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1.
Indian J Biochem Biophys ; 2010 Oct; 47(5): 265-271
Article in English | IMSEAR | ID: sea-135275

ABSTRACT

Microsomal glutathione transferase 1 (MGST1) is an integral homo-trimeric membrane protein with transferase and peroxidase activities. With glutathione as a co-substrate, it scavenges toxic compounds and may exert anti-apoptotic effect. We examined the effect of suppression of plasma membrane Ca2+-ATPase isoforms — PMCA2 or PMCA3 on MGST1 in PC12 cells. GSH level was significantly higher in PMCA2-reduced line, but similar GSSG/GSH ratios in all cell lines suggested an efficient protection or absence of oxidative stress. The ATP concentration decreased in both modified lines, although in PMCA2-suppressed cells the decrease was higher. Total GSTs activity in postmitochondrial fraction increased by 30% in the cells with reduced PMCA3. After treatment with MGST1 activator N-ethylmaleimide (NEM), the activity increased in both transfected lines by 30-40%. Real-time PCR also showed a higher mRNA expression of MGST1 in these lines. Staining with antibody recognizing all cytosolic and membrane-bound GSTs revealed the difference in oligomeric forms of GSTs, and specific anti-MGST1 antibody showed the presence of MGST1 hexamers in the transfected cells. Formation of similar hexamers was detected in the control line after treatment with peroxynitrite. Modification of MGST1 under reduced PMCAs amount may represent an adaptive mechanism that offers protection against the cytotoxicity mediated by increased Ca2+.


Subject(s)
Adaptation, Physiological/physiology , Animals , Enzyme Activation , Glutathione Transferase/metabolism , Microsomes/enzymology , PC12 Cells , Plasma Membrane Calcium-Transporting ATPases/metabolism , Rats
2.
Indian J Biochem Biophys ; 2010 Feb; 47(1): 44-48
Article in English | IMSEAR | ID: sea-135242

ABSTRACT

An SDS-PAGE analysis of renal microsomal fraction of albino mice was performed to study the involvement of proteins in dexamethasone-induced type-2 diabetes mellitus (DM) and their alterations by metformin, a widely accepted oral antidiabetic drug. In addition, changes in renal lipid peroxidation (LPO), activities of superoxide dismutase (SOD) and catalase (CAT), reduced glutathione (GSH) content, as well as renal somatic index (RSI) and daily rate of water consumption were also investigated. While dexamethasone administration (1.0 mg/kg for 21 days) expressed two renal proteins (43 kDa and 63.23 kDa), in addition to the increased fasting serum levels of glucose and insulin, renal LPO, RSI and daily rate of water consumption, a parallel decrease in renal SOD, CAT and GSH was also observed. Treatment with metformin normalized these alterations including the renal proteins and LPO, confirming its efficacy in ameliorating dexamethasone-induced type-2 DM and also the association of two proteins with type-2 DM.


Subject(s)
Animals , Catalase/metabolism , Dexamethasone , Diabetes Mellitus, Experimental/metabolism , Diabetes Mellitus, Type 2/metabolism , Glutathione/metabolism , Hypoglycemic Agents/pharmacology , Kidney/drug effects , Kidney/enzymology , Kidney/metabolism , Male , Metformin/pharmacology , Mice , Microsomes/drug effects , Microsomes/enzymology , Microsomes/metabolism , Radioimmunoassay , Superoxide Dismutase/metabolism
3.
Braz. j. med. biol. res ; 43(1): 52-56, Jan. 2010. tab, ilus
Article in English | LILACS | ID: lil-535636

ABSTRACT

Aldosterone concentrations vary in advanced chronic renal failure (CRF). The isozyme 11â-hydroxysteroid dehydrogenase 2 (11â-HSD2), which confers aldosterone specificity for mineralocorticoid receptors in distal tubules and collecting ducts, has been reported to be decreased or normal in patients with renal diseases. Our objective was to determine the role of aldosterone and 11â-HSD2 renal microsome activity, normalized for glomerular filtration rate (GFR), in maintaining K+ homeostasis in 5/6 nephrectomized rats. Male Wistar rats weighing 180-220 g at the beginning of the study were used. Rats with experimental CRF obtained by 5/6 nephrectomy (N = 9) and sham rats (N = 10) were maintained for 4 months. Systolic blood pressure and plasma creatinine (Pcr) concentration were measured at the end of the experiment. Sodium and potassium excretion and GFR were evaluated before and after spironolactone administration (10 mg·kg-1·day-1 for 7 days) and 11â-HSD2 activity on renal microsomes was determined. Systolic blood pressure (means ± SEM; Sham = 105 ± 8 and CRF = 149 ± 10 mmHg) and Pcr (Sham = 0.42 ± 0.03 and CRF = 2.53 ± 0.26 mg/dL) were higher (P < 0.05) while GFR (Sham = 1.46 ± 0.26 and CRF = 0.61 ± 0.06 mL/min) was lower (P < 0.05) in CRF, and plasma aldosterone (Pald) was the same in the two groups. Urinary sodium and potassium excretion was similar in the two groups under basal conditions but, after spironolactone treatment, only potassium excretion was decreased in CRF rats (sham = 0.95 ± 0.090 (before) vs 0.89 ± 0.09 µEq/min (after) and CRF = 1.05 ± 0.05 (before) vs 0.37 ± 0.07 µEq/min (after); P < 0.05). 11â-HSD2 activity on renal microsomes was lower in CRF rats (sham = 0.807 ± 0.09 and CRF = 0.217 ± 0.07 nmol·min-1·mg protein-1; P < 0.05), although when normalized for mL GFR it was similar in both groups. We conclude that K+ homeostasis is ...


