Biomedical subjects

E Mezey

Publications and source records attributed to E Mezey.

At least 109 records · Page 6Linked to original sources

PubMed1993

Two glycine transporter variants with distinct localization in the CNS and peripheral tissues are encoded by a common gene.

We have isolated a cDNA encoding a high affinity, Na+/Cl(-)-dependent glycine transporter, GLYT-2, which is distinct from another glycine transporter, GLYT-1. While the 3' sequences of these two cDNAs are identical, the 5' noncoding regions and the N-termini are completely different. GLYT-1 is found only in the white matter of the CNS, while GLYT-2 is found in the gray matter of the CNS as well as in macrophages and mast cells in peripheral tissues. Our findings suggest that tissue-specific alternative splicing or alternative promoter usage from a single gene results in two mRNA products encoding similar but distinct glycine transporters. The anatomic distribution of GLYT-2 mRNA supports the emerging status of glycine as a supraspinal neurotransmitter and suggests that glycine may function as a chemical messenger outside the CNS.

Amino Acid Sequence↗

PubMed1993

Treatment of alcoholic liver disease.

The treatment of alcoholic liver disease at present consists of abstinence from alcohol, bed rest, and dietary intake or administration of adequate amounts of calories and protein. Besides corticosteroids, which have been shown to improve hospital survival in severely ill patients with alcoholic hepatitis and liver transplantation in advanced cirrhosis, no successful specific therapy is available for alcoholic liver disease. Potential new therapeutic approaches include: (1) Treatment with specific dietary supplements such as polyunsaturated lecithin, which in baboons prevented the progression of the early stages of pericentral and interstitial fibrosis to septal fibrosis and cirrhosis; (2) antagonists to cytokines or antibodies to cytokine receptors for cytokines that have been shown to enhance hepatocellular necrosis or fibrosis; (3) substances that block pathways of oxygen radical formation or increase their metabolism or binding to form nonharmful compounds; (4) inhibition of collagen synthesis by proline analogues that increase intracellular collagen degradation or increase in collagen degradation by stimulation of collagenase or by insertion of exogenous DNA encoding amino or carboxyterminal peptides of procollagen into hepatocytes; and (5) stimulation of hepatic regeneration and recovery from alcohol-inducer liver injury.

Adrenal Cortex Hormones↗

PubMed1993

Effect of 4-(diethylamino)benzaldehyde on ethanol metabolism in mice.

The compound 4-(diethylamino)benzaldehyde (DEAB) is a potent inhibitor of cytosolic (class 1) aldehyde dehydrogenase (ALDH) in vitro and can overcome cyclophosphamide resistance in murine leukemia cells characterized by their high content of ALDH. In this study, we examined the in vivo effect of DEAB in mice on ethanol metabolism and on antipyrine clearance as a measure of the microsomal mixed function oxidase activity. DEAB administered in doses of 50 and 100 mg/kg increased the blood acetaldehyde concentration and decreased the plasma acetate concentration in mice treated with ethanol. A pharmacokinetic approach demonstrated that DEAB in doses of 50 and 100 mg/kg inhibited the fraction of ethanol converted to acetate by 32.5 and 67.5%, respectively. This inhibition was comparable with that produced by disulfiram. DEAB produced optimal inhibition of ALDH 10-15 min after administration. DEAB did not change the half-life or the total clearance of antipyrine. We conclude that DEAB is a potent inhibitor of ALDH in vivo and has no effect on the mixed function oxidase activity as determined by antipyrine clearance.

Acetaldehyde↗

PubMed1993

Gastric inhibitory polypeptide receptor, a member of the secretin-vasoactive intestinal peptide receptor family, is widely distributed in peripheral organs and the brain.

