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Biomedical subjects

M D Maines

Publications and source records attributed to M D Maines.

At least 127 records · Page 7Linked to original sources
PubMed1983

Biliverdin reductase: characterization in the rat kidney and the inhibition of activity by mercuric chloride.

The effects of metal ions on the activities of biliverdin reductase in the rat kidney and liver were examined; the pH optimum and the cofactor requirement for the enzyme activity in the kidney were also studied. The reduction of biliverdin IX alpha by biliverdin reductase in the rat kidney cytosol fraction could be supported by NADPH and NADH. The activity was optimal around pH 8.7 when NADPH was the cofactor. The activity with NADH was undetectable at this pH. NADH-dependent biliverdin reductase was optimal at pH 7.0, where the NADPH-dependent activity was negligible. Biliverdin reductase activity was not inducible in the kidney or liver in response to treatment of rats with metal ions--Co2+, Ni2+, Pb2+, Sn2+, Zn2+, Cd2+, and Cu2+ or sodium selenite. Rather, both NADPH- and NADH-dependent activities in the kidney were decreased markedly in a time- and dose-related manner following the administration of HgCl2 (10-30 mumoles/kg, 24 hr). The pretreatment of rats (30 min) with sodium selenite (5 mumoles/kg) effectively blocked the Hg2+ (20 mumoles/kg, 24 hr) inhibition of the kidney cytosol biliverdin reductase activity. Similarly, in vitro Hg2+ was an effective inhibitor of the kidney biliverdin reductase. In addition, highly purified biliverdin reductase also was extremely sensitive to Hg2+ and the thiol reagent, 5,5'-dithiobis-(2-nitrobenzoic acid). The inhibition of purified reductase by 5,5'-dithiobis-(2-nitrobenzoic acid), but not by Hg2+, could be reversed by dithiothreitol.

Animals↗
PubMed1982

Inhibition of the enzymes of glutathione metabolism by mercuric chloride in the rat kidney: reversal by selenium.

The treatment of rats with 10 mumoles/kg (s.c.) of mercuric chloride (Hg2+) caused time-dependent decreases in the activities of the enzymes of the glutathione (GSH) metabolism pathway in the kidney. Twenty-four hours after administration of Hg2+, the activities of gamma-glutamylcysteine synthetase and glutathione disulfide (GSSG)-reductase in the kidney were decreased by 50-60%, and the activities of the GSH catabolic enzymes, gamma-glutamyl transpeptidase and GSH-peroxidase, were decreased by 25-35%. In the liver, only the activity of GSSG-reductase was decreased at this time. The observed decreases in the enzyme activities were not accompanied by a depression in the cellular protein concentration. The same pattern of enzyme response was noted when rats were given 30 mumoles/kg Hg2+; however, the decreases in the specific activity of the enzymes were accompanied by great losses in the cellular protein concentrations in both the liver and the kidney (35-40%). This dose of Hg2+ also caused significant decreases in the concentration of GSH in both organs. In vitro, Hg2+ only inhibited the activity of GSSG-reductase. When rats were given sodium selenite (Na2SeO3; 5, 10 or 20 mumoles/kg, s.c.) 30 min after Hg2+ treatment (10 mumoles/kg), the Hg2+-related depressions in the activities of the enzymes of GSH metabolism in the liver and the kidney were blocked. Also, in rats treated with 30 mumoles/kg Hg2+, the administration of 10 mumoles/kg selenium significantly decreased the magnitude of depression in the concentration of GSH in the kidney.

Animals↗
PubMed1981

Purification and characterization of biliverdin reductase from rat liver.

