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S K Suneja

Publications and source records attributed to S K Suneja.

At least 19 recordsLinked to original sources
PubMed1992

Evidence that beta-hydroxyacyl-CoA dehydrase purified from rat liver microsomes is of peroxisomal origin.

The present study provides strong evidence that the previously isolated hepatic microsomal beta-hydroxyacyl-CoA dehydrase (EC 4.2.1.17), believed to be a component of the fatty acid chain-elongation system, is derived, not from the endoplasmic reticulum, but rather from the peroxisomes. The isolated dehydrase was purified over 3000-fold and showed optimal enzymic activity toward beta-hydroxyacyl-CoAs or trans-2-enoyl-CoAs with carbon chain lengths of 8-10. The purified preparation (VDH) displayed a pH optimum at 7.5 with beta-hydroxydecanoyl-CoA, and at 6.0 with beta-hydroxystearoyl-CoA. Competitive-inhibition studies suggested that VDH contained dehydrase isoforms, and SDS/PAGE showed three major bands at 47, 71 and 78 kDa, all of which reacted to antibody raised to the purified preparation. Immunocytochemical studies with anti-rabbit IgG to VDH unequivocally demonstrated gold particles randomly distributed throughout the peroxisomal matrix of liver sections from both untreated and di-(2-ethylhexyl) phthalate-treated rats. No labelling was associated with endoplasmic reticulum or with the microsomal fraction. Substrate-specificity studies and the use of antibodies to VDH and to the peroxisomal trifunctional protein indicated that VDH and the latter are separate enzymes. On the other hand, the VDH possesses biochemical characteristics similar to those of the D-beta-hydroxyacyl-CoA dehydrase recently isolated from rat liver peroxisomes [Li, Smeland & Schulz (1990) J. Biol. Chem. 265, 13629-13634; Hiltunen, Palosaari & Kunau (1989) J. Biol. Chem. 264, 13536-13540]. Neither enzyme utilizes crotonoyl-CoA or cis-2-enoyl-CoA as substrates, but both enzymes convert trans-2-enoyl substrates into the D-isomer only. In addition, the VDH also contained beta-oxoacyl-CoA reductase (beta-hydroxyacyl-CoA dehydrogenase) activity, which co-purified with the dehydrase.

Animals↗
PubMed1992

Depletion of rat hepatic glutathione and inhibition of microsomal trans-2-enoyl-CoA reductase activity following administration of a dec-2-ynol and dec-2-ynoic acid.

The effects of administration of dec-2-ynol and dec-2-ynoic acid on the hepatic glutathione (GSH) content and hepatic microsomal trans-2-enoyl-CoA reductase activity were examined in rat. Both compounds, when administered ip, caused a marked depletion of GSH levels and a corresponding inactivation of trans-2-enoyl-CoA reductase activity in both a time- and dose-dependent manner. The dec-2-ynoic acid caused greater hepatotoxicity than dec-2-ynol based on serum alanine transaminase activity. Based on the observations that (a) the alcohol did not interact with GSH in the presence or absence of cytosol, (b) the spectral manifestation of the interaction between GSH and the alcohol occurred only when NAD+ was added to the reaction mixture containing the cytosol and reactants, and (c) a similar absorbance spectrum was obtained following the interaction between aldehyde and GSH, it was concluded that dec-2-ynol is converted to an electrophile, dec-2-ynal, which causes depletion of GSH. The decrease in GSH content following administration of the acid appears to be due to activation of the acid to the electrophile, dec-2-ynoyl CoA, which then interacts with GSH, resulting in its depletion, based on the in vitro observations that (a) the acid did not interact with GSH in the presence or absence of cytosol, and (b) the spectral manifestation of interaction between GSH and dec-2-ynoyl CoA occurred both nonenzymatically and enzymatically in the presence of rat liver glutathione S-transferase (Sigma). Bovine serum albumin stimulated the enzymatic reaction. Comparable to the effects on GSH were the effects of dec-2-ynol, dec-2-ynal, dec-2-ynoic acid, and dec-2-ynoyl CoA on the microsomal trans-2-enoyl-CoA reductase activity in vitro. While the alcohol had no effect on the enzyme activity, its electrophilic product, the aldehyde, was a potent inhibitor. Similarly, the acid did not inhibit the enzyme activity unless the acid was present at high concentration; however, its electrophilic product, the CoA thioester, was a very potent inhibitor at very low concentration.

