烟酰胺腺嘌呤二核苷酸磷酸氧化酶与心房颤动的研究

心房颤动是最常见的心律失常,其发生和维持的分子机制尚未明确。近年来发现心房颤动存在氧化应激,并已从动物实验、临床试验等多方面证实其主要来源为烟酰胺腺嘌呤二核苷酸磷酸（NADPH）氧化酶,推测NADPH 氧化酶可能在心房颤动发生发展过程中起重要作用。以NADPH 氧化酶为靶点的心房颤动干预措施如肾素-血管紧张素系统抑制剂和他汀类等也愈受关注。     

烟酰胺腺嘌呤二核苷酸磷酸氧化酶2影响心房颤动发生的机制研究进展

心房颤动（atrial fibrillation,AF）简称房颤,是一种常见的心律失常疾病,其发生机制被认为与氧化应激密切相关。烟酰胺腺嘌呤二核苷酸磷酸（NADPH）被认为是体内心血管系统中活性氧（reactive oxygen species,ROS）及氧化应激的重要来源之一。研究表明,NADPH亚型中的NADPH氧化酶2(NOX2)是产生活性氧的主要来源,NOX2是一种跨膜蛋白,可以将电子从NADPH转移到氧分子中产生活性氧,而活性氧的产生将造成心房的电重构与结构重构,从而增加房颤的易感性。本文将对NOX2在房颤发生中所存在的机制及研究进展做一综述。     

软脂酸通过上调还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶表达促进血管内皮细胞凋亡

目的探讨软脂酸(PA)诱导的血管内皮细胞凋亡中还原型烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶的作用。方法人脐静脉内皮细胞(HUVEC)贴壁培养后分为对照组;PA(200、400、800μmol/L)浓度组(分别为PA200组、PA400组、PA800组);NADPH氧化酶抑制剂apocynin(50、100、200 mol/L)干预组(分别为apo50+PA400组、apo100+PA400组、apo200+PA400组)。采用流式细胞仪检测细胞凋亡率;提取细胞蛋白,采用Westernblot技术检测NADPH氧化酶亚基p47phox蛋白表达的水平。倒置共聚焦荧光显微镜检测活性氧的产生。结果 PA呈浓度依赖性诱导HUVEC凋亡;与对照组比较,PA400、PA800组HUVEC凋亡率明显升高,P47phox蛋白表达明显增强(P<0.05),PA400组活性氧产生明显升高(P<0.05);与PA400组比较,apo100+PA400组、apo200+PA400组HUVEC凋亡率明显降低,p47phox蛋白表达明显减弱,活性氧产生明显降低(P<0.05)。结论 PA呈浓度依赖性诱导HUVEC凋亡,apocynin能部分抑制PA的上述作用,其机制与下调NADPH氧化酶亚基p47phox蛋白表达及活性氧产生有关。     

慢性间歇低氧大鼠的血压变化及其颈动脉体中还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶的表达

目的 探讨慢性间歇低氧(chronic intermittent hypoxia,CIH)大鼠的血压变化及其颈动脉体中还原型烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶的表达情况,以明确CIH致血压升高的可能机制.方法 清洁级雄性SD大鼠30只按随机数字表法分为CIH组、慢性持续缺氧组及对照组.尾袖法测量大鼠尾动脉收缩压,采用RT-PCR法检测大鼠颈动脉体中NADPH氧化酶各亚基gp91 phox、p22phox及p47phox mRNA的表达,对颈动脉体行免疫组织化学染色,并对p22phox的表达情况行半定量分析.结果 CIH组大鼠的尾动脉收缩压为[(145±11)mm Hg,1 mm Hg=0.133kPa],较CIH组(129±9)mm Hg及对照组(124±7)mm Hg显著升高(F值为19.895,P＜0.01),CIH组gp91phox、p22phox及p47phox mRNA的表达(分别为2.82±0.51、2.74±0.45和2.88±0.47)较慢性持续缺氧组(分别为2.35±0.42、2.25±0.38和2.41±0.43)及对照组(分别为2.23±0.35、2.16±0.30和2.30±0.36)显著升高(F值分别为5.794、6.854和7.163,P＜0.01)免疫组织化学检查结果显示CIH组p22phox蛋白相对表达量(99±12)较其他两组(分别为38±7和34±8)增多.结论 CIH可刺激大鼠颈动脉体中NADPH氧化酶的表达上调并使血压升高,NADPH氧化酶在颈动脉体中的过度表达可能与OSAHS患者发生高血压病存在相关联系.

