Usefulness of urinary kidney injury molecule‐1 (Kim‐1) as a biomarker for cisplatin‐induced sub‐chronic kidney injury

We explored biomarkers suitable for monitoring sub‐chronic kidney injury using the three rat models of cisplatin (CDDP)‐induced kidney injury, which were designed to extend the current knowledge beyond the sub‐acute exposure period. In the pilot study, a single intravenous administration of 1.5 mg kg^–1 CDDP to rats was confirmed to result in no histopathological changes. Subsequently, CDDP was intravenously administered to rats at a dose of 1.5 mg kg^–1 for 4 days at 24‐h intervals (Experimental model 1) and for up to 10 weeks at weekly intervals (Experimental models 2 and 3), and the changes in blood and urine components, such as recently recommended urinary biomarkers (Kim‐1, clusterin and so on) and traditional blood biomarkers (blood urea nitrogen and serum creatinine), were examined together with the histopathological changes in renal tissues during the development of the kidney injury in each model. In these experimental models, a significant increase in urinary Kim‐1 was observed prior to the histopathological changes in renal tissues, and these changes were retained after the adverse histopathological changes. Significant changes in all of the other urinary biomarkers examined occurred along with the histopathological changes. In addition, the increase in urinary Kim‐1 after weekly treatment with CDDP for 4 weeks was reduced in a time‐dependent manner after cessation of the drug. The present findings indicate that urinary Kim‐1 is the most useful biomarker for CDDP‐induced rat sub‐chronic kidney injury among the biomarkers examined. Copyright © 2014 John Wiley & Sons, Ltd.     

N‐acetylcysteine improves renal hemodynamics in rats with cisplatin‐induced nephrotoxicity

This work investigated the effect of N‐acetylcysteine (NAC), on renal hemodynamics in cisplatin (CP)‐induced nephrotoxicity in Wistar–Kyoto (WKY) rats. The animals were divided into four groups (n = 5 or 6). The first and second groups received normal saline (control) and intraperitoneal (i.p.) N‐acetylcysteine (500 mg kg^?1 per day for 9 days), respectively. The third and fourth groups were given a single intraperitoneal (i.p.) injection of CP (5 mg kg^?1) and an i.p. injection of CP (5 mg kg^?1) together with i.p. NAC (500 mg kg^?1 per day for 9 days), respectively. At the end of the experiment, rats were anesthetized and blood pressure and renal blood flow were monitored, followed by intravenous (i.v.) injection of norepinephrine (NE) for measurement of renal vasoconstrictor responses. CP caused a significant reduction in renal blood flow but did not affect NE‐induced renal vasoconstriction. In addition, CP significantly increased plasma concentrations of urea and creatinine and urinary N‐acetyl‐β‐D ‐glucosaminidase (NAG) activity and kidney relative weight. CP decreased body weight and creatinine clearance. Histopathologically, CP caused remarkable renal damage compared with control. NAC alone did not produce any significant change in any of the variables measured. However, NAC significantly ameliorated CP‐induced hemodynamic, biochemical and histopathological changes. The concentration of platinum in the kidneys of CP ? NAC treated rats was less than in CP‐treated rats by 37%. The results show that administration of i.p. NAC (500 mg kg^?1 per day for 9 days) reversed the renal hemodynamic changes as well as the biochemical and histopathological indices of CP‐induced nephrotoxicity in WKY rats. Copyright © 2009 John Wiley & Sons, Ltd.     

The peroxisome proliferator‐activated receptor‐γ agonist pioglitazone protects against cisplatin‐induced renal damage in mice

