Urinary Thiobarbituric Acid Reactive Substances Research Articles

Exposure to Benzo[a]pyrene and 1-Nitropyrene in Particulate Matter Increases Oxidative Stress in the Human Body.

Polycyclic aromatic hydrocarbons (PAHs) have been reported to cause oxidative stress in metabolic processes. This study aimed to evaluate the relationship between exposure to PAHs, including benzo[a]pyrene (BaP) and 1-nitropyrene (1-NP), in the atmosphere and oxidative stress levels in the human body. This study included 44 Korean adults who lived in Cheongju, Republic of Korea. Atmospheric BaP and 1-NP concentrations and urinary 6-hydroxy-1-nitropyrene (6-OHNP), N-acetyl-1-aminopyrene (1-NAAP), and 1-hydroxypyrene (1-OHP) concentrations were measured. The oxidative stress level was assessed by measuring urinary thiobarbituric acid-reactive substances (TBARS) and 8-hydroxydeoxyguanosine (8-OHdG) concentrations. Urinary TBARS and 6-OHNP concentrations significantly differed between winter and summer. BaP exposure was significantly associated with urinary 8-OHdG concentrations in summer. However, atmospheric 1-NP did not show a significant correlation with oxidative stress marker concentrations. Urinary 1-NAAP concentration was a significant determinant for urinary 8-OHdG concentration in summer. Oxidative stress in the body increases in proportion to inhalation exposure to BaP, and more 8-OHdG is produced in the body as the amount of 1-NP, which is metabolized to 1-AP or 1-NAAP, increases.

MO079: Treatment with Nebivolol Ameliorates Renovascular Alterations and Oxidative Stress in Tenofovir-Induced Nephrotoxicity in Rats

Abstract BACKGROUND AND AIMS Human immunodeficiency virus (HIV) continues to be a major global public health issue, having claimed millions of lives so far. Tenofovir disoproxil fumarate (TDF), a widely prescribed component in antiretroviral regimens for HIV treatment, has been associated with nephrotoxicity. Nebivolol (NBV) is a third generation selective β-1 adrenergic receptor blocker currently used in the treatment of hypertension and cardiovascular diseases. The potential mechanisms by which NBV may protect renal structure and function include suppression of oxidative stress and enhancement of nitric oxide synthesis. The aim of this study was to investigate whether NBV could be an effective therapeutic option to mitigate TDF-induced nephrotoxicity. METHOD We divided Wistar rats into four groups: control (C, n = 8), received a standard diet for 30 days; TDF (n = 9), received a standard diet with the addition of TDF (300 mg/kg food) for 30 days; NBV (n = 8), received a standard diet for 30 days with the addition of NBV (100 mg/kg food) in the last 15 days; and TDF + NBV (n = 9), received a standard diet with the addition of TDF for 30 days and NBV in the last 15 days. We evaluated inulin clearance (GFR); tubular injury; mean arterial pressure (MAP); renal blood flow (RBF) and calculated renal vascular resistance (RVR); plasma and tissue angiotensin II (AIIp and AIIt, respectively); plasma aldosterone; plasma asymmetric dimethylarginine (ADMA); plasma and urinary thiobarbituric acid reactive substances (TBARSp and TBARSu, respectively); and whole blood glutathione (GSH). In renal tissue, we immunoblotted for endothelial nitric oxide synthase (eNOS), p47phox and p67phox. Data are expressed as mean ± SEM. RESULTS TDF + NBV rats showed an improvement in renal function, a normalization of MAP and attenuation in renal vasoconstriction, evidenced by increased RBF and diminished RVR. Administration of NBV also reduced AII and aldosterone levels in TDF-treated animals. Furthermore, the TDF + NBV group showed a lower plasma ADMA concentration and a higher protein expression of eNOS. Treatment with NBV was responsible for the maintenance of an adequate balance of TBARS and GSH levels, and it was associated with reductions in renal p47phox and p67phox. All results are described in Table 1. CONCLUSION Our data demonstrated the protective role of NBV in TDF-induced nephrotoxicity. The beneficial effects of NBV include a reduction in oxidative stress and RAAS activation, as well as a decrease in endothelial dysfunction, observed by an upregulation on the NO cascade. Therefore, NBV may be an effective therapeutic strategy in the treatment of patients with TDF-induced nephrotoxicity. Financial support: CAPES and FAPESP (2019/20840–0, 2018/12297–1, 2018/04930–6).

