Specificity of in vitro covalent binding of tienilic acid metabolites to human liver microsomes in relationship to the type of hepatotoxicity: comparison with two directly hepatotoxic drugs.

In order to better understand the first steps leading to drug-induced immunoallergic hepatitis, we studied the target of anti-LKM2 autoantibodies appearing in tienilic acid-induced hepatitis, and the target of tienilic acid-reactive metabolites. It was identified as cytochrome P450 2C9, (P450 2C9): indeed, anti-LKM2 specifically recognized P450 2C9, but none of the other P450s tested (including other 2C subfamily members, 2C8 and 2C18). Tienilic acid-reactive metabolite(s) specifically bound to P450 2C9, and experiments with yeast expressing active isolated P450s showed that P450 2C9 was responsible for tienilic acid-reactive metabolite(s) production. Results of qualitative and quantitative covalent binding of tienilic acid metabolite(s) to human liver microsomes were then compared to those obtained with two drugs leading to direct toxic hepatitis, namely, acetaminophen and chloroform. Kinetic constants (Km and Vmax) were measured, and the covalent binding profile of the metabolites to human liver microsomal proteins was studied. Tienilic acid had both the lowest Km and the highest covalent binding rate at pharmacological doses. For acetaminophen and chloroform, several microsomal proteins were covalently bound, while covalent binding was highly specific for tienilic acid and dihydralazine, another drug leading to immunoallergic hepatitis. Although low numbers of drugs were tested, these results led us to think that there may exist a relationship between the specificity of covalent binding and the type of hepatotoxicity.

Male rat hepatic UDP-glucuronosyltransferase activity toward thyroxine. Activation and induction properties--relation with thyroxine plasma disappearance rate.

Detergent-activation of UDP-glucuronosyltransferase (UGT) isoenzyme(s) involved in thyroxine (T4) glucuronidation in control, phenobarbital (PB)- and 3-methylcholanthrene (3-MC)-treated rats showed that between the four tested detergents, i.e. Triton X-100, Brij 58, Lubrol Px and 3-(3-cholamidopropyl)dimethylammonio-1-propanesulfonic acid (CHAPS), optimal activation of T4 UGT was displayed by the zwitterion CHAPS. "Native" versus optimal detergent-activated T4 UGT activity determination revealed that the latency of T4 UGT in microsomes from 3-MC-treated rats was decreased while the latency of T4 UGT in microsomes from PB-treated rats was increased compared to control, and suggest that the UGT isoenzyme(s) involved in the hepatic glucuronidation of T4 is (are) different in PB-treated rats than in 3-MC-treated rats. After a 7-day treatment with 20 mg/kg 3-MC, the activity of T4 UGT was increased 5-fold when determined in "native" and 4-fold when determined in optimal detergent-activated microsomes compared to controls. After a 7-day treatment with 75 mg/kg PB, T4 UGT was equivalent to the control when determined in "native", and increased 1.3-fold when determined in optimal detergent-activated microsomes. The results thus extend evidence that both 3-MC and PB induce the synthesis of UGT protein(s) involved in the glucuronidation of T4, 3-MC being a strong and PB a weak inducer. Hyperthyroid and hypothyroid status, achieved respectively by a 7-day treatment with 100 microns/kg T4 or a 7-day treatment with 10 mg/kg of one of the antithyroid drugs propylthiouracile or methymazole, did not modify T4 UGT activity, suggesting that the isoenzyme(s) conjugating T4 in microsomes from control rats is (are) unlikely to be either 4-nitrophenol or bilirubin UGT isoenzymes. After 14 days of treatment with 75 mg/kg PB, the hepatic glucuronidation rate of T4 was not different from the control when enzyme activity was expressed per mg microsomal protein but was significantly increased 1.4-fold when expressed per whole liver. A significant (1.5-fold) increase in the 125I-T4 plasma elimination rate was also observed in PB-treated rats compared to controls. A strong (3.6-fold) increase in the T4 glucuronidation rate was observed in rats treated with 5 mg/kg 3-MC for 14 days while the 125I-T4 plasma elimination rate was equivalent to the controls. These results demonstrate that there is no direct relation between T4 UGT activity (and subsequent biliary secretion of T4-glucuronides) and T4 plasma clearance and suggest an important contribution of the intestinal exchangeable thyroid hormone pool to the maintenance of blood thyroid hormone levels.

