p. 460-469.
43. Geiszt, M., et al., Dual oxidases represent novel hydrogen peroxide sources supporting mucosal surface host defense. FASEB J.,2003.17(11):p. 1502-1504.
44. Johnson, K.R., et al., Congenital Hypothyroidism, Dwarfism, and Hearing Impairment Caused by a Missense Mutation in the Mouse Dual Oxidase 2 Gene, Duox2. Mol Endocrinol,2007.21(7):p.1593-1602.
45. Yano, M., et al., Short-term exposure of high glucose concentration induces generation of reactive oxygen species in endothelial cells:implication for the oxidative stress associated with postprandial hyperglycemia. Redox Rep,2004. 9(2):p.111-6.
46. Touyz, R.M., et al., Expression of a Functionally Active gp91phox-Containing Neutrophil-Type NAD(P)H Oxidase in Smooth Muscle Cells From Human Resistance Arteries:Regulation by Angiotensin Ⅱ. Circ Res,2002.90(11):p. 1205-1213.
47. Lee, S.B., et al., Serum withdrawal kills U937 cells by inducing a positive mutual interaction between reactive oxygen species and phosphoinositide 3-kinase. Cell Signal,2005.17(2):p.197-204.
48. Neumann, M. and M. Naumann, Beyond I{kappa}Bs:alternative regulation of NF-{kappa}B activity. FASEB J.,2007.21(11):p.2642-2654.
49. Moynagh, P.N., The NF-{kappa}B pathway. J Cell Sci,2005.118(20):p. 4589-4592.
50. Hayden, M.S. and S. Ghosh, Signaling to NF-{kappa}B. Genes Dev.,2004. 18(18):p.2195-2224.
51. Jobin, C. and R.B. Sartor, The Ikappa B/NF-kappa B system:a key determinant of mucosal inflammation and protection. Am J Physiol Cell Physiol,2000.278(3):p. C451-462.
52. Liu, S.F. and A.B. Malik, NF-{kappa}B activation as a pathological mechanism of septic shock and inflammation. Am J Physiol Lung Cell Mol Physiol,2006.290(4):p. L622-645.
53. Somwar, R., et al., A Dominant-negative p38 MAPK Mutant and Novel Selective Inhibitors of p38 MAPK Reduce Insulin-stimulated Glucose Uptake in 3T3-L1 Adipocytes without Affecting GLUT4 Translocation. J. Biol. Chem., 2002.277(52):p.50386-50395.
54. Fujishiro, M., et al., MKK6/3 and p38 MAPK Pathway Activation Is Not Necessary for Insulin-induced Glucose Uptake but Regulates Glucose Transporter Expression. J. Biol. Chem.,2001.276(23):p.19800-19806.
55. Gonzalez-Yanes, C., et al., Oleylethanolamide impairs glucose tolerance and inhibits insulin-stimulated glucose uptake in rat adipocytes through p38 and JNK MAPK pathways. Am J Physiol Endocrinol Metab,2005.289(5):p. E923-929.
56. Montessuit, C., et al., Regulation of glucose transporter expression in cardiac myocytes:p38 MAPK is a strong inducer of GLUT4. Cardiovasc Res,2004. 64(1):p.94-104.
57. Tsiani, E., et al., High glucose-enhanced activation of mesangial cell p38 MAPK by ET-1, ANG Ⅱ, and platelet-derived growth factor. Am J Physiol Endocrinol Metab,2002.282(1):p. E161-169.
58. Kawano, Y., et al., Evidence against high glucose as a mediator of ERK1/2 or p38 MAPK phosphorylation in rat skeletal muscle. Am J Physiol Endocrinol Metab,2001.281(6):p. E1255-1259.
59. Begum, N. and L. Ragolia, High glucose and insulin inhibit VSMC MKP-1 expression by blocking iNOS via p38 MAPK activation. Am J Physiol Cell Physiol,2000.278(1):p. C81-91.
60. Patel, T.N., M.H. Shishehbor, and D.L. Bhatt, A review of high-dose statin therapy:targeting cholesterol and inflammation in atherosclerosis. Eur Heart J,2007.28(6):p.664-672.
61. Maron, D.J., S. Fazio, and M.F. Linton, Current Perspectives on Statins. Circulation,2000.101(2):p.207-213.
62. Suh, Y.A., et al., Cell transformation by the superoxide-generating oxidase Moxl. Nature,1999.401(6748):p.79-82.
63. Wassmann, S., et al., Cellular Antioxidant Effects of Atorvastatin In Vitro and In Vivo. Arterioscler Thromb Vase Biol,2002.22(2):p.300-305.
1.王镜岩,朱圣庚,and徐长法,生物化学(下册).2006：高等教育出版社.177.
2. Christian, W., et al., The natural history of insulin secretory dysfunction and insulin resistance in the pathogenesis of type 2 diabetes mellitus. J. Clin. Invest.,1999.104(6):p.787-794.
3. Zimmet, P., K.G.M.M. Alberti, and J. Shaw, Global and societal implications of the diabetes epidemic. Nature,2001.414(6865):p.782-787.
4. Salmi, M., et al., Insulin-Regulated Increase of Soluble Vascular Adhesion Protein-1 in Diabetes. Am J Pathol,2002.161(6):p.2255-2262.
5. Ceriello, A., et al., Evidence for an Independent and Cumulative Effect of Postprandial Hypertriglyceridemia and Hyperglycemia on Endothelial Dysfunction and Oxidative Stress Generation:Effects of Short-and Long-Term Simvastatin Treatment. Circulation,2002.106(10):p.1211-1218.
6. Otero, K., et al., Albumin-derived advanced glycation end-products trigger the disruption of the vascular endothelial cadherin complex in cultured human and murine endothelial cells. Biochem J,2001.359(Pt 3):p.567-74.
7. Stratton, I.M., et al., Association of glycaemia with macrovascular and microvascular complications of type 2 diabetes (UKPDS 35):prospective observational study. BMJ,2000.321(7258):p.405-412.
8. Fonseca, V.A., Management of diabetes mellitus and insulin resistance in patients with cardiovascular disease. Am J Cardiol,2003.92(4A):p.50J-60J.
