miR­148a­3p regulates alcoholic liver fibrosis through targeting ERBB3.

Alcoholic liver disease greatly affects human health. Previous studies have identified that microRNAs (miRNAs) are associated with the pathogenesis of alcoholic liver fibrosis (ALF). Therefore, the present study explored the regulatory mechanism of miR­148a­3p in ALF. An ALF model was established in rats by alcohol gavage, followed by treatment with miR­148a­3p. Reverse transcription­quantitative (RT­q) PCR was performed to detect miR­148a­3p expression in the rat liver tissues. The levels of lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine transaminase (ALT) and alkaline phosphatase (ALP) were determined by enzyme­labeled colorimetry. Liver damage was evaluated by liver indices and histology. The direct target gene of miR­148a­3p was predicted by a dual luciferase reporter assay. The effects of miR­148a­3p and miR­148a­3p in combination with receptor tyrosine­protein kinase erbB­3 (ERBB3) on HSC­T6 cell viability and apoptosis were detected by MTT and flow cytometry assays, respectively. Western blotting and RT­qPCR assays were performed to detect the expression levels of proteins and mRNA associated with fibrosis and apoptosis. The data showed that miR­148a­3p mimics inhibited the expression levels of AST, ALT, ALP, LDH, α­SMA and type I collagen in the model, decreased the liver indices, and improved the liver damage caused by alcohol. ERBB3, which was predicted as the direct target gene of miR­148a­3p, reversed the effects of ERBB3 on promoting cell viability and inhibiting apoptosis. Concomitantly, miR­148a­3p reversed the increased expression of Bcl­2 and inhibited the expression levels of Bax and c­cleaved­3 caused by ERBB3. These data suggested that miR­148a­3p regulated ALF and the viability and apoptosis of hepatic stellate cells through targeting ERBB3.

Identification and expression analyses of the alanine aminotransferase (AlaAT) gene family in poplar seedlings.

Alanine aminotransferase (AlaAT, E.C.2.6.1.2) catalyzes the reversible conversion of pyruvate and glutamate to alanine and α-oxoglutarate. The AlaAT gene family has been well studied in some herbaceous plants, but has not been well characterized in woody plants. In this study, we identified four alanine aminotransferase homologues in Populus trichocarpa, which could be classified into two subgroups, A and B. AlaAT3 and AlaAT4 in subgroup A encode AlaAT, while AlaAT1 and AlaAT2 in subgroup B encode glutamate:glyoxylate aminotransferase (GGAT), which catalyzes the reaction of glutamate and glyoxylate to α-oxoglutarate and glycine. Four AlaAT genes were cloned from P. simonii × P. nigra. PnAlaAT1 and PnAlaAT2 were expressed predominantly in leaves and induced by exogenous nitrogen and exhibited a diurnal fluctuation in leaves, but was inhibited in roots. PnAlaAT3 and PnAlaAT4 were mainly expressed in roots, stems and leaves, and was induced by exogenous nitrogen. The expression of PnAlaAT3 gene could be regulated by glutamine or its related metabolites in roots. Our results suggest that PnAlaAT3 gene may play an important role in nitrogen metabolism and is regulated by glutamine or its related metabolites in the roots of P. simonii × P. nigra.

Determination of enzymatic source of alanine aminotransferase activity in serum from dogs with liver injury.

INTRODUCTION: Increase of serum alanine aminotransferase (ALT) activity is widely used as a surrogate marker for tissue damage. Two ALT isoforms, ALT1 and ALT2, have been cloned recently in mammals. The study investigated the source of elevated ALT activity in serum of dogs treated with a hepatotoxic compound. METHODS: ALT activity was measured by enzyme assay. Immunoblot analysis was performed using generated specific peptide antibodies against dog ALTs. LC-MS/MS-based proteomics analysis was conducted to independently identify dog ALT peptides. Serum samples immunodepleted of major serum components by Seppro IgY-D11 microbead spin column were evaluated by the immunoblot analysis, and compared with those of the ALT activity. RESULTS: Involvement of ALT enzyme(s) is consistent with the following observations: 1) all the substrates (L-alanine and alpha-ketoglutarate) were required for serum ALT activity as purified porcine ALT1 needed for activity, 2) serum ALT activity was inhibited by L-cycloserine, a known ALT inhibitor, and 3) apparent Km value for the ALT reaction catalyzed by the serum, liver, and skeletal muscle was roughly similar. Immunoblot analysis showed that ALT1 was detected in liver and both ALTs were detected in the skeletal muscle. The relative expression level of ALTs was -: liver ALT1>skeletal muscle ALT1>skeletal muscle ALT2. LC-MS/MS-based proteomics analysis gave similar results. Immunoblot analysis of the depleted serum samples revealed the presence of ALT1 in compound-treated dogs. Intensity of the ALT1 band detected in the sera correlated well with the ALT activity measured by the enzyme assay. DISCUSSION: Based on these findings, we conclude that the elevation of serum ALT activity in dogs with liver injury is attributed to elevation of ALT1 protein level in serum. The methodology to directly detect ALT proteins in serum could be a tool to facilitate our understanding of biological and toxicological significance of the ALT isoenzymes.

