Methotrexate Causes Oxidative Stress in Rat Kidney Tissues

Aim. Methotrexate (MTX), a folic acid antagonist, is one of the chemotherapeutic agents widely used in the treatment of some types of cancers. Nephrotoxicity is one of the complications of MTX treatment. The aim of this study was to investigate possible effects of MTX treatment on the oxidant/antioxidant status in rat kidney tissues and enzymatic mechanisms leading to nephrotoxicity. Methods. For this aim, 10 Sprague-Dawley type female rats of 4 weeks old were used in the study. The animals were divided into two groups randomly. Five of them were used as control, and the others were treated with MTX intravenously (60 mg/m² of body surface area per week) for 7 weeks. At the end of this period, they were sacrificed, and kidney tissues were removed to be used in the analyses of malondialdehyde (MDA) levels, antioxidant potential (AOP) values, and superoxide dismutase, catalase, glutathione peroxidase, xanthine oxidase, adenosine deaminase, and 5′ nucleotidase enzyme activities. Results. There was significant increase in the MDA level in the MTX group compared with the control group (1.74 ± 0.23 nmol/mg vs. 1.04 ± 0.30 nmol/mg; p < 0.05, respectively). There were however no meaningful differences between enzyme activities and AOP values of the groups. Conclusion. It has been suggested that MTX leads to oxidative stress in rat kidney tissues, which might be one of the reasons for MTX-induced nephrotoxicity.     

Effects of Pomegranate Juice on Hyperoxaluria-Induced Oxidative Stress in the Rat Kidneys

To evaluate the role of the inducible nitric oxide synthase (iNOS), selective nuclear factor-kB (NF-kB), and p38-mitogene-activated protein kinase (p38-MAPK) on hyperoxaluria-induced oxidative stress and stone formation in rat kidneys. The rats were divided into five groups: group 1, control group; group 2: ethylene glycol (EG) group; group 3: EG?+?pomegranate juice (PJ)-low group; group 4: EG?+?PJ-middle group; group 5: EG?+?PJ-high group. Rats were sacrificed on 7, 15, and 45 days. The iNOS expression, p65-NF-kB and p38-MAPK activity, and oxidative stress markers were evaluated in the kidney. Crystal depositions were evident on day 7, and mild and severe crystallization were observed on day 15 and 45 in EG group, respectively. There was limited or no crystal formation in rats in both middle- and high-dose PJ groups when compared to low-dose PJ group. Crystal depositions, iNOS, p38-MAPK and p65-NF-kB activity, and oxidative stress markers were found to be decreased by middle- and high-dose PJ treatment. PJ was found to have inhibitory effects on renal tubular cell injury and oxidative stress caused by oxalate crystals by reducing ROS, iNOS, p38-MAPK, and NF-kB expression.     

Effect of Intravenous Iron Sucrose on Oxidative Stress in Peritoneal Dialysis Patients

Aim. Intravenous iron therapy is an accepted treatment for patients receiving hemodialysis and continuous ambulatory peritoneal dialysis (CAPD). Studies have found enhanced oxidative stress in hemodialysis patients receiving intravenous iron, but there are no clinical data for CAPD patients. The aim of the current study was to investigate the effect of 100 mg of intravenous iron-sucrose on the erythrocyte (RBC) antioxidant enzymes (namely, superoxide dismutase [SOD], catalase [CAT], and glutathione peroxidase [GSHPx]) and plasma malondialdehyde (MDA), an oxidant molecule, in CAPD patients. Methods. Twelve CAPD patients receiving maintenance intravenous iron-sucrose were recruited. After a 12-hour fast, blood samples were taken for hemoglobin, iron, ferritin, and high-sensitivity C-reactive protein (hsCRP), and for baseline activities of erythrocyte antioxidant enzymes (i.e., SOD, CAT, GSHPx) and the plasma oxidant molecule, MDA. 100 mg iron-sucrose was infused over 30 minutes. Blood samples taken during (i.e., 15 minutes after commencement of infusion) and after (i.e., at 30 minutes, 60 minutes, and 6 hours after commencement) the infusion were taken for measurement of plasma iron, ferritin, TSAT, RBC SOD, CAT, GSHPx, and plasma MDA. Results. Plasma iron and transferrin saturation elevated significantly during infusion (p < 0.05). There was no significant change in erythrocyte SOD, CAT, GSHPx, or in MDA activities. There was a reduction of GSHPx activity at the 30th minute (from 153.69 ± 66.69 to 123.68 ± 25.50 mU/mL), but it was not statistically significant. The patients were grouped according to baseline ferritin (100–400 and 400–800 ng/mL); 60th-minute MDA was significantly higher in the latter group (p < 0.05). There was no correlation between hsCRP and oxidant-antioxidant balance. No correlation was noted between RBC antioxidant enzymes or plasma oxidant molecule and ferritin levels. Conclusion. There are no acute deteriorating effects from a 100 mg of intravenous iron-sucrose in CAPD patients with optimal iron stores. This dose may be applied safely in CAPD patients.     