Acyl Coenzyme A↗
PubMed1992

The fatty acid chain elongation system of mammalian endoplasmic reticulum.

Much has been learned about FACES of the endoplasmic reticulum since its discovery in the early 1960s. FACES consists of four component reactions, requires the fatty acid to be activated in the form of a CoA derivative, utilizes reducing equivalents in the form of NADH or NADPH, is induced by a fat-free diet, resides on the cytoplasmic surface of the endoplasmic reticulum, appears to function in concert with the desaturase system and appears to exist in multiple forms (either multiple condensing enzymes connected to a single pathway or multiple pathways). FACES has been found in all tissues investigated, namely, liver, brain, kidney, lung, adrenals, retina, testis, small intestine, blood cells (lymphocytes and neutrophils) and fibroblasts, with one exception--the heart has no measurable activity. Yet, much more needs to be learned. The critical, inducible and rate-limiting condensing enzyme has resisted solubilization and purification; the purification of the other components has met with limited success. We know nothing about the site of synthesis of each component of FACES. How is each component enzyme integrated into the endoplasmic reticulum membrane? Is there a single mRNA directing synthesis of all four components or are there four separate mRNAs? How are elongation and desaturation coordinated? What is (are) the physiological regulator(s) of FACES--ADP, AMP, IP3, G-proteins, phosphorylation, CoA, Ca2+, cAMP, none of these? The molecular biology of FACES is only in the fetal stage of development. We are only scratching the surface--it is an undiscovered country.

Animals↗
PubMed1991

Do rat kidney cortex microsomes possess the enzymatic machinery to desaturate and chain elongate fatty acyl-CoA derivatives?

Rat kidney cortex microsomal preparations were unable to catalyze delta 9, delta 6 and delta 5 desaturation of stearoyl-coenzyme A (CoA), linoleoyl-CoA and dihomo-gamma-linolenoyl-CoA, respectively. The kidney cortex microsomal fraction, however, did catalyze the malonyl-CoA dependent fatty acyl-CoA elongation. The biochemical properties of palmitoyl-CoA elongation were studied as a function of protein concentration, time, reduced nicotinamide adenine dinucleotide phosphate (NADPH), malonyl-CoA and substrate concentrations; of the substrates investigated, delta 6,9,12-18:3 was the most active. Unlike what was observed in the hepatic system, a high-carbohydrate, fat-free diet did not induce kidney fatty acid chain elongation. All intermediate kidney cortex microsomal reactions, i.e., beta-ketoacyl-CoA reductase, beta-hydroxyacyl-CoA dehydrase and trans-2-enoyl-CoA reductase activities, were significantly higher (greater than one order of magnitude) than the condensing enzyme activity, suggesting that the rate-limiting step in total elongation is the initial condensation reaction. Contrary to other reports, the results suggest that the kidney cannot synthesize arachidonic acid needed for eicosanoid production.

Acyl Coenzyme A↗
PubMed1991

Decreased long-chain fatty acyl CoA elongation activity in quaking and jimpy mouse brain: deficiency in one enzyme or multiple enzyme activities?

Using long-chain fatty acyl CoAs (arachidoyl CoA and behenoyl CoA), a decrease in overall fatty acid chain elongation activity was observed in the quaking and jimpy mouse brain microsomes relative to controls. Arachidoyl CoA (20:0) and behenoyl CoA (22:0) elongation activities were depressed to about 50% and 80% of control values in quaking and jimpy mice, respectively. Measurement of the individual enzymatic activities of the elongation system revealed a single deficiency in enzyme activity; only the condensation activity was reduced to the same extent as total elongation in both quaking and jimpy mice. The activities of the other three enzymes, beta-ketoacyl CoA reductase, beta-hydroxyacyl CoA dehydrase, and trans-2-enoyl CoA reductase, in both mutants were similar to the activities present in the control mouse. In addition, the activities of these three enzymes were more than two to three orders of magnitude greater than the condensing enzyme activity in all three groups, establishing that the condensing enzyme catalyzes the rate-limiting reaction step of total elongation. When the elongation of palmitoyl CoA was measured, only a 25% decrease in total elongation occurred in both mutants; a similar percent decrease in the condensation of palmitoyl CoA also was observed. The activities of the other three enzymes were unaffected. These results support the concept of either multiple elongation pathways or multiple condensing enzymes.