9. Cheng, A.Y.Y. and I.G. Fantus, Oral antihyperglycemic therapy for type 2 diabetes mellitus. CMAJ,2005.172(2):p.213-226.
10. Bouche, C., et al., The Cellular Fate of Glucose and Its Relevance in Type 2 Diabetes. Endocr Rev,2004.25(5):p.807-830.
11. Fryer, L.G.D., A. Parbu-Patel, and D. Carling, The Anti-diabetic Drugs Rosiglitazone and Metformin Stimulate AMP-activated Protein Kinase through Distinct Signaling Pathways. J. Biol. Chem.,2002.277(28):p.25226-25232.
12. Inzucchi, S.E., et al., Efficacy and Metabolic Effects of Metformin and Troglitazone in Type Ⅱ Diabetes Mellitus. N Engl J Med,1998.338(13):p. 867-873.
13. Petersen, K.F., et al., Mechanism of troglitazone action in type 2 diabetes. Diabetes,2000.49(5):p.827-831.
14. Vogt, C., et al., Regulation of hexokinase Ⅱ expression in human skeletal muscle in vivo. Metabolism,2000.49(6):p.814-8.
15. Robey, R.B., et al., Regulation of Mesangial Cell Hexokinase Activity and Expression by Heparin-binding Epidermal Growth Factor-like Growth Factor. J. Biol. Chem.,2002.277(17):p.14370-14378.
16. Ducluzeau, P.-H., et al., Regulation by Insulin of Gene Expression in Human Skeletal Muscle and Adipose Tissue:Evidence for Specific Defects in Type 2 Diabetes. Diabetes,2001.50(5):p.1134-1142.
17. Basu, A., et al., Effects of type 2 diabetes on the ability of insulin and glucose to regulate splanchnic and muscle glucose metabolism:evidence for a defect in hepatic glucokinase activity. Diabetes,2000.49(2):p.272-283.
18. Sato, F., et al., Effects of Diet-Induced Moderate Weight Reduction on Intrahepatic and Intramyocellular Triglycerides and Glucose Metabolism in Obese Subjects. J Clin Endocrinol Metab,2007.92(8):p.3326-3329.
19. Kelley, D.E., et al., Dysfunction of Mitochondria in Human Skeletal Muscle in Type 2 Diabetes. Diabetes,2002.51(10):p.2944-2950.
20. Mootha, V.K., et al., PGC-1 [alpha]-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet,2003.34(3):p.267-273.
21. Patti, M.E., et al., Coordinated reduction of genes of oxidative metabolism in humans with insulin resistance and diabetes:Potential role of PGC1 and NRF1. Proceedings of the National Academy of Sciences,2003.100(14):p. 8466-8471.
22. Gastaldelli, A., et al., Effect of Physiological Hyperinsulinemia on Gluconeogenesis in Nondiabetic Subjects and in Type 2 Diabetic Patients. Diabetes,2001.50(8):p.1807-1812.
23. Basu, R., et al., Comparison of the Effects of Pioglitazone and Metformin on Hepatic and Extra-Hepatic Insulin Action in People With Type 2 Diabetes. Diabetes,2008.57(1):p.24-31.
24. Jin, E.S., et al., Role of Excess Glycogenolysis in Fasting Hyperglycemia Among Pre-Diabetic and Diabetic Zucker (fa/fa) Rats. Diabetes,2007.56(3): p.777-785.
25. Lachin, J.M., et al., Factors Associated With Diabetes Onset During Metformin Versus Placebo Therapy in the Diabetes Prevention Program. Diabetes,2007.56(4):p.1153-1159.
26. Ortmeyer, H.K., et al., Skeletal muscle glycogen synthase subcellular localization:effects of insulin and PPAR-{alpha} agonist (K-111) administration in rhesus monkeys. Am J Physiol Regul Integr Comp Physiol, 2005.288(6):p. R1509-1517.
27. Prats, C., et al., Phosphorylation-dependent Translocation of Glycogen Synthase to a Novel Structure during Glycogen Resynthesis. J. Biol. Chem., 2005.280(24):p.23165-23172.
28. Taylor, A.J., J.M. Ye, and C. Schmitz-Peiffer, Inhibition of glycogen synthesis by increased lipid availability is associated with subcellular redistribution of glycogen synthase. J Endocrinol,2006.188(1):p.11-23.
29. Cline, G.W., et al., Impaired Glucose Transport as a Cause of Decreased Insulin-Stimulated Muscle Glycogen Synthesis in Type 2 Diabetes. N Engl J Med,1999.341(4):p.240-246.
30. Pratipanawatr, T., et al., Normalization of Plasma Glucose Concentration by Insulin Therapy Improves Insulin-Stimulated Glycogen Synthesis in Type 2 Diabetes. Diabetes,2002.51(2):p.462-468.
31. M, M., et al., Regulation of endogenous glucose production by glucose per se is impaired in type 2 diabetes mellitus. J Clin Invest.,1998.102(4):p. 744-753.
32. Shah, P., et al., Lack of Suppression of Glucagon Contributes to Postprandial Hyperglycemia in Subjects with Type 2 Diabetes Mellitus. J Clin Endocrinol Metab,2000.85(11):p.4053-4059.
33. Dunning, B.E. and J.E. Gerich, The Role of {alpha}-Cell Dysregulation in Fasting and Postprandial Hyperglycemia in Type 2 Diabetes and Therapeutic Implications. Endocr Rev,2007.28(3):p.253-283.
34. Singh, V., et al., Somatostatin Receptor Subtype-2-Deficient Mice with Diet-Induced Obesity Have Hyperglycemia, Nonfasting Hyperglucagonemia, and Decreased Hepatic Glycogen Deposition. Endocrinology,2007.148(8):p. 3887-3899.
35. Groemping, Y, et al., Molecular basis of phosphorylation-induced activation of the NADPH oxidase. Cell,2003.113(3):p.343-55.
