Sexually differentiated response to choline in choline deficiency and ethionine intoxication.

A sex difference exists in the response of rats to a choline deficient diet and to ethionine intoxication. Female rats are less susceptible than males to the acute effects of choline deficiency, such as fatty liver and impaired secretion of triglycerides into blood plasma, while they are more susceptible to inhibition of liver protein synthesis and triglyceride accumulation by ethionine. These differences have been ascribed to sex differences in the biosynthesis of phosphatidylcholine in the liver of rats. The available data indicate that females are more dependent than males on the stepwise methylation of phosphatidylethanolamine rather than the direct incorporation of preformed choline. Continuous prefeeding with choline for three weeks was able to shift the female pattern of response to choline deficiency and ethionine intoxication towards that observed in males; thus, choline caused accumulation of hepatic triglycerides and a decrease in plasma triglycerides after choline deficiency, while it protected against ethionine induced triglyceride accumulation and protein synthesis inhibition in the liver. These results suggest that choline prefeeding in females makes them more dependent on choline availability and, thus, more susceptible to a choline deficient diet and less sensitive to ethionine intoxication, as are males. No effect of choline was observed in either choline deficient or ethionine intoxicated male rats.

Relation between blood clearance and hepatic uptake of 99mTc-phytate in rats with hepatic injury.

The relation among the blood clearance of 99mTc-phytate (99mTc-P), the hepatic uptake of 99mTc-P and the severity of hepatic injury was investigated by using the rats with carbon tetrachloride (CCl4), D-galactosamine (Gal N), alpha-naphthylisothiocyanate (ANIT) or DL-ethionine (EthN) induced hepatic injury. After the administration of CCl4, GalN or ANIT, serum GPT activity increased significantly with the increase of dose level, and the degree of this increase was in the order: GalN greater than CCl4 greater than ANIT. However, the mild increase in serum GPT activity was observed after EthN administration. The blood clearance rate of 99mTc-P and the hepatic uptake ratio of 99mTc-P decreased with the increase of dose level after CCl4, GalN or ANIT administration, but significant changes were not found after EthN administration. The degree of decrease in the blood clearance rate of 99mTc-P was in the order: GalN not equal to CCl4 greater than ANIT, and the degree of decrease in the hepatic uptake ratio of 99mTc-P was in the order: GalN not equal to CCl4 greater than ANIT. These results suggest that the disorder in the hepatocytes may be one of causes for inducing the decrease in the hepatic uptake of 99mTc-P, and the consequence of this decrease may induce the decrease in the blood clearance of 99mTc-P.

Genetic analysis of L-ethionine-mediated induction of alpha-fetoprotein in mice.

Two genetic loci regulate hepatic alpha-fetoprotein (AFP) mRNA levels in adult mice. The raf locus controls basal levels and the Rif locus determines the degree of induction during liver regeneration. We have investigated the function of each locus during L-ethionine-mediated AFP induction using adult female mice with different Rif/raf genotypes. A single intraperitoneal injection of L-ethionine (0.5 mg/g body weight) resulted in significant triglyceride accumulation in hepatic parenchymal cells and increased AFP synthesis 48-96 h following injection. Hepatic AFP mRNA levels in Balb/cJ mice (high basal level/high induction level during regeneration) were 10- to 30-fold higher than Balb/cCRBL or C3H/He mice (low basal level/high induction level) following ethionine injection, indicating that raf-mediated differences persisted throughout the course of acute ethionine poisoning. The magnitude of this induction was similar to that seen during carbon tetrachloride-induced regeneration. In contrast, C57BL/6 mice (low basal level/low induction level during regeneration) contained hepatic AFP mRNA levels similar to Balb/cCRBL and C3H/He mice following ethionine injection. Thus, Rif-dependent differences seen during liver regeneration were not seen during acute ethionine poisoning. This leads us to conclude that (1) hepatic AFP mRNA induction by ethionine may not be mediated by the Rif locus if Rif is a transcriptional inducer, or (2) if Rif is a transcriptional repressor, it is inactivated equally in all strains during acute ethionine poisoning, unlike during liver regeneration. Hepatic albumin mRNA levels were not affected by ethionine treatment in vivo. L-Ethionine elevated AFP mRNA levels in primary mouse hepatocyte cultures; however, ethionine treatment also increased albumin mRNA levels in vitro.

Function of reticuloendothelial system on ethionine induced liver injury in mice.

Phagocytic activity as a function of the reticuloendothelial system (RES) has been studied in ethionine induced liver injury by using the carbon clearance test. Liver damage in male and female mice was induced by DL- and L-ethionine injections (1000 mg/kg/day, i.p.). In both female and male mice, a single dose or three injections of DL- or L-ethionine caused increases in liver/body weight ratio, A/G ratio, GOT and GPT levels, and BSP retention. There was the decrease of the total protein levels in the serum. The degree of liver injury was more severe after three injections of DL- and L-ethionine than after a single injection of them. After a single injection of ethionine, the L-isomer induced a slightly greater response than the racemic mixture, except for BSP retention. On the other hand, phagocytic activities by the carbon clearance test were increased after a single injection or three injections of DL- and L-ethionine. That is, the K value was increased in all ethionine treated mice except for females with three injections of DL-ethionine. The alpha value was increased after three injections in DL- and L-ethionine treated males and DL-ethionine treated females. In addition, the increase in carbon uptake by Kupffer cells can be seen by light microscopy after a single injection or three injections of DL- or L-ethionine. These findings indicate that ethionine injections induce the enhancement of RES phagocytosis, although the biochemical parameters indicating liver injury are changed severely. These results support the data indicating no correlation between the alteration of RES activity and the degree of liver injury.