Brain Death Is Associated With Endoplasmic Reticulum Stress and Apoptosis in Rat Liver

Cell death pathways initiated by stress on the endoplasmic reticulum (ER) have been implicated in a variety of common diseases, such as ischemia/reperfusion injury, diabetes, heart disease, and neurodegenerative disorders. However, the contribution of ER stress to apoptosis and liver injury after brain death is not known. In the present study, we found that brain death induces a variety of signature ER stress markers, including ER stress–specific X box–binding protein 1 and up-regulation of glucose-regulated protein 78. Furthermore, brain death causes up-regulation of C/EBP homologous protein and caspase-12. Consistent with this, terminal deoxynucleotidyl transferase–mediated 2′-deoxyuridine 5′-triphosphate nick-end labeling assay and transmission electron microscopy confirmed apoptosis in the liver after brain death. Taken together, the present study provides strong evidence supporting the presence and importance of ER stress and response in mediating brain death–induced apoptosis and liver injury.     

Oxidative Stress in 5/6 Nephrectomized Rat Model: Effect of Alpha-Lipoic Acid

Alpha-lipoic acid (ALA) is a powerful antioxidant, and its effect in ameliorating complications of diabetes mellitus has been widely documented. The aim of this study was to investigate the role of ALA in the disease progression of remnant kidneys (RK). Systolic blood pressure (SBp), hemoglobin, renal function, kidney malondialdehyde (MDA), glutathione (GSH), vitamin E (Vit E) concentrations, p65 nuclear factor (NF)-κB activity, and macrophage infiltration were analyzed in sham and RK rats supplemented with ALA (100 mg/kg body weight, i.p., every other day) or vehicle for 12 weeks. RK rats exhibited increases in SBp, kidney MDA concentration, p65 NF-κB activity, and macrophage infiltration, which were prominent in weeks 4 and 8 and decreased at week 12. RK rats also showed anemia, microalbuminuria, and decreased renal function and kidney GSH and Vit E concentrations. These changes were all attenuated by 8 weeks of ALA treatment compared to RK vehicle group. But the continued ALA treatment after week 8 reversed the beneficial effect on SBp, kidney MDA concentration, p65 NF-κB activity, macrophage infiltration, anemia, microalbuminuria, and renal function, while the beneficial effect was maintained if the treatment was discontinued after week 8. Furthermore, ALA increased albuminuria and kidney MDA concentration in sham animals. In conclusion, ALA administration attenuates oxidative stress, inflammation, and hypertension and delayed the deterioration of kidney function in RK rats with enhanced oxidative stress, while in healthy animals or RK rats with a well-balanced redox state, ALA may act as a pro-oxidant, contributing to renal dysfunction.     

氧化应激反应导致大鼠自体移植静脉内膜增生的作用

目的探讨氧化应激反应在自体移植静脉术后内膜增生中的作用。方法将雌性Sprague-Dawley（SD）大鼠70只按随机分组方法分为两组：实验组（60只）,分为移植术后1d、1周、2周、4周、6周和2个月6个时间点,每个时间点10只,建立大鼠自体移植静脉模型,以上6个时间切取移植静脉;对照组（10只）,直接切取同样长度的颈外静脉。应用计算机图象分析仪计算新生内膜面积/中膜面积（neointima to tunica media area,I/M）;应用免疫组织化学方法检测核转录因子-κB（nuclear factor-kappa B,NF-κB）和铜锌超氧化物歧化酶（copper zinc superoxidedismutase,CuZnSOD）;用比色法检测大鼠血清中丙二醛（malondialdehyde,MDA）的含量。结果对照组大鼠NF-κB、CuZnSOD有少量表达。实验组大鼠移植术后NF-κB的表达随着时间的增加逐渐增加,移植术后2周达到高峰,维持1个月左右后逐渐下降,2个月时接近基础水平;CuZnSOD表达逐渐升高,移植术后1周左右达到高峰,持续2～3周逐渐下降至正常水平。对照组血清中MDA含量为4.966±1.346nmol/ml,实验组移植术后大鼠血清中MDA的含量急剧升高,移植术后1d达到高峰（21.161±2.174nmol/ml）,然后逐渐下降,2个月时接近正常水平（6.208±2.908nmol/ml）,两组比较差异有统计学意义（P〈0.05）;对照组I/M为0.2096±0.0253,实验组移植术后1周I/M为0.6806±0.0737,4周达高峰（1.4527±0.0824）,6周为1.0353±0.0656,2个月时为0.9583±0.0516,两组差异有统计学意义（P〈0.05）。结论自体静脉移植后内皮细胞损伤导致氧化应激反应,通过激活NF-κB放大炎症反应,在引发移植静脉再狭窄过程中起重要作用     