36. Xia, L., et al., Mesangial cell NADPH oxidase upregulation in high glucose is protein kinase C dependent and required for collagen IV expression. Am J Physiol Renal Physiol,2006.290(2):p. F345-356.
37. Vecchione, C., et al., Selective Rac-1 Inhibition Protects From Diabetes-Induced Vascular Injury. Circ Res,2006.98(2):p.218-225.
38. Hayaishi-Okano, R., et al., NAD (P) H oxidase p22 phox C242T polymorphism affects LDL particle size and insulin resistance in Japanese subjects. J Atheroscler Thromb,2002.9(4):p.200-5.
39. Weidig, P., D. McMaster, and U. Bayraktutan, High glucose mediates pro-oxidant and antioxidant enzyme activities in coronary endothelial cells. Diabetes Obes Metab,2004.6(6):p.432-41.
40. Matsushita, H., K.H. Lee, and P.S. Tsao, Cyclic strain induces reactive oxygen species production via an endothelial NAD(P)H oxidase. J Cell Biochem Suppl,2001. Suppl 36:p.99-106.
41. Gorlach, A., et al., Oxidative stress and expression of p22phox are involved in the up-regulation of tissue factor in vascular smooth muscle cells in response to activated platelets. FASEB J.,2000.14(11):p.1518-1528.
42. Hink, U., et al., Mechanisms Underlying Endothelial Dysfunction in Diabetes Mellitus. Circ Res,2001.88(2):p. e14-22.
43. Guzik, T.J., et al., Mechanisms of Increased Vascular Superoxide Production in Human Diabetes Mellitus:Role of NAD(P)H Oxidase and Endothelial Nitric Oxide Synthase. Circulation,2002.105(14):p.1656-1662.
44. Nishikawa, T., et al., Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature,2000.404(6779):p. 787-790.
45. Du, X.-L., et al., Hyperglycemia-induced mitochondrial superoxide overproduction activates the hexosamine pathway and induces plasminogen activator inhibitor-1 expression by increasing Spl glycosylation. Proceedings of the National Academy of Sciences,2000.97(22):p.12222-12226.
46. El-Remessy, A.B., et al., High Glucose-Induced Tyrosine Nitration in Endothelial Cells:Role of eNOS Uncoupling and Aldose Reductase Activation. Invest. Ophthalmol. Vis. Sci.,2003.44(7):p.3135-3143.
47. Zhang, Z., et al., High Glucose Inhibits Glucose-6-phosphate Dehydrogenase via cAMP in Aortic Endothelial Cells. J. Biol. Chem.,2000.275(51):p. 40042-40047.
48. Leopold, J.A., et al., Glucose-6-Phosphate Dehydrogenase Overexpression Decreases Endothelial Cell Oxidant Stress and Increases Bioavailable Nitric Oxide. Arterioscler Thromb Vase Biol,2003.23(3):p.411-417.
49. Leopold, J.A., et al., Glucose-6-phosphate dehydrogenase deficiency promotes endothelial oxidant stress and decreases endothelial nitric oxide bioavailability. FASEB J.,2001:p.00-0893fje.
50. Dandona, P., et al., Insulin Inhibits Intranuclear Nuclear Factor {{kappajj B and Stimulates Ⅰ {{kappa}}B in Mononuclear Cells in Obese Subjects: Evidence for an Anti-inflammatory Effect? J Clin Endocrinol Metab,2001. 86(7):p.3257-3265.
51. Sheu, M.L., et al., High Glucose Induces Human Endothelial Cell Apoptosis Through a Phosphoinositide 3-Kinase-Regulated Cyclooxygenase-2 Pathway. Arterioscler Thromb Vase Biol,2005.25(3):p.539-545.
52. Cosentino, F., et al., High Glucose Causes Upregulation of Cyclooxygenase-2 and Alters Prostanoid Profile in Human Endothelial Cells:Role of Protein Kinase C and Reactive Oxygen Species. Circulation,2003.107(7):p. 1017-1023.
53. Ha, H. and H.B. Lee, Reactive oxygen species amplify glucose signalling in renal cells cultured under high glucose and in diabetic kidney. Nephrology (Carlton),2005.10 Suppl:p. S7-10.
54. Piconi, L., et al., Intermittent high glucose enhances ICAM-1, VCAM-1, E-selectin and interleukin-6 expression in human umbilical endothelial cells in culture:the role of poly(ADP-ribose) polymerase. J Thromb Haemost,2004. 2(8):p.1453-9.
55. Srinivasan, S., et al., Glucose Regulates Interleukin-8 Production in Aortic Endothelial Cells through Activation of the p38 Mitogen-activated Protein Kinase Pathway in Diabetes. J. Biol. Chem.,2004.279(30):p.31930-31936.
56. Takaishi, H., et al., High glucose accelerates MCP-1 production via p38 MAPK in vascular endothelial cells. Biochem Biophys Res Commun,2003. 305(1):p.122-8.
57. Kyriakis, J.M. and J. Avruch, Mammalian Mitogen-Activated Protein Kinase Signal Transduction Pathways Activated by Stress and Inflammation. Physiol. Rev.,2001.81(2):p.807-869.
58. Liu, W., A. Schoenkerman, and W.L. Lowe, Jr., Activation of members of the mitogen-activated protein kinase family by glucose in endothelial cells. Am J Physiol Endocrinol Metab,2000.279(4):p. E782-790.
59. Ho, F.M., et al., High glucose-induced apoptosis in human vascular endothelial cells is mediated through NF-kappaB and c-Jun NH2-terminal kinase pathway and prevented by PI3K/Akt/eNOS pathway. Cell Signal,2006. 18(3):p.391-9.
60. Kwan, J., et al., In high glucose protein kinase C-zeta activation is required for mesangial cell generation of reactive oxygen species. Kidney Int,2005.68(6): p.2526-41.
61. Ceolotto, G, et al., Rosiglitazone Reduces Glucose-Induced Oxidative Stress Mediated by NAD(P)H Oxidase via AMPK-Dependent Mechanism. Arterioscler Thromb Vase Biol,2007.27(12):p.2627-2633.