Abstract：

Objective Chronic intermittent hypoxia (CIH) occurs in patients with obstructive sleep apnea-hyponea syndrome (OSAHS) and has adverse effects on multiple physiological functions. Previous studies have shown that reflexes arising from carotid bodies mediate CIH evoked circulation-respiratory responses, and reactive oxygen species (ROS) play important roles in eliciting systemic responses to CIH.But very little is known about the molecular mechanisms underlying CIH. NADPH oxidase is the most important sources of ROS. In the present study we examined changes of blood pressure and expression of NADPH oxidase in carotid body in rats exposed to intermittent hypoxia. Methods Thirty healthy male SD rats were randomly divided into 3 groups, a CIH group, a chronic continuous hypoxia group and a control group. The systolic blood pressure (SBP) was measured with tail-cuff method. RT-PCR was used to examine mRNA expressions of NADPH oxidase subunit gp91phox, p22phox, p47phox. Immunohistochemistry and semiquantitative analysis of NADPH oxidase subunits p22phox were done in the carotid body sections of all rats. Results Compared with normal group [ ( 124 ± 7 ) mm Hg, 1 mm Hg = 0. 133 kPa ] and chronic continuous hvpoxia group[ (129 ± 9) mm Hg], the SBP in CIH group [( 145 ± 11 ) mm Hg] was significantly higher( F = 19. 895, P ＜0. 01 ＝, and the expression of NADPH oxidase subunits gp91phox,p22phox,p47phox mRNA in CIH group ( 2. 82 ± 0. 51, 2. 74 ± 0. 45, 2. 88 ± 0. 47, respectively ) were significantly higher than those in chronic continuous hypoxia group ( 2. 35 ± 0. 42, 2. 25 ± 0. 38, 2. 41 ±0. 43, respectively)and normal group(2. 23 ±0. 35, 2. 16 ±0. 30, 2. 30 ±0. 36, respectively) ( F =5.794,6. 854, 7. 163, respectively, P ＜ 0. 01 ). The Immunohistochemistry and semiquantitative analysis showed that the expression of NADPH oxidase subunit p22phox in the carotid body in CIH group ( 99 ± 12 ) were more than those in chronic continuous hypoxia and control groups ( 38 ± 7 and 34 ± 8, P ＜ 0. 05 ).Conclusion CIH upregulates the expression of NADPH oxidase in rat carotid body and elevates the rat SBP. These results indicate that NADPH oxidase up-expression is closely associated with OSAHS patients with hypertension.     

阻断肾素-血管紧张素系统对糖尿病大鼠内脏脂肪组织NADPH氧化酶和iNOS表达的影响

阻断糖尿病大鼠肾素血管紧张素系统，可通过下调内脏脂肪组织还原型辅酶Ⅱ氧化酶和诱导型一氧化氮合酶的表达，减少内脏脂肪的氧化应激，改善糖脂代谢和胰岛素抵抗。     

还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶在脂多糖诱导的小鼠急性心肌损伤组织中的表达及意义

目的探讨还原型烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶的非吞噬细胞氧化酶(Nox)家族在脂多糖(LPS)诱导的小鼠急性心肌损伤组织中的表达及意义。方法选取8~10周龄无特定病原体C57/BL6雄性小鼠30只,随机分为对照组和LPS组,每组15只。LPS组小鼠通过腹腔注射大剂量LPS(10 mg/kg)诱导急性心肌损伤模型,对照组腹腔注射等量生理盐水,观察6 h后取材,逆转录-聚合酶链反应(RT-PCR)检测Nox、B细胞淋巴瘤/白血病蛋白-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、半胱氨酸天门冬氨酸特异性蛋白酶3(Caspase 3)基因表达,Western blotting检测Nox 2、Nox 4、Bax、Bcl-2和Caspase 3蛋白表达,免疫组织化学法检测4羟基壬烯醛(4-HNE)表达,末端脱氧核糖核酸转移酶(Td T)介导的脱氧尿苷三磷酸(dUTP)切口末端标记技术(TUNEL)法检测心肌组织细胞凋亡,比较两组结果差异。使用SPSS 13.0统计软件对数据进行分析,两组比较采用t检验。结果相比对照组,LPS组小鼠心肌Nox 2、Nox 4、Bax基因和蛋白以及Caspase 3蛋白表达水平明显升高,Bcl-2基因和蛋白表达水平明显降低,差异具有统计学意义(P0.05)。免疫组织化学检测结果表明相比对照组,LPS组心肌组织4-HNE表达明显增加,差异具有统计学意义(P0.05)。TUNEL检测结果表明相比对照组,LPS组心肌组织心肌凋亡细胞明显增加,差异具有统计学意义(P0.05)。结论 NADPH氧化酶通过调控氧化应激和细胞凋亡参与急性心肌损伤的发生发展。     

