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Biomedical subjects

A Bellocchio

Publications and source records attributed to A Bellocchio.

9 recordsLinked to original sources
PubMed2000

Effects of vitamin E on dolichol content of rats acutely treated with 1,2-dichloroethane.

Previous investigations have demonstrated that 1,2-dichloroethane (DCE) poisoning affects dolichol (Dol) concentration in rat liver. Dol, a long-chain polyprenol, is considered an important membrane component: as dolichyl phosphate, it is rate limiting for the synthesis of glycoprotein; as free or fatty acid, it is highly concentrated in the Golgi apparatus (GA) where it can increase membrane fluidity and permeability, required glycoprotein maturation and secretion. DCE biotransformation may stimulate pro-oxidant events through hepatocellular glutathione depletion. Since the molecules of Dol are susceptible to oxidative degradation, the aim of this investigation is to verify whether vitamin E (vit. E) supplementation in rats is able to prevent Dol breakdown during acute DCE treatment. Before acute DCE administration (628 mg/kg body weight), a group of male Wistar rats were pretreated with vit. E (33 mg/kg body weight) for 3 days. High-performance liquid chromatography analysis has shown that within 5-60 min after DCE administration, the Dol concentration decreased in liver homogenate, cytosol, microsomes and GA. Particularly, 60 min after the treatment, Dol levels in the trans Golgi fraction were 71% lower than in controls. Rat pre-treatment with vit. E prevented the DCE-induced decrease in Dol concentrations of all liver fractions considered, in particular the reduction of total-Dol observed in the trans Golgi fraction 60 min after treatment was only 40%. These data suggest that hepatic metabolism of DCE is able to promote peroxidative attacks which lead to the degradation of Dol molecules. The pre-treatment of rats with vit. E results in a good, although not complete, prevention of total-Dol depletion after DCE poisoning.

Animals↗
PubMed2000

Pseudodominance of lipoprotein lipase (LPL) deficiency due to a nonsense mutation (Tyr302>Term) in exon 6 of LPL gene in an Italian family from Sardinia (LPL(Olbia)).

We analyzed the molecular defect in the lipoprotein lipase (LPL) gene of a young boy from Sardinia who had primary hyperchylomicronemia, pancreatitis, and a complete LPL deficiency in post-heparin plasma. Analysis of LPL gene was performed by using single strand conformation polymorphism (SSCP) and direct sequencing of SSCP-positive region. The proband was homozygous for a C > A transversion in exon 6, which converts the codon for tyrosine at position 302 into a termination codon and eliminates an RsaI restriction site; this allowed the rapid screening of the proband's family members, among whom nine heterozygotes and one additional homozygote were identified. The homozygote was the proband's paternal grandmother who had shown the first clinical manifestation (recurrent pancreatitis) of LPL deficiency at the age of 54 years. LPL mutation carriers showed a mild dyslipidemic phenotype characterized by a reduction of high density lipoprotein-cholesterol (HDL-C) levels, HDL-C/total cholesterol ratio, and low density lipoprotein (LDL) size, associated with a variable increase of triglyceride levels. Five of these carriers were also heterozygotes for beta-thalassemia (Q39X mutation). In these double mutation carriers, plasma HDL-C levels were higher and plasma triglycerides tended to be lower than in carriers of LPL mutation alone. The Tyr302 > Term mutation encodes a truncated protein of 301 amino acids that is probably not secreted by the LPL producing cells. This is the first mutation of LPL gene found in Sardinians.

Apolipoproteins E↗
PubMed1998

1,1,2,2-Tetrachloroethane-induced early decrease of dolichol levels in rat liver microsomes and Golgi apparatus.

Dolichols are long-chain polyprenols containing 14-22 isoprene units, present in mammalian tissues as free dolichol (Free-Dol), fatty acyl dolichyl esters (Dol-FA), and dolichyl phosphate (Dol-P). The hepatic level of Dol-P seems to be a rate-limiting factor for glycosylation processes. Previous studies from our laboratory demonstrated the susceptibility of the dolichol molecule to undergo radical attacks. Since the toxicity of 1,1,2,2-tetrachloroethane (TTCE)is dependent on the free-radical production during hepatic biotrasformation, it was of interest to determine whether this haloalkane might affect glycosylation mechanisms by changing dolichol levels and distribution in rat liver microsomes and Golgi apparatus (GA). Male Sprague-Dawley rats received a single dose of TTCE (574 mg/kg body weight) and were then sacrificed at different times (5, 15, 30, or 60 min). In the TTCE-treated rats both serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities and hepatic triglycerides (TG) were significantly higher than control, while microsomal glucose 6-phosphatase (G6Pase) activity was decreased. In total microsomes Dol-P levels considered rate-limiting for the biosynthesis of the N-glycosylated proteins were significantly lower than in the control group 15 min after TTCE treatment. In normal rat liver, F1 secretory fraction of CA is 60-fold enriched in total dolichol content with respect to microsomes. In this compartment the total dolichol content, essential for the increase in membrane fluidity and permeability required for glycoprotein maturation and secretion, decreased significantly 5 min after TTCE treatment. Our results suggest that TTCE may affect dolichol functions in rat liver.

Animals↗
PubMed1998

Effect of ethanol administration on the level of dolichol in rat liver microsomes and Golgi apparatus.

