The capacities of urinary trefoil factor 3 (TFF3) and urinary albumin to detect acute renal tubular injury have never been evaluated with sufficient statistical rigor to permit their use in regulated drug development instead of the current preclinical biomarkers serum creatinine (SCr) and blood urea nitrogen (BUN). Working with rats, we found that urinary TFF3 protein levels were markedly reduced, and urinary albumin were markedly increased in response to renal tubular injury. Urinary TFF3 levels did not respond to nonrenal toxicants, and urinary albumin faithfully reflected alterations in renal function. In situ hybridization localized TFF3 expression in tubules of the outer stripe of the outer medulla. Albumin outperformed either SCr or BUN for detecting kidney tubule injury and TFF3 augmented the potential of BUN and SCr to detect kidney damage. Use of urinary TFF3 and albumin will enable more sensitive and robust diagnosis of acute renal tubular injury than traditional biomarkers.

Mercuric ions are highly reactive and form a variety of organic complexes or conjugates in vivo. The renal proximal tubule is a primary target for mercury uptake and toxicity, and circumstantial evidence implicates organic anion transporters in these processes. To test this hypothesis directly, the transport and toxicity of mercuric-thiol conjugates were characterized in a Madin-Darby canine kidney cell line stably transfected with the human organic anion transporter 1 (hOAT1). 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl-terazolium bromide assays (for mitochondrial dehydrogenase) confirmed that mercuric conjugates of the thiols N-acetylcysteine (NAC), cysteine, or glutathione were more toxic in hOAT1-transfected cells than in the nontransfected cells. The NAC-Hg 2ϩ conjugate was most cytotoxic, inducing greater than 50% cellular death over 18 h at a concentration of 100 M. The cytotoxic effects were fully reversed by probenecid (an OAT1 inhibitor) and partially reversed by p-aminohippurate (an OAT1 substrate). Toxicity of this conjugate was reduced by the OAT1-exchangeable dicarboxylates ␣-ketoglutarate, glutarate, and adipate, but not by succinate, a nonexchangeable dicarboxylate.203 Hg-uptake studies showed probenecid-sensitive uptake of mercury-thiol conjugates in the hOAT1-transfected cells. The apparent K m for the NAC-Hg 2ϩ conjugate was 44 Ϯ 9 M. Uptake of the NAC-Hg 2ϩ conjugate was cis-inhibited by glutarate, but not by methylsuccinate, paralleling their effects on toxicity. Probenecid-sensitive transport of the NAC-Hg 2ϩ conjugate was also shown to occur in Xenopus laevis oocytes expressing the hOAT1 or the rOAT3 transporters, suggesting that OAT3 may also transport thiolHg 2ϩ conjugates. Thus, renal accumulation and toxicity of thiolHg 2ϩ conjugates may depend in part on the activity of the organic transport system.

In this study, approximately 40 endogenous metabolites were identified and quantified by (1)H NMR in urine samples from male rats dosed with two proximal tubule toxicants, cisplatin and gentamicin. The excreted amount of a majority of those metabolites in urine was found to be dose-dependent and exhibited a strong correlation with histopathology scores of overall proximal tubule damage. MetaCore pathway analysis software (GeneGo Inc.) was employed to identify nephrotoxicant-associated biochemical changes via an integrated quantitative analysis of both urine metabolomic and kidney transcriptomic profiles. Correlation analysis was applied to establish quantitative linkages between pairs of individual metabolite and gene transcript profiles in both cisplatin and gentamicin studies. This analysis revealed that cisplatin and gentamicin treatments were strongly linked to declines in mRNA transcripts for several luminal membrane transporters that handle each of the respective elevated urinary metabolites, such as glucose, amino acids, and monocarboxylic acids. The integrated pathway analysis performed on these studies indicates that cisplatin- or gentamicin-induced renal Fanconi-like syndromes manifested by glucosuria, hyperaminoaciduria, lactic aciduria, and ketonuria might be better explained by the reduction of functional proximal tubule transporters rather than by the perturbation of metabolic pathways inside kidney cells. Furthermore, this analysis suggests that renal transcription factors HNF1alpha, HNF1beta, and HIF-1 might be the central mediators of drug-induced kidney injury and adaptive response pathways.

Epithelial cells of the gut, antennal glands, integument, and gills of crustaceans regulate the movements of ions into and across these structures and thereby influence the concentrations of ions in the hemolymph. Specific transport proteins serving cations and anions are found on apical and basolateral cell membranes of epithelia in these tissues. In recent years, a considerable research effort has been directed at elucidating their physiological and molecular properties and relating these characteristics to the overall biology of the organisms. Efforts to describe ion transport in crustaceans have focused on the membrane transfer properties of Na+/H+ exchange, calcium uptake as it relates to the molt cycle, heavy metal sequestration and detoxification, and anion movements into and across epithelial cells. In addition to defining the properties and mechanisms of cation movements across specific cell borders, work over the past 5 yr has also centered on defining the molecular nature of certain transport proteins such as the Na+/H+ exchanger in gill and gut tissues. Monovalent anion transport proteins of the gills and gut have received attention as they relate to osmotic and ionic balance in euryhaline species. Divalent anion secretion events of the gut have been defined relative to potential roles they may have in hyporegulation of the blood and in hepatopancreatic detoxification events involving complexation with cationic metals.