62. Pricci, F., et al., Oxidative stress in diabetes-induced endothelial dysfunction involvement of nitric oxide and protein kinase C. Free Radic Biol Med,2003. 35(6):p.683-94.
63. Shaw, S., et al., High Glucose Augments the Angiotensin Ⅱ-induced Activation of JAK2 in Vascular Smooth Muscle Cells via the Polyol Pathway. J. Biol. Chem.,2003.278(33):p.30634-30641.
64. The Diabetes Prevention Program Research, G., Effects of Withdrawal From Metformin on the Development of Diabetes in the Diabetes Prevention Program. Diabetes Care,2003.26(4):p.977-980.
65. Heine, R.J., et al., Management of hyperglycaemia in type 2 diabetes:the end of recurrent failure? BMJ,2006.333(7580):p.1200-1204.
66. Committee, T.C.D.A.C.P.GE., Canadian Diabetes Association 2003 Clinical Practice Guidelines for the Prevention and Management of Diabetes in Canada. Can J Diabetes,2003.27:p. S1-S152.
67. Welschen, L.M.C., et al., Self-Monitoring of Blood Glucose in Patients With Type 2 Diabetes Who Are Not Using Insulin:A systematic review. Diabetes Care,2005.28(6):p.1510-1517.
68. van Dam, R.M. and F.B. Hu, Coffee Consumption and Risk of Type 2 Diabetes: A Systematic Review. JAMA,2005.294(1):p.97-104.
2. Ago, T., et al., Nox4 as the Major Catalytic Component of an Endothelial NAD(P)H Oxidase. Circulation,2004.109(2):p.227-233.
3. Kwan, J., et al., In high glucose protein kinase C-zeta activation is required for mesangial cell generation of reactive oxygen species. Kidney Int,2005.68(6): p.2526-41.
4. Pricci, F., et al., Oxidative stress in diabetes-induced endothelial dysfunction involvement of nitric oxide and protein kinase C. Free Radic Biol Med,2003. 35(6):p.683-94.
5. Lee, H.B., et al., Reactive oxygen species amplify protein kinase C signaling in high glucose-induced fibronectin expression by human peritoneal mesothelial cells. Kidney Int,2004.65(4):p.1170-1179.
6. Jiang, C., et al., Methyl selenium-induced vascular endothelial apoptosis is executed by caspases and principally mediated by p38 MAPK pathway. Nutr Cancer,2004.49(2):p.174-83.
7. Osone, S., et al., Fenretinide induces sustained-activation of JNK/p38 MAPK and apoptosis in α reactive oxygen species-dependent manner in neuroblastoma cells. Int J Cancer,2004.112(2):p.219-24.
8. Pearl-Yafe, M., et al., The p38 pathway partially mediates caspase-3 activation induced by reactive oxygen species in Fanconi anemia C cells. Biochem Pharmacol,2004.67(3):p.539-46.
9. Ho, F.M., et al., High Glucose-Induced Apoptosis in Human Endothelial Cells Is Mediated by Sequential Activations of c-Jun NH2-Terminal Kinase and Caspase-3. Circulation,2000.101(22):p.2618-2624.
10. Nakagami, H., et al., Phosphorylation of p38 Mitogen-Activated Protein Kinase Downstream of Bax-Caspase-3 Pathway Leads to Cell Death Induced by High D-Glucose in Human Endothelial Cells. Diabetes,2001.50(6):p. 1472-1481.
11. Jiang, D.J., et al., Asymmetric dimethylarginine induces apoptosis via p38 MAPK/caspase-3-dependent signaling pathway in endothelial cells. J Mol Cell Cardiol,2006.40(4):p.529-39.
12. Quagliaro, L., et al., Intermittent High Glucose Enhances Apoptosis Related to Oxidative Stress in Human Umbilical Vein Endothelial Cells:The Role of Protein Kinase C and NAD(P)H-Oxidase Activation. Diabetes,2003.52(11):p. 2795-2804.
13. Bedard, K. and K.-H. Krause, The NOX Family of ROS-Generating NADPH Oxidases:Physiology and Pathophysiology. Physiol. Rev.,2007.87(1):p. 245-313.
14. Riganti, C., et al., Diphenyleneiodonium Inhibits the Cell Redox Metabolism and Induces Oxidative Stress. J. Biol. Chem.,2004.279(46):p.47726-47731.
15. Balcerczyk, A., et al., Induction of apoptosis and modulation of production of reactive oxygen species in human endothelial cells by diphenyleneiodonium. Biochem Pharmacol,2005.69(8):p.1263-73.
16. Guzik, T.J., et al., Mechanisms of Increased Vascular Superoxide Production in Human Diabetes Mellitus:Role of NAD(P)H Oxidase and Endothelial Nitric Oxide Synthase. Circulation,2002.105(14):p.1656-1662.
17. Zhang, Z., et al., High Glucose Inhibits Glucose-6-phosphate Dehydrogenase via cAMP in Aortic Endothelial Cells. J. Biol. Chem.,2000.275(51):p. 40042-40047.
18. Inoguchi, T., et al., Protein Kinase C-Dependent Increase in Reactive Oxygen Species (ROS) Production in Vascular Tissues of Diabetes:Role of Vascular NAD(P)H Oxidase. J Am Soc Nephrol,2003.14(90003):p. S227-232.
19. Babior, B.M., NADPH Oxidase:An Update. Blood,1999.93(5):p. 1464-1476.
20. DeLeo, F.R., et al., NADPH Oxidase Activation and Assembly During Phagocytosis. J Immunol,1999.163(12):p.6732-6740.
21. Bissonnette, S.A., et al., Phosphatidylinositol 3-Phosphate-dependent and-independent Functions of p40phox in Activation of the Neutrophil NADPH Oxidase. J. Biol. Chem.,2008.283(4):p.2108-2119.
22. Li, J.-M., et al., Essential Role of the NADPH Oxidase Subunit p47phox in Endothelial Cell Superoxide Production in Response to Phorbol Ester and Tumor Necrosis Factor-(alpha). Circ Res,2002.90(2):p.143-150.
