Some aspects of functioning of the chain of electron transport in rat liver mitochondria in acute and chronic experimental hepatites

164

Poster Session 3B. Organ Toxicity

into the stomach through a gastric tube. Six, 12 and 24 hrs and 2.5 and 7 days after administration of methanol morphological picture and histochemical changes in the liver sections were screened. The lesions observed in the liver varied from dilated sinusoids with blood stasis through vacuolar degeneration in hepatocytes to microfocal changes in necrotic parenchymal cells. Phagocytic cells exhibited enhanced reaction. The intensity of the above changes depended on the time lapse since methanol administration. Increased acid phosphatase activity and reduced glycogen pool (pronounced most significantly 24 hrs after intoxication) seem to confirm temporary destruction of the liver cells.

D. Pach 1 , T. Gawlikowski *, B. Huszno 1 , D. Targosz. 1Department ofEndocrinology CM UJ; Department of Clinical Toxicology Collegium Medicum Jagiellonian University, Krakow; POLAND

MITOCHONDRIAL INJURYIN THE RATLIVER

G. Blaich *, A. Wey. Drug Toxicology, Knoll AG, D-67061 Ludwigshafen, Germany Valproic acid (VA), paraquat (PA) and methapyrilene (MP) were all shown to affect rat liver mitochondrial function. The objective of this study was to characterise these alterations in isolated rat liver mitochondria after in vivo treatment of male and female Wistar rats (4/group/sex) with VA (1% in the diet, 10 weeks), with PA (100 mg/kg bwt, single ip application) and MP (100 mg/kg bwt/day, po/gavage for 4 weeks). a-ketoglutarate (a-KG, site 1)and succinate (Sue, site Ilj-driven respiration from male animals are summarised in the following table <* = p ~ 0.05, Dunnett test; t/-l- = in-/decrease). PA treatment did not affect Sue-dependent respiration but a-KG-driven, ADP-stimulated and uncoupler-induced respiration was reduced. MP treatment resulted in a partial uncoupling of mitochondria (t of pretest and state 4, -l- of state 3, ADP/O ratio and acceptor control ratio (ACR) for site I and site H substrates). VA treatment disclosed only slight effects on respiration (-l- of ACR and t of uncoupling). VA a-KG

Pretest State 3 State 4 ACR

Uncoupling ADP/O-ratio

.J,

t

MP

PA Sue

a-KG

(.J,)

t'

.J,

.J, .J,'

t

Sue

IP3B52 I THE EVALUATION OF CARBOFURAN HEPATOTOXICITY IN ACUTE POISONING

IP3B50 liN VIVO STUDYON XENOBIOTICA-INDUCED

Parameter

and concentration dependent increase in fatty acyl CoA-oxidase (FACO). We also observed an increase in the number of peroxisomes In in vivo experiments, at doses of 58-465 mg/kg, and after treatment for 15, 30, 60 and 90 days, fomesafen induced an increase in FACO activity, in the number of hepatocyte peroxisomes, and in liver size; we also observed liver hyperplasia. Our results demonstrate that fomesafen behaves as a peroxisome proliferator, both in vivo and in vitro, same as other phenoxyacetic acid derivatives used as herbicides.

a-KG

Sue

r

t' t'

t' t

.J,' {' {'

t

't' " "

,j.'

Conclusion: PA-induced injury of the respiratory chain is located at complex I (eg only a-KG sensitive) whereas chronic MP administration resulted in disruption of energy supply due to uncoupling of oxidative phosphorylation. Chronic lO-week treatment with VA uncoupled only very slightly the respiratory chain.

I P3B51 liN VITROAND IN VIVOEFFECTS OF FOMESAFEN ON FATTY ACYL CoA OXIDASE (FACO) ACTIVITY IN RAT HEPATOCYTES

L. Orfila *, M.M. Salazar-Bookaman. Unidad de Cultivo Celular. Facultad de Fannacia, Universidad Central de Venezuela, Caracas, Venezuela

A large variety of agents of diverse chemical structure cause peroxisome proliferation. Fomesafen (5-(2-chloro-4-(trifiuormethyl)-phenoxy)-N-methylsulfonyl-2-nitrobenzamide) is a compound structurally related to herbicides of the phenoxyacetic acid family, with properties of peroxisome proliferator. After treatment with peroxisomes proliferation inducers several biochemical and histological changes are observed. We studied fomesafen effects in SpragueDawley hepatocytes primary culture and in H411EC3 hepatoma cells. At concentrations of 0.001-0. 1 mM, fomesafen induced a significant