Gastric inhibitory polypeptide (GIP), or glucose-dependent insulinotropic peptide, is released from endocrine cells in the small intestine after meals. It is involved in several facets of the anabolic response and is thought to be particularly important in stimulating insulin secretion. We have cloned, functionally expressed, and mapped the distribution of the receptor for GIP. It is a member of the secretin-vasoactive intestinal polypeptide family of G-protein-coupled receptors. When expressed in tissue culture cells, it stimulates cAMP production (EC50 0.3 nM) and also increases intracellular calcium accumulation. GIP receptor mRNA is present in the pancreas as well as the gut, adipose tissue, heart, pituitary, and inner layers of the adrenal cortex, whereas it is not found in kidney, spleen, or liver. It is also expressed in several brain regions, including the cerebral cortex, hippocampus, and olfactory bulb. These results suggest that GIP may have previously undescribed actions. GIP receptor localization in the adrenal cortex suggests that it may have effects on glucocorticoid metabolism. Neither GIP nor its effects have been described in the central nervous system, and mRNA for the known peptide ligand for the receptor cannot be detected in the brain by in situ hybridization or polymerase chain reaction. This suggests that a novel peptide may be present in the brain.

Amino Acid Sequence↗

PubMed1993

Hormonal regulation of the rat class I alcohol dehydrogenase gene.

Expression of the rat liver (class I) alcohol dehydrogenase gene is regulated by hormones. Potential nuclear transcription factors of the rat class I alcohol dehydrogenase gene were identified. Proteins from rat liver nuclear extracts were found to bind to the alcohol dehydrogenase promoter at regions 1, 2, and 3, corresponding to positions -2 to -18, -36 to -44, and -52 to -60, relative to the start site of transcription. Region 1 was found to be bipartite and to interact with both the CCAAT/enhancer binding protein (C/EBP) and the liver activator protein (LAP) between position -10 and -22, while a different protein designated EDBP (for Enhancer-site Downstream Binding Protein) bound between position -1 and -10, a sequence previously not known to interact with any transcription factors. Region 2 has the sequence 5' CACCC-'3; binding to this sequence has been shown to act cooperatively with binding of the glucocorticoid binding site in the activation of various genes. The upstream stimulatory factor (USF) binds to region 3. C/EBP, LAP, and USF were all found to activate the alcohol dehydrogenase promoter in cotransfection experiments. It is postulated that one or more of these nuclear factors may mediate hormonal effects on the alcohol dehydrogenase gene.

Alcohol Dehydrogenase↗

PubMed1992

Localization of targets for anti-ulcer drugs in cells of the immune system.

The gastric mucosa consists of the epithelium, which lines the lumen, the lamina propria, and the muscularis mucosae. The targets of drugs used to treat stomach and duodenal ulcers are thought to be the acid-secreting parietal cells of the epithelium. However, immune cells in the lamina propria are the only cells that showed detectable messenger RNAs for histamine, muscarinic, gastrin, and dopamine receptors by in situ hybridization histochemistry. None of the epithelial cells expressed any of these messenger RNAs. Thus, the targets of antiulcer drugs seem to be cells of the immune system in the gut and not parietal cells, as generally believed. This conclusion may revise the thinking about ulcer formation and may shed light on the etiology of such chronic small intestinal diseases as Crohn's disease.

Animals↗

PubMed1992

Enhancer-site downstream binding protein activity is enriched in rat tissues that express the class I alcohol dehydrogenase gene.

The activity of the rat class I alcohol dehydrogenase (ADH) is enriched in certain tissues including the liver, intestine and testis. The tissue-specific expression of the gene encoding ADH in the rat was studied and found to closely correlate with tissue isozymic activity. A factor designated enhancer-site downstream binding protein (EDBP) was recently identified in the rat liver and found to interact with the proximal promoter of the class I ADH gene. The distribution of EDBP in nuclear extracts obtained from various tissues was examined based on its sequence-specific DNA binding property and found to correlate with tissue ADH expression. These findings suggest that EDBP is potentially a positive regulatory factor which is involved in controlling the tissue-specific expression of the ADH gene.

Alcohol Dehydrogenase↗

PubMed1992

Ethanol enhances ADP-ribosylation of protein in rat hepatocytes.