23. Xia, L., et al., Mesangial cell NADPH oxidase upregulation in high glucose is protein kinase C dependent and required for collagen TV expression. Am J Physiol Renal Physiol,2006.290(2):p. F345-356.
24. Silver, A.E., et al., Overweight and Obese Humans Demonstrate Increased Vascular Endothelial NAD(P)H Oxidase-p47phox Expression and Evidence of Endothelial Oxidative Stress. Circulation,2007.115(5):p.627-637.
25. Baumer, A.T., et al., Phosphatidylinositol 3-Kinase-dependent Membrane Recruitment of Rac-1 and p47phox Is Critical for{alpha}-Platelet-derived Growth Factor Receptor-induced Production of Reactive Oxygen Species. J. Biol. Chem.,2008.283(12):p.7864-7876.
26. Usatyuk, P.V., et al., Regulation of Hyperoxia-induced NADPH Oxidase Activation in Human Lung Endothelial Cells by the Actin Cytoskeleton and Cortactin. J. Biol. Chem.,2007.282(32):p.23284-23295.
27. Kitada, M., et al., Translocation of Glomerular p47phox and p67phox by Protein Kinase C-{beta} Activation Is Required for Oxidative Stress in Diabetic Nephropathy. Diabetes,2003.52(10):p.2603-2614.
28. Xia, L., et al., Reactive oxygen species, PKC-betal, and PKC-{zeta} mediate high-glucose-induced vascular endothelial growth factor expression in mesangial cells. Am J Physiol Endocrinol Metab,2007.293(5):p. E1280-1288.
29. Lassegue, B. and R.E. Clempus, Vascular NAD(P)H oxidases:specific features, expression, and regulation. Am J Physiol Regul Integr Comp Physiol, 2003.285(2):p. R277-297.
30. Iwashima, F., et al., Aldosterone Induces Superoxide Generation via Racl Activation in Endothelial Cells. Endocrinology,2008.149(3):p.1009-1014.
31. Krotz, F., et al., Mycophenolate Acid Inhibits Endothelial NAD(P)H Oxidase Activity and Superoxide Formation by a Racl-Dependent Mechanism. Hypertension,2007.49(1):p.201-208.
32. Cheng, G., et al., Noxl-dependent Reactive Oxygen Generation Is Regulated by Racl. J. Biol. Chem.,2006.281(26):p.17718-17726.
33. Ueyama, T., M. Geiszt, and T.L. Leto, Involvement of Racl in Activation of Multicomponent Noxl-and Nox3-Based NADPH Oxidases. Mol. Cell. Biol., 2006.26(6):p.2160-2174.
34. Furst, R., et al., Atrial Natriuretic Peptide Induces Mitogen-Activated Protein Kinase Phosphatase-1 in Human Endothelial Cells via Racl and NAD(P)H Oxidase/Nox2-Activation. Circ Res,2005.96(1):p.43-53.
35. Vecchione, C., et al., Selective Rac-1 Inhibition Protects From Diabetes-Induced Vascular Injury. Circ Res,2006.98(2):p.218-225.
36. Pagano, P.J., et al., Localization of a constitutively active, phagocyte-like NADPH oxidase in rabbit aortic adventitia:Enhancement by angiotensin 营Ⅰ. Proceedings of the National Academy of Sciences,1997.94(26):p. 14483-14488.
37. Li, X.J., et al., Leu505 of Nox2 is crucial for optimal p67phox-dependent activation of the flavocytochrome b558 during phagocytic NADPH oxidase assembly. J Leukoc Biol,2007.81(1):p.238-249.
38. Weaver, M., et al., Adventitial delivery of dominant-negative p67phox attenuates neointimal hyperplasia of the rat carotid artery. Am J Physiol Heart Circ Physiol,2006.290(5):p. H1933-1941.
39. Yamakawa, T., et al., Lysophosphatidylcholine Activates Extracellular Signal-Regulated Kinases 1/2 Through Reactive Oxygen Species in Rat Vascular Smooth Muscle Cells. Arterioscler Thromb Vase Biol,2002.22(5):p. 752-758.
40. Frey, R.S., et al., PKC{zeta} Regulates TNF-{alpha}-Induced Activation of NADPH Oxidase in Endothelial Cells. Circ Res,2002.90(9):p.1012-1019.
41. Banfi, B., et al., A Ca2+ -activated NADPH Oxidase in Testis, Spleen, and Lymph Nodes. J. Biol. Chem.,2001.276(40):p.37594-37601.
42. Thallas-Bonke, V., et al., Inhibition of NADPH Oxidase Prevents Advanced Glycation End Product-Mediated Damage in Diabetic Nephropathy Through a Protein Kinase C-(alpha)-Dependent Pathway. Diabetes,2008.57(2):p. 460-469.
43. Geiszt, M., et al., Dual oxidases represent novel hydrogen peroxide sources supporting mucosal surface host defense. FASEB J.,2003.17(11):p. 1502-1504.
44. Johnson, K.R., et al., Congenital Hypothyroidism, Dwarfism, and Hearing Impairment Caused by a Missense Mutation in the Mouse Dual Oxidase 2 Gene, Duox2. Mol Endocrinol,2007.21(7):p.1593-1602.
45. Yano, M., et al., Short-term exposure of high glucose concentration induces generation of reactive oxygen species in endothelial cells:implication for the oxidative stress associated with postprandial hyperglycemia. Redox Rep,2004. 9(2):p.111-6.
46. Touyz, R.M., et al., Expression of a Functionally Active gp91phox-Containing Neutrophil-Type NAD(P)H Oxidase in Smooth Muscle Cells From Human Resistance Arteries:Regulation by Angiotensin Ⅱ. Circ Res,2002.90(11):p. 1205-1213.
47. Lee, S.B., et al., Serum withdrawal kills U937 cells by inducing a positive mutual interaction between reactive oxygen species and phosphoinositide 3-kinase. Cell Signal,2005.17(2):p.197-204.
48. Neumann, M. and M. Naumann, Beyond I{kappa}Bs:alternative regulation of NF-{kappa}B activity. FASEB J.,2007.21(11):p.2642-2654.