An attempt to evaluate morphological and functional liver status in acute oral carbofuran poisoning using static scintigraphy, hepatography and measurement of chosen enzymes activity was undertaken in the study. Under analysis there were 24 carbofuran orally poisoned patients 4 women (16.7%) and 20 men (83.3%) treated in 1992-1997. Patients with liver or biliary tract diseases, chronic circulatory insufficiency symptoms, HBs antigen carriers and heavy drinkers were excluded from the study. The reference group comprised 29 healthy men not exposed to hepatotoxic agents. Results: Statistically higher activities of AST, ALT, AP, GTP and higher concentration of bilirubin were noted in the group of poisoned patients compared with the control group. The pathological scintigraphy and hepatohraphy results, dependent on age and poisoning severity, were found in all poisoned patients and their frequency was significantly higher than in the control group. Control scintigraphic examination revealed an improvement in the liver scintigraphic picture in 36.4% of patients. Deterioration was noted only in 1 (9.1%) patient. More than 61% of the examined patients showed serious liver lesion in hepatography. Analyzing the individual rates of the liver lesion an improvement of the liver function revaled by control hepatography was noted in 90.9% of examined patients. Considering arbitrary criteria for the degree of the liver lesion, the improvement in the liver function was noted in 36.4% of the examined patients. The same level of the liver lesion was noted in 1 person. Deterioration was not observed in any case. Conclusion: simultaneous liver scintigraphic examination (static and dynamic scintigraphy) and determination of marker enzymes activity allow to evaluate the liver morphological and functional condition in the course of carbofuran acute poisoning.

I P3B53 I SOMEASPECTS OF FUNCTIONING OF THE CHAIN OF ELECTRON TRANSPORT IN RAT LIVER MITOCHONDRIA IN ACUTE AND CHRONIC EXPERIMENTAL HEPATITES

V.N. Kovalenko *, LS. Blazhchuk. Institute ofPharmacology and Toxicology, Academy ofMedical Sciences, UA Influence of duration of action of hepatotoxic xenobiotics on rat liver mitochondrial respiratory processes were studied. It was shown that both acute poisoning with carbon tetrachloride, acetaminophen, tetracycline and prolonged treatment with carbon tetrachloride for 60 days and its combination with alcohol induced the increase of rate of ascorbate- and NADPH-dependent production of malonic dialdehyde in the endoplasmic reticulum and liver mitochondria. Acute liver injury was accompanied with the decrease of both succinate- and NADH cytochrome c oxidoreductase activities in liver mitochondria. Repeated treatment with carbon tetrachloride and its combination with alcohol induced the inhibition of NADH-dependent enzymes and the increase of the activity of succinate-related way of the chain

Poster Session 38. Organ Toxicity

of electron transport in mitochondria. That supposed metabolically adaptive conversion of respiratory chain in chronic chemically induced liver injury. Corrective effects oftwo a-tocopherol derivatives on those processes were shown.

IP3B54 I DEVELOPMENT OF HEPATIC FIBROSIS(CIRRHOSIS) MODEL IN RATS AND SCREENING OFTHE ANTIFIBROTIC AGENTS

Y.S. Lee *1, K.Y. Kim2 , B.S. Yu2 , K.S. Ki13 , J.Y. Jeong-', J.1. Cho 1 , J.H. Che1 . 1 Seoul National University; 2Wonkwang University; 3 Korea Food & Drug Administration, Republic of Korea

In the present study, we developed hepatic fibrosis(cirrhosis) model in Sprague-Dawley (SD) rats. Rats were subjected to common bile duct ligation/scission (BDUs) by orally administered with sodium diatrizoate (ethibloc) and were injected intraperitoneally 10 mglkg dimethylnitrosamine (DMN) three consecutive days a week for three weeks, respectively. Serum and liver tissues were prepared for measurement of procollagen type III propeptide (PIIINP) using enzyme liked immunosorbent assay (ELISA), total liver collagen (as hydroxyproline[HYP]), histopathological changes. BDUs rats were induced biliary fibrosis without inflammation, as characterized by the proliferation of bile ductule from portal triad, within two weeks and DMN treated rats were induced nodular fibrosis at five weeks, as characterized by the hyperaccumulation of fibrous components, infiltration of mononuclear leukocytes. Total liver HYP and PIIINP were significant increased and were found good correlations between total liver HYP and PIIINP. Therefore, we developed hepatic fibrosis (cirrhosis) model in SD rats. Next, to elucidate antifibrotic effect of herbal extract, we used BDUs with sodium diatrizoate rat hepatic fibrosis model. GB and SF have been used as hepatoprotective agents in Korea. But, its antifibrotic potential in chronic liver diseases has not been explored yet. Therefore, we applied GB and SF to BDUs with sodium diatrizoate rats for four weeks. Serum enzyme activity and total liver HYP in GB treated rats were less than those in untreated rats, whereas PIIINP value of GB treated rats were more increased than untreated rats. Histopathological changes were similar to that of untreated rats. However, SF treated rats caused 50% decrease in total liver HYP and PIIINP. At the same time, proliferation of bile ductule was decreased in SF treated rats. We therefore suggest that (1) BDUs with sodium diatrizoate is a suitable model to test the effect of antifibrotic drugs because it induces progressive rat secondary biliary fibrosis without major inflammation, (2) oral SF can ameliorate hepatic collagen accumulation in biliary fibrosis, and (3) PIIINP and HYP appear to be a suitable marker to monitor the inhibition of hepatic fibrogenesis in this model of biliary fibrosis.