Data obtained in our laboratory had suggested that acute ethanol administration (6 g/kg body weight) selectively and rapidly affects the intracellular system of protein glycosylation at the level of the Golgi apparatus. Dolichols are important membrane components, and dolichyl phosphate is a glycosyl sugar carrier for N-glycosylation of proteins in endoplasmic reticulum and is considered rate-limiting for this process. In this study, modifications in the concentration and distribution of liver microsomal dolichols after acute ethanol administration were investigated. Between 3 and 24 hr after ethanol administration, the microsomal dolichyl phosphate concentration was significantly lower than in control animals. The highest reduction was observed at 12 hr (-52%). An earlier and more marked reduction of total dolichol was observed in the Golgi apparatus, and, in particular, in the secretory fraction F1 (-70% at 6 hr). Ethanol treatment of isolated hepatocytes led to a significant reduction of the de novo synthesis of both dolichyl phosphate and free dolichol. Moreover, in vitro experiments have demonstrated that pro-oxidant agents lead to a significant decrease of both free dolichol and dolichyl phosphate. Our results suggest that acute ethanol administration induces a marked decrease of dolichols, probably by increasing the degradation and impairing the biosynthetic pathway of these molecules.

Animals↗
PubMed1997

Different efficacy of iodoacetic acid and N-ethylmaleimide in high-performance liquid chromatographic measurement of liver glutathione.

The widely used high-performance liquid chromatography (HPLC) procedure to determine glutathione in biological samples utilizing iodoacetic acid as thiol quenching agent and 1-fluoro-2,4-dinitrobenzene for derivatization has been modified regarding tissue sample processing and storage of the working solutions. The modified procedure compared with the original method reduces artifactual oxidation in rat liver glutathione measurement (1.47+/-0.8% vs. 2.84+/-0.69%, respectively). In both HPLC procedures, an increase in artifactual oxidation was found in both standard glutathione solutions and hepatic samples when N-ethylmaleimide instead of iodoacetic acid was used for thiol trapping.

Animals↗
PubMed1997

Effects of acute glutathione depletion induced by L-buthionine-(S,R)-sulfoximine on rat liver glucose-6-phosphatase activity.

The effect of acute glutathione (GSH) depletion induced by GSH-depleting agent L-buthionine-(S,R)-sulfoximine (BSO) on hepatic microsomal glucose-6-phosphatase (G6Pase) activity in male Wistar rats was investigated. Liver GSH evaluated in high-performance liquid chromatography after administration of 4 mmol.Kg-1 BSO i.p. was decreased by 19% and 50% at the time-points of 1.5 h and 3 h, respectively. In these conditions, a significant decrease in Vmax and an increasing trend in K(m) of hepatic G6Pase activity were observed, especially in 3 h BSO-rats. Alterations in kinetic parameters of G6Pase were calculated in both intact and detergent-treated microsomes, using glucose-6-phosphate and pyrophosphate as substrate. A little increase in thiobarbituric acid-reactive substances and a limited decrease in 5,5'-dithiobis(2-nitrobenzoate)-reactive protein thiols were also noted. The results of this study show that acute GSH depletion induced by BSO is able to affect hepatic microsomal G6Pase activity. A possible explanation to account for the effect of BSO-induced GSH depletion on hepatic G6Pase system is discussed.

Animals↗
PubMed1996

Effect of chronic ethanol consumption on glycosylation processes in rat liver microsomes and Golgi apparatus.

Previous studies have demonstrated that acute ethanol intoxication affects various steps of protein glycosylation at the level of rat liver endoplasmic reticulum and Golgi apparatus. The aim of this investigation was to demonstrate whether chronic ethanol intake can induce definitive changes of liver glycoprotein processing. Rats were given ethanol by liquid diet for 8 weeks. At the end of this period the triglyceride levels in liver homogenate and microsomes were significantly higher than in controls. Isolated hepatocytes prelabelled with [3H]Na palmitate and [14C]glucosamine showed a significant storage of the lipid and carbohydrate radioactivity in microsomes and Golgi apparatus and a significant impairment of labelled glycolipoprotein secretion. Changes of the glycosylation steps were observed both in endoplasmic reticulum and in Golgi apparatus: in the former the levels of dolichyl phosphate, which is rate-limiting for the synthesis of glycoprotein, showed a significant reduction; in the latter the activity of the main enzymes responsible for the terminal glycosylation process was significantly decreased. These data suggest that an impairment of glycoprotein maturation may be involved in the pathogenesis of liver injury induced by chronic ethanol intake.

Alcoholism↗
PubMed1993

Modulation of rat liver protein kinase C during "in vivo" CC14-induced oxidative stress.

Rat intoxication with a single dose of the hepatotoxin carbon tetrachloride induces a significant modification of liver protein kinase C total activity which depends on the degree of the intrahepatocyte oxidative unbalance provoked by various concentrations of the haloalkane. Low carbon tetrachloride amounts stimulate total protein kinase C activity, while one order of magnitude higher amounts exert strong enzyme inhibition. The latter effect is due to an early inactivation followed with progress of time by a proteolytic degradation of the enzyme. A pathological recruitment of the calcium-dependent protein kinase C regulatory enzymes calpain and calpastatin appears responsible for protein kinase C loss. The prolonged excess of cytosolic calcium which characterizes the single high dose carbon tetrachloride poisoning also leads to inactivation of calpain II and calpastatin in a time-dependent manner.

Animals↗