非洛地平对载脂蛋白E基因敲除小鼠动脉粥样硬化斑块及烟酰胺腺嘌呤二核苷酸氧化酶的影响

目的:探讨钙通道阻断剂(CCB)非洛地平对载脂蛋白E基因敲除(apoEKO)小鼠动脉粥样硬化斑块及烟酰胺腺嘌呤二核苷酸[NAD(P)H]氧化酶的影响。方法:apoEKO小鼠随机分为普通饮食组、高胆固醇饮食组、高胆固醇饮食加非洛地平组。无创血压系统测小鼠血压;内眦动脉取血检测血清TC和TG水平;冷冻切片光镜下定位主动脉根部,油红O染色评估斑块大小;实时定量PCR和Western blot方法检测主动脉中NAD(P)H氧化酶亚基p47phox和Rac-1表达。结果:高胆固醇饮食组小鼠血压没有明显变化,血脂明显升高(P<0.01),且斑块面积明显高于普食组(P<0.01);非洛地平可以明显减小斑块面积(P<0.01),同时还可以降低NAD(P)H氧化酶亚基p47phox和Rac-1表达(P<0.01)。结论:非洛地平可能通过阻断氧化应激反应抑制动脉粥样硬化发生发展。     

胰高血糖素样肽1对1型糖尿病大鼠心肌烟酰胺腺嘌呤二核苷酸氧化酶亚单位表达的影响

目的 探讨胰高血糖素样肽1 （GLP-1）对1型糖尿病大鼠心肌组织烟酰胺腺嘌呤二核苷酸（NADPH）氧化酶亚单位p22phox和Nox4表达的影响.方法 将42只雄性SD大鼠用随机数字表法分为正常对照组（NC组,n=7）和糖尿病造模组（n=35）.采用链脲佐菌素（STZ）对糖尿病造模组制备1型糖尿病大鼠模型.将造模成功的29只1型糖尿病大鼠用随机数字表法分为糖尿病组（DM组,n=10）,糖尿病GLP-1低剂量治疗组（DL组,n=10）和糖尿病GLP-1高剂量治疗组（DH组,n=9）.DL组予以艾塞那肽1μg/kg,2次/d皮下注射,DH组予以艾塞那肽5μg/kg,2次/d皮下注射.艾塞那肽治疗8周后处死动物.用实时荧光定量聚合酶链反应测定4组大鼠心肌p22phox和Nox4 mRNA的表达,免疫组化法检测心肌铜-锌-超氧化物歧化酶（Cu-Zn-SOD）蛋白表达.多组比较采用单因素方差分析.结果 与NC组比较,DM组大鼠心肌p22phox和Nox4 mRNA表达显著升高（t=5.77、5.36,均P＜0.05）,心肌Cu-Zn-SOD蛋白表达显著升高（t=59.91,P＜0.05）.艾塞那肽治疗8周后,与DM组比较,DL组和DH组大鼠心肌p22phox和Nox4 mRNA表达均显著降低（t=16.86、7.66和16.11、7.59,均P＜0.05）,心肌Cu-Zn-SOD蛋白表达均显著降低（t=56.00、47.05,均P＜0.05）.与DL组比较,DH组大鼠心肌p22phox和Nox4 mRNA表达显著降低（t=10.14、8.67,均P ＜0.05）,而心肌Cu-Zn-SOD表达无明显变化（=81.91,P＞0.05）.结论 GLP-1可剂量依赖性地下调1型糖尿病大鼠心肌p22phox和Nox4的表达,非剂量依赖性地上调心肌Cu-Zn-SOD表达,减轻氧化应激对心肌的损害,对糖尿病大鼠心肌产生保护作用.     