Acetyltransferases↗
PubMed1990

Enzyme site-specific changes in hepatic microsomal fatty acid chain elongation in streptozotocin-induced diabetic rats.

The hepatic microsomal fatty acid chain elongation of palmitoyl-CoA and gamma-linolenoyl-CoA was diminished by 40-50% in male Sprague-Dawley rats made diabetic for 2 and 4 weeks following the intravenous administration of a single dose (65 mg/kg) of streptozotocin. Analysis of the activities of the four enzymatic components showed that only one enzyme, the condensing enzyme, which catalyzes the initial and rate-limiting step in chain elongation, was altered by the diabetic state. Both chain elongation and condensation activities were depressed to the same extent, whereas beta-ketoacyl-CoA reductase, beta-hydroxyacyl-CoA dehydrase and trans-2-enoyl-CoA reductase activities were the same as the values obtained with non-diabetic controls. 2 week administration of 10 units of insulin per day to rats which were diabetic for a 2-week period resulted in the reversal of the reduced palmitoyl-CoA elongation and condensation activities to control values. However, neither the condensation nor the elongation of gamma-linolenoyl was reversed by the insulin treatment. These results support the notion of multiple condensing enzymes or chain elongation systems.

Acyl Coenzyme A↗
PubMed1989

Evidence for two separate beta-ketoacyl CoA reductase components of the hepatic microsomal fatty acid chain elongation system in the rat.

The hepatic microsomal fatty acid chain elongation system can utilize either NADPH or NADH. Elongation activity, measured as the rate of malonyl CoA incorporation into palmitoyl CoA, was enhanced by a fat-free diet and by bovine serum albumin (BSA) when either cofactor was employed. When the intermediate products were determined, it was observed that in the presence of BSA and NADPH, the predominant product was the saturated elongated fatty acid, whereas in the presence of BSA and NADH, the major intermediate was the beta-ketoacyl derivative. Employing beta-ketostearoyl CoA as substrate, BSA markedly inhibited NADH-supported beta-ketoacyl CoA reductase activity and stimulated NADPH-supported activity. Furthermore, the sum of the NADH-dependent and NADPH-dependent beta-ketoreductase activities approximated the activity obtained when both cofactors were present in the incubation medium, suggesting the existence of two beta-ketoacyl CoA reductases, one using NADH and the other NADPH.

Acyl Coenzyme A↗
PubMed1989

Evidence for two separate beta-ketoacyl CoA reductase components of the hepatic microsomal fatty acid chain elongation system in the rat.

The hepatic microsomal fatty acid chain elongation system can utilize either NADPH or NADH. Elongation activity, measured as the rate of malonyl CoA incorporation into palmitoyl CoA, was enhanced by a fat-free diet and by bovine serum albumin (BSA) when either cofactor was employed. When the intermediate products were determined, it was observed that in the presence of BSA and NADPH, the predominant product was the saturated elongated fatty acid, whereas in the presence of BSA and NADH, the major intermediate was the beta-ketoacyl derivative. Employing beta-ketostearoyl CoA as substrate, BSA markedly inhibited NADH-supported beta-ketoacyl CoA reductase activity and stimulated NADPH-supported activity. Furthermore, the sum of the NADH-dependent and NADPH-dependent beta-ketoreductase activities approximated the activity obtained when both cofactors were present in the incubation medium, suggesting the existence of two beta-ketoacyl CoA reductases, one using NADH and the other, NADPH.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1989

Topography of rat hepatic microsomal enzymatic components of the fatty acid chain elongation system.

The orientation of the condensing enzyme, the beta-hydroxyacyl-CoA dehydrase, and the trans-2-enoyl CoA reductase within the rat liver microsomal membrane was investigated by the use of impermeant inhibitors of enzyme activity: trypsin, chymotrypsin, subtilisin, mercury-dextran, and anti-beta-hydroxyacyl-CoA dehydrase IgG. The activity of the condensing enzyme was inhibited more than 70% by various proteases and was completely inhibited by 80 microM mercury-dextran. Similar results were obtained for the trans-2-enoyl-CoA reductase activity. On the other hand, in the absence of detergent, proteases inhibited beta-hydroxyacyl-CoA dehydrase activity by 25-40%, while in the presence of detergent the inhibition increased to 65-90%. Furthermore, anti-beta-hydroxyacyl-CoA dehydrase IgG, which in the absence of detergent produced no inhibition, in the presence of detergent inhibited beta-hydroxyacyl-CoA dehydrase activity by more than 80%; under identical conditions, preimmune IgG caused a 13% inhibition. Microsomes used throughout this study displayed greater than 90% latency with respect to mannose-6-phosphatase activity, indicating that the microsomes were intact. Latency was not affected by the proteases, by mercury-dextran, or by the presence of the enzyme assay components. These results suggest that both the condensing enzyme and the reductase are present on the cytoplasmic surface of the membrane, whereas the beta-hydroxyacyl-CoA dehydrase is embedded in the microsomal membrane.