49. Moynagh, P.N., The NF-{kappa}B pathway. J Cell Sci,2005.118(20):p. 4589-4592.
50. Hayden, M.S. and S. Ghosh, Signaling to NF-{kappa}B. Genes Dev.,2004. 18(18):p.2195-2224.
51. Jobin, C. and R.B. Sartor, The Ikappa B/NF-kappa B system:a key determinant of mucosal inflammation and protection. Am J Physiol Cell Physiol,2000.278(3):p. C451-462.
52. Liu, S.F. and A.B. Malik, NF-{kappa}B activation as a pathological mechanism of septic shock and inflammation. Am J Physiol Lung Cell Mol Physiol,2006.290(4):p. L622-645.
53. Somwar, R., et al., A Dominant-negative p38 MAPK Mutant and Novel Selective Inhibitors of p38 MAPK Reduce Insulin-stimulated Glucose Uptake in 3T3-L1 Adipocytes without Affecting GLUT4 Translocation. J. Biol. Chem., 2002.277(52):p.50386-50395.
54. Fujishiro, M., et al., MKK6/3 and p38 MAPK Pathway Activation Is Not Necessary for Insulin-induced Glucose Uptake but Regulates Glucose Transporter Expression. J. Biol. Chem.,2001.276(23):p.19800-19806.
55. Gonzalez-Yanes, C., et al., Oleylethanolamide impairs glucose tolerance and inhibits insulin-stimulated glucose uptake in rat adipocytes through p38 and JNK MAPK pathways. Am J Physiol Endocrinol Metab,2005.289(5):p. E923-929.
56. Montessuit, C., et al., Regulation of glucose transporter expression in cardiac myocytes:p38 MAPK is a strong inducer of GLUT4. Cardiovasc Res,2004. 64(1):p.94-104.
57. Tsiani, E., et al., High glucose-enhanced activation of mesangial cell p38 MAPK by ET-1, ANG Ⅱ, and platelet-derived growth factor. Am J Physiol Endocrinol Metab,2002.282(1):p. E161-169.
58. Kawano, Y., et al., Evidence against high glucose as a mediator of ERK1/2 or p38 MAPK phosphorylation in rat skeletal muscle. Am J Physiol Endocrinol Metab,2001.281(6):p. E1255-1259.
59. Begum, N. and L. Ragolia, High glucose and insulin inhibit VSMC MKP-1 expression by blocking iNOS via p38 MAPK activation. Am J Physiol Cell Physiol,2000.278(1):p. C81-91.
60. Patel, T.N., M.H. Shishehbor, and D.L. Bhatt, A review of high-dose statin therapy:targeting cholesterol and inflammation in atherosclerosis. Eur Heart J,2007.28(6):p.664-672.
61. Maron, D.J., S. Fazio, and M.F. Linton, Current Perspectives on Statins. Circulation,2000.101(2):p.207-213.
62. Suh, Y.A., et al., Cell transformation by the superoxide-generating oxidase Moxl. Nature,1999.401(6748):p.79-82.
63. Wassmann, S., et al., Cellular Antioxidant Effects of Atorvastatin In Vitro and In Vivo. Arterioscler Thromb Vase Biol,2002.22(2):p.300-305.
1.王镜岩,朱圣庚,and徐长法,生物化学(下册).2006：高等教育出版社.177.
2. Christian, W., et al., The natural history of insulin secretory dysfunction and insulin resistance in the pathogenesis of type 2 diabetes mellitus. J. Clin. Invest.,1999.104(6):p.787-794.
3. Zimmet, P., K.G.M.M. Alberti, and J. Shaw, Global and societal implications of the diabetes epidemic. Nature,2001.414(6865):p.782-787.
4. Salmi, M., et al., Insulin-Regulated Increase of Soluble Vascular Adhesion Protein-1 in Diabetes. Am J Pathol,2002.161(6):p.2255-2262.
5. Ceriello, A., et al., Evidence for an Independent and Cumulative Effect of Postprandial Hypertriglyceridemia and Hyperglycemia on Endothelial Dysfunction and Oxidative Stress Generation:Effects of Short-and Long-Term Simvastatin Treatment. Circulation,2002.106(10):p.1211-1218.
6. Otero, K., et al., Albumin-derived advanced glycation end-products trigger the disruption of the vascular endothelial cadherin complex in cultured human and murine endothelial cells. Biochem J,2001.359(Pt 3):p.567-74.
7. Stratton, I.M., et al., Association of glycaemia with macrovascular and microvascular complications of type 2 diabetes (UKPDS 35):prospective observational study. BMJ,2000.321(7258):p.405-412.
8. Fonseca, V.A., Management of diabetes mellitus and insulin resistance in patients with cardiovascular disease. Am J Cardiol,2003.92(4A):p.50J-60J.
9. Cheng, A.Y.Y. and I.G. Fantus, Oral antihyperglycemic therapy for type 2 diabetes mellitus. CMAJ,2005.172(2):p.213-226.
10. Bouche, C., et al., The Cellular Fate of Glucose and Its Relevance in Type 2 Diabetes. Endocr Rev,2004.25(5):p.807-830.
11. Fryer, L.G.D., A. Parbu-Patel, and D. Carling, The Anti-diabetic Drugs Rosiglitazone and Metformin Stimulate AMP-activated Protein Kinase through Distinct Signaling Pathways. J. Biol. Chem.,2002.277(28):p.25226-25232.
12. Inzucchi, S.E., et al., Efficacy and Metabolic Effects of Metformin and Troglitazone in Type Ⅱ Diabetes Mellitus. N Engl J Med,1998.338(13):p. 867-873.
13. Petersen, K.F., et al., Mechanism of troglitazone action in type 2 diabetes. Diabetes,2000.49(5):p.827-831.
14. Vogt, C., et al., Regulation of hexokinase Ⅱ expression in human skeletal muscle in vivo. Metabolism,2000.49(6):p.814-8.
15. Robey, R.B., et al., Regulation of Mesangial Cell Hexokinase Activity and Expression by Heparin-binding Epidermal Growth Factor-like Growth Factor. J. Biol. Chem.,2002.277(17):p.14370-14378.