Oxidative Stress and Inflammation Contribute to Kidney Injury Risk During Prolonged Passive Extreme Heat Exposure

IntroductionKidney injury is a top cause of hospitalization during extreme heat events. Currently, wet bulb temperatures (Twet) during extreme heat events rarely exceed 31°C. However, it is predicted that Twet during future extreme heat events will exceed 34˚C, the limit of compensability in humans. We previously reported that, compared to a Twet of 31˚C, an 8 h resting exposure to a Twet of 34˚C elevated the risk of kidney injury risk, as quantified via the FDA‐approved kidney injury risk biomarker – the product of urine insulin‐like growth factor binding‐protein 7 and urine tissue inhibitor of metalloproteinases 2 ([IGFBP7·TIMP‐2]). This kidney injury risk is likely of tubular origin, and while the mechanisms underlying this extreme heat mediated tubular injury are unknown, an inflammatory etiology occurring secondary to oxidative stress has been speculated.PurposeTest the hypothesis that kidney injury risk during prolonged passive extreme heat exposure is contributed to by oxidative stress and inflammation.MethodsFifteen healthy men underwent 8 h resting exposures in Twet of 31°C and 34°C. Blood and urine samples were collected at 0 and 8 h. Following observations of elevations in [IGFBP7·TIMP‐2] in 34°C vs. 31°C (p<0.01), assays were conducted for urine liver‐type fatty acid binding‐protein (uL‐FABP ‐ a marker of tubular injury caused by oxidative stress), urine thiobarbituric acid reactive substances (uTBARS ‐ a marker of renal oxidative stress), urine interleukin‐18 (uIL‐18 ‐ a marker of tubular injury and inflammation), and plasma interleukin‐17a (pIL‐17a ‐ a circulating marker of inflammation, n=10). Serum TBARS (sTBARS) was measured to assess systemic oxidative stress. Data are reported as a change from 0 h (mean ± SD). Urine markers were normalized to urine flow rate.ResultsuL‐FABP increased in 34°C (+42 ± 64 ng/min, p<0.01) but not 31°C (+11 ± 20 ng/min, p=0.75) and was different between trials at 8 h (p<0.01). uTBARS increased in 34°C (+43 ± 37 nmol/min, p<0.01) but not 31°C (+14 ± 13 nmol/min, p=0.27) and was different between trials at 8 h (p<0.01). sTBARS did not change in 34°C (+1.2 ± 4.0 nmol/mL, p=0.08) nor 31°C (+0.3 ± 2.9 nmol/mL, p=0.92) and did not differ between trials at 8 h (p=0.16). uIL‐18 increased in 34°C (+27 ± 19 pg/min, p<0.01) and 31°C (+20 ± 25 pg/min, p<0.01) and was higher in 34°C at 8 h (p=0.05). pIL‐17a increased in 34°C (+199 ± 90 fg/dL, p<0.01) but not 31°C (+9 ± 96 fg/dL, p=0.94) and was different between trials at 8 h (p<0.01).ConclusionIncreases in uL‐FABP and uTBARS, in the absence of differences in sTBARS, support that renal tubular injury in 34°C is likely due to oxidative stress. pIL‐17a is selectively elevated in ischemic kidney injury. Thus, elevations in pIL‐17a in 34°C indicates inflammation of renal origins and this was translated to greater increases in uIL‐18 in 34°C. These findings support that kidney injury risk during prolonged passive extreme heat exposure is contributed to by oxidative stress and inflammation.

Enhancing kidney DDAH-1 expression by adenovirus delivery reduces ADMA and ameliorates diabetic nephropathy.

Endothelial dysfunction, characterized by reduced bioavailability of nitric oxide and increased oxidative stress, is a hallmark characteristic in diabetes and diabetic nephropathy (DN). High levels of asymmetric dimethylarginine (ADMA) are observed in several diseases including DN and are a strong prognostic marker for cardiovascular events in patients with diabetes and end-stage renal disease. ADMA, an endogenous endothelial nitric oxide synthase (NOS3) inhibitor, is selectively metabolized by dimethylarginine dimethylaminohydrolase (DDAH). Low DDAH levels have been associated with cardiac and renal dysfunction, but its effects on DN are unknown. We hypothesized that enhanced renal DDAH-1 expression would improve DN by reducing ADMA and restoring NOS3 levels. DBA/2J mice injected with multiple low doses of vehicle or streptozotocin were subsequently injected intrarenally with adenovirus expressing DDAH-1 (Ad-h-DDAH-1) or vector control [Ad-green fluorescent protein (GFP)], and mice were followed for 6 wk. Diabetes was associated with increased kidney ADMA and reduced kidney DDAH activity and DDAH-1 expression but had no effect on kidney DDAH-2 expression. Ad-GFP-treated diabetic mice showed significant increases in albuminuria, histological changes, glomerular macrophage recruitment, inflammatory cytokine and fibrotic markers, kidney ADMA levels, and urinary thiobarbituric acid reactive substances excretion as an indicator of oxidative stress, along with a significant reduction in kidney DDAH activity and kidney NOS3 mRNA compared with normal mice. In contrast, Ad-h-DDAH-1 treatment of diabetic mice reversed these effects. These data indicate, for the first time, that DDAH-1 mediates renal tissue protection in DN via the ADMA-NOS3-interaction. Enhanced renal DDAH-1 activity could be a novel therapeutic tool for treating patients with diabetes.