Biliverdin reductase in a stable form was purified to homogeneity from rat liver cytosol. The purified enzyme showed 3700-fold increase in specific activity when compared with the crude preparation, and the extent of recovery was 30-35%. The molecular weight was estimated at 34,000-36,000. The amino acid analysis of the purified preparation revealed the presence of 3 cysteine residues/mol of enzyme. The reductase utilized NADPH and NADH as electron donors. The NADPH-dependent biliverdin reductase activity was extremely sensitive to SH reagents, including 5,5'-dithiobis(2-nitrobenzoic acid), N-ethylmaleimide, p-chloromercuribenzoic acid, and iodoacetamide. However, the pretreatment of the enzyme with NADPH and biliverdin fully protected the reductase from inactivation by these reagents. The enzyme activity was irreversibly inhibited by HgCl2. The addition of dithiothreitol to the enzyme inhibited by 5,5'-dithiobis(2-nitrobenzoic acid) promoted the full reversal of inhibition. The enzyme exhibited different pH optima for activity with NADPH (pH 8.7) and NADH (pH 7.0). The apparent Km for biliverdin was established to be 5.0 microM with NADH and 3.0 microM with NADPH. The apparent Km for NADPH was 3.0 microM, while that of NADH was 270 microM. The enzyme activity was inhibited by the substrate when the concentration exceeded 4.0-5.0 microM. The product, bilirubin, inhibited the enzyme activity in a competitive manner. In addition, the reductase was inhibited by hematin and zinc-protoporphyrin. Dilution produced instability in the enzyme, but the presence of exogenous proteins, such as serum albumin, beta-lactoglobulin, and lysozyme, stabilized the enzyme protein.

Amino Acids↗
PubMed1981

Effect of allylisopropylacetamide on glutathione metabolism in the rat liver. The possible role of glutathione in the induction of 5-aminolaevulinate synthase.

Administration of allylisopropylacetamide to rats caused a marked decline in the concentrations of reduced and oxidized glutathione in the liver. However, this decrease occurred in the presence of uninhibited activities of gamma-glutamylcysteine synthase and glutathione reductase, and unaltered activities of glutathione transferases A, B and C. The administration of cysteine, the rate-limiting precursor of glutathione formation, to rats treated with allylisopropylacetamide potentiated the inductive effects of the agent on 5-aminolaevulinate synthase, and markedly decreased the extent of decrease in glutathione concentrations by the agent. Conversely, the administration of diethyl maleate, which depletes the hepatic glutathione concentrations, to allylisopropylacetamide-pretreated rats (1h) diminished the extent of 5-aminolaevulinate synthase induction and the production of porphyrins by nearly 50%, when measured at 16h. This treatment did not alter the extent of non-enzymic degradation of liver haem by allylisopropylacetamide. When diethyl maleate was administered to the animals possessing high 5-aminolaevulinate synthase activity (at 3, 7 and 15h after allylisopropylacetamide), in 1h the enzyme activity was markedly decreased. Diethyl maleate had no effect on induction of 5-aminolaevulinate synthase by 3,5-diethoxycarbonyl-1,4-dihydrocollidine, also a potent porphyrinogenic agent. Diethyl maleate alone neither inhibited 5-aminolaevulinate synthase activity nor decreased the cellular content of porphyrins and haem. The data suggest that the decreases observed in the glutathione concentrations after allylisopropylacetamide administration are not the result of decreased production of the tripeptide. Rather, they most likely reflect the increased utilization of glutathione. The findings further suggest that the inhibition by diethyl maleate of allylisopropylacetamide-stimulated 5-aminolaevulinate synthase involves the inhibition of induction processes.

5-Aminolevulinate Synthetase↗
PubMed1981

Zinc . protoporphyrin is a selective inhibitor of heme oxygenase activity in the neonatal rat.

The present study was undertaken to examine the liver, spleen and kidney heme oxygenase activity in the rat, and also to investigate the response of the enzyme to a variety of metalloporphyrin complexes. The enzyme activity in the liver and the kidney of 3--4 day-old rats was several-fold greater than the corresponding values in the adult animals; however, the splenic enzyme activity was markedly depressed in comparison to that of adult rats. During the first 2--3 weeks post-parturation period, the activity of heme oxygenase in the spleen progressively increased, and in 4 weeks approached the adult values. The treatment of the newborn animals with the metalloporphyrin complex. Zn . protoporphyrin-IX, inhibited heme oxygenase activity in the spleen, liver and the kidney. Sn . protoporphyrin treatment also inhibited the activity of the enzyme in the liver and the spleen. The mechanism of the inhibition appeared to be competitive in nature. In contrast, the treatment of the newborn animals with Co . protoporphyrin increased the activity of the enzyme in the tested organs. The treatment of newborn animals with Fe . protoporphyrin (heme) also increased heme oxygenase activity in the spleen and the kidney. In addition, Co . and Fe . protoporphyrin complexes inhibited the activity of delta-aminolevulinate synthetase in the spleen; Sn . protoporphyrin and Zn . protoporphyrin, however, did not alter the activity of this enzyme. The effects of Co . protoporphyrin and Zn. protoporphyrin on the microsomal contents of cytochromes P-450, b5, the total heme, and the microsomal drug metabolism activity in the liver were compared. Zn . protoporphyrin was ineffective in altering the indicated cellular variables. According to these findings Zn . protoporphyrin may be useful as an experimental tool for the selective suppression of heme degradation activity.