Combination treatment of IFNalpha2b and ribavirin in patients with chronic hepatitis C and persistently normal ALTs.

Combination therapy of interferon-alpha2b and ribavirin was prospectively evaluated in 20 patients with chronic replicative hepatitis and persistently normal ALTs. Patients with normal ALTs on three or more occasions within 6 months received interferon-alpha2b 3 MU three times a week with ribavirin 1000-1200 mg everyday for 12 months and had a follow-up of 6 months. HCV genotype 1 was found in 16, and HCV genotype 2 or 3 in 4 patients. No patient experienced an ALT elevation during therapy. Ten of 20 patients (50%) cleared virus at the end of treatment. In an intent-to-treat analysis, a sustained virological response (SR) was achieved in 8 of 20 patients (40%). Nonresponse occurred in 5 patients. Relapse and breakthrough were seen in 2 patients each. Treatment was discontinued in 3 patients due to side effects. Interferon (IFN) ribavirin combination therapy is effective in patients with normal ALTs and appears superior to IFN monotherapy.

[National study of screening practices for hepatitis C among hemodialysis patients. Hepatitis Group]. / Etude nationale des pratiques de dépistage de l'hépatite C chez les patients hémodialysés. Hepatitis Group.

French and American consensus conferences on hepatitis C confirmed the burden of that disease, especially in high risk populations. In France, the seroprevalence of HCV is about 20% among haemodialysed patients. This study aimed at describing the French screening practices in haemodialysed patients. In 1995, 1213 self-administered questionnaires were sent to nephrologists working in 715 dialysis units. The response rate was 48% (585/1213) and 485 questionnaires were analysed. In 98% of questionnaires nephrologists answered that they prescribed screening test. Routine screening with alanine amino-transferase (ALT) was reported in 98% of questionnaires, usually once a month (57%) or four times a year (23%). Routine anti-HCV serology was reported by 96%, usually once (28%) or twice (46%) a year. The two main annual strategies combining ALT and anti-HCV serology were 12 ALT and 2 serologies (21%), or 12 ALT and 1 serology (14%) per year. HCV RNA detection was reported mainly in the case of positive anti-HCV serology (70%). The study suggested heterogeneity in screening practices and revealed the need to determine the cost-effectiveness ratios of the various strategies.

Serum amino acids, liver status, and antiepileptic drug therapy in epilepsy.

Serum amino acid (AA) profiles are altered in epilepsy. It is not clear whether this is due to the disease process itself or to other variables such as seizure type, seizure frequency, duration of illness, medication, or altered liver function. We investigated serum AA profiles and liver enzymes in 73 epileptic patients and 90 healthy subjects and evaluated the data by analysis of variance to discriminate between age, sex, seizure type, duration of illness, seizure frequency, antiepileptic drug (AED) and increased serum liver enzyme levels, and their putative interaction with the serum AA profile. There was no correlation between the changes in the AA profile and age, duration of illness, seizure frequency, and seizure type. Seventy-two percent of the AED-treated patients and 33% of the unmedicated patients showed an increase in one or several serum liver enzymes [alanine aminotransferase (ALT), aspartate aminotransferase (AST), and/or gamma-glutamyl transferase (gamma-GT)]; particularly gamma-GT. We observed a significant increase in serum concentrations of glutamine and glycine and decreased levels of taurine, threonine, serine, valine, methionine, isoleucine, leucine, phenylalanine, histidine, tryptophan, and arginine in AED-treated patients but not in unmedicated patients. These results show that the changes in the serum AA profiles of epileptic patients treated with AEDs occur in patients with alteration of serum liver enzymes; whether this implies a causal relation is still uncertain.