晚期糖基化终产物对大鼠血管外膜成纤维细胞中烟酰胺腺嘌呤二核苷酸磷酸氧化酶p22phox亚基及活性氧表达的影响

目的:探讨晚期糖基化终产物(AGE)对大鼠血管外膜成纤维细胞(AF)中烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶p22phox亚基及活性氧表达的影响。方法:用组织贴块法培养SD大鼠的血管外膜成纤维细胞,用逆转录—聚合酶链反应、蛋白质印迹检测不同浓度的糖基化人血清白蛋白(AGE-HSA,分为对照组、100μg/ml AGE-HSA组,200μg/ml AGE-HSA组、300μg/ml AGE-HSA组)对NADPH氧化酶p22phox亚基信使核糖核酸(mRNA)及蛋白的表达的影响,并观察不同干预因素[分为对照组1、200μg/ml AGE-HSA组(200μg/ml处理组1)、200μg/ml AGE-HSA+50μg/ml抗RAGE中和抗体组(抗RAGE中和抗体组1)及200μg/ml AGE-HSA+30 nmol/L坎地沙坦组(坎地沙坦组1)对AGE-HSA上调NADPH氧化酶p22phox亚基mRNA和蛋白表达的影响。用2’,7’-二氯荧光黄双乙酸盐检测不同干预因素[空白对照组、AGE-HSA 200μg/ml组(200μg/ml处理组2)、200μg/ml AGE-HSA+50μg/ml抗RAGE中和抗体组(抗RAGE中和抗体组2)、200μg/ml AGE-HSA+30μmol/L夹竹桃素组(夹竹桃组),200μg/ml AGE-HSA+30 nmol/L坎地沙坦组(坎地沙坦组2)对血管外膜成纤维细胞内活性氧表达的影响。结果:3个浓度(100μg/ml、200μg/ml和300μg/ml)的AGE-HSA组均与对照组比较,血管外膜成纤维细胞NADPH氧化酶p22phox亚基mRNA及蛋白的表达随AGE-HSA增加呈浓度依赖性上调;抗RAGE中和抗体组1、坎地沙坦组1比200μg/ml处理组1的p22phox mRNA及蛋白表达降低;抗RAGE中和抗体组2、夹竹桃素组2及坎地沙坦组2比200μg/mlAGE-HSA处理组2血管外膜成纤维细胞内活性氧相对荧光强度降低,上述比较差异均有统计学意义(P<0.05)。结论:AGE-HSA经由RAGE影响活性氧与NADPH氧化酶p22phox亚基的表达上调,活性氧的上调与NADPH氧化酶p22phox亚基表达相关。NADPH氧化酶抑制剂及坎地沙坦可通过下调NADPH氧化酶p22phox亚基的表达减少血管外膜成纤维细胞内活性氧的产生。     

烟酰胺腺嘌呤二核苷酸磷酸氧化酶在大鼠心肌梗死后心室肌中的改变及夹竹桃麻素的干预效果

目的 探讨烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶在大鼠心肌梗死后左心室心肌中的改变及夹竹桃麻素对NADPH氧化酶的干预效果.方法 通过结扎雄性SD大鼠左冠状动脉前降支制作大鼠心肌梗死模型,假手术组缝合线只穿过前降支而不结扎.将两组分别随机分为药物干预组和安慰剂组两个亚组[心肌梗死安慰剂亚组(n=7),其余3组(n...     

Albuminuria, expression of nicotinamide adenine dinucleotide phosphate oxidase and monocyte chemoattractant protein-1 in the renal tubules of hypertensive Dahl salt-sensitive rats.