Acyl-CoA Dehydrogenases↗
PubMed1989

Spectrophotometric assay for the condensing enzyme activity of the microsomal fatty acid chain elongation system.

A rapid and simple spectrophotometric method was developed to measure the activity of the condensing enzyme component of the microsomal fatty acid chain elongation system. The intermediate product of the condensation reaction is the beta-ketoacyl CoA which exists in two tautomeric forms, i.e., keto and enol. The addition of bovine serum albumin (BSA) to a cuvette cell containing a beta-ketoacyl CoA derivative resulted in the formation of a 303-nm absorbance peak, characteristic of enolate formation. The beta-ketoacyl CoAs with carbon chain length of 6 to 18 interacted with BSA to produce the 303-nm peak; acetoacetyl CoA was the only beta-keto compound tested which did not interact with BSA to produce the peak. Other compounds which were unaffected by BSA included CoA, free beta-keto acid, beta-hydroxyacyl CoA, acyl CoA, trans-2-enoyl CoA, and malonyl CoA. BSA could not be replaced by ovalbumin; furthermore, denatured (boiling) BSA could not induce the 303-nm peak. The specific activity of the condensing enzyme measured by the spectrophotometric method compares favorably with the activity obtained by the radioactive method. The apparent extinction coefficient (epsilon) for the absorbance peak generated by the beta-keto thioester varied from 5 to 30 mM-1 cm-1 depending on the beta-keto derivative. The spectrophotometric procedure can be used in the determination of the condensing enzyme activity in not only hepatic microsomes but also in kidney and brain microsomes both of which have significantly lower activity. The advantages of the novel method over the radioactive method are that (i) it does not involve the use of radioactive compounds, (ii) it is much less cumbersome and significantly less costly, and (iii) it is rapid and easy to perform.

Animals↗
PubMed1989

Effect of oral administration of T-2 toxin on glutathione shuttle enzymes, microsomal reductases and lipid peroxidation in rat liver.

Effects of T-2 toxin on liver lipid peroxidation, glutathione shuttle enzymes and microsomal reductases have been studied in rats at 8, 16 and 24 hr after feeding a single dose of toxin (2.0 mg/kg) and at 7, 14 and 21 days after feeding of toxin (0.75 mg/kg) daily. Feeding of a single dose of T-2 toxin caused significant increase in liver lipid peroxidation in rats at 8, 16 and 24 hr post treatment. The liver lipid peroxidation was also significantly increased at 14 and 21 days after feeding of 0.75 mg/kg of T-2 toxin daily to rats. The activities of liver GSH-shuttle enzymes, i.e. glutathione peroxidase, glutathione reductase and glucose-6-phosphate dehydrogenase, were significantly higher in rats after both feeding schedules of T-2 toxin. NADPH-cytochrome c reductase activity was significantly lower at 8, 16 and 24 hr in liver of rats fed a single dose of T-2 toxin, whereas NADH-cytochrome b5 reductase was significantly higher until 16 hr and then declined below normal at 24 hr post treatment. In rats fed multiple doses of T-2 toxin, both liver microsomal reductases were significantly reduced. These results suggest that T-2 toxin/or its metabolites in the liver may be involved in the generation of free radicals which cause the observed increase in lipid peroxidation.

Administration, Oral↗
PubMed1989

Scintigraphy in evaluation of the hypoplastic right hepatic lobe: a rare variant.

A rare variant of right hepatic lobe hypoplasia associated with high gallbladder position in the right upper abdomen is described. Pain is frequent and may be due to cholelithiasis. It is important to recognize this variant because an associated hypertrophic left hepatic lobe can clinically masquerade as an abdominal mass. Radionuclide studies and abdominal computerized tomography are useful in defining the hypertrophied left hepatic lobe and ectopic gallbladder. The duodenum and hepatic flexure are positioned high due to space left by the hypoplastic right lobe.