Peroxisome proliferator‐activated receptor‐γ (PPAR‐γ) agonists not only improve metabolic abnormalities of diabetes and consequent diabetic nephropathy, but they also protect against non‐diabetic kidney disease in experimental models. Here, we investigated the effect of PPAR‐γ agonist pioglitazone against acute renal injury on a cisplatin model in mice. Nephrotoxicity was induced by a single intraperitoneal (i.p.) injection of cisplatin (10 mg kg^–1). Pioglitazone was administered for six consecutive days in doses of 15 or 30 mg kg^–1 day^–1, per os (p.o.), starting 3 days before cisplatin injection. Cisplatin treatment to mice induced a marked renal failure, characterized by a significant increase in serum urea and creatinine levels and alterations in renal tissue architecture. Cisplatin exposure induced oxidative stress as indicated by decreased levels of non‐enzymatic antioxidant defenses [glutathione (GSH) and ascorbic acid levels] and components of the enzymatic antioxidant defenses [superoxide dismutase (SOD), catalase (CAT) glutathione peroxidase (GPx), glutathione reductase (GR) and and glutathione S‐transferase(GST) activities)] in renal tissue. Administration of pioglitazone markedly protected against the increase in urea and creatinine levels and histological alterations in kidney induced by cisplatin treatment. Pioglitazone administration ameliorated GSH and ascorbic acid levels decreased by cisplatin exposure in mice. Pioglitazone protected against the inhibition of CAT, SOD, GPx, GR and GST activities induced by cisplatin in the kidneys of mice. These results indicated that pioglitazone has a protective effect against cisplatin‐induced renal damage in mice. The protection is mediated by preventing the decline of antioxidant status. The results have implications in use of PPAR‐γ agonists in human application for protecting against drugs‐induced nephrotoxicity. Copyright © 2012 John Wiley & Sons, Ltd.     

Age‐related susceptibility to brain ischemia in mice

In this study, we evaluated the effect of age on survival and on neurological alterations produced by bilateral sequential common carotid artery (CCA) sectioning in mice. Male mice, 3–4 (young), 20–30 (adult), and 40–60 (aged) weeks old, underwent sequential CCA sectioning at an interval of 32 days. After the second surgery, mortality was registered continuously over a period of 24 h and thereafter every 24 h up to 3 days. Neurological assessment was performed 24 h after the second surgery. The influence of age on survival was analyzed using a log‐rank test, and survival curves were generated by the Kaplan‐Meier method. In mice with previous left CCA sectioning, sectioning of the contralateral artery induced a wide range of neurological alterations and a high mortality rate. Young and aged animals had a significantly lower (P < 0.05) survival rate as compared to adults (3, 6, 9, and 12 h after the second surgery), thus confirming that young and aged animals are, in fact, less resistant to brain ischemia. In contrast, the percentage of mice showing severe neurological alterations increased with age. This observation coincides with clinical evidence showing that elderly tend to have a worse outcome than younger patients do. These findings indicate that, besides morphological changes, studies of influence of age on the susceptibility to brain ischemia should include mortality and functional endpoints. Drug Dev. Res. 57:161–166, 2002. © 2003 Wiley‐Liss, Inc.     

Age-related differences in susceptibility to toxic effects of valproic acid in rats

A multi-age rat model was evaluated as a means to identify a potential age-related difference in liver injury following exposure to valproic acid (VPA), a known pediatric hepatotoxic agent. Different age groups of Sprague-Dawley (SD) rats (10-, 25-, 40-, 80-day-old) were administered VPA at doses of 160, 320, 500 or 650 mg kg(-1) (i.p.) for 4 days. Animals from all age groups developed toxicity after treatment with VPA; however, the patterns of toxicity were dissimilar within each age group. The high dose of VPA caused significant lethality in 10- and 25-day-old rats. All doses of VPA caused decrease in the platelet counts (10-, 25-day-old rats) and the rate of growth (40-day-old rats) and increases in the urine creatine concentration (high dose, 80-day-old rats). VPA induced hepatic and splenic alterations in all age groups. The most severe lesions were found mostly in 10- and 80-day-old rats. Significant changes in blood urea nitrogen, alanine aminotransferase and alkaline phosphatase were observed in 10-day-old pups after treatment with low doses of VPA. The highest VPA dose caused significant decreases in the levels of serum total protein (40- and 80-day-old rats). Principal component analysis of spectra derived from terminal urine samples of all age groups showed that each age group clusters separately. In conclusion, this study showed that the vulnerability profile of each age group was different indicating that a multi-age pediatric animal model is appropriate to assess more completely age-dependent changes in drug toxicity.     