In chronic renal diseases, experimental and human data suggest that excess albumin filtered through the glomerular capillary barrier is over-reabsorbed by proximal tubular cells, thereby activating these cells and upregulating the expression of chemokines. On the other hand, a high-salt diet has been shown to induce proteinuria in hypertensive Dahl salt-sensitive (DSS) rats, accompanied with the expression of reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in the kidney. In the current study, we therefore examined albuminuria and the expressions of NADPH oxidase and monocyte chemoattractant protein-1 (MCP-1) in the renal tubular cells in hypertensive DSS rats, as well as the effects of the antioxidant N-acetylcysteine (NAC) on each of these parameters. DSS rats were fed a normal-salt diet (0.24% NaCl), a high-salt diet (8% NaCl), or a high-salt diet plus NAC supplementation (15 mg/mL drinking water) for 4 weeks. The high-salt diet provoked an increase in glomerular injuries accompanied with albuminuria and in urinary H2O2 and MCP-1 excretion. Immunohistochemical analysis showed the prominent expression of MCP-1 in the dilated tubular cells, where the NADPH oxidase subunit p47phox was also expressed. The current results suggest that albuminuria caused expression of NADPH oxidase and MCP-1 in the dilated renal tubules, resulting in interstitial inflammation and migration of mononuclear cells in DSS rats, because blockade of albuminuria by NAC counteracted the p47phox and MCP-1 expression.     

Transient association of the nicotinamide adenine dinucleotide phosphate oxidase subunits p47phox and p67phox with phagosomes in neutrophils from patients with X-linked chronic granulomatous disease

Optimal microbicidal activity of polymorphonuclear leukocytes (PMNs) requires recruitment of a functional nicotinamide adenine dinucleotide phosphate (NADPH) oxidase to the phagosome. In this study, we used a synchronized phagocytosis assay and immunofluorescence microscopy (IFM) to examine the association of cytosolic NADPH oxidase subunits with phagosomes containing opsonized zymosan (OpZ). Ingestion of OpZ began within 30 seconds of particle binding and forming phagosomes were enriched for both F-actin and the actin-binding protein p57. NADPH oxidase subunits p47phox and p67phox were also recruited to forming phagosomes and were retained on mature phagosomes for at least 15 minutes. Colocalization of F-actin, p57, and p47phox on phagosomes was confirmed by immunoblotting. Translocation of p67phox, but not p57, to forming phagosomes was deficient in PMNs lacking p47phox. Surprisingly, we found that in PMNs from six individuals with X-linked chronic granulomatous disease (CGD), p47phox and p67phox accumulated in the periphagosomal area during ingestion of OpZ. However, in marked contrast to normal PMNs, p47phox and p67phox were shed from nascent phagosomes along with F-actin and p57 once OpZ was internalized (approximately 5 minutes). These data support a model in which flavocytochrome b is required for stable membrane binding of p47phox and p67phox, but not their association with the cytoskeleton or transport to the cell periphery.     

Increased adiponectin synthesis in the visceral adipose tissue in men with coronary artery disease treated with pravastatin: a role of the attenuation of oxidative stress

Pravastatin is reported to increase the adiponectin level in humans, but the mechanism remains unclear. We examined plasma and gene expressions of adiponectin, tumor necrosis factor (TNF)-α, interleukin (IL)-6 and protein carbonyl level, an indicator of oxidative stress, in visceral and subcutaneous adipose tissue from 32 patients with coronary artery disease undergoing coronary artery bypass grafting (CABG). Fourteen patients with serum LDL-cholesterol level >100 mg/dl were treated with pravastatin at 10 mg/day for 2 months before CABG (Statin), and the other 18 with LDL-cholesterol ≤100 mg/dl were not (Control). The plasma adiponectin level was higher in the Statin than the Control group (P < 0.05), but TNF-α and IL-6 levels were not different. Adiponectin gene expression in visceral tissue was 3-fold higher in the Statin than the Control group (P < 0.01), but was not different in subcutaneous tissue. TNF-α and IL-6 gene expressions in each tissue were not different between the 2 groups. Protein carbonyl levels in plasma and visceral tissue were lower in the Statin than the Control group (both, P < 0.05). Thus, adiponectin expression and generation in visceral adipose tissue is increased in men with coronary artery disease treated with pravastatin. Pravastatin-initiated attenuation of oxidative stress could be involved.     