5-Aminolevulinate Synthetase↗
PubMed1981

Host heme biosynthesis and degradation in schistosomiasis.

In mice infected with Schistosoma mansoni, hepatic heme metabolism is markedly altered. The production of the immediate precursor, delta-aminolevulinic acid, is diminished, while the activity of the catabolic enzyme, heme oxygenase, is greatly increased. These changes are accompanied by a reduction in the cellular content of hemoproteins in various organs. Specifically, cytochrome levels in myocardial mitochondria are reduced, and liver cytochromes P-450 and b5 are also diminished. As a consequence of the latter, the microsomal oxidative enzyme activities, which are mediated by P-450, such as ethylmorphine N-demethylase and aniline hydroxylase, are considerably impaired. Barbiturate-induced sleeping time in mice heavily infected with schistosomes was found to be significantly prolonged. A green discoloration of the liver and spleen seen in advanced murine schistosomiasis is not likely to be due to the production of abnormal pyrrolic pigments, since hemoglobin heme was found to be degraded via the usual catabolic pathways to physiological bile pigments. Total serum iron was found to be increased by 100% in schistosome-infected mice. Serum unsaturated iron binding capacity was, however, not increased significantly. Demonstration that the activities of enzymes of heme metabolism, which are known to be regulated by heme and metal ions, are altered in the host as a consequence of the parasitism suggests that these perturbations may be mediated by heme or its iron released by the digestion of erythrocytes by schistosomes.

5-Aminolevulinate Synthetase↗
PubMed1980

Regional distribution of the enzymes of haem biosynthesis and the inhibition of 5-aminolaevulinate synthase by manganese in the rat brain.

The activity of 5-aminolaevulinate synthase, the rate-limiting enzyme of haem biosynthesis, is differentially distributed in various regions of the rat brain. The cerebellum possessed the highest enzyme activity of the eight regions studied. The cerebral cortex and the midbrain also exhibited high 5-aminolaevulinate synthase activity; the septum, hypothalamus, thalamus, amygdala and the hippocampus possessed much lower enzyme activity. However, the total porphyrin and haem contents of the different brain segments did not vary greatly. Mn(2+), when administered subcutaneously to rats, effectively inhibited the activity of 5-aminolaevulinate synthase in the cerebellum, midbrain and cerebral cortex; however, repeated injections of the metal ion neither decreased the haem and porphyrin contents of the brain nor induced haem oxygenase activity. Mn(2+) was not an effective inhibitor of 5-aminolaevulinate synthase activity in vitro. On the other hand, studies carried out with the liver in vivo suggested that Mn(2+) may alter the turnover rate of cellular haem and haemoproteins. In that event, it is likely that the inhibition of 5-aminolaevulinate synthase by Mn(2+) was in part a result of the inhibition of protein synthesis by the metal ion. It is postulated that the haem and porphyrin contents of the brain are maintained at a steady-state level, due in part to the refractoriness to inducers of the regulatory mechanism for haem catabolic enzymes and in part to the ability of the organ to utilize haem precursors derived from extraneuronal sources.

5-Aminolevulinate Synthetase↗
PubMed1977

Metals as regulators of heme metabolism.