Female↗
PubMed1989

Discrepant sulfur colloid and radioparticle liver uptake in superior vena cava obstruction: case report.

The presence of collateral venous channels connecting the upper extremity veins and portal vein via the paraumbilical veins is considered the probable explanation for the observed scintigraphic hepatic "hot spot". This is seen in [99mTc]sulfur colloid liver imaging and perfusion lung imaging with 99mTc radiolabeled particles injected into an antecubital vein in the presence of superior vena caval (SVC) obstruction. The typical distribution is one of focal uptake centrally, anteriorly, and inferiorly. An unusual pattern is described in this report and mechanisms proposed for the "diffuse homogeneous" hepatic uptake also observed in a patient with SVC obstruction undergoing a perfusion lung scan.

Aged↗
PubMed1987

Liver lipid metabolism in T-2 toxicosis. I. Effects of a single dose feeding of T-2 toxin to rats.

The acute effects of oral administration of a single dose of T-2 toxin (2.0 mg/kg body wt) to rats on whole liver lipid metabolism were studied at 8, 16 and 24 h post-treatment. Administration of T-2 toxin significantly increased liver and microsomal total lipids, free cholesterol, esterified cholesterol and triglycerides initially at 8 h, which subsequently returned to control values at 24 h. However, no significant alterations were observed in the contents of whole liver and liver microsomal total phospholipids and phosphatidyl choline, except that phosphatidyl ethanolamine and sphingomyelin + lysophosphatidyl ethanolamine contents in liver at 16 and 24 h and sphingomyelin + lysophosphatidyl ethanolamine content in liver microsomes at all three periods were significantly lower. The incorporation of 1-14C-acetate into whole liver and liver microsomal total lipids was reduced at 16 and 24 h post feeding. However, the incorporation of 1-14C-acetate into liver and microsomal free cholesterol, esterified cholesterol and triglycerides was significantly higher at 8 h, subsequently returning to the control value at 24 h; incorporation was significantly lower even into microsomal triglycerides. The incorporation of 1-14C-acetate into liver and its microsomal total phospholipids, phosphatidyl choline, phosphatidyl ethanolamine and sphingomyelin + lysophosphatidyl ethanolamine, was significantly decreased at all three periods post toxin treatment. The results suggested that T-2 toxin inhibited the incorporation of 14C-acetate mainly into liver and its microsomal phospholipids and their subfractions in rats.

Animals↗
PubMed1987

T-2 toxin induced changes in liver and serum enzymes of rats.

The effects on liver and serum enzymes of feeding a single dose (2 mg/kg) and daily doses (0.75 mg/kg) of T-2 toxin were studied in young male rats. Sample times were 8, 16 and 24 hr for single dose administration and 7, 14 and 21 days for daily dose administration. T-2 toxin in single and daily doses significantly reduced activities of hepatic glutamate pyruvate transaminase (GPT) and alkaline and acid phosphatases at all the sampling periods. In both feeding trials, levels of serum GPT increased, while that of acid and alkaline phosphatases significantly decreased at all the sampling times. This study indicates that the liver is affected by feeding T-2 toxin to rats.

Acid Phosphatase↗
PubMed1987

Effects of T-2 toxin gavage on the synthesis and contents of rat-liver macromolecules.

Effects of oral administration of T-2 toxin (0.75 mg/kg body weight/day) for 7, 14 or 21 days on the liver and plasma of young male rats were studied. A significant decrease in body weight and an increase in liver weight were observed in rats treated with T-2 toxin. Liver protein and glycogen levels were significantly lower than in controls after 21 days of treatment, but no significant differences were observed after 7 or 14 days. Levels of RNA in liver were significantly increased after 7, 14 and 21 days of treatment whereas liver DNA levels were significantly lower than in controls at each time interval. Liver microsomal protein was significantly decreased after 14 and 21 days, but microsomal RNA contents were significantly increased at 7 days and significantly decreased at 21 days. The levels of serum protein at 7, 14 and 21 days and of blood glucose at 14 and 21 days were significantly lower in T-2 toxin-treated rats. The levels of incorporation of [14C]leucine and [3H]uridine into liver protein and RNA, and into liver microsomal protein and RNA, were higher than in controls at 7 days, but then decreased. The incorporation of [3H]thymidine into liver DNA was not significantly altered in animals treated with the toxin.

Animals↗