Decreases in hepatocyte NAD+ produced by ethanol are only partially explained by the increased conversion of NAD+ to NADH and NADP+. The purpose of this study was to determine whether a mechanism for the ethanol-induced decrease in NAD+ is its increased use in ADP-ribosylation. Exposure of hepatocytes in culture for 2 hr to 100 mmol/L ethanol increased the incorporation of 14C-ribose from prelabeled NAD+ into 14C-ribosylated proteins. Poly (ADP-ribose) polymerase activity was increased by exposure of isolated hepatocytes to 100 mmol/L ethanol for 10 min. In hepatocyte culture, increases in poly (ADP-ribose) polymerase were not detected after exposure to 100 mmol/L ethanol for 10 min or 2 hr but rather occurred at 24 hr. Ethanol exposure of hepatocytes in culture for 2 hr, however, decreased the Km for NAD+ of poly (ADP-ribose) polymerase. Both nicotinamide and 5-aminobenzamide, which are inhibitors of poly (ADP-ribose) polymerase, prevented the decrease in NAD+ produced by 2-hr exposure of hepatocytes in culture to 100 mmol/L ethanol. The effect of ethanol in decreasing DNA synthesis on days 3 and 4 of culture was not reversed by the inhibitors of poly (ADP-ribose) polymerase. These results indicate that increased ADP-ribosylation of hepatocyte proteins is a mechanism for the effect of ethanol in decreasing NAD+.

Adenosine Diphosphate Ribose↗

PubMed1992

The first 22 base pairs of the proximal promoter of the rat class I alcohol dehydrogenase gene is bipartite and interacts with multiple DNA-binding proteins.

The rat class I alcohol dehydrogenase (ADH) gene is primarily expressed in the liver. We previously showed that the liver-enriched transcription factor, the CCAAT/enhancer binding protein (C/EBP), binds to the proximal promoter of the rat class I ADH gene between positions -11 and -22 relative to the start site of transcription. We now demonstrate that another transcription factor, the liver activator protein (LAP), also interacts with the same region of the promoter based on the following observations: (1) LAP synthesized by in vitro transcription and translation of cloned cDNA sequence forms complexes with an oligonucleotide containing the C/EBP-binding sequence within the ADH promoter as determined by the electrophoretic mobility shift assay (EMSA), (2) purified LAP interacts with the proximal ADH promoter when analyzed by the DNase I protection assay, and (3) an ADH promoter-reporter gene construct containing the C/EBP-binding site is transactivated by an eukaryotic expression vector containing the LAP sequence. EMSA of an oligonucleotide containing the first 22 base pairs (between positions -1 and -22) of the ADH promoter with rat liver nuclear extracts (RLNE) resulted in the formation of two major complexes. Complex 1 was competed away by a heterologous oligonucleotide containing a C/EBP-binding site within the promoter of the adipocyte 422 (aP2) gene, while complex 2 was not. Additional competition experiments with the ADH or 422 (aP2) oligonucleotide using either RLNE or extracts from 3T3-L1 adipocytes demonstrated that complex 1 contains either C/EBP or LAP, while complex 2 contains a DNA-binding protein that binds to a novel sequence 5'-TGGCCCAGTT-3' between positions -1 and -10 of the ADH promoter. Ultraviolet cross-linking between RLNE and a labeled oligonucleotide containing the above sequence indicates that this protein, designated EDBP (for enhancer-site downstream binding protein), has an estimated molecular weight of 47 kDa, which is larger than that reported for either C/EBP (42 kDa) or LAP (36 kDa).

3T3 Cells↗

PubMed1992

Sex differences in gastric alcohol dehydrogenase activity in Sprague-Dawley rats.

The effects of sex differences and of fasting on gastric alcohol dehydrogenase activity were determined in Sprague-Dawley rats. Gastric alcohol dehydrogenase activity and enzyme protein levels were higher in female than in male rats. Ovariectomy and orchiectomy had no effect on alcohol dehydrogenase and did not alter the sex difference in enzyme activity. Fasting decreased the enzyme activity more in female than in male rats, abolishing the sex difference. Serum gastrin levels measured in female rats decreased on fasting and returned to normal levels within 24 hours of refeeding. Short- and long-term administration of pentagastrin to fasted and fed female rats did not affect the enzyme activity or enzyme protein level, except for a transient increase in enzyme activity but not in enzyme protein level 12 hours after administration to fasted fats. Omeprazole, which increased serum gastrin levels and decreased enzyme activity but not enzyme protein levels, was found to be a competitive inhibitor of the enzyme with a Ki of 0.40 mmol/L. The mechanisms for the sex differences and changes with fasting in rat gastric alcohol dehydrogenase activity remain unknown.