16. Ducluzeau, P.-H., et al., Regulation by Insulin of Gene Expression in Human Skeletal Muscle and Adipose Tissue:Evidence for Specific Defects in Type 2 Diabetes. Diabetes,2001.50(5):p.1134-1142.
17. Basu, A., et al., Effects of type 2 diabetes on the ability of insulin and glucose to regulate splanchnic and muscle glucose metabolism:evidence for a defect in hepatic glucokinase activity. Diabetes,2000.49(2):p.272-283.
18. Sato, F., et al., Effects of Diet-Induced Moderate Weight Reduction on Intrahepatic and Intramyocellular Triglycerides and Glucose Metabolism in Obese Subjects. J Clin Endocrinol Metab,2007.92(8):p.3326-3329.
19. Kelley, D.E., et al., Dysfunction of Mitochondria in Human Skeletal Muscle in Type 2 Diabetes. Diabetes,2002.51(10):p.2944-2950.
20. Mootha, V.K., et al., PGC-1 [alpha]-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet,2003.34(3):p.267-273.
21. Patti, M.E., et al., Coordinated reduction of genes of oxidative metabolism in humans with insulin resistance and diabetes:Potential role of PGC1 and NRF1. Proceedings of the National Academy of Sciences,2003.100(14):p. 8466-8471.
22. Gastaldelli, A., et al., Effect of Physiological Hyperinsulinemia on Gluconeogenesis in Nondiabetic Subjects and in Type 2 Diabetic Patients. Diabetes,2001.50(8):p.1807-1812.
23. Basu, R., et al., Comparison of the Effects of Pioglitazone and Metformin on Hepatic and Extra-Hepatic Insulin Action in People With Type 2 Diabetes. Diabetes,2008.57(1):p.24-31.
24. Jin, E.S., et al., Role of Excess Glycogenolysis in Fasting Hyperglycemia Among Pre-Diabetic and Diabetic Zucker (fa/fa) Rats. Diabetes,2007.56(3): p.777-785.
25. Lachin, J.M., et al., Factors Associated With Diabetes Onset During Metformin Versus Placebo Therapy in the Diabetes Prevention Program. Diabetes,2007.56(4):p.1153-1159.
26. Ortmeyer, H.K., et al., Skeletal muscle glycogen synthase subcellular localization:effects of insulin and PPAR-{alpha} agonist (K-111) administration in rhesus monkeys. Am J Physiol Regul Integr Comp Physiol, 2005.288(6):p. R1509-1517.
27. Prats, C., et al., Phosphorylation-dependent Translocation of Glycogen Synthase to a Novel Structure during Glycogen Resynthesis. J. Biol. Chem., 2005.280(24):p.23165-23172.
28. Taylor, A.J., J.M. Ye, and C. Schmitz-Peiffer, Inhibition of glycogen synthesis by increased lipid availability is associated with subcellular redistribution of glycogen synthase. J Endocrinol,2006.188(1):p.11-23.
29. Cline, G.W., et al., Impaired Glucose Transport as a Cause of Decreased Insulin-Stimulated Muscle Glycogen Synthesis in Type 2 Diabetes. N Engl J Med,1999.341(4):p.240-246.
30. Pratipanawatr, T., et al., Normalization of Plasma Glucose Concentration by Insulin Therapy Improves Insulin-Stimulated Glycogen Synthesis in Type 2 Diabetes. Diabetes,2002.51(2):p.462-468.
31. M, M., et al., Regulation of endogenous glucose production by glucose per se is impaired in type 2 diabetes mellitus. J Clin Invest.,1998.102(4):p. 744-753.
32. Shah, P., et al., Lack of Suppression of Glucagon Contributes to Postprandial Hyperglycemia in Subjects with Type 2 Diabetes Mellitus. J Clin Endocrinol Metab,2000.85(11):p.4053-4059.
33. Dunning, B.E. and J.E. Gerich, The Role of {alpha}-Cell Dysregulation in Fasting and Postprandial Hyperglycemia in Type 2 Diabetes and Therapeutic Implications. Endocr Rev,2007.28(3):p.253-283.
34. Singh, V., et al., Somatostatin Receptor Subtype-2-Deficient Mice with Diet-Induced Obesity Have Hyperglycemia, Nonfasting Hyperglucagonemia, and Decreased Hepatic Glycogen Deposition. Endocrinology,2007.148(8):p. 3887-3899.
35. Groemping, Y, et al., Molecular basis of phosphorylation-induced activation of the NADPH oxidase. Cell,2003.113(3):p.343-55.
36. Xia, L., et al., Mesangial cell NADPH oxidase upregulation in high glucose is protein kinase C dependent and required for collagen IV expression. Am J Physiol Renal Physiol,2006.290(2):p. F345-356.
37. Vecchione, C., et al., Selective Rac-1 Inhibition Protects From Diabetes-Induced Vascular Injury. Circ Res,2006.98(2):p.218-225.
38. Hayaishi-Okano, R., et al., NAD (P) H oxidase p22 phox C242T polymorphism affects LDL particle size and insulin resistance in Japanese subjects. J Atheroscler Thromb,2002.9(4):p.200-5.
39. Weidig, P., D. McMaster, and U. Bayraktutan, High glucose mediates pro-oxidant and antioxidant enzyme activities in coronary endothelial cells. Diabetes Obes Metab,2004.6(6):p.432-41.
40. Matsushita, H., K.H. Lee, and P.S. Tsao, Cyclic strain induces reactive oxygen species production via an endothelial NAD(P)H oxidase. J Cell Biochem Suppl,2001. Suppl 36:p.99-106.
41. Gorlach, A., et al., Oxidative stress and expression of p22phox are involved in the up-regulation of tissue factor in vascular smooth muscle cells in response to activated platelets. FASEB J.,2000.14(11):p.1518-1528.
