Induction Of Δ-aminolevulinic Acid Synthetase Research Articles

Relative Potencies of Polychlorinated Dibenzo-p-dioxins, Dibenzofurans, and Biphenyls Derived from Hepatic Porphyrin Accumulation in Mice

Hepatic porphyrin accumulation was studied after subchronic dosing of female B6C3F1 mice by gavage with single congeners of polychlorinated or polybrominated dibenzo-p-dioxins (PCDDs, PBDDs), polychlorinated dibenzofurans (PCDFs), and polychlorinated biphenyls (PCBs). Quantitative hepatic porphyrin profile analyses in selected samples showed uroporphyrin and heptacarboxylporphyrin as the main porphyrins detected. Dose-dependent increases in total hepatic porphyrins were found for all congeners tested. At lower dose levels, relative potencies, based on administered dose as well as target tissue dose, of PCDDs, PCDFs, and coplanar PCBs, using 2,3,7,8-tetrachlorodibenzo-p-dioxin as a reference compound, were in the same range as those previously derived from the induction of hepatic CYP1A1 and CYP1A2 enzyme activities. CYP1A2 has been reported to be involved in the oxidation of uroporphyringen III to uroporphyrin III. All these facts suggest the involvement of an aryl hydrocarbon receptor-mediated mechanism in hepatic porphyrin accumulation, possibly via CYP1A2. However, the relative potencies of the mono-ortho-substituted PCBs were higher for hepatic porphyrin accumulation than for hepatic CYP1A1 and CYP1A2 induction. In addition, hepatic porphyrin accumulation was the highest after exposure to mono-ortho-PCBs. Since mono-ortho-substituted PCBs induce the phenobarbital-inducible CYP2B isoforms of cytochrome P450, an additional induction of δ-aminolevulinic acid synthetase may also contribute to hepatic porphyrin accumulation following subchronic exposure to these particular congeners. Relative potencies derived from hepatic porphyrin accumulation after PCDD, PCDF, or coplanar PCB administration are a useful tool in risk assessment. However, the higher relative potencies of the mono-ortho-substituted PCBs have important implications for risk assessment of these compounds.

Role of inhibition of uroporphyrinogen decarboxylase in PCB-induced porphyria in mice

The oral administration of 3,4,5,3′,4′,5′-hexachlorobiphenyl for 3 weeks to mice caused a marked accumulation of porphyrins in the liver of C57BL 6 and C57BL 10 mice but not in the liver of ddY mice. The time course of induction of δ-aminolevulinic acid synthetase (ALA-S), cytochrome P-450, and mixed function oxidases and inhibition of uroporphyrinogen decarboxylase (URO-D) in the liver of C57BL 6 mice and ddY mice fed a diet containing 500 ppm of a commercial PCB (Kanechlor-500) were investigated to clarify the sole factor in inducing porphyria. The activity of URO-D in the liver of C57BL 6 mice was depressed approximately 80% at 3 weeks when a large amount of uroporphyrin accumulated. Male ddY mice showed only a slight increase in uroporphyrin accumulation in the liver and a moderate decrease of URO-D activity even at the 10th week. ALA-S, cytochrome P-450, and mixed function oxidases were induced in both strains of mice, although the magnitude of these inductions in C57BL 6 mice was greater than that in ddY mice. No differences were detected between the two strains in the content and gas chromatographic pattern of PCB remaining in liver cytosol (6 weeks). In addition there was no relationship between the time of onset of porphyria and that of the maximal induction of drug-metabolizing function in C57BL 6 mice. These results indicate that the development of porphyria is causally related to the inhibition of URO-D rather than the induction of drug-metabolizing function. The hypothesis that porphyria first develops when the ratio of hepatic URO-D and ALA-S activities decreases to less than 1.0 is presented.