番茄红素对血液透析患者静脉铁剂诱导氧化应激的影响

目的：观察维持性血液透析（MHD）患者由静脉铁剂诱导的氧化应激状态，探讨番茄红素对其的干预作用。方法：30例MHD患者，随机分为对照组和试验组，各15例。对照组在患者透析时给予蔗糖铁注射液100mg，2次／周，共10次，观察时间8周；试验组除蔗糖铁注射液使用外，同时口服番茄红素胶囊，2粒／次，2次／d。用药8周，观察时间8周，观察并比较两组患者治疗前及治疗8周后的血红蛋白（m）、红细胞比容（Hct）、血清铁（SI）、铁蛋白（SF）、转铁蛋白饱和度（TSAT）以及超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH—px）、丙二醛（MDA）等指标的变化。结果：两组患者治疗后SOD和GSH—px水平较治疗前均有显著下降（P〈0．01）；但是试验组下降幅度显著性小于对照组（P〈0．01）。两组患者治疗后MDA水平较治疗前有显著性升高（P〈0．01）；但是试验组升高幅度显著性小于对照组（P〈0.01）。治疗后两组血清MDA水平与SF呈正相关（P〈0.01）。治疗后两组Hb、Hct、SI、SF、TSAT较治疗前均有显著性升高（P〈0．01），并且两组升高幅度相似，无统计学差异。结论：静脉铁剂治疗加剧了MHD患者的氧化应激状态。番茄红素可减轻这种氧化应激状态。     

缬沙坦对单侧输尿管梗阻大鼠肾脏氧化应激的影响

目的：探讨血管紧张素Ⅱ-1型受体拮抗剂（AT1Ra）缬沙坦对单侧输尿管梗阻（UUO）大鼠肾脏氧化应激的影响。方法：Wistar大鼠行左侧输尿管结扎术,分为UUO模型组（n=11）,缬沙坦治疗组（n=11）,同时设假手术对照组（n=7）。术后第14天处死各组大鼠,进行HE和Masson染色,观察肾脏病理变化;比色法测定肾组织丙二醛（MDA）和超氧化物歧化酶（SOD）含量;免疫组织化学方法测定α-平滑肌肌动蛋白（α-SMA）和转化生长因子-β1（TGF-β1）的表达。结果：UUO组与假手术组大鼠比较,肾脏病理改变加重,肾组织MDA含量明显升高（P〈0.01）,SOD含量下降（P〈0.05）,肾组织α-SMA和TGF-β1的表达显著增加（P〈0.01）。缬沙坦治疗组与UUO组大鼠比较,肾间质纤维化减轻（P〈0.05）,肾组织MDA含量降低（P〈0.05）,SOD含量升高（P〈0.05）;同时肾组织α-SMA和TGF-β1的表达降低（P〈0.05）。结论：缬沙坦可通过减少UUO组肾组织脂质过氧化物的产生,增加抗氧化酶的含量,下调TGF-β1的表达,从而显著改善UUO所致的肾间质纤维化。     

Early Events in Kidney Donation: Progression of Endothelial Activation, Oxidative Stress and Tubular Injury After Brain Death

