formazans and Kidney-Diseases

The kidney is one of the main targets of xenobiotic-induced toxicity, but early detection of renal damage is difficult. Recently, several novel biomarkers of nephrotoxicity have been identified by transcription profiling, including kidney injury molecule-1 (Kim-1), lipocalin-2, tissue inhibitor of metalloproteinases-1 (Timp-1), clusterin, osteopontin (OPN), and vimentin, and suggested as sensitive endpoints for acute kidney injury in vivo. However, it is not known if these cellular marker molecules may also be useful to predict chronic nephrotoxicity or to detect nephrotoxic effects in vitro. In this study, a panel of new biomarkers of renal toxicity was assessed via quantitative real-time PCR, immunohistochemistry, and immunoblotting in rats treated with the nephrotoxin ochratoxin A (OTA) for up to 90 days and in rat proximal tubule cells (NRK-52E) treated with OTA in vitro. Repeated administration of OTA to male F344/N rats for 14, 28, or 90 days resulted in a dose- and time-dependent increase in the expression of Kim-1, Timp-1, lipocalin-2, OPN, clusterin, and vimentin. Changes in gene expression were found to correlate with the progressive histopathological alterations and preceded effects on traditional clinical parameters indicative of impaired kidney function. Induction of Kim-1 messenger RNA expression was the earliest and most prominent response observed, supporting the use of this marker as sensitive indicator of chronic kidney injury. In contrast, no significant increase in the expression of putative marker genes and proteins were evident in NRK-52E cells after exposure to OTA for up to 48 h, suggesting that they may not be suitable endpoints for sensitive detection of nephrotoxic effects in vitro.

Topics: Animals; Biomarkers; Carcinogens; Cell Adhesion Molecules; Cell Line; Cell Survival; Dose-Response Relationship, Drug; Formazans; Gene Expression; Immunoenzyme Techniques; Kidney Diseases; Kidney Tubules, Proximal; Male; Membrane Proteins; Mycotoxins; No-Observed-Adverse-Effect Level; Ochratoxins; Rats; Rats, Inbred F344; RNA, Messenger; Tetrazolium Salts

Platinum (II) complexes with cyclobutanespiro-5'-hydantoin and cycloheptanespiro-5'-hydantoin were synthesized and evaluated by means of general physicochemical methods. The data from the elemental analysis, IR and NMR spectra suggested the formation of cis-[Pt(C6H8N2O2)2(NH3)2](NO3)2 x 4H2O (PtCBH), when cyclobutanespiro-5'-hydantoin was used as a ligand and cis-[Pt(C9H14N2O2)(NH3)2](NO3)2 x 4H2O (PtCHTH), when cycloheptanespiro-5'-hydantoin was used, respectively. The novel complexes exerted cytotoxic effects at micromolar concentrations against a panel of human tumor cell lines. They were found to trigger apoptosis in HL-60 and BV-173 cells as evidenced by DNA-laddering detection. The evaluation of the effects of PtCBH, PtCHTH and the antineoplastic drugs cisplatin and oxaliplatin against cultured murine kidney epithelial cells revealed that the hydantoin complexes were far less nephrotoxic in vitro.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cell Survival; Cyclobutanes; Cycloheptanes; Epithelial Cells; Formazans; HL-60 Cells; Humans; Hydantoins; Kidney Diseases; Magnetic Resonance Spectroscopy; Mice; Molecular Structure; Organoplatinum Compounds; Platinum; Spectrophotometry, Infrared; Spiro Compounds; Tetrazolium Salts

Fluoromethyl-2,2-difluoro-1-(trifluoromethyl)vinyl ether (FDVE) is a fluorinated alkene formed by degradation of the volatile anesthetic sevoflurane in anesthesia machines. FDVE is nephrotoxic in rats but not humans. Rat FDVE nephrotoxicity is attributed to FDVE glutathione conjugation and bioactivation of subsequent FDVE-cysteine S-conjugates, in part by renal beta-lyase. Although FDVE conjugation and metabolism occur in both rats and humans, the mechanism for selective toxicity in rats and lack of effect in humans is incompletely elucidated. This investigation measured FDVE S-conjugate cytotoxicity in cultured human proximal tubular HK-2 cells, and compared this with known cytotoxic S-conjugates. HK-2 cells were incubated with FDVE and its GSH, cysteine S-mercapturic acid, cysteine S-sulfoxide, and mercapturic acid sulfoxide conjugates (0.1-2.7 mM) for 24 h. Cytotoxicity was determined by lactate dehydrogenase (LDH) release, total LDH, and the ability of viable cells to reduce a tetrazolium-based compound (MTT). FDVE was cytotoxic only at concentrations >/=0.9 mM. No increase in LDH release was observed with either FDVE-GSH conjugate. The FDVE-cysteine conjugates S-(1,1-difluoro-2-fluoromethoxy-2-(trifluoromethyl) ethyl)-L-cysteine (DFEC) and (Z)-S-(1-fluoro-2-fluoromethoxy-2-(trifluoromethyl) vinyl)-L-cysteine ((Z)-FFVC) caused significant differences in LDH release and MTT reduction only at 2.7 mM; (Z)-FFVC was slightly more cytotoxic. Both S-(1,1-difluoro-2-fluoromethoxy-2-(trifluoromethyl) ethyl)-L-cysteine sulfoxide (DFEC-SO) and (Z)-N-acetyl-S-(1-fluoro-2-fluoromethoxy-2-(trifluoromethyl) vinyl)-L-cysteine sulfoxide ((Z)-N-Ac-FFVC-SO) caused slightly greater changes in LDH release or total LDH than the corresponding equimolar DFEC and (Z)-N-acetyl-S-(1-fluoro-2-fluoromethoxy-2-(trifluoromethyl) vinyl)-L-cysteine ((Z)-N-Ac-FFVC) conjugates. In contrast to FDVE S-conjugates, S-(1,2-dichlorovinyl)-L-cysteine was markedly cytotoxic, at concentrations as low as 0.1 mM. These results show that human proximal tubular cells are relatively resistant to FDVE and FDVE S-conjugate cytotoxicity. This may partially explain the lack of FDVE nephrotoxicity in humans.

Topics: Anesthetics, Inhalation; Cell Line; Cysteine; Ethers; Formazans; Glutathione; Humans; Hydrocarbons, Fluorinated; Kidney Diseases; Kidney Tubules, Proximal; L-Lactate Dehydrogenase; Methyl Ethers; Sevoflurane; Tetrazolium Salts