Protective effects of apocynin on cisplatin‐induced ototoxicity in an auditory cell line and in zebrafish

Cisplatin is a very effective anticancer drug and generates reactive oxygen species (ROS) such as superoxide anions that can deplete antioxidant protective molecules in the cochlea. These processes result in the death of cochlear hair cells by induction of apoptosis. Apocynin, which is used as a specific nicotinamide adenine dinucleotide phosphate oxidase inhibitor, has a preventive effect for intracellular ROS generation. In this study, the effect of apocynin was investigated in a cochlear organ of Corti‐derived cell line, HEI‐OC1 cells, and in transgenic zebrafish (Brn3C: EGFP). To investigate the protective effects of apocynin, HEI‐OC1 cells were treated with various concentrations of apocynin and a 20 µm concentration of cisplatin, simultaneously. An in vivo study of transgenic zebrafish (Brn3C: EGFP) was used to investigate the protective effects of apocynin on cisplatin‐induced hair cell death. In an in vitro study, apocynin appeared to protect against cisplatin‐induced apoptotic features on Hoechst 33258 staining in the HEI‐OC1 cells. Treatment of the HEI‐OC1 cells with 100 µm of apocynin, significantly decreased caspase‐3 activity. Treatment of the cells with a 100 µm concentration of apocynin and a 20 µm concentration of cisplatin significantly decreased the intracellular ROS production. In the in vivo study, apocynin significantly decreased the TUNEL reaction and prevented cisplatin‐induced hair cell loss of the neuromasts in the transgenic zebrafish at low concentrations (125 and 250 µm ). These findings suggest that apocynin has antioxidative effects and prevents cisplatin‐induced apoptotic cell death in HEI‐OC1 cells as well as in zebrafish. Copyright © 2011 John Wiley & Sons, Ltd.     

Dexamethasone exacerbates cisplatin‐induced muscle atrophy

Dexamethasone for antiemetic therapy is typically administered with anticancer drugs such as cisplatin. We previously reported that cisplatin upregulates the muscle‐specific E3 ubiquitin ligase genes, namely muscle ring‐finger protein 1 (MuRF1) and atrophy gene‐1 (atrogin‐1), and promotes muscle atrophy in mice. It is well known that dexamethasone causes upregulation of MuRF1 and Atrogin‐1 expression in skeletal muscles. Although it is speculated that a combination of dexamethasone and cisplatin worsens muscle atrophy, there are no reports based on research. We thereby investigated the effects of cisplatin and dexamethasone, alone or in combination, on the expression of MuRF1 and Atrogin‐1 in murine skeletal muscles and C2C12 myotubes. Mice were intraperitoneally injected with cisplatin or the vehicle control once daily for 4 days. Dexamethasone or the vehicle control was subcutaneously administered 30 minutes prior to the administration of cisplatin. Dexamethasone enhanced MuRF1 and Atrogin‐1 gene expression upregulated by cisplatin in murine quadriceps muscles and C2C12 myotubes. Cisplatin‐caused upregulation of myostatin and downregulation of IGF‐1 gene expression were also enhanced by co‐administration of dexamethasone in murine quadriceps muscles and C2C12 myotubes. This study shows that the combination treatment of cisplatin and dexamethasone exacerbated muscle atrophy in mice. Therefore, this treatment regimen might exacerbate muscle atrophy in cancer patients.     

Diallyl sulfide alleviates cisplatin‐induced nephrotoxicity in rats via suppressing NF‐κB downstream inflammatory proteins and p53/Puma signalling pathway

Despite being a potent anticancer drug, nephrotoxicity is an adverse effect which renders the clinical use of cisplatin (Cis) limited. The protective role of diallyl sulfide (DAS); a naturally occurring organo‐sulfide, present in garlic, in cisplatin‐induced nephrotoxicity has been reported earlier. However, the mechanism through which DAS exerts its nephroprotective activity remains elusive. The aim of the current study was to elucidate the possible mechanisms underlying the reno‐protective effect of DAS in cisplatin‐induced nephrotoxicity in rats. DAS was given at 2 dose levels; 50 and 100 mg/kg, orally for 4 consecutive days, starting 1 hour after administration of single dose of cisplatin (3.5 mg/kg, intraperitoneally [i.p.]). The Cis‐induced elevation in serum urea and creatinine, degree of histopathological alterations was significantly ameliorated in cisplatin groups co‐treated with DAS. In addition, DAS significantly restored Cis‐depleted glutathione (GSH) content and superoxide dismutase (SOD) activity and attenuated Cis‐elevated Malondialdehyde (MDA) level. Also, DAS significantly reduced Cis‐increased renal expression of nuclear factor kappa B (NF‐κB) and subsequent pro‐inflammatory mediators; tumour necrosis factor alpha (TNF‐α), interleukin‐1β (IL‐1β), intercellular adhesion molecule‐1 (ICAM‐1) and inducible nitric oxide synthase (iNOS) in kidney tissues. Moreover, co‐treatment with DAS significantly inhibited Cis‐increased caspase‐8 and ‐9 levels. Additionally, DAS significantly mitigated Cis‐induced protein expression of p53, Puma, and Bax while, it significantly restored Cis‐reduced protein expression of Bcl‐xL compared to the Cis group. In conclusion, these results demonstrate that DAS ameliorates cisplatin‐induced nephrotoxicity in rats through enhancement of antioxidant defense, reduction of inflammatory cytokine tissue levels as well as inhibition of apoptosis via p53/Puma signalling pathway.     

