Differential effects of cisplatin on mouse hepatic and renal mitochrondrial DNA

The objective of this study was to determine if the nephrotoxic effects induced by cisplatin were correlated to mitochondrial DNA damage. Comparisons were made with the liver since hepatotoxicity is rarely observed. Cisplatin doses of 10, 20 and 40 mg/kg were administered intraperitoneally to C57BL/6J mice. Mitochrondrial DNA was isolated from both the hepatic and renal tissues and quantitated by hybridization with a specific mitochondrial probe. Cisplatin caused differential effects on mouse hepatic and renal mitochondrial DNA. The 10 and 20 mg/kg dose caused an elevation in mitochondrial DNA levels in the hepatic, but no increase in the renal tissue was observed. This is the first study demonstrating an organ specific effect of cisplatin at the DNA level.     

Induction of repairable DNA damage in Escherichia coli and interaction with DNA in vitro by the radical cation of chlorpromazine

Studies were performed to determine the DNA interactions of and the induction of cytotoxic effects by the radical cation (CPZ+.) formed enzymatically from chlorpromazine (CPZ): in the presence of native DNA the lifetime of CPZ+. is markedly increased. The decreased reactivity of CPZ+. in the presence of native DNA and the concomitant increased viscosity of CPZ+.-DNA complexes strongly support the assumption that CPZ+. does form intercalation complexes with DNA. The relative strong bacteriotoxicity of CPZ+. hindered the accurate determination of mutagenesis in various Salmonella indicator strains, but a test for repairable DNA damage in Escherichia coli using various repair-deficient strains indicated that the cytotoxic action of CPZ+. is in part due to DNA alterations which can be excised in wild-type DNA repair-proficient strains. After activation of CPZ with long wavelength UV light, genetic effects are observed in S. typhimurium strain TA98, as well as in the E. coli tester strains. The possible role of CPZ+. in the photosensitization of CPZ is discussed.     

Fibronectin turnover in human mesangial cell cultures as affected by Adriamycin

Fibronectin (FN) turnover and turnover changes induced by the anticancer drug Adriamycin (ADR) were measured in human mesangial cells (HMC) in vitro. HMC cultures synthesize cellular FN (2.2+-0.3% of totalprotein synthesis; n = 12) which is secreted and incorporated into a fibrillar extracellular matrix (ECM). A 24 hr incubation of HMC with ADR (0.5–5 g/ml) resulted in an accumulation of FN in the culture medium, with a maximum increase following 5 pglml(7.3+-2.3pg/cell vs. controls: 4.4+-1.9pg/cell; n= 10). Correspondingly, radioactively labeled immunoprecipitable FN was increased in a dosage-dependent manner in the culture medium up to 50% vs. controls. The incorporation of radioactively labeled FN into ECM was significantly increased following 2 g ADR/ml. In accordance, immunofZuorescence staining revealed an expansion ofpericellular FNfibers in cultures exposed to 2 g ADR/ml. Concomitant with the accumulation of extracelhlar FN, radioactively labeled FN in the cells was reduced by 22%. Qualitative characterization of FN patterns revealed a diminished number of degradation products in the culture medium ofADR-treated HMC. These data suggest thatADR interferes with the turnover of FN secreted by HMC in vitro in such a way that FN accumulates extracellularly. This in turn leads to a reduced FN synthesis. These findings are compatible with a loss of urinary FN degradation products accompanying the onset ofproteinuria in ADR-treated rats.Abbreviations ADR adriamycin - BSA bovine serum albumin - DTT dithiothreitol - ECM extracellular matrix - EDTA ethylenediamine tetraacetic acid disodium salt - ELISA enzyme-linked immunosorbent assay - FCS fetal calf serum - FITC fluorescein isothiocyanate - FN fibronectin - HMC human mesangial cell - PBS phosphate buffered saline - PMSF phenylmethylsulfonyl fluoride - SDS sodium dodecyl sulfate - SDS-PAGE SDS-polyacrylamide gel electrophoresis     