Cerebral injury leading to brain death (BD) causes major physiologic derangements in potential organ donors, which may result in vascular-endothelial activation and affect posttransplant graft function. We investigated the kinetic of pro-coagulatory and pro-inflammatory endothelial activation and the subsequent oxidative stress and renal tubular injury, early after BD declaration. BD was induced by slowly inflating a balloon-catheter inserted in the extradural space over a period of 30 min. Rats (n = 30) were sacrificed 0.5, 1, 2 or 4 h after BD-induction and compared with sham-controls. This study demonstrates immediate pro-coagulatory and pro-inflammatory activation of vascular endothelium after BD in kidney donor rats, proportional with the duration of BD. E- and P-Selectins, Aα/Bβ-fibrinogen mRNA were abruptly and progressively up-regulated from 0.5 h BD onwards; P-Selectin membrane protein expression was increased; fibrinogen was primarily visualized in the peritubular capillaries. Plasma von Willebrand factor was significantly higher after 2 h and 4 h BD. Urine heart-fatty-acid-binding-protein and N-acetyl-glucosaminidase, used as new specific and sensitive markers of proximal and distal tubular damage, were found significantly increased after 0.5 h, with a maximum at 4 h. Unexpectedly, oxidative stress was detectable only late, after the installation of tubular injury, suggesting only a secondary role for hypoxia in triggering these injuries.     

精索静脉曲张与氧化应激的研究

目的 :研究精索静脉曲张 (VC)时氧化应激的损害机制。　方法 :2 8例VC不育男性 ,行精索内静脉结扎术 ,采精索内静脉血和外周静脉血 ;建立大鼠左侧VC模型 (n =12 ) ,以假手术大鼠为对照组 (n =8) ,术后 3个月取睾丸组织。采用分光光度法检测VC男性血浆及大鼠睾丸组织中一氧化氮 (NO)、一氧化氮合酶 (NOS)、黄嘌呤氧化酶(XO)、乳酸 (Lac)和乳酸脱氢酶 (LDH)含量。　结果 :VC患者精索内静脉血浆中NO、NOS、XO、Lac含量明显高于外周静脉血浆 (P <0 .0 1,P <0 .0 5 ) ,LDH明显降低 (P <0 .0 1)。实验性VC大鼠左侧睾丸组织NO、XO高于对照组 (P <0 .0 1) ,Lac明显降低 (P <0 .0 1)。　结论 :VC时 ,可由于曲张精索静脉血浆中NO、NOS和XO及睾丸组织中NO和XO产生增加 ,以及Lac和LDH含量的变化 ,而导致精子生成障碍或 /和精子活力下降 ,引起男性不育。     

糖尿病氧化应激与勃起功能障碍的关系

ED是糖尿病进程中发病率较高的并发症。糖尿病可引发氧化应激,氧化应激通过对血管内皮、周围神经、阴茎平滑肌功能的影响和引发细胞凋亡等机制在糖尿病性ED的发病进程中起重要作用。现就氧化应激与糖尿病性ED关系的研究进展作一综述。     

缬草油对2型糖尿病大鼠肾脏氧化应激和蛋白激酶C活性的影响

目的：现察缬草油对2型糖尿病大鼠肾脏乳化应激(OS)和蛋白激酶C（PKC)活性的影响。方法：将实验动物分为正方对照组、糖尿病组及缬草油治疗组。检测各组第4、5、11、17周的血糖、血胰岛素、血脂(TG、TC)，11、17用的血肌酐(Scr)、尿素氮(BUN)、尿微白蛋白排泄率(UAE)、肾组织中丙二醛(NDA)含量、铜，锌—超氧化物歧化酶（Cu，Zn—SOD)、过氧化氨酶(CALT)、谷光甘肽过氧化酶(GSH—Px)活性及PTC活性，同时留取肾脏作HE染色行病理检查。结果：与糖尿病组相比，缬草油治疗组血TG、TC、Scr、BUN、UAE、肾脏MDA含量、肾细胞膜PKC均明显下降，而肾脏抗氧化酶活性(Cu，Zn—SOD、CAT、GSH—Px)则明显上升，病理检查发现缬草油治疗组较糖尿病组肾小球体积略有缄小，系膜增生明显浅轻。结论：缬草油可以明显改善2型DM大鼠肾脏内的OS，抑制PKC的激活，减少蛋白尿，延缓肾功能损害的进展。     

氧化应激与糖尿病性勃起功能障碍

勃起功能障碍(ED)是糖尿病进程中常见的并发症,其发病机制十分复杂,涉及到神经及神经递质、血管及血管活性物质、内分泌、代谢等多方面因素。糖尿病状态下,自由基增多引发氧化应激损伤,影响到糖尿病性ED发生发展中的各个环节。本文针对氧化应激在糖尿病性勃起功能障碍进程中的作用作一综述。     