L-Homoarginine supplementation prevents diabetic kidney damage.

l‐homoarginine is an endogenous, non‐proteinogenic amino acid that has emerged as a new player in health and disease. Specifically, low l‐homoarginine levels are associated with cardiovascular diseases, stroke, and reduced kidney function. However, the role of l‐homoarginine in the pathogenesis of diabetic nephropathy (DN) is not known. Experiments were conducted in 6‐week‐old Ins2Akita mice supplemented with l‐homoarginine via drinking water or mini osmotic pump for 12 weeks. Both plasma and kidney l‐homoarginine levels were significantly reduced in diabetic mice compared to nondiabetic controls. Untreated Ins2Akita mice showed significant increases in urinary albumin excretion, histological changes, glomerular macrophage recruitment, the inflammatory cytokine KC‐GRO/CXCL1, and urinary thiobarbituric acid reactive substances (TBARS) excretion as an indicator of oxidative stress, along with a significant reduction in kidney nitrate + nitrite levels compared to control mice at 18 weeks of age. In contrast, l‐homoarginine supplementation for 12 weeks in Ins2Akita mice, via either drinking water or mini osmotic pump, significantly reduced albuminuria, renal histological changes, glomerular macrophage recruitment, KC‐GRO/CXCL1 levels, urinary TBARS excretion, and largely restored kidney nitrate + nitrite levels. These data demonstrate that l‐homoarginine supplementation attenuates specific features of DN in mice and could be a potential new therapeutic tool for treating diabetic patients.

Increased arterial pressure in mice with overexpression of the ADHD candidate gene calcyon in forebrain.

The link between blood pressure (BP) and cerebral function is well established. However, it is not clear whether a common mechanism could underlie the relationship between elevated BP and cognitive deficits. The expression of calcyon, a gene abundant in catecholaminergic and hypothalamic nuclei along with other forebrain regions, is increased in the brain of the spontaneously hypertensive rat (SHR) which is a widely accepted animal model of essential hypertension and attention deficit hyperactivity disorder (ADHD). Previous studies demonstrated that mice with up-regulation of calcyon in forebrain (CalOE) exhibit deficits in working memory. To date, there is no evidence directly connecting calcyon to BP regulation. Here, we investigated whether forebrain up-regulation of calcyon alters BP using radiotelemetry. We found that CalOE mice exhibited higher mean arterial pressure (MAP) compared to tTA controls. Plasma norepinephrine levels were significantly higher in CalOE mice compared to tTA controls. Silencing the transgene with doxycycline normalized BP in CalOE mice, whereas challenging the mice with 4% high salt diet for 12 days exacerbated the MAP differences between CalOE and tTA mice. High salt diet challenge also increased proteinuria and urinary thiobarbituric acid reactive substances (TBARs) in tTA and CalOE; and the increases were more prominent in CalOE mice. Taken together, our data suggest that upregulation of calcyon in forebrain could increase BP via alterations in noradrenergic transmission and increased oxidative stress during high salt challenge. Overall, this study reveals that calcyon could be a novel neural regulator of BP raising the possibility that it could play a role in the development of vascular abnormalities.

Moderate Effect of Flavonoids on Vascular and Renal Function in Spontaneously Hypertensive Rats

Many studies have shown that flavonoids are effective as antihypertensive drugs in arterial hypertension. In the present work, we have analyzed the effects of some flavonoid extracts in the spontaneous hypertensive rat model (SHR). An important feature of this study is that we have used a low dose, far from those that are usually applied in human therapy or experimental animals, a dose that responded to the criterion of a potential future commercial use in human subjects. Treatments were carried out for 6 and 12 weeks in two groups of SHR rats, which received apigenin, lemon extract, grapefruit + bitter orange (GBO) extracts, and cocoa extract. Captopril was used as a positive control in the SHR group treated for 6 weeks (SHR6) and Diosmin was used as the industry reference in the SHR group treated for 12 weeks (SHR12). Captopril and GBO extracts lowered the high arterial pressure of the SHR6 animals, but none of the extracts were effective in the SHR12 group. Apigenin, lemon extract (LE), GBO, and captopril also improved aortic vascular relaxation and increased plasma and urinary excretion of nitrites, but only in the SHR6 group. Kidney and urinary thiobarbituric acid reactive substances (TBARS) were also significantly reduced by GBO in the SHR6 rats. Apigenin also improved vascular relaxation in the SHR12 group and all the flavonoids studied reduced urinary thiobarbituric acid reactive substances (TBARS) excretion and proteinuria. Vascular abnormalities, such as lumen/wall ratio in heart arteries and thoracic aorta, were moderately improved by these treatments in the SHR6 group. In conclusion, the flavonoid-rich extracts included in this study, especially apigenin, LE and GBO improved vascular vasodilatory function of young adult SHRs but only the GBO-treated rats benefited from a reduction in blood pressure. These extracts may be used as functional food ingredients with a moderate therapeutic benefit, especially in the early phases of arterial hypertension.

Influence of feeding thermally peroxidized soybean oil on growth performance, digestibility, and gut integrity in finishing pigs.