Characteristics of microsomal reduction of benzo[a]pyrene 4,5-oxide

NADPH-reduction of benzo[a]pyrene 4,5-oxide (BP-4,5-oxide) to BP required four components from rat liver: cytochrome P-450, NADPH cytochrome P-450 reductase, phosphatidylcholine and a soluble, heat-sensitive factor which was present in 105 000 × g supernatant and was also released from microsomes by sonication. The requirement for this factor contrasts with recently reported results from Sugiura et al. (Cancer Res., 40 (1980) 2910). Oxide-reduction was 40 times faster under anaerobic conditions, but oxygen did not affect the stimulation factor. This stimulation was highest (× 15) at low concentrations of microsomal protein (<0.1 mg/ml) and was almost absent at high concentrations of microsomal protein (>1 mg/ml). Oxide-reduction activity was proportional to microsomal protein concentration in the presence of added 105 000 × g supernatant, but for microsomes alone (>0.1 mg/ml) exhibited a parallel plot with an intercept at 0.08 mg/ml microsomal protein. Stimulation was highest at high concentrations of BP-4,5-oxide and a linear plot of V⁻¹ vs. [BP-4,5-oxide]⁻¹ was only obtained in the presence of 105 000 × g supernatant (K_m = 3 μM, V_max = 3.3 nmol/mg/min). Microsomal hydration of BP-4,5-oxide (inhibited in reductase assays) was unaffected by 105 000 × g supernatant, suggesting that stimulation of oxide-reduction did not derive from solubilization of BP-4,5-oxide. Stimulation was observed in the initial rate of reaction and was independent of incubation time. Inhibition of lipid peroxidation, removal of peroxides and deoxygenation were all excluded as explanations of the stimulatory effect.     

D(+) galactosamine induced oxidative and nitrosative stress-mediated renal damage in rats via NF-κB and inducible nitric oxide synthase (iNOS) pathways is ameliorated by a polyphenol xanthone,mangiferin

The present study investigated the possible protective effect of mangiferin against D(+) galactosamine (DGal)-induced nephrotoxicity. DGal intoxication increased reactive oxygen species (ROS), reactive nitrogen species and tumor necrosis factor-α (TNF-α) production and disturbed the antioxidant machineries in the kidney tissue. Mangiferin treatment post to DGal exposure reduced all these DGal-induced adverse effects. Signal transduction studies showed that DGal significantly increased the protein expression of Bax, cytochrome c, caspase 3/9 and inducible nitric oxide synthase (iNOS) in the cytosol and NF-κB in nuclear fraction. The same exposure, on the other hand, reduced the protein expression of Bcl-2 in the cytosol. Mangiferin treatment could, however, reduce the DGal-induced up-regulation of cytochrome c, NF-κB, iNOS, caspase 3/9 and alter the reciprocal regulation of Bcl-2 family proteins. Histological studies also revealed the nephroprotective effect of mangiferin against DGal induced nephrotoxicity. Combining, results suggest that mangiferin protects rat's kidney in DGal-induced oxidative/nitrosative stress and acute nephrotoxicity via its antioxidant activities.     

Reduction of carbon tetrachloride-induced nephropathy by melatonin administration

The aim of this study was to investigate possible protective effects of melatonin on carbon tetrachloride (CCl4)-induced renal damage in rats. A total of 24 animals were divided into three equal groups: the control rats received pure olive oil subcutaneously, rats in the second group were injected with CCl4 (0.5 ml kg-1, s.c. in olive oil) and rats in the third group were injected with CCl4 (0.5 ml kg-1) plus melatonin (25 mg kg-1, s.c. in 10% ethanol) every other day for 1 month. At the end of the experimental period, the animals were sacrificed and blood samples were collected. The kidneys were removed and weighed. Urea and creatinine levels were determined in blood samples. Histopathological examination of the kidney was performed using light microscopic methods. Administration of CCl4 significantly increased relative kidney weight (g per 100 g body weight) and decreased serum urea levels compared to controls (p<0.01). Melatonin treatment significantly (p<0.01) reduced relative kidney weight, and it produced a statistically equal (p=0.268) relative weight with the kidneys of control rats. CCl4 administration alone also caused histopathologically prominent damage in the kidney compared to the control group. Glomerular and tubular degeneration, interstitial mononuclear cell infiltration and fibrosis, vascular congestion around the tubules, and interstitial haemorrhage in perivascular areas were observed in the renal cortex and cortico-medullary border. However, the affect of CCl4 on the medulla was limited. Melatonin provided protection against CCl4-induced renal toxicity as was evident by histopathological evaluation. In view of the present findings, it is suggested that melatonin protects kidneys against CCl4 toxicity.     

Enantioselective metabolism of ifosfamide by the kidney

Ifosfamide (IF), a potent chemotherapeutic agent for solid tumors, is known to cause high rates of nephrotoxicity, which is most likely due to the renal production of the metabolite chloroacetaldehyde. Enantioselective oxidation of IF has been shown in the liver but has never been reported in the kidney. Using porcine and human kidney samples, as well as the renal porcine cell line LLCPK-1, we document enantioselective metabolism of IF with prevalent production of the N-dechloroethylifosfamide (DCEIF) metabolites from the (S)-IF enantiomer compared to the amount of N-DCEIF metabolites produced from the (R)-IF enantiomers. Since IF enantiomers appear to be equally effective in chemotherapy, these results suggest that replacing the clinically standard racemic mixture of IF with (R)-IF may decrease renal metabolism of the drug and hence may decrease nephrotoxicity.     

Tubular stress proteins and nitric oxide synthase expression in rat kidney exposed to mercuric chloride and melatonin.