氧化胁迫与精子功能损伤

人类精子对氧化胁迫特别敏感 ,损伤精子功能的氧化胁迫有两个细胞来源 :精子自身和白细胞。由于精子膜含有高浓度的不饱和脂肪酸 ,而且精子自身的抗氧化能力很弱 ,在过量活性氧的攻击下 ,易发生脂类过氧化反应 ,使精子膜的流动性和完整性受到损伤 ,进而破坏精子功能 ,并最终引起男性不育。此外 ,过量的活性氧还可造成精子核DNA的损伤 ,而父代精子DNA的损伤又与子代儿童癌症的发生有密切的关联。     

Diabetes,Oxidative Stress and Physical Exercise

Oxidative stress, an imbalance between the generation of reactive oxygen species and antioxidant defense capacity of the body, is closely associated with aging and a number of diseases including cancer, cardiovascular diseases, diabetes and diabetic complications. Several mechanisms may cause oxidative insult in diabetes, although their exact contributions are not entirely clear. Accumulating evidence points to many interrelated mechanisms that increase production of reactive oxygen and nitrogen species or decrease antioxidant protection in diabetic patients. In modern medicine, regular physical exercise is an important tool in the prevention and treatment of diseases including diabetes. Although acute exhaustive exercise increases oxidative stress, exercise training has been shown to up regulate antioxidant protection. This review aims to summarize the mechanisms of increased oxidative stress in diabetes and with respect to acute and chronic exercise.Key Words: Diabetes, physical activity, antioxidants, reactive oxygen species     

Oxidative Stress in Children with Acute Glomerulonephritis

Background. Oxidative stress has not been adequately investigated in acute glomerulonephritis (AGN); therefore, we aimed to evaluate the oxidative stress (OS) status in children with AGN both at acute and remission stages. Patients and methods. Seventeen children (mean ± SEM, age 9.0 ± 0.5 years) with AGN and 17 healthy controls were included. In addition to routine laboratory investigations, two blood samples were obtained from patients, at admission and after 6–10 weeks, to measure erythrocyte superoxide dismutase (SOD) activity and plasma malondialdehyde (MDA) level. Results. Significantly elevated MDA levels (5.11 ± 0.28 vs. 3.15 ± 0.25 nmol/mL; p < 0.001) were found in acute stage of AGN compared with the controls; however there was no significant difference in SOD activities (3732 ± 193 vs. 4035 ± 142 U/gHb; p > 0.05) between acute stage-AGN and control subjects. Significantly elevated SOD activities (3985 ± 195 U/gHb, p?=?0.034) and decreased MDA levels (4.01 ± 0.38 nmol/mL, p?=?0.001) were found at remission stage when compared with the acute stage. MDA levels and SOD activities of remission phase were similar to those of controls (p > 0.05). A significantly positive correlation was found between MDA levels and SOD activities in remission period (r?=?0.654, p?=?0.004). Patients with and without impaired renal functions had similar MDA levels and SOD activities (p > 0.05). No significant correlation was found between glomerular filtration rates (GFR) and MDA levels (p > 0.05) and between GFR and SOD (p > 0.05) activities in acute stage-AGN. Conclusions. Oxidative stress may play important role in the pathogenesis of AGN and not be correlated with renal functions. Further research is needed to determine magnitude of OS and indications for antioxidants in other glomerulopathies.     

Oxidative Stress and Lipid Peroxidation in the Ischemic Small Intestine: Pathological and Biochemical Evaluation in a Rat model of Superior Mesenteric Ischemia