Consumption of peroxidized lipids has been shown to reduce pig performance and energy and lipid digestibility. Objectives of the current study were to evaluate the effect of feeding soybean oil (SO) with different levels of peroxidation on growth performance, lipid, N, and GE digestibility, plasma Trp, and gut integrity in finishing pigs. Fifty-six barrows (46.7 ± 5.1 kg initial BW) were randomly assigned to one of four diets in each of two dietary phases, containing either 10% fresh SO (22.5 °C) or thermally processed SO (45 °C for 288 h, 90 °C for 72 h, or 180 °C for 6 h), each infused with of 15 L/min of air. Peroxide values were 2.0, 17.4, 123.6, and 19.4 mEq/kg; 2,4-decadienal values were 2.07, 1.90, 912.15, and 915.49 mg/kg; and 4-hydroxynonenal concentrations were 0.66, 1.49, 170.48, and 82.80 mg/kg, for the 22.5, 45, 90, and 180 °C processed SO, respectively. Pigs were individually housed and fed ad libitum for 81 d to measure growth performance, including a metabolism period to collect urine and feces for determination of GE, lipid, N digestibility, and N retention. Following the last day of fecal and urine collection when pigs were in the metabolism crates, lactulose and mannitol were fed and subsequently measured in the urine to evaluate gut permeability, while markers of oxidative stress were evaluated in plasma, urine, and liver. There were no differences observed in ADFI (P = 0.91), but average daily gain (ADG) and gain:feed G:F were decreased in pigs fed 90 °C SO diet (P ≤ 0.07) compared to pigs fed the other SO diets. Pigs fed the 90 and 180 °C SO had the lowest (P = 0.05) DE as a % of GE compared to pigs fed the 22.5 °C SO, with pigs fed the 45 °C SO being intermediate. Lipid digestibility was similarly affected (P = 0.01) as energy digestibility, but ME as a % of DE was not affected by dietary treatment (P = 0.16). There were no effects of lipid peroxidation on N digested, N retained, or the urinary lactulose:mannitol ratio (P ≥ 0.25). Pigs fed the SO processed at 90 and 180 °1C had lower concentrations (P < 0.01) of plasma Trp compared to pigs fed the 22.5 and 45 °C SO treatments. Pigs fed 90 °C SO had the greatest (P < 0.01) concentrations of F2-isoprostane in plasma and urine thiobarbituric acid reactive substances compared to the other SO treatments. These results indicate that the change in FA composition and/or the presence of lipid peroxidation products in peroxidized SO may reduce ADG, G:F, and digestibility of GE and ether extract, but has little impact on N digestibility and balance or on gut permeability.

Abstract P048: Effects of Aerobic Exercise on Oxidative Stress in the Kidneys of Type2 Diabetic Rats

Objective: Aerobic exercise is known to have multiple beneficial effects including renal protection in type 2 diabetes mellitus and obesity. However, the mechanisms regulating these actions of aerobic exercise remain unclear. The present study evaluated the effects of chronic aerobic exercise on early stage of diabetic nephropathy focusing on renal oxidative stress and nitric oxide (NO), using Zucker diabetic fatty (ZDF) rats as an animal model of obese type 2 diabetes. Methods: Male ZDF rats (6 weeks old) underwent forced treadmill exercise for 8 weeks (Ex-ZDF). Sedentary ZDF (Sed-ZDF) and Zucker lean (Sed-ZL) rats served as controls. After the last exercise session, all rats were subjected to an intra-peritoneal glucose tolerance test. Result: Exercise ameliorated hyperglycemia with increased insulin secretion, and raised the values of homeostasis model assessment for β-cell function (HOMA-β) in ZDF rats. Exercise also reduced albumin excretion and normalized creatinine clearance in ZDF rats. Endothelial (e) and neuronal (n) NO synthase (NOS) protein expression, and NOS activity in the kidneys of Sed-ZDF rats were lower compared with Sed-ZL rats, while both NOS expression and NOS activity were upregulated by exercise in ZDF rats. Although there was no significant difference in plasma levels of adiponectin between the groups, plasma levels of leptin were significantly higher in the Ex-ZDF group compared with the Sed-ZL and Sed-ZDF groups. Exercise decreased plasma and urinary thiobarbituric acid-reactive substances (TBARS) as an index of lipid peroxidation in ZDF rats. Additionally, renal nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity and expression of p47phox protein were significantly higher in the kidneys of the Sed-ZDF group compared with the Sed-ZL group, and there was no significant difference between the Ex-ZDF and Sed-ZL groups. Immunoblots for Nox4, Nox2, namely gp91phox and p22phox protein were undetected in the kidneys of the Sed-ZDF and Ex-ZDF group as well as the Sed-ZL group. In addition, expression of nitrotyrosine as an index of peroxynitrite (ONOO–) formation was significantly lower in the kidneys of the Sed-ZDF group compared with the Sed-ZL group, and was significantly higher in the Ex-ZDF group compared with the Sed-ZDF group. Further, morphometric evidence of renal damage was alleviated in response to exercise. Conclusion: Upregulated NOS activity and NOS protein, and ameliorated NADPH oxidase activity and p47phox protein in the kidneys may be potential mechanisms by which chronic aerobic exercise can reduced early diabetic nephropathy in ZDF rats. Chronic aerobic exercise does have beneficial effects and may be a novel therapeutic approach for preventing the development of renal dysfunction in patients with type 2 diabetes mellitus and obesity.

Do teas rich in antioxidants reduce the physicochemical and peroxidative risk factors for calcium oxalate nephrolithiasis in humans? Pilot studies with Rooibos herbal tea and Japanese green tea.