Subject(s)
Animals , Male , Rats , /physiology , Homeostasis/physiology , Kidney Failure, Chronic/metabolism , Microsomes/enzymology , Potassium/metabolism , /metabolism , Aldosterone/blood , Blood Pressure/physiology , Kidney Failure, Chronic/enzymology , Nephrectomy , Rats, Wistar
4.
Mem. Inst. Oswaldo Cruz ; 103(6): 549-553, Sept. 2008. ilus, tab
Article in English | LILACS | ID: lil-495743

ABSTRACT

Benznidazole (Bz) and Nifurtimox (Nfx) have been used to treat Chagas disease. As recent studies have de-monstrated cardiotoxic effects of Nfx, we attempted to determine whether Bz behaves similarly. Bz reached the heart tissue of male rats after intragastric administration. No cytosolic Bz nitroreductases were detected, although microsomal NADPH-dependent Bz nitroreductase activity was observed, and appeared to be mediated by P450 reductase. No ultrastructurally observable deleterious effects of Bz were detected, in contrast to the overt cardiac effects previously reported for Nfx. In conclusion, when these drugs are used in chagasic patients, Bz may pose a lesser risk to heart function than Nfx when any cardiopathy is present.


Subject(s)
Animals , Male , Rats , Heart/drug effects , Myocardium/metabolism , Nifurtimox/pharmacokinetics , Nitroimidazoles/pharmacokinetics , Trypanocidal Agents/pharmacokinetics , Biotransformation , Drug Evaluation, Preclinical , Microscopy, Electron, Transmission , Microsomes/enzymology , Nifurtimox/adverse effects , Nitroimidazoles/adverse effects , Nitroreductases/analysis , Rats, Sprague-Dawley , Time Factors , Trypanocidal Agents/adverse effects
5.
Indian J Exp Biol ; 2007 Apr; 45(4): 390-7
Article in English | IMSEAR | ID: sea-62507

ABSTRACT

A genomic DNA sequence (fad2-1) encoding seed specific microsomal 0-6 desaturase was isolated from soybean (Glycine max. L cv. Pusa-9702). A positive genomic clone of 1852 nucleotides containing a single uninterrupted 3' end exonic region with an ORF of 1140 bp encoding a peptide of 379 amino acids, a complete 3' UTR of 206 bp and 86 bp of 5' UTR interrupted by a single intron of 420 bp was obtained on screening the sub-genomic library of soybean. Southern blots revealed at least two copies of the gene per haploid genome. Analysis of the translated product showed the presence of three histidine boxes, with the general sequence HXXXH and five probable transmembrane segments reported to be involved in substrate specificity.


Subject(s)
Amino Acid Sequence , Base Sequence , Blotting, Southern , DNA, Plant/analysis , Fatty Acid Desaturases/classification , Gene Dosage , Genes, Plant , Genome, Plant/genetics , Microsomes/enzymology , Molecular Sequence Data , Phylogeny , Soybeans/enzymology
6.
Article in English | WPRIM | ID: wpr-72669

ABSTRACT

Cyclosporin A (CsA)-induced hyperkalemia is caused by alterations in transepithelial K(+) secretion resulting from the inhibition of renal tubular Na(+), K(+) -ATPase activity. Thyroxine enhances renal cortical Na(+), K(+) -ATPase activity. This study investigated the effect of thyroxine on CsA-induced hyperkalemia. Sprague-Dawley rats were treated with either CsA, thyroxine, CsA and thyroxine, or olive-oil vehicle. CsA resulted in an increase in BUN and serum K(+), along with a decrease in creatinine clearance, fractional excretion of potassium, and renal cortical Na(+), K(+) -ATPase activity, as compared with oil vehicle administration. Histochemical study showed reduced Na(+), K(+) -ATPase activity in the proximal tubular epithelial cells of the CsA-treated compared with the oil-treated rats. Histologically, isometric intracytoplasmic vacuolation, disruption of the arrangement and swelling of the mitochondria, and a large number of lysosomes in the tubular epithelium were characteristic of the CsA-treated rats. Co-administration of thyroxine prevented CsA-induced hyperkalemia and reduced creatinine clearance, Na(+), K(+) -ATPase activity, and severity of the histologic changes in the renal tubular cells when compared with the CsA-treated rats. Thyroxine increased the fractional excretion of potassium via the preservation of Na(+), K(+) -ATPase activity in the renal tubular cells. Thus, the beneficial effects of thyroxine may be suited to treatment modalities for CsA-induced hyperkalemia.