Age‐related changes in hepatic expression and activity of drug metabolizing enzymes in male wild‐type and breast cancer resistance protein knockout mice

This study aimed to reveal age‐related changes in the expression and activity of seven hepatic drug metabolizing enzymes (DMEs) in male wild‐type and breast cancer resistance protein knockout (Bcrp1^?/?) FVB mice. The protein expression of four cytochrome P450 (Cyps) (Cyp3a11, 2d22, 2e1, and 1a2), and three UDP‐glucuronosyltransferases (Ugts) (Ugt1a1, 1a6a, and 1a9) in liver microsomes of wild‐type and Bcrp1^?/? FVB mice at different ages were determined using a validated ultra high performance liquid chromatography with tandem mass spectrometry (UHPLC–MS/MS) method. The activities and mRNA levels of these DMEs were measured using the probe substrates method and real‐time PCR, respectively. In the liver of wild‐type FVB mice, Cyp3a11, 2d22, 2e1, 1a2, Ugt1a1, and 1a6a displayed maximum protein levels at 6–9 weeks of age. Cyp1a2, Ugt1a1, 1a6a, and 1a9 showed maximum activities at 6–9 weeks of age, whereas Cyp3a11, 2d22, and 2e1 showed maximum activities in 1–3‐week‐old mice. Additionally, most of the DMEs showed maximum mRNA levels in 17‐week‐old mice liver. Compared with wild‐type FVB mice, the protein levels of these DMEs showed no significant changes in Bcrp1^?/? FVB mice liver. However, the activity of Cyp2e1 was increased and that of Cyp2d22 was decreased. In conclusion, t he seven hepatic DMEs in FVB mice liver showed significant alterations in an isoform‐specific manner with increased age. Although the protein levels of these DMEs showed no significant changes, the activities of Cyp2e1 and 2d22 were changed in Bcrp1^?/? mice.     

Novel synthetic protective compound,KR‐22335, against cisplatin‐induced auditory cell death

Cisplatin [cis‐diammine‐dichloroplatinum (II)] is a widely used chemotherapeutic agent, and one of its most severe side effects is ototoxicity. In the course of developing a new protective agent against cisplatin‐induced ototoxicity, we have been interested in a novel synthetic compound, 3‐Amino‐3‐(4‐fluoro‐phenyl)‐1H‐quinoline‐2,4‐dione (KR‐22335). We evaluated the effectiveness of KR‐22335 as an otoprotective agent against cisplatin‐induced toxicity. The otoprotective effect of KR‐22335 against cisplatin was tested in vitro in cochlear organs of Corti‐derived cell lines, HEI‐OC1, and in vivo in a zebrafish (Danio rerio) model. Cisplatin‐induced apoptosis, cell cycle arrest and an increase in intracellular reactive oxygen species (ROS) generation were demonstrated in HEI‐OC1 cells. KR‐22335 inhibited cisplatin‐induced apoptosis and mitochondrial injury in HEI‐OC1 cells. KR‐22335 inhibited cisplatin‐induced activation of JNK, p‐38, caspase‐3 and PARP in HEI‐OC1 cells. Scanning and transmission electron micrographs showed that KR‐22335 prevented cisplatin‐induced destruction of kinocilium and stereocilia in zebrafish neuromasts. Tissue TUNEL of neuromasts in zebrafish demonstrated that KR‐22335 blocked cisplatin‐induced TUNEL positive hair cells in neuromasts. The results of this study suggest that KR‐22335 may prevent ototoxicity caused by the administration of cisplatin through the inhibition of mitochondrial dysfunction and suppression of ROS generation. KR‐22335 may be considered as a potential candidate for protective agents against cisplatin‐induced ototoxicity. Copyright © 2013 John Wiley & Sons, Ltd.     