Stress proteins such as HSP70 members (HSP72 and GRP75) and metallothionein (MT) protect the kidney against oxidative damage and harmful metals, whereas inducible nitric oxide synthase (iNOS) regulates tubular functions. A single dose of mercuric chloride (HgCl(2)) can cause acute renal failure in rats, its main target being the proximal tubule. Oxidative damage has been proposed as one of its pathogenic mechanisms. In this study we tested whether melatonin (MEL), a powerful antioxidant compound, is effective against HgCl(2) nephrotoxicity. Rats were treated with saline, HgCl(2) (3.5 mg/kg), MEL (5 mg/kg), and MEL + HgCl(2) and examined after 24 hr for HSP72, GRP75, MT, and iNOS by immunohistochemistry and immunoblotting. Tubular effects of the treatment were then characterized by ultrastructure. In the HgCl(2) group, all markers were overexpressed in convoluted proximal tubules and sometimes in distal tubules. In the MEL + HgCl(2) group, GRP75 and iNOS decreased in convoluted and straight proximal tubules, whereas HSP72 and MT persisted more than the saline and MEL-only groups. Tubular damage and mitochondrial morphometry were improved by MEL pretreatment. In conclusion, the beneficial effect of MEL against HgCl(2) nephrotoxicity was outlined morphologically and by the reduction of the tubular expression of stress proteins and iNOS. These markers could represent sensitive recovery index against mercury damage.     

Calcineurin regulation of cytoskeleton organization: a new paradigm to analyse the effects of calcineurin inhibitors on the kidney

Calcineurin is a serine/threonine phosphatase originally involved in the immune response but is also known for its role as a central mediator in various non-immunological intracellular signals. The nuclear factor of activated T cell (NFAT) proteins are the most widely described substrates of calcineurin, but ongoing work has uncovered other substrates among which are the cytoskeleton organizing proteins (i.e. cofilin, synaptopodin, WAVE-1). Control over cytoskeletal proteins is of outmost interest because the phenotypic properties of cells are dependent on cytoskeleton architecture integrity, while rearrangements of the cytoskeleton are implicated in both physiological and pathological processes. Previous works investigating the role of calcineurin on the cytoskeleton have focused on neurite elongation, myocyte hypertrophic response and recently in kidney cells structure. Nuclear factor of activated T cell activation is expectedly identified in the signalling pathways for calcineurin-induced cytoskeleton organization, however new NFAT-independent pathways have also been uncovered. The aim of this review is to summarize the current knowledge on the effects of calcineurin on cytoskeletal proteins and related intracellular pathways. These newly described properties of calcineurin on cytoskeletal proteins may explain some of the beneficial or deleterious effects observed in kidney cells associated with the use of the calcineurin inhibitors, cyclosporine and tacrolimus.     

Rosmarinus officinalis essential oil modulates renal toxicity and oxidative stress induced by potassium dichromate in rats

BACKGROUNDChromium hexavalent (CrVI) is known as a toxic contaminant that induced oxidative stress and nephrotoxicity in humans and animals. Rosmarinus officinalis is a perennial herb rich in biologically active constituents that have powerful antioxidant properties. So, the current work evaluated the effectiveness of Rosmarinus officinalis essential oil (REO) against alterations induced by potassium dichromate in the kidney of male rats.METHODSGC-MS analysis, in vitro total phenol contents, and DPPH scavenging activity of REO were estimated. Thirty-five Wistar male rats were categorized into 5 groups. The first group was the control, the second one was orally administered rosemary essential oil (REO; 0.5 mL/kg BW), the third group was injected intraperitoneally with hexavalent chromium (CrVI; 2 mg/kg BW) for 14 days, the fourth group used as the protective group (REO was administrated 30 min before i.p. injection of CrVI) and the fifth group applied as the therapeutic group (rats injected with CrVI 30 min followed by oral administration of REO), respectively.RESULTSTwenty-nine components were detected with high total phenolic contents and high DPPH scavenging activity. Results revealed that CrVI- intoxicated rats showed a valuable increase in oxidative stress profile (TBARS and H₂O₂) and a notable decline in glutathione (GSH), total protein content, and enzymatic antioxidants (SOD, CAT, GPx, and GST). Furthermore, serum kidney functions biomarkers (urea, creatinine, and uric acid) were increased significantly. Also, the administration of CrVI showed histological and immunohistochemical (PCNA-ir) changes in rat kidney tissue. Otherwise, administration of REO pre or post-treatment with CrVI significantly restored most of the biochemical parameters in addition to improvement in kidney tissue architecture. Moreover, individual intake with REO exhibited an amendment in oxidative stress markers.CONCLUSIONConclusively, REO had a potential antioxidant capacity in ameliorating K₂Cr₂O₇-induced nephrotoxicity, especially in the protection group.