Several experimental and animal studies have demonstrated that substances rich in antioxidants can reduce the physicochemical and peroxidative risk factors for calcium oxalate (CaOx) renal stone formation in urine and blood. However, there are very few such investigations in humans. In the present pilot study, two varieties of tea, a green one from Japan (JGT) and a herbal one from South Africa (Rooibos) (RT), both rich in antioxidants, were administered to a group of CaOx stone formers (SF) (n=8) for 30days. Both teas were analysed for polyphenols by high-performance liquid chromatography and for minerals by plasma atomic and optical emission spectroscopy. 24h urines (baseline and day 30) were analysed for lithogenic factors. CaOx metastable limits and crystal nucleation and growth kinetics were also determined in each urine sample. Deposited crystals were inspected by scanning electron microscopy. Blood samples were collected (baseline and day 30). Biomarkers of oxidative stress including plasma and urinary thiobarbituric acid reactive substances (TBARS) and urinary N-acetyl-β-D-glucosaminidase (NAG) were also determined. Urinary physicochemical risk factors were also investigated after ingestion of RT for 30days in two control groups (CG1 and CG2), the latter one of which consisted of habitual JGT drinkers. Statistical analyses were performed using Wilcoxon signed rank tests and Mann-Whitney tests for paired and independent measurements, respectively. Several flavonoids and catechins were quantified in RT and JGT, respectively, confirming that both teas are rich sources of antioxidants. Mineral content was found to be far below dietary reference intakes. There were no significant changes in any of the urinary physicochemical or peroxidative risk factors in the control groups or in SF, except for the supersaturation (SS) of brushite (Bru) which decreased in the latter group after ingestion of JGT. Crystal morphology showed a tendency to change from mixed CaOx mono- and di-hydrate to monohydrate after ingestion of each tea. Since the latter form has a stronger binding affinity for epithelial cells, this effect is not protective. Analysis of the physicochemical and peroxidative risk factors in CG1 and CG2 did not reveal any evidence of a synergistic effect between the two teas. Paradoxically, baseline risk factors in the habitual JGT control group were significantly raised relative to those in CG1. Our preliminary results suggest that ingestion of RT and JGT does not reduce the risk factors for CaOx stone formation in humans, but these findings need to be tested in further studies involving much larger sample sizes.

Pharmacogenetics may Influence Tacrolimus Daily Dose, but not Urinary Tubular Damage Markers in the Long-Term Period after Renal Transplantation.

SummaryBackgroundThe primary goal of this study was to evaluate the influence of cytochrome P450 (CYP) 3A5 (6986A>G) and ABCB1 (3435C>T) polymorphisms on tacrolimus (TAC) dosage regimen and exposure. Second, we evaluated the influence of TAC dosage regimen and the tested polymorphisms on renal oxidative injury, as well as the urinary activities of tubular ectoenzymes in a long-term period after transplantation. Also, we aimed to determine the association between renal oxidative stress and tubular damage markers in the renal transplant patients.MethodsThe study included 72 patients who were on TAC based immunosuppression. Allele-specific PCR was used for polymorphism determination. We measured the urinary thiobarbituric acid reactive substances (TBARS) and reactive carbonyl derivates (RCD) in order to evaluate oxidative injury, as well as the urinary activities of ectoenzymes (N-acetyl-β-D-glucosaminidase, aminopeptidase N and dipeptidyl peptidase IV) to evaluate tubular damage.ResultsThe carriers of CYP 3A5*1 allele required statistically higher daily doses of TAC than CYP *3/*3 carriers, as well as the carriers of C allele of ABCB1 gene compared to those with TT genotype. Also, there were no differences in TBARS, RCD and the activities of ectoenzymes between the patients’ genotypes. Our results showed significant correlations between urinary TBARS and RCD and the ectoenzymes’ activities.ConclusionsOur findings suggest that CYP 3A5 and ABCB1 3435 polymorphism may affect TAC daily doses, but not the drug’s tubular toxicity. Furthermore, tubular damage may be associated with increased renal oxidative stress.

Arginase inhibition: a new treatment for preventing progression of established diabetic nephropathy

Our previous publication showed that inhibition of arginase prevents the development of diabetic nephropathy (DN). However, identification of targets that retard the progression of established DN-which is more clinically relevant-is lacking. Therefore, we tested the hypothesis that arginase inhibition would prevent the progression of established DN. Effects of arginase inhibition were compared with treatment with the angiotensin-converting enzyme inhibitor captopril, a current standard of care in DN. Experiments were conducted in Ins2(Akita) mice treated with the arginase inhibitor S-(2-boronoethyl)-l-cysteine (BEC) or captopril starting at 6 wk of age for 12 wk (early treatment) or starting at 12 wk of age for 6 wk (late treatment). Early and late treatment with BEC resulted in protection from DN as indicated by reduced albuminuria, histological changes, kidney macrophage infiltration, urinary thiobarbituric acid-reactive substances, and restored nephrin expression, kidney nitrate/nitrite, kidney endothelial nitric oxide synthase phosphorylation, and renal medullary blood flow compared with vehicle-treated Ins2(Akita) mice at 18 wk of age. Interestingly, early treatment with captopril reduced albuminuria, histological changes, and kidney macrophage infiltration without affecting the other parameters, but late treatment with captopril was ineffective. These findings highlight the importance of arginase inhibition as a new potential therapeutic intervention in both early and late stages of diabetic renal injury.