[The forest, the land, and man]. / Le bois, la terre et l'homme.

PIP: The faulty perception of the natural environment and cultural practices of the Sahel have led to inappropriate responses to the problems of providing firewood and of deforestation. The public rapidly became aware of damage caused by deforestation of the Sahel, but the tendency to attribute the damage to collection of firewood was overly simplistic. When desertification began to attract notice, there was little scientific, technical, or statistical knowledge of the dry zone forest. Research efforts were concentrated on the humid zone forests which were perceived to be potentially more lucrative. No systematic inventories of the dry zone forests or studies of their regeneration were available. In fact, under traditional cultivation techniques the clearing of the land did not cause destruction of the ecosystem because of its capacity for regeneration so long as the land was allowed to lie fallow. Most of the tree plantations installed by governments and nongovernmental organizations have failed because they were planted in overly dry zones, the species chosen where not appropriate for the climate, or the local populations were unable to care for them at times when their own time requirements for agriculture were greatest. In addition, the tree plantings did not provide the other foods and forest products utilized in cooking, curing, and trade. In traditional agriculture, a family plot of hectare or more is cleared, cultivated for 2-3 years, allowed to lie fallow for 8-9 years, and cultivated again for 2-3 years before being abandoned to a longterm fallow when the family moves on. Demographic growth in the region has led to a general shortening of fallow periods and cultivation of increasingly fragile and marginal zones. This factor, and pasturing of excessive numbers of animals, have been the main causes of desertification. New cultivation patterns and more appropriate tree plantings are urgently needed. Field studies should be conducted to help identify ecologically appropriate and socially accepted measures. If the current system is allowed to continue, catastrophic deforestation will ensue.^ieng

Effects on rats of subacute intoxication with deltamethrin via an osmotic pump.

To explore the possibility of liver enzyme induction by deltamethrin, subacute intoxication was carried out in rats for 28 days, by administration 7.2 mg.Kg-1.day-1 of deltamethrin i.p. delivered by an osmotic pump inserted in the peritoneal cavity. The body weight curve of the treated rats increased slightly but not significantly compared to the controls. No neurotoxic effect was observed. Blood parameters were unchanged, except for eosinophilia and an increase in the plasma Na+ level. Cytochrome P-450, cytochrome b5, NADPH-cytochrome c reductase, esterases and the activities of six mixed function oxidases were assayed. No variation was noted. Ultrastructural study of the liver, more specially in midlobular region, showed that deltamethrin increased the number of mitochondria and altered their shape which became irregular. These findings were consistent with morphometric results. Succinate cytochrome c reductase, citrate synthase and cytochrome c oxidase were essayed, only this last showed a significant enhancement in deltamethrin treated rats.

In vitro covalent binding of the pyrethroids cismethrin, cypermethrin and deltamethrin to rat liver homogenate and microsomes.

Phenobarbital-induced rat liver homogenate and microsomes were used to study covalent binding of 14C-labelled (at the alcohol moiety) cismethrin, 14C-labelled (at the alcohol and acid moieties) cypermethrin, and 14C-labelled (at the alcohol and acid moieties) deltamethrin. Covalent binding was dependent on pyrethroid concentration. With liver homogenate, inhibition of esterases by tetraethylpyrophosphate and of mitochondrial respiration by rotenone or potassium cyanide only slightly altered the covalent binding level. With microsomes, inhibition of cytochrome P-450 and mixed function oxidases by carbon monoxide and piperonyl butoxide reduced the covalent binding so far as to be nearly absent. Eighty percent inhibition of epoxide hydrolase decreased the covalent binding by 50%. The comparison of data between alcohol and acid labelling of the same pyrethroid suggested that, in vitro, the whole molecule is bound to proteins and that hydrolysis can occur afterwards. The experiments stress the role of cytochrome P-450-dependent monoxygenases in the covalent binding process.