The effect of trichloroethylene metabolites on the hepatic vitamin B12-dependent methionine salvage pathway and its relevance to increased excretion of formic acid in the rat.

The industrial solvent trichloroethylene (TCE) and its two major metabolites trichloroethanol (TCE-OH) and trichloroacetic acid (TCA) cause formic aciduria in male F344 rats. Prior treatment of male F344 rats with 1-aminobenzotriazole a cytochrome P450 inhibitor, followed by TCE (16mk/kg, po), completely prevented formic aciduria, but had no effect on formic acid excretion produced by TCA (8 or 16 mg/kg, po), suggesting TCA may be the proximate metabolite producing this response. Dow and Green reported an increase in the concentration of 5-methyltetrahydrofolate (5-MTHF) in the plasma of rats treated with TCE-OH, suggesting a block in the cycling of 5-MTHF to tetrahydrofolate (THF). This pathway is under the control of the vitamin B12-dependent methionine salvage pathway. We therefore treated rats with three daily doses of methylcobalamin (CH3Cbl) or hydroxocobalamin (OHCbl), a cofactor for methionine synthase, or L-methionine, followed by TCE (16 mg/kg) to determine if they could alleviate the formic aciduria. These pretreatments only partially reduced the excretion of formic acid in the urine. Although prior treatment with S-adenosyl-L-methionine had no effect on formic acid excretion. Consistent with these findings, the activity of methionine synthase in the liver of TCE-treated rats was not inhibited. Transcriptomic analysis of the liver-identified nine differential expressed genes, of note, was downregulation of Lmbrd1 involved in the conversion of vitamin B12 into CH3Cbl, a cofactor for methionine synthase. Our findings indicate that the formic aciduria produced by TCE-OH and TCA may be the result of a block in the recycling of 5-MTHF to THF, the effect on the methionine salvage pathway being a secondary response following acute exposure.

Trichloroethylene and trichloroethanol-induced formic aciduria and renal injury in male F-344 rats following 12 weeks exposure.

Trichloroethylene (TCE) is widely used as a cleaning and decreasing agent and has been shown to cause liver tumours in rodents and a small incidence of renal tubule tumours in male rats. The basis for the renal tubule injury is believed to be related to metabolism of TCE via glutathione conjugation to yield the cysteine conjugate that can be activated by the enzyme cysteine conjugate ß-lyase in the kidney. More recently TCE and its major metabolite trichloroethanol (TCE-OH) have been shown to cause formic aciduria which can cause renal injury after chronic exposure in rats. In this study we have compared the renal toxicity of TCE and TCE-OH in rats to try and ascertain whether the glutathione pathway or formic aciduria can account for the toxicity. Male rats were given TCE (500mg/kg/day) or TCE-OH at (100mg/kg/day) for 12 weeks and the extent of renal injury measured at several time points using biomarkers of nephrotoxicity and prior to termination assessing renal tubule cell proliferation. The extent of formic aciduria was also determined at several time points, while renal pathology and plasma urea and creatinine were determined at the end of the study. TCE produced a very mild increase in biomarkers of renal injury, total protein, and glucose over the first two weeks of exposure and increased Kim-1 and NAG in urine after 1 and 5 weeks exposure, while TCE-OH did not produce a consistent increase in these biomarkers in urine. However, both chemicals produced a marked and sustained increase in the excretion of formic acid in urine to a very similar extent. The activity of methionine synthase in the liver of TCE and TCE-OH treated rats was inhibited by about 50% indicative of a block in folate synthesis. Both renal pathology and renal tubule cell proliferation were reduced after TCE and TCE-OH treatment compared to controls. Our findings do not clearly identify the pathway which is responsible for the renal toxicity of TCE but do provide some support for metabolism via glutathione conjugation.

Trichloroethylene-induced formic aciduria: effect of dose, sex and strain of rat.

The industrial solvent trichloroethylene (TCE) has been reported to increase the excretion of formic acid in the urine of male Fischer 344 (F-344) rats following large oral doses. We have examined the dose-response relationship for formic aciduria in male and female Fischer 344 rats, the effect of some known metabolites of TCE and examined the response in male Wistar rats to help understand its relevance to renal toxicity. We report that doses of TCE as low as 8 mg/kg for 3 days to both male and female F344 rats produced formic aciduria. The formic aciduria was time-dependent being more marked after 3 doses compared to one dose in male F344 rats and to a lesser extent in female F344 rats. TCE administration to male Wistar rats produced less formic aciduria than in male F344 rats, indicating a strain difference in response. As TCE is primarily metabolised by cytochrome P450 2E1, Wistar rats were administered inducers of cytochrome P450 2E1 followed by TCE, this increased formic acid excretion to a concentration similar to that observed in male F344 rats, indicating a role for P450. Administration of the major metabolites of TCE, trichloroethanol and trichloroacetic acid to male F344 rats also produced a marked and sustained formic aciduria, while the metabolite of TCE formed via glutathione conjugation had no effect on formic acid excretion. The mechanism whereby this response occurs is currently not understood, but the formic acid excreted is not a metabolite of TCE, but appears to be due to interference with the metabolic utilisation of formate by a down stream metabolite of TCE. Over the three days of the studies no histopathological evidence of kidney toxicity was observed in F344 rats given TCE, indicating that the perturbation of formate metabolism does not lead to acute renal injury.

Hyperglycemia and renal mass ablation synergistically augment albuminuria in the diabetic subtotally nephrectomized rat: implications for modeling diabetic nephropathy.

UNLABELLED: BACKGROUND/AIMSBACKGROUND/AIMS: While experimental models that emulate diabetic nephropathy are valuable tools for elucidating pathogenetic mechanisms and developing novel therapies, existing models imperfectly recapitulate human disease. In diabetes, hyperglycemia and hemodynamic forces act in concert to induce renal injury. Accordingly, in the present study, we combined streptozotocin-induced diabetes with surgical ablation of 5/6 of the kidney mass with the aim of evaluating their additive effects on renal function and glomerular morphology. METHODS: Female F344 rats were randomized to undergo subtotal nephrectomy (SNx) either at baseline or following 4 weeks of diabetes. RESULTS: In comparison to sham rats, rats with diabetes or rats after SNx surgery, diabetic subtotally nephrectomized (DM-SNx) rats demonstrated an increase in systolic blood pressure, glomerular volume and mesangial matrix. Albuminuria was synergistically increased by hyperglycemia and renal mass ablation associated with decreased nephrin expression. In contrast, glomerular capillary rarefaction and glomerular filtration rate were similarly reduced in SNx and DM-SNx rats. CONCLUSION: The DM-SNx rat recapitulates some of the features of human disease, most notably augmented albuminuria. Since this model avoids the deletion or overexpression of gene(s) linked to the pathogenesis of nephropathy, the DM-SNx rat model represents a complementary tool for the trial of novel therapies.