The lipid peroxidation and antioxidant status of type 2 diabetic patients in Kashmir (India)

The present study was to evaluate the redox balance in type 2 diabetes mellitus by measuring oxidative stress and antioxidant status along with lipid profile parameters in Kashmiri population. So far, no study has been undertaken on this aspect in Kashmiri population. We, therefore, aimed this maiden study in Kashmiri population to examine redox balance by measuring oxidative stress and antioxidant status along with lipid profile in 50 patients of diabetes mellitus type 2 and 30 matched normal subjects. Fifty patients of diabetes mellitus type 2 without any complications (mean age = 59.8 ± 10.4 years) and 30 normal subjects (mean age = 52.8 ± 12.8 years) were included in this study. Body mass index and waist/hip ratio was measured. Fasting blood sample was collected for the analysis of total antioxidant activity, plasma and urinary thiobarbituric acid reactive substances and lipid profile by standard procedures in both the groups. Total cholesterol, triglyceride, LDL cholesterol, plasma, and urinary thiobarbituric acid reactive substances (raised oxidative stress) were significantly raised, whereas plasma total antioxidant activity was significantly reduced in diabetes mellitus type 2 patients as compared to controls. The comparison of old and fresh cases revealed that though total antioxidant activity was lower and plasma thiobarbituric acid reactive substances and urinary thiobarbituric acid reactive substances were higher in patients, but did not attain the level of significance. W/H ratio is significantly higher in patients compared to normal subjects. But, no significant correlation of body mass index and waist/hip with lipid profile is observed in both control and patients. Raised oxidative stress in type 2 diabetes mellitus patients, along with deranged lipid profile, increased W/H ratio and decreased antioxidant activity could be the risk factors in the development of complications associated with diabetes mellitus.

Influence of thermally oxidized vegetable oils and animal fats on intestinal barrier function and immune variables in young pigs.

To evaluate the effect of feeding thermally oxidized lipids on metabolic oxidative status, gut barrier function, and immune response of young pigs, 108 barrows (6.67 ± 0.03 kg BW) were assigned to 12 dietary treatments in a 4 × 3 factorial arrangement in addition to a corn-soybean meal control diet. Main effects were 4 lipid sources (corn oil [CN], canola oil [CA], poultry fat [PF], and tallow [TL]) and 3 oxidation levels (original lipids [OL], slow oxidation [SO] of lipids heated for 72 h at 95°C, or rapid oxidation [RO] of lipids heated for 7 h at 185°C). Pigs were provided ad libitum access to diets for 28 d followed by controlled feed intake for 10 d. After a 24-h fast on d 38, serum was collected and analyzed for α-tocopherol (α-T), thiobarbituric acid reactive substances (TBARS), endotoxin, haptoglobin, IgA, and IgG. On the same day following serum collection, lactulose and mannitol were fed and subsequently measured in the urine to evaluate gut permeability. There was a source × peroxidation interaction for serum α-T concentration where pigs fed SO or RO had decreased (P < 0.05) serum α-T concentration compared with pigs fed OL in CA and CN diets but not in pigs fed PF and TL diets. There was no source × peroxidation interaction for serum TBARS, but among all lipid sources, pigs fed SO or RO lipids had increased (P < 0.05) serum TBARS compared with pigs fed OL. In addition, pigs fed CN or CA had greater (P < 0.05) serum TBARS compared with pigs fed PF or TL diets. There were no lipid source × peroxidation level interaction or lipid source or peroxidation level effects on serum endotoxin, haptoglobin, IgA, or IgG. Pigs fed lipid supplemented diets tended to have increased serum endotoxin (P = 0.06), IgA (P = 0.10), and IgG (P = 0.09) compared with pigs fed the control diet. There were no lipid source × peroxidation level interaction or lipid source or peroxidation level effects on urinary TBARS and lactulose to mannitol ratio. Compared with pigs fed the control diet, pigs fed diets containing lipids had a lower lactulose to mannitol ratio (P < 0.01). In conclusion, feeding weaning pigs diets containing 10% thermally oxidized lipids for 38 d, especially vegetable oils containing greater concentrations of PUFA, appeared to impair oxidative status but had little influence on gut barrier function or serum immunity parameters.