Subject(s)
Animals , Cyclosporine/antagonists & inhibitors , Hyperkalemia/chemically induced , Immunosuppressive Agents/antagonists & inhibitors , Kidney Cortex/drug effects , Male , Microsomes/enzymology , Potassium/blood , Rats , Rats, Sprague-Dawley , Sodium-Potassium-Exchanging ATPase/metabolism , Thyroxine/pharmacology
7.
Acta physiol. pharmacol. ther. latinoam ; 49(3): 134-40, 1999. tab, graf
Article in English | LILACS | ID: lil-246051

ABSTRACT

The specific activity of long-chain acyl-CoA synthetase in microsomes from various tissues of control and calcium-deficient rats was determined. It was found that the saturated acids, palmitic and stearic, were preferential substrates compared to the non-saturated linoleic, alpha-linolenic and eicosa-8,11,14-trienoic acids. All of them showed similar Vm values with different affinity constants. After 60-day treatment on a calcium-deficient diet (0.5 g Ca/kg diet), a significant increase in the acyl-CoA synthetase activity was observed for all the tested fatty acids in liver and kidney microsomes. These changes were e voked without any modification in the substrate selectivities shown for the control microsomes, and they were well-correlated with calcium level in both tissues. Under the calcium deficient state an increase in Vm values was observed for palmitic and eicosatrienoic acids with no changes in the corresponding Km, suggesting an increment in the number of active enzyme molecules within the microsomal membrane.


Subject(s)
Animals , Rats , Female , Calcium/deficiency , Coenzyme A Ligases/metabolism , Kidney/cytology , Liver/cytology , Microsomes/enzymology , Brain/cytology , Myocardium/cytology , Rats, Wistar
8.
Indian J Biochem Biophys ; 1992 Oct; 29(5): 418-24
Article in English | IMSEAR | ID: sea-27787

ABSTRACT

The cytoplasmic localisation of cinnamic acid 4-hydroxylase (CA4H) has been shown by isolation and subcellular fractionation of the enzyme in Hepes buffer. The enzyme was purified by ammonium sulphate fractionation followed by AcA-34 molecular sieve chromatography. The enzyme existed as a high molecular mass which dissociated to a lower form on dilution on the column. The pH optimum, sulphydryl requirement, molecular and preliminary kinetic characteristics were investigated.


Subject(s)
Cations, Divalent , Cell Fractionation , Cytochrome P-450 Enzyme System/isolation & purification , Hydrogen-Ion Concentration , Kinetics , Microsomes/enzymology , Mixed Function Oxygenases/isolation & purification , Solanum tuberosum/enzymology , Subcellular Fractions/enzymology , Trans-Cinnamate 4-Monooxygenase
9.
Indian J Biochem Biophys ; 1991 Apr; 28(2): 136-9
Article in English | IMSEAR | ID: sea-26495

ABSTRACT

The microsomal membranes isolated from rat testes have been found to contain a Mg(2+)-dependent and a Mg(2+)-independent Ca(2+)-ATPase. The enzyme activities were inhibited by two contraceptive drugs--gossypol and chlorpromazine. The inhibition by the former was affected by the presence of ligand(s) and not the substrate in the incubation medium, whereas ligand(s)/substrate did not affect the inhibition by chlorpromazine. This may be explained from the fact that the binding of chlorpromazine and ligand(s)/substrate to the enzyme are independent of each other whereas in case of gossypol the ligand(s) compete with the drug at the binding site of the enzyme.


Subject(s)
Animals , Ca(2+) Mg(2+)-ATPase/metabolism , Calcium-Transporting ATPases/metabolism , Chlorpromazine/pharmacology , Gossypol/pharmacology , Intracellular Membranes/enzymology , Kinetics , Male , Microsomes/enzymology , Rats , Testis/enzymology
10.
Article in English | IMSEAR | ID: sea-20350

ABSTRACT

The influence of nutritional factors on aflatoxin B1 (AFB1)-induced liver tumours was investigated in rats. When a dose of 500 micrograms AFB1/kg body weight was given to rats in the absence of any anticarcinogen, 80 per cent of the rats developed liver tumours as compared to 0 to 40 per cent in those which received anticarcinogens. While beta-carotene totally inhibited the development of liver tumours ascorbic acid, selenium, and uric acid reduced the percentages of tumour-bearing rats to 13 per cent each. GSH and vitamin E also reduced these percentages to 20 and 40 per cent respectively. The reduction of tumour incidence by each anticarcinogen was associated with induction of increased microsomal enzyme activity. Inhibition of AFB1-induced liver cancer development thus seems to occur through microsomal enzyme induction and AFB1 activation.