IP3B55 I DISTURBANCE IN THE LIVERSTRUCTURE AND XENOBIOTIC METABOLIZING FUNCTION IN CHOLESTATIC INJURY: MECHANISMS AND CORRECTION L.F. Legonkova *, M.l. Bushma, V.E. Karpovich, LV. Zverinsky, N.G. Melnichenko, V.S. Nikitin. Institute ofBiochemistry Belarus, Grodno, Belarus

The common bile duct was ligated in anesthetized animals and they were administered with essentiale (I ml/kg, intragastrically, in starch mucilage) individually or in combination with nikethamide and a-tocopherol (50 mg/kg each) daily over 35 days. Control rats (laparotomy with and without common bile duct ligation) were injected with the same volume of starch mucilage. It was found that 36 days following the ligation of the bile duct the rat liver showed intensive regeneration of cholangia and cholangiols, microlobular cirrhosis and hepatocyte vacuolar and granular dystrophy. The cytochrome P450

165

content, the rates of NADPH oxidation and amidopyrine demethylation as well as the NADPH-cytochrome P450 reductase activity were decreased by 30-46%. The hemilurninescence "rapid burst" in liver microsomes was increased by 75%, where as the antioxidative activity - decreased by 41%. The administration of essentiale was accompanied by a reduced number of necroses and the intensity hepatocyte vacuolar dystrophy as well as increased glutathione S-transferase activity and normalized microsomal antioxidative activity. Following the administration of drug combination the liver structure was close to that in essentiale-treated rats, the metabolizing ability was considerably improved. The activities of microsomal and cytosolic glutathione S-transferases were in excess of the values in intact rats by 86 and 49% respectively. In cholestasis, the main role in the mechanisms of disturbances of the liver structure and function was played by the bile acids activation of lipid peroxidation and disturbed absorption of food hydrophobic antioxidants. The normalizing effect of the drug complex on the activities of the xenobiotic biotransformation enzyme systems and lipid peroxidation in cholestasis may be due to nikethamide enzyme-activiting effect the antioxidative properties of the drug components.

IP3B56 I MORPHOLOGICAL AND ULTRASTRUCTURAL PICTURE OF RATHEPATOCYTES IN FLUOR INTOXICATION

Ewa Dabrowska, Department of stomatology Medical Academy of Bialystok. Poland The aim of the study was to evaluate histopatological changes in the rat liver caused by the application of various fluor doses. The effect of fluor on liver tissue and the changes observed after discontinuation of fluor administration were investigated. The experiment used 115 Wistar rats. For 12 weeks the animals divided into 3 experimental groups received the following fluor (F) doses with drinking water. Fluor administration in aqueous solutions of sodium fluoride produces pathological changes in the rat liver directly proportional to the F dose applied.

IP3B58 I p-AMINOHIPPURIC ACID(PAH) ACCUMULATION IMPAIRMENT AFTERTREATMENT WITH SEGMENT-8PECIFIC NEPHROTOXICANTS

A. Trevisan *, P. Cristofori' , G. Fanelli, F. Bicciato, E. Stocco, M. Giraldo. Institute of Occupational Health, University of Padua. Padua; 1 Glaxo- Wellcome, Verona, Italy Hexachloro-l :3-butadiene (HCBD) and potassium dichromate (Cr6+) are chemicals toxic for the pars recta and the pars convoluta, respectively. PAH accumulation takes place in the basolateral membrane of the proximal tubule and its impairment is a sign of damage of the membrane. Segmentary localization of PAH accumulation is discussed, because different authors localize it in the pars convoluta only or in both. HCBD 100 mg/kg i.p. or Cr6+ 25 mglkg s.c, were injected in two months old male Wistar rats. Twenty four and 48 hours after treatment, rats were sacrificed and kidneys were removed and prepared to determine PAH accumulation in renal cortical slices; at the same time, histological evaluation of the kidney was performed. Table shows percent variation and statistical significance to the respects of the controls of PAH accumulation after treatment with both chemicals: 24 hours after treatment

48 hours after treatment

%

p

%

p

HCBD

92

n.s.

cr6+

68

< 0.01

64 41

<0.01 < 0.001