Adenosine kinase inhibition protects the kidney against streptozotocin-induced diabetes through anti-inflammatory and anti-oxidant mechanisms

Adenosine provides anti-inflammatory effects in cardiovascular disease via the activation of adenosine A2A receptors; however, the physiological effect of adenosine could be limited due to its phosphorylation by adenosine kinase. We hypothesized that inhibition of adenosine kinase exacerbates extracellular adenosine levels to reduce renal inflammation and injury in streptozotocin-induced diabetes. Diabetes was induced in male C57BL/6 mice by daily injection of streptozotocin (50mg/kg/day, i.p. for 5 days). Control and diabetic mice were then treated with the adenosine kinase inhibitor ABT702 (1.5mg/kg, i.p. two times a week for 8 weeks, n=7–8/group) or the vehicle (5% DMSO). ABT702 treatment reduced blood glucose level in diabetic mice (∼20%; P<0.05). ABT702 also reduced albuminuria and markers of glomerular injury, nephrinuria and podocalyxin excretion levels, in diabetic mice. Renal NADPH oxidase activity and urinary thiobarbituric acid reactive substances (TBARS) excretion, indices of oxidative stress, were also elevated in diabetic mice and ABT702 significantly reduced these changes. ABT702 increased renal endothelial nitric oxide synthase expression (eNOS) and nitrate/nitrite excretion levels in diabetic mice. In addition, the diabetic mice displayed an increase in renal macrophage infiltration, in association with increased renal NFκB activation. Importantly, treatment with ABT702 significantly reduced all these inflammatory parameters (P<0.05). Furthermore, ABT702 decreased glomerular permeability and inflammation and restored the decrease in glomerular occludin expression in vitro in high glucose treated human glomerular endothelial cells. Collectively, the results suggest that the reno-protective effects of ABT702 could be attributed to the reduction in renal inflammation and oxidative stress in diabetic mice.

Antioxidant effect of imperatorin from Angelica dahurica in hypertension via inhibiting NADPH oxidase activation and MAPK pathway

Imperatorin (IMP) is an active furocoumarin in the traditional Chinese medicine Angelica dahurica and has been demonstrated to have vasodilatory activity. In the present study, we investigated the effect of IMP on blood pressure (BP) and antioxidant effects in spontaneously hypertensive rats (SHR) and human embryonic kidney 293 cells. SHR were administered IMP (6.25, 12.5, and 25 mg/kg/d) or tempol (18 mg/kg/d) daily by gavage for 12 weeks. Thiobarbituric acid–reactive substances, proteinuria levels, and superoxide dismutase activity were evaluated with commercial kits. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits of the renal cortical tissues were determined by reverse transcriptase polymerase chain reaction and Western blot. Twenty-four hour urinary 8-Iso-prostaglandin F2α was measured by enzyme linked immunosorbent assay. Systolic BP and diastolic BP were significantly reduced by treatment with IMP (6.25, 12.5, and 25 mg/kg/d) in SHR. Meanwhile, we found that renal cortical superoxide dismutase activities were significantly increased in IMP-treated groups. Renal cortical and urinary thiobarbituric acid–reactive substances' levels, the 24-hour urinary excretion of 8-Iso-prostaglandin F2α, and proteinuria in the IMP-treated group, were lower than SHR group. After that, we found the messenger RNA expressions and protein levels of NADPH oxidase subunits were markedly reduced after IMP treated in SHR. IMP also reduced the phosphorylation of protein kinase B, extracellular signal-regulated kinase1/2, p38 mitogen-activated protein kinase (MAPK), and c-Jun N-terminal kinase in renal cortical in SHR. In addition, H2O2-induced ROS production in human embryonic kidney 293 cells was markedly attenuated by IMP. H2O2-induced activation of MAPK, protein kinase B, and expression of NADPH oxidase were also attenuated by pretreatment of IMP. In summary, IMP showed antihypertensive effect via prevention of renal injury not only by reducing NADPH oxidase but also by inhibiting of MAPK pathway.

Nitro-oleic acid protects against adriamycin-induced nephropathy in mice

Adriamycin (ADR) administration in susceptible rodents such as the BALB/c mouse strain produces injury to the glomerulus mimicking human focal glomerular sclerosis. The goal of the present study was to use this model to investigate antiproteinuric action of nitro-oleic acid (OA-NO2), a nitric oxide-derived endogenous lipid product, which has exhibited multiple attractive signaling properties particularly in the kidney. BALB/c mice were pretreated for 2 days with OA-NO2 at 5 mg·kg(-1)·day(-1) via an osmotic minipump, followed by a single injection of vehicle or adriamycin (10 mg/kg) via the tail vein. Albuminuria and renal function were analyzed at 1 wk post-ADR treatment. ADR mice developed prominent albuminuria, hypoalbuminemia, hyperlipidemia, and severe ascites. In contrast, the symptoms of nephrotic syndrome were greatly improved by OA-NO2 treatment. In parallel, plasma creatinine and plasma urea nitrogen were elevated in the ADR group, and the severity was less in the ADR+OA-NO2 group. OA-NO2 attenuates ADR-induced glomerulosclerosis, podocyte loss, and tubulointerstitial fibrosis. Indices of oxidative stress, including plasma and urinary thiobarbituric acid-reactive substances and renal expression of NAD(P)H oxidase p47(phox) and gp91(phox), and inflammation, including renal expression of TNF-α, IL-1β, and MCP-1 in response to ADR, were all similarly suppressed. Together, these findings suggest that OA-NO2 exerts renoprotective action against ADR nephropathy likely via its anti-inflammatory and antioxidant properties.