Alcohol Dehydrogenase↗

PubMed1991

Cloning of the cocaine-sensitive bovine dopamine transporter.

A cDNA encoding the dopamine transporter from bovine brain substantia nigra was identified on the basis of its structural homology to other, recently cloned, neurotransmitter transporters. The sequence of the 693-amino acid protein is quite similar to those of the rat gamma-aminobutyric acid, human norepinephrine, and rat serotonin transporters. Dopamine transporter mRNA was detected by in situ hybridization in the substantia nigra but not in the locus coeruleus, raphe, caudate, or other brain areas. [3H]Dopamine accumulation in tissue culture cells transfected with the cDNA was inhibited by amphetamine, cocaine, and specific inhibitors of dopamine transport, including GBR12909.

Amino Acid Sequence↗

PubMed1991

Cloning of a serotonin transporter affected by antidepressants.

A complementary DNA clone for a serotonin (5HT) transporter has been isolated from rat basophilic leukemia cells. The complementary DNA sequence predicts a 653-amino acid protein with 12 to 13 putative transmembrane domains. The 5HT transporter has significant homology to the gamma-aminobutyric acid, dopamine, and norepinephrine transporters. Uptake by CV-1 cells expressing the transporter complementary DNA resembles 5HT uptake by platelets and brain synaptosomes; it is sensitive to antidepressants, amphetamine derivatives, and cocaine.

Amino Acid Sequence↗

PubMed1991

The upstream stimulatory factor binds to and activates the promoter of the rat class I alcohol dehydrogenase gene.

The gene encoding rat class I alcohol dehydrogenase (ADH) is expressed primarily in the liver. Recent studies in our laboratories indicate that multiple cellular factors present in the rat liver interact with various regions of the promoter of this gene. One of the regions contains the sequence 5'-CACATG-3' that has an "E box" homology to which a number of transcription factors containing the basic helix-loop-helix motif bind. We now demonstrate that the human transcription factor, upstream stimulatory factor (USF), a basic helix-loop-helix-containing protein, binds to and activates the promoter of the rat class I ADH gene. Electrophoretic mobility shift assays of labeled oligonucleotide containing the 5'-CACATG-3' sequence within the ADH promoter revealed the formation of multiple DNA-protein complexes when nuclear extracts obtained from adult rat liver were used. The binding of proteins to the DNA could be competed away with an oligonucleotide specifying a sequence within the adenovirus major late promoter (MLP) that had previously been shown to bind USF. Similar complexes were observed when electrophoretic mobility shift assays of labeled MLP oligonucleotide were performed with rat liver nuclear extracts. Conversely, nuclear extracts isolated from HeLa cells, cells known to have abundant USF, contain factors that interact with the sequence present in the ADH promoter. This interaction could be competed efficiently by the MLP oligonucleotide. USF synthesized in an in vitro transcription and translation system also binds to the ADH promoter as well as to the MLP. In addition, antiserum directed against USF recognizes factors present in the rat liver nuclear extracts that interact with the ADH promoter. Furthermore, transcription directed from both the ADH and the adenovirus major late promoters was inhibited by an oligonucleotide representing the USF-binding site within the ADH promoter in a cell-free in vitro transcription system. Lastly, an ADH promoter-reporter gene construct was transactivated by an eukaryotic expression vector containing USF in HepG2 cells co-transfected with the two constructs. These experiments demonstrate that USF is present in the rat liver and that it binds to and activates the promoter of the rat class I ADH gene in a sequence-specific manner.

Alcohol Dehydrogenase↗

PubMed1991

The origin of somatostatin-containing nerve fibers innervating the hypothalamic supraoptic nucleus.