Heme is essential for cell respiration, energy generation, and oxidative biotransformations. The latter function is exemplified by the oxidative metabolism of various endogenous and exogenous chemicals catalyzed by the heme protein cytochrome P-450. Recent studies have established that metal ions directly regulate cellular content of heme, and thus of heme proteins by controlling production of delta-aminolevulinate synthetase and heme oxygenase, the rate-limiting enzymes for heme synthesis and degradation, respectively. Metal ions also alter cellular content of glutathione. In excess amounts, metal ions greatly accelerate the turnover and degradation of heme and substantially impair the oxidative functions of cells--particularly those dependent on cytochrone P-450. As a result, the biological impact of chemicals which are detoxified or metabolically transformed by the P-450 system is greatly altered.

5-Aminolevulinate Synthetase↗
PubMed1977

Enzymes of heme metabolism in the kidney: regulation by trace metals which do not form heme complexes.

The in vivo regulation by metal ions of the enzymes of heme metabolism in kidney-particularly of ALAS, the rate-limiting enzyme in heine formation- was investigated. Ni(2+) and Pt(4+), metals which do not enzymatically form metalloporphyrins, were found to regulate ALAS in kidney as they do in liver. The pattern of this regulation was generally similar to that observed with heme and metal ions in liver, i.e., a late increase in enzyme activity after an early period in which ALAS activity was unaltered or inhibited. The metals did not interact with the enzyme in vitro to alter its activity. In this study no direct reciprocal relationship between ALAS activity and total cellular heine content was demonstrated. The metal ions, particularly Pt(4+), also altered the activity of other enzymes of heme biosynthesis in kidney. Pt(4+) severely inhibited the activity of ALAD and UROS. Ni(2+) and Pt(4+) were potent inducers of heme oxygenase, the initial and rate-limiting enzyme in heine degradation. It is proposed that the physiological regulation of ALAS is mediated through the action of metal ions, rather than by the cellular content of heine, and that the regulation of ALAS by heine reflects the action of the central metal ion of heme rather than that of the entire metalloporphyrin complex. In this proposed mechanism for metal ion regulation of ALAS, the tetrapyrrole moiety of heine is considered to function principally as an efficient carrier of metal to the regulatory site for ALAS production, inasmuch as the tetrapyrrole ring itself has been shown in earlier studies not to have any effect on ALAS activity. The production of heine oxygenase is believed to be similarly regulated.

5-Aminolevulinate Synthetase↗
PubMed1977

Solubilization and partial purification of heme oxygenase from rat liver.

Hepatic microsomal heme oxygenase was solubilized, partially purified, and characterized from Co2+-treated rats. The enzyme on sodium dodecyl sulfate-polyacrylamide gel electrophoresis exhibited a minimum molecular weight of greater than or equal to 68,000. The solubilized enzyme was totally devoid of contamination with cytochrome P-450 or b5. The requirement for reduced pyridine nucleotides was absolute, and ascorbate could not support heme oxidative activity. However, both TPNH and DPNH could serve as electron donors, with TPNH being more effective. The presence of an appropriate flavoprotein reductase was essential for heme oxidation. The enzyme had an apparent Km of 40 micrometer, a pH optimum of 7.5, and lost substantial activity upon freezing and thawing. Methemoglobin was 30% as effective a substrate for the enzyme as was heme. Free porphyrins could not serve as substrates for the enzyme. The activity of the enzyme was inhibited by HgCl2, p-chloromercuribenzoate, iodoacetamide, mercaptoethanol, and dithiothrietol indicating that free -SH group(s) is necessary for enzyme activity.

Animals↗
PubMed1977

Bile pigment formation by skin heme oxygenase: studies on the response of the enzyme to heme compounds and tissue injury.

Skin heme oxygenase is locally elevated by stimuli such as tissue injury and injections of whole blood, myoglobin, and hematin. The enzyme activity is also increased at the proximity of the injection site of chemicals such as cobalt and cobalt-protoporphyrin-IX (cobalt-heme). Protoporphyrin-IX, the tetrapyrrole nucleus of type-b heme compounds, was ineffective in altering the enzyme activity in vivo. The developmental pattern of heme oxygenase in skin was compared to that of the enzyme in liver. The enzyme activity in both organs was greatest during the 1st postpartum wk and declined to adult levels after 2 wk. The physiological implications of the increased activity of skin heme oxygenase are discussed, and it is concluded that the activity of the hepatic heme oxygenase system and that of the skin are regulated by the same mechanism.

Animals↗