Abstract 440: Female Spontaneously Hypertensive Rats are More Dependent on Nitric Oxide in Modulating Blood Pressure and Renal Injury Than Males

We recently demonstrated that female spontaneously hypertensive rats (SHR) have higher nitric oxide (NO) bioavailability in the kidney compared to males. Therefore, we hypothesize that female SHR are more dependent on the NO synthase (NOS) system to modulate their blood pressure, and as a result will have a greater rise in blood presure and renal injury in response to chronic NOS inhibition compared to males. Mean arterial pressure in SHR was very sensitive to NOS inhibition, with both sexes exhibiting a significant increase in blood pressure to the non-specific NOS inhibitor N G -nitro-L-arginine methyl ester (L-NAME) at a dose of 2 mg/kg/day. However, there was not a sex difference in the percent increase in mean arterial pressure. Treatment of female and male SHR with 7 mg/kg/day L-NAME for 2 weeks significantly increased mean arterial pressure and indices of renal injury in both females and males; the percent increases in mean arterial pressure and injury were greater in females during the last three days of L-NAME treatment (% increase in mean arterial pressure was 44± 1.3 in females vs. 35± 2.9 in males, P&lt;0.05). L-NAME treatment also increased indices of inflammation and oxidative stress as urinary monocyte chemoattractant protein-1 (MCP-1) and thiobarbituric acid reactive substances (TBARs) excretion levels were increased by L-NAME; however, the increases were greater in females. Renal cortical macrophage and T cell infiltrations and soluble intracellular adhesion molecule-1 (sICAM-1) were also elevated following L-NAME treatment (cortical sICAM-1 levels increased from 4.7± 0.2 to 9± 0.1 ng/mg in males vs. 4.6± 0.1 to 7.9± 0.3 ng/mg in females, P&lt;0.05). Although renal cortical macrophage infiltration was greater in female SHR vs. males following L-NAME treatment, the increase in cortical T cell infiltration and sICAM-1 were greater in males. These data suggest that the NO system plays an important role in modulating blood pressure, renal injury and inflammation in SHR, with females being more dependent on the NOS system than males.

Role of Heme Oxygenase-1 in Polymyxin B-Induced Nephrotoxicity in Rats

Polymyxin B (PMB) is a cationic polypeptide antibiotic with activity against multidrug-resistant Gram-negative bacteria. PMB-induced nephrotoxicity consists of direct toxicity to the renal tubules and the release of reactive oxygen species (ROS) with oxidative damage. This study evaluated the nephroprotective effect of heme oxygenase-1 (HO-1) against PMB-induced nephrotoxicity in rats. Adult male Wistar rats, weighing 286 ± 12 g, were treated intraperitoneally once a day for 5 days with saline, hemin (HO-1 inducer; 10 mg/kg), zinc protoporphyrin (ZnPP) (HO-1 inhibitor; 50 μmol/kg, administered before PMB on day 5), PMB (4 mg/kg), PMB plus hemin, and PMB plus ZnPP. Renal function (creatinine clearance, Jaffe method), urinary peroxides (ferrous oxidation of xylenol orange version 2 [FOX-2]), urinary thiobarbituric acid-reactive substances (TBARS), renal tissue thiols, catalase activity, and renal tissue histology were analyzed. The results showed that PMB reduced creatinine clearance (P < 0.05), with an increase in urinary peroxides and TBARS. The PMB toxicity caused a reduction in catalase activity and thiols (P < 0.05). Hemin attenuated PMB nephrotoxicity by increasing the catalase antioxidant activity (P < 0.05). The combination of PMB and ZnPP incremented the fractional interstitial area of renal tissue (P < 0.05), and acute tubular necrosis in the cortex area was also observed. This is the first study demonstrating the protective effect of HO-1 against PMB-induced nephrotoxicity.

Response of Urinary Biomarkers of Systemic Oxidation to Oral Iron Supplementation in Healthy Men

Urinary biomarkers are used in assessment of severe, clinical oxidative stress. Little is known, however, about their diagnostic value within the normative range. To evaluate the response of urinary thiobarbituric acid reactive substances (TBARS) and 8-hydroxy-2-deoxyguanosine (8-OHdG) as indicators of systemic oxidation in response to short-term oral iron and antioxidant supplementation. Five healthy adult men participated in the pilot study phase and 12 in the definitive intervention trial. For 7 days each, separated by 12-day washouts, the subjects received different treatment regimens, consisting of 120 mg of iron, 120 mg of iron in refined palm oil, and 120 mg of iron in palm oil combined with one of the two doses of Carotino Tocotrienol Carotene Mixed Concentrate (CTCMC). Creatinine-normalized urinary TBARS and 8-OHdG concentrations were quantified in samples taken from subjects with and without active supplementation. Temporal and correlative associations between TBARS and 8-OHdG were explored. Daily intake of supplemental iron failed to produce any increment in urinary excretion of TBARS or 8-OHdG. However, a significant within-individual correlation between the urinary biomarkers was observed (Spearman r = 0.697, p < .0001, n = 466). Both doses of CTCMC significantly lowered urinary excretion of both oxidation indicators. Despite the lack of effect of oral iron on the biomarkers of systemic oxidation, they show a strong and significant mutual association within the nonpathological range of oxidative stress in healthy male adults.