Light and electron microscopic studies were performed to study the connections between somatostatin (SOS)-containing nerve terminals and vasopressin (VP)-containing neurons in the rat supraoptic nucleus (SON). SOS-positive fibers innervate the SON in both the oxytocinergic and vasopressinergic areas. Using double immunostaining symmetric synaptic contacts were visualized between SOS immunoreactive boutons and the soma of VP immunopositive neurons. Surgical transection deafferentiating the SON from all possible directions do not effect the presence of SOS immunopositive fibers. These results suggest a local origin of the SOS fibers. Somatostatin-containing perikarya can indeed be found at the dorsal border of the SON at the rostral and caudal pole of the nucleus--we suggest that these cells innervate the SON. The presence of synaptic contacts between SOS fibers and VP neurons as well as the lack of these fibers in the VP deficient Brattleboro rats indicate a role for SOS in the synthesis and/or release of vasopressin in the SON.

Animals↗

PubMed1991

CCAAT/enhancer binding protein binds and activates the promoter of the rat class I alcohol dehydrogenase gene.

CCAAT/enhancer binding protein (C/EBP), a DNA binding protein originally isolated from rat liver, activates the transcription of a number of liver-specific genes. We studied the effect of C/EBP on the transcription of the rat class I alcohol dehydrogenase gene. Purified C/EBP was shown to bind to a segment of the rat class I alcohol dehydrogenase gene promoter between position -10 and -22. Nuclear extracts isolated from rat liver contained a heat stable factor(s) that also bound to this sequence. Cotransfection experiments in HepG2 cells showed transactivation of alcohol dehydrogenase promoter-CAT fusion constructs by an expression vector containing wild-type C/EBP. Expression vector containing a mutated C/EBP that failed to bind DNA also failed to activate the promoter. These experiments demonstrate that C/EBP expression can activate the rat class I alcohol dehydrogenase gene promoter.

Alcohol Dehydrogenase↗

PubMed1991

Effect of acute intravenous alcohol on plasma lipoproteins in man.

The effect of acute administration of ethanol was determined on plasma lipoproteins and post-heparin lipolytic activities in six normal subjects. Ethanol was administered intravenously (IV) in a dose of 0.6 g/kg body weight over 1 hour and blood samples were obtained for 4 hours thereafter. In four subjects, the determinations were repeated after the infusion of an isovolumetric amount of saline. Ethanol resulted in a rapid increase in plasma triglycerides, which persisted throughout the study. Ethanol did not result in changes in high-density lipoprotein (HDL)-cholesterol, low-density lipoprotein (LDL)-cholesterol, or apolipoprotein administration of heparin. The increase in triglycerides without changes in HDL-cholesterol after acute ethanol administration may contribute to an increased severity of cardiovascular disease in binge drinkers.

Adult↗

PubMed1991

Time dependent changes in CRF and its mRNA in the neurons of the inferior olive following surgical transection of the olivocerebellar tract in the rat.

Changes in corticotropin-releasing factor (CRF) immunostaining and CRF mRNA in neurons of the inferior olive were compared following unilateral surgical transections of the olivocerebellar tract. Alterations in CRF immunoreactivity could not be observed earlier than 24 h after surgery. The difference--an accumulation in the contralateral side--was most pronounced at 3 days, and disappeared by the 7th postoperative day. On the other hand, changes in mRNA could be observed as early as 5-30 min after the transection. The most significant accumulation was present at 3 h after the cut and no difference could be observed from the 3rd day following surgery. The results suggest that changes in mRNA levels by axotomy may occur at an earlier time point than previously thought.

Animals↗

PubMed1991

Activity of the beta-retinoic acid receptor promoter in transgenic mice.

LacZ reporter gene constructs were used to analyze the murine retinoic acid receptor beta (mRAR beta) gene promoter in transgenic mice. LacZ expression in transgenic mouse embryos with 250 bp of promoter sequences closely parallels that of RAR beta between embryonic days 8.5 and 12.5. This indicates that the -1 to -250 promoter region contains most regulatory elements required for tissue specific expression. Additional elements in the -250 to -625 region are required for high expression levels after day 12.5. Elements in the -625 to -3100 region are necessary to reproduce the RAR beta expression in the meninges and the eye mesenchyme. The expression pattern of the transgene and the endogenous RAR beta, as revealed by in situ hybridization, suggests an important role of the RAR beta in the developing nervous system.

Amino Acid Sequence↗