The purpose of our study was to evaluate the role of oxidative stress and lipid peroxidation in acute mesenteric ischemia. Thirty male Wistar albino rats weighing 240–260 g were randomized into control (no operation), sham (operation without ischemia), and ischemia groups. To induce ischemia, the superior mesenteric artery was sutured. Total antioxidant and oxidant capacity and lipid peroxidase activity were measured in blood samples collected at 0 min, 60 min, and 240 min, and the pathology of ileum segments resected at 240 min was evaluated. Total oxidant status did not differ among the groups. Total antioxidant status increased significantly with time in the ischemia group compared to the control and sham groups (P < 0.001). Although basal arylesterase activity was lower in the ischemia group than controls (P < 0.05), post-ischemia values were similar among the groups. Similarly, basal and stimulated paraoxonase activity in blood samples did not differ among the groups. In conclusion, oxidative stress and lipid peroxidation have no significant role in the pathophysiology of acute mesenteric ischemia.Key words: Mesenteric ischemia, Oxidative stress, Lipid peroxidationOxygen is both vital and damaging, as it is involved in the pathogenesis of many types of tissue injury.^1,2 Oxidative stress, which is an imbalance between reactive oxygen species and antioxidant defense systems, can be caused by several factors such as ischemia, heavy exercise, and inflammation.^3,4 Oxidative stress has been linked to some neurodegenerative and cardiovascular diseases, preeclampsia, gene mutations, and aging.⁵⁻⁹The role of oxidative stress and oxygen-derived free radicals in the pathophysiology of ischemic diseases was postulated first in the mid-1950s.¹⁰ After the development of advanced biochemical techniques to measure oxidant and antioxidant capacities in tissues, the relationship between ischemia and oxidative stress has been studied extensively.¹⁰Biochemical indicators of oxidative stress include total oxidant and antioxidant status and the activity of antioxidant enzymes such as serum dismutase, glutathione peroxidase, and stimulated paraoxonase. Specifically, paraoxonase, an ester hydrolase antioxidant with both arylesterase and paraoxonase activities, acts as an enzymatic defense against lipid hydroperoxides.^11,12 Serum paraoxonase and arylesterase activities recently have been found to be reduced in gastrointestinal diseases such as chronic liver disease.^13,14Oxidative stress has been proposed as an underlying mechanism of many chronic diseases, and reactive oxygen species are known to play a critical role in gut epithelial cell damage induced by ischemia.¹⁵ Intestinal ischemia includes a variety of potentially life-threatening conditions such as ischemic colitis and acute or chronic mesenteric ischemia.¹⁶ To determine the role of oxidative stress and lipid peroxidation in the pathophysiology of intestinal ischemia could lead to new approaches to treat and prevent these clinical conditions.In this experimental animal study, we aimed to evaluate the effect of arterial ischemia on both oxidant/antioxidant capacities and lipid peroxidation in a rat model of superior mesenteric ischemia.     

Hyperhomocysteinemia and Oxidative Stress During Dialysis Treatment

Background/Aims.?The concomitant presence of hyperhomocysteinemia and oxidative stress may represent a determinant factor for the occurrence of vascular alterations and cardiac diseases, the main cause of death among dialysis patients. The aim was to analyze the occurrence of hyperhomocysteinemia and oxidative stress and their possible relationship in dialysis patients. Methods.?Antioxidant substances, homocysteine, folate, and vitamin B12 were determined in blood from 32 patients on hemodialysis (HD), 21 patients on peritoneal dialysis (PD), and 12 healthy individuals. Results.?Different degrees of hyperhomocysteinemia were observed in all HD patients and in 95% of the PD patients (45.30 ± 24.89 µM in HD and 35.50 ± 26.53 µM in PD). Oxidative stress defined as an imbalance between oxidant and antioxidant forces was observed in all dialysis patients, but was more intense in HD individuals. In this group, lipoperoxidation and protein oxidation were associated with lower concentrations of antioxidants such as erythrocyte vitamin E and vitamin C. Conclusions.?Hyperhomocysteinemia and oxidative stress occur in both types of dialysis treatment, possibly contributing to the establishment of complications in these patients.     

Epithelial-to-Mesenchymal Transition and Oxidative Stress in Chronic Allograft Nephropathy

Epithelial-to-mesenchymal transition (EMT) and oxidative stress contribute to kidney tissue fibrosis in various forms of native kidney disease. However, their role in chronic allograft nephropathy (CAN) remains somewhat uncertain. To address this question, kidney transplants were performed in 3-month-old rats, using the Fisher 344 --> Lewis model of CAN. Six-month posttransplant, kidney allografts displayed significant tubular atrophy, interstitial fibrosis and vascular wall thickening. Allograft recipients had significantly higher levels of serum creatinine (4.7 +/- 1.3 versus 0.59 +/- 0.08 mg/dL, p = 0.03) and proteinuria (380 +/- 102 versus 30.2 +/- 8 mg/dL, p = 0.04) compared to syngeneic grafts. Semiquantitative PCR, immunoblot and immunohistochemical analyses demonstrated increased alpha-smooth muscle actin (alpha-SMA) mRNA and protein levels coupled with reduced E-cadherin mRNA and protein immunoreactivity, confirming the presence of CAN-associated EMT. Allograft alpha-SMA levels were increased as early as 1-2 weeks posttransplant. Immunohistochemical studies for collagen type I and III, superoxide anion (O(2) (-)), inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) confirmed that tubular O(2) (-), eNOS and iNOS, and interstitial collagen I, III and O(2) (-) levels were significantly increased in CAN-associated EMT. In conclusion, these observations suggest that CAN-associated EMT may be a link between oxidative stress and allograft fibrosis.