大黄酸上调糖尿病大鼠脂肪组织PPAR-γ及GluT-4表达并改善胰岛素敏感性

目的探讨大黄酸改善糖尿病大鼠脂肪组织胰岛素敏感性及降血糖的作用机制。方法雄性Wistar大鼠随机分为对照组（NC，n=15）及糖尿病造模组（DM，n=40），DM组成模后随机分为糖尿病模型组（DM—C，n=15）和糖尿病大黄酸治疗组（DM—T，n=15）。DM—T组予大黄酸100mg·kg^-1·d^-1灌胃11周。实验末检测各组大鼠FBG、FIns、TG、TC、胰岛素敏感指数（ISI）、过氧化物酶体增殖物激活受体7（PPAR-γ）及葡萄糖转运蛋白4（GluT-4）在脂肪组织的表达水平。结果17周末DM—C组较NC组FBG、TG明显升高，ISI明显降低，脂肪组织PPAR-γ及GluT-4蛋白表达明显降低；DM—T组较DM—C组FBG明显降低；ISI明显升高；脂肪组织PPAR-γ蛋白表达明显升高，积分光密度有统计学差异；GluT-4蛋白表达明显升高，积分密度（ID）有统计学差异。结论大黄酸可上调糖尿病大鼠脂肪组织PPAR-γ及GluT-4蛋白表达，降血糖并改善胰岛素敏感性。     

Serum prolactin and luteinizing hormone levels and the activities of hypothalamic monoamine oxidase A and B and phenylethanolamine-N-methyl transferase are changed during sexual maturation in male rats treated neonatally with melatonin

Male rat pups were given a single dose of melatonin on day 5 of age. On days 30, 45, and 60, prolactin (PRL), luteinizing hormone (LH), follicle-stimulating hormone (FSH), and testosterone (T) were measured in serum and monoamine oxidase A (MAO A) activity, monoamine oxidase B (MAO B), and phenylethanolamine-N-methyl transferase (PNMT) activity were measured in the hypothalamus. Melatonin administration increased serum PRL levels at all ages studied. Serum LH levels were decreased in the melatonin treated group on day 30, but levels were elevated on days 45 and 60 of age as compared to controls. LH response to luteinizing hormone-releasing hormone (LHRH) only increased in melatonin treated animals at 30 days of age. Serum T levels decreased with melatonin treatment at 30 days of age, but increased on days 45 and 60 of age. T response to human chorionic gonadotropin (hCG) was blunted by melatonin treatment at 30 days of life. Hypothalamic MAO A activity increased, after neonatal melatonin administration, at 30 and 45 days of age and decreased at 60 days of life. Early neonatal melatonin administration increased MAO B and PNMT activities on day 45. These findings suggest that neonatal melatonin administration induces an earlier sexual maturation in male rats, possibly related to PRL, LH, MAO, and PNMT increases.     

Increased expression of NAD(P)H oxidase subunits,NOX4 and p22phox,in the kidney of streptozotocin-induced diabetic rats and its reversibity by interventive insulin treatment

Aim/hypothesis An increased production of reactive oxygen species (ROS) could contribute to the development of diabetic nephropathy. NAD(P)H oxidase might be an important source of ROS production in kidney as reported in blood vessels. In this study, we show the increased expression of essential subunits of NAD(P)H oxidase, NOX4 and p22phox, in the kidney of diabetic rats.Methods The levels of mRNA of both NOX4 and p22phox were evaluated in kidney from streptozotocin-induced diabetic rats and age-matched control rats at 4 and 8 weeks after onset of diabetes by Northern blot analysis. The localization and expression levels of these components and 8-hydroxy-deoxyguanosine (8-OHdG), which is a marker of ROS-induced DNA damage, were also evaluated by immunostaining.Results The levels of both NOX4 and p22phox mRNA were increased in the kidney of diabetic rats as compared with control rats. Immunostaining analysis showed that the expression levels of NOX4 and p22phox were clearly increased in both distal tubular cells and glomeruli from diabetic rats. Both the localization and the expression levels of these components were in parallel with those of 8-OHdG. Interventive insulin treatment for 2 weeks completely restored the increased levels of these components in the diabetic kidney to control levels in parallel with those of 8-OHdG.Conclusions/interpretation This study provides evidence that NAD(P)H oxidase subunits, NOX4 and p22phox, were increased in the kidney of diabetic rats. Thus, NAD(P)H-dependent overproduction of ROS could cause renal tissue damage in diabetes. This might contribute to the development of diabetic nephropathy.Abbreviations ROS reactive oxygen species - NAD(P)H oxidase nicotinamide adenine dinucleotide phosphate oxidase - 8-OHdG 8-hydroxy-deoxyguanosine - STZ streptozotocin - mtDNA mitochondrial DNA - PKC protein kinase C