Subject(s)
Aflatoxin B1 , Aflatoxins , Animals , Antioxidants/pharmacology , Carcinogens , Liver Neoplasms/chemically induced , Male , Microsomes/enzymology , Rats , Rats, Inbred Strains
11.
Indian J Biochem Biophys ; 1990 Apr; 27(2): 76-80
Article in English | IMSEAR | ID: sea-28845

ABSTRACT

Prostaglandin synthetase was immobilized by adsorption of goat vesicular microsomes on silica gel containing CaSO4 (silica gel G). Repeated cycles of enzymatic conversion of arachidonic acid to prostaglandin by the immobilized microsomes increased the product yield by 1.5 fold, in comparison to the same by free microsomal particles. The presence of Ca2+ in silica gel is responsible for this improved yield of prostaglandin as the divalent metal ion stabilized prostaglandin synthetase activity in a remarkable way. Microsomal particles immobilized on solid supports like alumina G and controlled pore glass were not very effective.


Subject(s)
Animals , Enzyme Stability , Enzymes, Immobilized/metabolism , Gels , Goats , Male , Microsomes/enzymology , Prostaglandin-Endoperoxide Synthases/metabolism , Semen , Silicon Dioxide
12.
Indian J Exp Biol ; 1989 Mar; 27(3): 217-9
Article in English | IMSEAR | ID: sea-61033

ABSTRACT

Effects of prostatic inhibin and thyroid releasing hormone (TRH) on lipid peroxidation in rat prostate was studied in an in vitro system. It was found that both inhibited the lipid peroxidase activity thus having a protective role in the prostate.


Subject(s)
Animals , Humans , Inhibins/pharmacology , Lipid Peroxidation/drug effects , Male , Microsomes/enzymology , Mitochondria/enzymology , Peroxidases/antagonists & inhibitors , Prostate/drug effects , Rats , Thyrotropin-Releasing Hormone/pharmacology
14.
Acta physiol. pharmacol. latinoam ; 38(1): 49-58, ene.-mar. 1988. tab
Article in English | LILACS | ID: lil-96488

ABSTRACT

La administración dietas hiperglucídicas e hiperproteicas suministradas a ratas durante 3 días produce respectivamente una disminución y un aumento en el cociente araquidonato/linoleato en los lípidos totales de microsomas de pulmón, riñon e hígado. En el hígado y el riñon este efecto está correlacionado con un significativo descenso de la actividad de la delta6 desaturasa para el caso de la dieta hiperflucídica y con un aumento de la misma actividad enzimática en la dieta hiperproteica. La actividad de la delta6 desaturasa, medida a través de la conversión del ácido 1-14**C linoleico a ácido alfa-linolénico, no se detectó en los microsomas de pulmón debido probablemente a la poca capacidad de este tejido para producir el éster de CoA del sustrato usado, y a que el cociente 20:4/18:2 en este tejido fue similar al del hígado bajo las condiciones dietéticas analizadas. La anisotropía de fluorescencia (r) del definilhexatrieno mostró diferencias significativas entre los tres tejidos analizados, efecto que se correlacionó con sus respectivos cocientes colesterol/fosfolípidos. Ambos parámetros fueron inferiores en los microsomas hepáticos que en los de los otros tejidos y permanecieron sin modificarse bajo los diferentes regímenes estudiados. Los resultados indican que el efecto de las dietas hiperhidrocarbonada e hiperproteica sobre la delta6 desaturasa no conduce a alteraciones aparentes en las propiedades físicas de las membranas microsomales


Subject(s)
Rats , Animals , Female , Fatty Acid Desaturases/metabolism , Fatty Acids, Unsaturated/metabolism , Dietary Carbohydrates/pharmacology , Liver/metabolism , Kidney/metabolism , Microsomes/metabolism , Dietary Proteins/pharmacology , Lung/metabolism , Fatty Acids, Unsaturated/analysis , Liver , Liver/enzymology , Kidney/drug effects , Kidney/enzymology , Lipids/analysis , Microsomes/drug effects , Microsomes/enzymology , Microsomes, Liver , Microsomes, Liver/enzymology , Microsomes, Liver/metabolism , Lung , Lung/enzymology , Rats, Inbred Strains
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