42. Hink, U., et al., Mechanisms Underlying Endothelial Dysfunction in Diabetes Mellitus. Circ Res,2001.88(2):p. e14-22.
43. Guzik, T.J., et al., Mechanisms of Increased Vascular Superoxide Production in Human Diabetes Mellitus:Role of NAD(P)H Oxidase and Endothelial Nitric Oxide Synthase. Circulation,2002.105(14):p.1656-1662.
44. Nishikawa, T., et al., Normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage. Nature,2000.404(6779):p. 787-790.
45. Du, X.-L., et al., Hyperglycemia-induced mitochondrial superoxide overproduction activates the hexosamine pathway and induces plasminogen activator inhibitor-1 expression by increasing Spl glycosylation. Proceedings of the National Academy of Sciences,2000.97(22):p.12222-12226.
46. El-Remessy, A.B., et al., High Glucose-Induced Tyrosine Nitration in Endothelial Cells:Role of eNOS Uncoupling and Aldose Reductase Activation. Invest. Ophthalmol. Vis. Sci.,2003.44(7):p.3135-3143.
47. Zhang, Z., et al., High Glucose Inhibits Glucose-6-phosphate Dehydrogenase via cAMP in Aortic Endothelial Cells. J. Biol. Chem.,2000.275(51):p. 40042-40047.
48. Leopold, J.A., et al., Glucose-6-Phosphate Dehydrogenase Overexpression Decreases Endothelial Cell Oxidant Stress and Increases Bioavailable Nitric Oxide. Arterioscler Thromb Vase Biol,2003.23(3):p.411-417.
49. Leopold, J.A., et al., Glucose-6-phosphate dehydrogenase deficiency promotes endothelial oxidant stress and decreases endothelial nitric oxide bioavailability. FASEB J.,2001:p.00-0893fje.
50. Dandona, P., et al., Insulin Inhibits Intranuclear Nuclear Factor {{kappajj B and Stimulates Ⅰ {{kappa}}B in Mononuclear Cells in Obese Subjects: Evidence for an Anti-inflammatory Effect? J Clin Endocrinol Metab,2001. 86(7):p.3257-3265.
51. Sheu, M.L., et al., High Glucose Induces Human Endothelial Cell Apoptosis Through a Phosphoinositide 3-Kinase-Regulated Cyclooxygenase-2 Pathway. Arterioscler Thromb Vase Biol,2005.25(3):p.539-545.
52. Cosentino, F., et al., High Glucose Causes Upregulation of Cyclooxygenase-2 and Alters Prostanoid Profile in Human Endothelial Cells:Role of Protein Kinase C and Reactive Oxygen Species. Circulation,2003.107(7):p. 1017-1023.
53. Ha, H. and H.B. Lee, Reactive oxygen species amplify glucose signalling in renal cells cultured under high glucose and in diabetic kidney. Nephrology (Carlton),2005.10 Suppl:p. S7-10.
54. Piconi, L., et al., Intermittent high glucose enhances ICAM-1, VCAM-1, E-selectin and interleukin-6 expression in human umbilical endothelial cells in culture:the role of poly(ADP-ribose) polymerase. J Thromb Haemost,2004. 2(8):p.1453-9.
55. Srinivasan, S., et al., Glucose Regulates Interleukin-8 Production in Aortic Endothelial Cells through Activation of the p38 Mitogen-activated Protein Kinase Pathway in Diabetes. J. Biol. Chem.,2004.279(30):p.31930-31936.
56. Takaishi, H., et al., High glucose accelerates MCP-1 production via p38 MAPK in vascular endothelial cells. Biochem Biophys Res Commun,2003. 305(1):p.122-8.
57. Kyriakis, J.M. and J. Avruch, Mammalian Mitogen-Activated Protein Kinase Signal Transduction Pathways Activated by Stress and Inflammation. Physiol. Rev.,2001.81(2):p.807-869.
58. Liu, W., A. Schoenkerman, and W.L. Lowe, Jr., Activation of members of the mitogen-activated protein kinase family by glucose in endothelial cells. Am J Physiol Endocrinol Metab,2000.279(4):p. E782-790.
59. Ho, F.M., et al., High glucose-induced apoptosis in human vascular endothelial cells is mediated through NF-kappaB and c-Jun NH2-terminal kinase pathway and prevented by PI3K/Akt/eNOS pathway. Cell Signal,2006. 18(3):p.391-9.
60. Kwan, J., et al., In high glucose protein kinase C-zeta activation is required for mesangial cell generation of reactive oxygen species. Kidney Int,2005.68(6): p.2526-41.
61. Ceolotto, G, et al., Rosiglitazone Reduces Glucose-Induced Oxidative Stress Mediated by NAD(P)H Oxidase via AMPK-Dependent Mechanism. Arterioscler Thromb Vase Biol,2007.27(12):p.2627-2633.
62. Pricci, F., et al., Oxidative stress in diabetes-induced endothelial dysfunction involvement of nitric oxide and protein kinase C. Free Radic Biol Med,2003. 35(6):p.683-94.
63. Shaw, S., et al., High Glucose Augments the Angiotensin Ⅱ-induced Activation of JAK2 in Vascular Smooth Muscle Cells via the Polyol Pathway. J. Biol. Chem.,2003.278(33):p.30634-30641.
64. The Diabetes Prevention Program Research, G., Effects of Withdrawal From Metformin on the Development of Diabetes in the Diabetes Prevention Program. Diabetes Care,2003.26(4):p.977-980.
65. Heine, R.J., et al., Management of hyperglycaemia in type 2 diabetes:the end of recurrent failure? BMJ,2006.333(7580):p.1200-1204.
66. Committee, T.C.D.A.C.P.GE., Canadian Diabetes Association 2003 Clinical Practice Guidelines for the Prevention and Management of Diabetes in Canada. Can J Diabetes,2003.27:p. S1-S152.
67. Welschen, L.M.C., et al., Self-Monitoring of Blood Glucose in Patients With Type 2 Diabetes Who Are Not Using Insulin:A systematic review. Diabetes Care,2005.28(6):p.1510-1517.
68. van Dam, R.M. and F.B. Hu, Coffee Consumption and Risk of Type 2 Diabetes: A Systematic Review. JAMA,2005.294(1):p.97-104.
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