Increase in hepatic microsomal lipid peroxidation mediated by α-adrenergic system under cold stress and noradrenaline treatment

A membrane protein recognized by monoclonal antibody SQM1 was identified in human squamous carcinomas, including those originating in the head and neck (SqCHN), lung and cervix. Cell lines derived from SqCHN of previously untreated patients expressed high amounts of this protein. In contrast, many cell lines established from SqCHN of patients previously treated with chemotherapy and/or radiation showed diminished amounts of this SQM1 protein. The expression of SQM1 antigen was determined in several SgCHN cell lines made resistant by exposure to methotrexate (MTX) in vitro. The parent cell lines all exhibited strong binding to SQM1 antibody. The MTX-resistant sublines showed much lower membrane binding of SQM1. The lowest SQM1 reactivity was found in cell lines with high resistance to MTX and with diminished rate of MTX transport. Some highly MTX-resistant cell lines which had high levels of dihydrofolate reductase, but which retained a high rate of MTX transport, also retained high levels of SQM1 binding. Reduced SQM1 protein was also found in SgCHN cells which developed resistance to the alkylating drug cis-platinum (CDDP) and which showed reduced membrane transport of CDDP. Cell growth kinetics and non-specific antigenic shifts were not responsible for the differences in SQM1 binding between the parent cell lines and their drug-resistant sublines. The finding of a novel protein which is reduced in cells resistant to MTX and CDDP could contribute to our understanding of the basic mechanisms of drug resistance. By detecting SQM1 protein in clinical specimens, it may be possible to monitor the development of drug resistance in tumors.Abbreviations SqCHN Squamous Carcinoma of the Head and Neck - MTX Methotrexate - CDDP Cis-Platinum - DHFR Dihydrofolate Reductase     

Inhibition of mitochondrial oxidative phosphorylation by adriamycin

The antitumour antibiotic, adriamycin, inhibited oxidative phosphorylation in freshly prepared mitochondria from the heart, liver and kidney of the rat. It abolished respiratory control and stimulated ATPase activity. Succinate oxidation by heart mitochondria was extremely sensitive to the drug when hexokinase was present in the reaction medium. The sensitive site has been identified to lie in the region between the succinate dehydrogenase flavoprotein and ubiquinone of the respiratory chain.     

The metabolism of phenolic acids in the rat

Some of the enzyme systems in the formation of p-hydroxybenzoate from tyrosine have been studied in the rat liver in vitro. The conversion of p-hydroxycinnamate into p-hydroxybenzoate, which was found in rat liver mitochondria showed a number of differences when compared with the beta-oxidation of fatty acids. Studies with p-hydroxy[U-(14)C]cinnamate indicated that (14)CO(2) was released during the formation of p-hydroxybenzoate. The formation of p-hydroxycinnamate from tyrosine of p-hydroxyphenyl-lactate could not be demonstrated in vitro. The interconversion of p-hydroxycinnamate and p-hydroxyphenylpropionate was demonstrated in rat liver mitochondria.     

Ethanol-dependent oxygen consumption and acetaldehyde formation during vanadyl oxidation by H2O2

Sequential addition of vanadyl sulfate to a phosphate-buffered solution of H₂O₂ released oxygen only after the second batch of vanadyl. Ethanol added to such reaction mixtures progressively decreased oxygen release and increased oxygen consumption during oxidation of vanadyl by H₂O₂. Inclusion of ethanol after any of the three batches of vanadyl resulted in varying amounts of oxygen consumption, a property also shared by other alcohols (methanol, propanol and octanol). On increasing the concentration of ethanol, vanadyl sulfate or H₂O₂, both oxygen consumption and acetaldehyde formation increased progressively. Formation of acetaldehyde decreased with increase in the ratio of vanadyl:H₂O₂ above 2:1 and was undetectable with ethanol at 0.1 mM. The reaction mixture which was acidic in the absence of phosphate buffer (pH 7.0), released oxygen immediately after the first addition of vanadyl and also in presence of ethanol soon after initial rapid consumption of oxygen, with no accompanying acetaldehyde formation. The results underscore the importance of some vanadium complexes formed during vanadyl oxidation in the accompanying oxygen-transfer reactions.     

Vanadate stimulates oxidation of NADH to generate hydrogen peroxide

Vanadate in the polymeric form of decavanadate, but not other forms, stimulated oxidation of NADH to NAD⁺ NADPH was also oxidized with comparable rates. This oxidation of NADH was accompanied by uptake of oxygen and generated hydrogen peroxide with the following stoichiometry: NADH + H⁺ + O₂ → NAD⁺ + H₂O₂. The reaction followed second-order kinetics. The rate was dependent on the concentration of both NADH and vanadate and increased with decreasing pH. The reaction had an obligatory requirement for phosphate ions. Esr studies in the presence of the spin trap dimethyl pyrroline N oxide indicated the involvement of Superoxide anion as an intermediate. The reaction was sensitive to Superoxide dismutase and other scavengers of superoxide anions.     

Decrease of oxidative activities in brown adipose tissue mitochondria of cold acclimated rats on short term exposure to heat stress

Exposure of rats to the cold (4-5 degrees C) caused large (2-3-fold) increases in the mass of interscapular brown adipose tissue (BAT), its mitochondrial content and the basal metabolic rate of the animals. The rate of substrate oxidation by BAT mitochondria also increased about 3-fold. When cold-acclimated animals were exposed to heat (37 degrees C), the BMR decreased by half in 3 h, the earliest time interval tested. Mitochondrial substrate oxidation, as well as substrate-dependent H2O2 generation, showed a proportionate decrease in rates. In these mitochondria, activities of cytochrome c reductases, but not dehydrogenases with NADH, alpha-glycerophosphate and succinate as substrates, also showed a significant decrease. The concentration of cytochromes aa3 and b, but not cytochrome c, also decreased in BAT mitochondria from 12-h heat-exposed animals, while the change in concentration of cytochrome b alone was found as early as 3 h of heat exposure. These results identify the change in cytochromes as a mechanism of regulation of oxidative activities in BAT mitochondria under conditions of acute heat stress.     

Desensitization to ATP-Mg inhibition of 3-hydroxy-3-methylglutarylCoA reductase by heat treatment of microsomes

HMGCoA reductase is found to be inhibited by palmitylCoA and free CoA. The inhibition of this enzyme by ATP-Mg, but not by palmityl CoA, is lost on preincubation of microsomes at 50°C for 15 min.     

NADH-dependent decavanadate reductase, an alternative activity of NADP-specific isocitrate dehydrogenase protein

The well known NADP-specific isocitrate dehydrogenase (IDH) obtained from pig heart was found to oxidize NADH with accompanying consumption of oxygen (NADH:O(2)=1:1) in presence of polyvanadate. This activity of the soluble IDH-protein has the following features common with the previously described membrane-enzymes: heat-sensitive, active only with NADH but not NADPH, increased rates in acidic pH, dependence on concentrations of the enzyme, NADH, decavanadate and metavanadate (the two constituents of polyvanadate), and sensitivity to SOD and EDTA. Utilizing NADH as the electron source the IDH protein was able to reduce decavanadate but not metavanadate. This reduced form of vanadyl (V(IV)) was similar in its eight-band electron spin resonance spectrum to vanadyl sulfate but had a 20-fold higher absorbance at its 700 nm peak. This decavanadate reductase activity of the protein was sensitive to heat and was not inhibited by SOD and EDTA. The IDH protein has the additional enzymic activity of NADH-dependent decavanadate reductase and is an example of "one protein--many functions".