肼引发损伤相关小分子代谢物的研究进展

肼是化学合成工业及制药领域的重要中间产物,也是肼类药物的共同代谢产物。肼具有肝、肾、中枢神经等多器官毒性,严重威胁人类健康。随着生命科学技术的发展,代谢组学能够全面、整体地获得机体受到外源性刺激后体内内源性小分子代谢物的扰动信息。代谢组学已用于发现与肼损伤相关的具有特殊生物功能的可能生物标志物。该文总结了近年来基于代谢组学方法对肼引发损伤相关的小分子代谢物的研究进展,以期为肼损伤的机制研究、早期发现及治疗提供研究思路。     

Evaluation of aging influence on renal toxicity caused by segment‐specific nephrotoxicants of the proximal tubule in rat†

Little is known concerning the sensitivity of aged rats to xenobiotics inducing kidney damage. To increase this knowledge, the age‐dependent response of the kidney to hexachloro‐1 : 3‐butadiene (HCBD) or potassium dichromate (chromate) was investigated. Rats were treated at different ages with a single dose of segment‐specific nephrotoxicants of the proximal tubule, chosen on the basis of their specificity for S₃ and for S₁–S₂ segments, respectively. The toxicological impact of these xenobiotics has been evaluated through biochemical and genomic markers, and histopathological investigation of kidney samples. HCBD treatment induced tubular necrosis of the S₃ segment of the proximal tubule associated with changes of toxicological markers unrelated to the age. In contrast, chromate treatment induced an increased kidney damage related to the rat age. In fact, histopathological investigation revealed that at 1 month of age tubular vacuolar degeneration was seen affecting S₁–S₂ segments of the proximal tubule, whereas at 3 months of age tubular necrosis occurred in the same segments associated with tubular dilation of the distal portions. Consistently, biochemical analysis confirmed a direct correlation among genomic and biochemical marker variability and animal age. Altogether, the results show that during aging there is an increased sensitivity of kidney to chromate but not to HCBD‐induced damage and evidence differential age‐related selectivity of rats for nephrotoxic compounds. Significance for human risk assessment is discussed. Copyright © 2009 John Wiley & Sons, Ltd.     

Genetic overexpressing of GPx‐1 attenuates cocaine‐induced renal toxicity via induction of anti‐apoptotic factors

The present study investigates the role of the glutathione peroxidase (GPx)‐1 gene in cocaine‐induced renal damage in mice. Multiple doses of cocaine increased lipid peroxidation, protein oxidation, and glutathione oxidation in the kidney of the non‐transgenic mice (non‐TG mice). The enzymatic activities of GPx and glutathione reductase were significantly decreased in non‐TG mice, whereas superoxide dismutase was increased in the early phase of cocaine exposure. Treatment with cocaine resulted in significant decreases in expression of Bcl‐2 and Bcl‐xl in the kidney of non‐TG mice, which resulted in significant increases in Bax and cleaved‐caspase 3. Consistently, cocaine‐induced tubular epithelial vacuolization and focal tubular necrosis were mainly observed in the proximal tubules in the kidneys of non‐TG mice. These renal pathologic changes were much less pronounced in GPx‐1 TG than in non‐TG mice. These results suggest that the GPx‐1 gene is a protective factor against nephrotoxicity induced by cocaine via interactive modulations between antioxidant and cell survival signaling processes.     

Protective effects of emodin against cisplatin‐induced oxidative stress in cultured human kidney (HEK 293) cells

Emodin (a rhubarb anthraquinone) has strong antioxidant and anticancer actions, and recent studies indicated that it reduces cellular oxidative stress induced by various insults and drugs. Cisplatin is an anticancer drug that is associated with nephrotoxicity and induces oxidative stress in cultured human kidney (HEK 293) cells. This study aimed to assess the in‐vitro antioxidant properties of the emodin against cisplatin‐induced oxidative stress in HEK 293 cells. Our study revealed that emodin acted as a potent free radical scavenger and provided nephroprotection against cisplatin‐induced oxidative stress. Emodin as low as 0.5 µm did not decrease cell viability and restored the cisplatin‐induced glutathione depletion and total antioxidant capacity in a dose‐dependent manner. Emodin augmented the cisplatin‐induced inhibition of antioxidant enzymes (catalase, glutathione peroxidase, glutathione S‐transferase, glutathione reductase and superoxide dismutase). These results suggest that emodin has the potential to be used as an adjunct therapeutic agent in patients receiving cisplatin treatment. Copyright © 2012 John Wiley & Sons, Ltd.