[Methionine derivatives with liver protecting activity]. / Derivati della metionina ad attività epatoprotettrice.

Some methionine derivatives with the amine group acylated with an aliphatic group bearing a free, esterified or etherified thiol group in the omega position were prepared and tested for protection against CCl4, paracetamol, ethyl alcohol and ethionine intoxication. Acid (XIV) (MG 28013) (Table I) and some of its alkyl derivatives (XVIII) (MG 28228) and (XIX) (MG 28226) proved to have significant activity in these tests. A more in-depth study on MG 28226 however indicated a slight intolerance when subacute toxicity was examined (90 gg).

Therapeutic properties of dihydroxy-dibutylether on sub-acute liver damage induced by several hepatotoxic agents in rats.

The data presently reported show that repeated exposure of rats to allyl alcohol, ethionine or alpha-naphthyl isothiocyanate (ANIT) impaired some plasmatic parameters mainly by means of different mechanisms which involve the liver function. In particular, four administrations of allyl alcohol, given every other day, increased glutamate oxaloacetate transaminase (GOT) for at least 48 hours from the last dose; ethionine reduced the bromsulphthalein (BSP) clearance even after 48 hours from the 1st, 2nd, or 3rd administration given on the 1st, 5th, or 12th day; ANIT, administered daily for seven days, increased glutamate pyruvate transaminase (GPT), alkaline phosphatase (AP), bilirubin, triglycerides (TG) and cholesterol (CH) while it decreased the body-weight and retarded growth for at least six to seven days after intoxication. Twice daily administrations of dihydroxy-dibutylether (DHBE), a strong choleretic agent, brought to normality the parameters impaired by the three hepatointoxicating agents even when the intoxication was already established. In fact, DHBE reduced the plasma GOT levels increased by allyl alcohol, improved the BSP clearance impaired by ethionine and tended to normalize the parameters modified by ANIT by lowering GPT, AP, CH, TG and bilirubin plasma levels and by enhancing body growth. The curative activity of DHBE does not seem to be related only to a membrane stabilizing action since silymarin, a known cell membrane stabilizer, does not significantly influence the parameters described above in all the experimental conditions. Even the choleretic activity of DHBE alone might not be sufficient to explain its hepatoprotective action since fenipentol (a known choleretic agent) is inactive at least after ANIT intoxication.

Morphological and cytochemical alterations of the Golgi apparatus and GERL in rat parotid acinar cells during ethionine intoxication and recovery.

The present electron microscopic cytochemical investigation was undertaken to characterize the alterations in the golgi apparatus and GERL of rat parotid acinar cells during ethionine intoxication and recovery. Although the Golgi apparatus and GERL were reduced in size, and some broadening of the Golgi saccules occurred as the result of ethionine treatment, the relative localization of thiamine pyrophosphatase (TPPase) activity in the Golgi saccules, and acid phosphatase activity (AcPase) in GERL, remained unchanged. Shortly after ethionine treatment was stopped, a dramatic redistribution of enzyme activities was noted. Within the first 24 hours of recovery, the Golgi apparatus began to enlarge, and the content of secretory granules increased. By day 3 of recovery, cisternae morphologically identifiable as GERL and forming secretory granules possessed TPPase activity, while AcPase activity was virtually undetectable. After seven days of recovery, the Golgi apparatus and GERL appeared both morphologically and cytochemically normal. The enzyme modulation observed during recovery may be correlated with increased secretory granule production. Furthermore, the presence of TPPase activity in GERL and forming secretory granules lends support to the suggestion that GERL may be derived from the trans Golgi saccule.

Regulatory mechanisms of alpha-fetoprotein production in injured rat liver.

The role of alpha-fetoprotein (AFP)-suppressing factor(s) in the increased production of AFP in liver injury was investigated with carbon tetrachloride and ethionine-treated rats. Elevated concentrations of serum AFP induced by these treatments decreased upon administration of prednisolone. However, injections of methionine and adenosine 5'-triphosphate (ATP), which significantly accelerated the rapid fall of serum AFP levels after birth, produced no such effects on increased AFP levels in hepatic injury. This ineffectiveness of methionine and ATP may suggests that their metabolisms are impaired in the injured liver.

alpha-Fetoprotein in toxic liver injury.

The temporal sequence of alpha-fetoprotein appearance in serum was determined in both necrogenic and nonnecrogenic liver injury. Ethionine, thioacetamide, and CCl4 were used to intoxicate male and female rats for evaluating serum enzyme levels, mitotic indices, and morphological reflections of impairment. Thioacetamide- and CCl4-induced cell death preceded the mitotic wave in residual hepatocytes, and, in the case of both agents, this intoxicant-mediated necrosis preceded the emergence of alpha-fetoprotein. Yet, although there was no evidence of either cell destruction or significant mitotic activity in ethionine-poisoned animals, serum alpha-fetoprotein levels progressively increased. Thus the temporal sequence of alpha-fetoprotein synthesis and/or release and cellular reorganization for regeneration suggests that reappearance of the protein macro-molecule is an expression of the altered phenotype observed during the "step-down" phase of liver regeneration.

Rat liver isozymes in acute carbon tetrachloride and ethionine poisoning.

Changes in protein elution patterns and among others in the distribution profiles of some isozymes, as the lysosomal acid phosphatase, the microsomal alkaline phosphatase, the cytoplasmic fraction of aspartate aminotransferase and some fractions of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase, have been found in liver experimental fatty change induced in rats by carbon tetrachloride and ethionine. The possible meaning of these changes is discussed.