Effects of succinylacetone on dimethylsulfoxide-mediated induction of heme pathway enzymes in mouse Friend virus-transformed erythroleukemia cells

Heme has been reported to exert a control over its own biosynthesis and to affect the erythroid differentiation process at different sites. In this study, succinylacetone, a powerful inhibitor of δ-aminolevulinic acid dehydrase was used to block heme synthesis and to study the effects of heme depletion on the dimethylsulfoxide (DMSO)-mediated induction of the heme pathway enzymes in Friend virus-transformed erythroleukemia cells. The presence of succinylacetone in the medium during the DMSO treatment (1) potentiates the induction of δ-aminolevulinic acid synthetase (the first enzyme of the pathway) and this effect is reversed by the addition of exogenous hemin; (2) does not affect the induction of δ-aminolevulinic acid dehydrase (the second enzyme); (3) prevents the induction of porphobilinogen deaminase (the third enzyme), since no increase could be detected in either the enzyme activity or the immunoreactive protein and this effect could not be reversed by the addition of exogenous hemin; (4) does not affect the induction of ferrochelatase. The possible role of heme or of intermediate metabolites of the pathway on the induction of these enzymes during the erythroid differentiation process is discussed.

Induction of δ-aminolevulinic acid synthetase in chick embryo kidney

Induction of δ-aminolevulinic acid synthetase in chick embryo kidney

Induction of porphyrin synthesis in chick embryo liver cell culture by synthetic polychlorobiphenyl isomers

The effects of a series of synthetic di-tetra- and hexachlorobiphenyl isomers and commercial polychlorinated biphenyls on the porphyrin biosynthesis in chick embryo liver cells in culture were examined. It was found that 3,4,3′,4′-tetra- and 3,4,5,3′,4′,5′-hexachlorobiphenyl isomers were the most active inducers, which were approximately 20 times as active as 1,4-dihydro-3,5-dicarbethoxy-2,4,6-trimethylpyridine (DDC) in porphyrin production. 3,5,3′,5′-Tetra- and 2,3,4,2′,3′,4′-hexachlorobiphenyl isomers were moderate inducers, which were approximately 2.0 to 2.5 times as active as DDC. 2,4,6,2′,4′,6′-Hexachlorobiphenyl showed the same activity as DCC. Compounds such as 4,4′-di-, 2,3,2′,3′-, 2,4,2′,4′- and 2,6,2′,6′-tetrachlorobiphenyl were weak inducers and 2,5,2′,5′-tetrachloro- and decachlorobiphenyl isomers were found to be inactive. Kanechlor-400 was the strongest inducer among the commercial polychlorinated biphenyls investigated. The structural requirements for potent porphyrin-inducing activity of chlorobiphenyl isomers were found to be the para and meta substituted structure causing a more highly conjugated and nearly coplanar conformation. It was found that induction caused by some chlorobiphenyls was subject to feed-back repression by end-product heme. In addition, the metabolism of chlorobiphenyls in mice was influenced by the unsubstituted pairs of carbon atoms in the molecule. These results lead us to postulate the following hypothesis, namely, that strong inducers may displace heme directly and incorporate into a hydrophobic pocket of the apo-represor protein, thus causing an induction of δ-aminolevulinic acid synthetase.

Induction of delta-aminolevulinic acid synthetase by flutamide, a non-steroidal antiandrogen.

Abstract Flutamide, a nonsteroidal antiandrogen, was found to be an effective inducer of δ-aminolevulinate synthetase, the rate-limiting enzyme in the heme biosynthetic pathway, in cultured chick embryo liver cells. This antiandrogen increased the activity of the enzyme 24-fold at a concentration of 1 μM and maximally stimulated the enzyme (62-fold) at a concentration of 50 μM. The steroidal antiandrogen, 6α-bromo-17α-methyl-17β-hydroxy-4-oxa-5α-androstan-3-one, did not stimulate aminolevulinate synthetase activity in these cells. Studies with inhibitors suggest that the induction of aminolevulinate synthetase by flutamide requires continued RNA and protein synthesis. In addition, the induction of the enzyme was inhibited (85%) by 15 μM hemin, the physiological repressor of aminolevulinate synthetase.

Cytochrome P-450 heme and the regulation of δ-aminolevulinic acid synthetase in the liver

Hepatic δ-aminolevulinic acid synthetase was induced in rats injected with allylisopropylacetamide. The induction process was studied in relation to experimental perturbation of cytochrome P-450 in the liver. Animals were treated with either administered endotoxin or exogenous heme, both of which accelerate degradation of cytochrome P-450 heme. These manipulations were effective in blocking induction of δ-aminolevulinic acid synthetase, and the effect of each compound was proportional to its ability to stimulate degradation of cytochrome P-450 heme. The findings suggest that the heme moiety of cytochrome P-450 dissociates reversibly from its apoprotein and, prior to its degradation, mixes with endogenously synthesized heme to form a pool that regulates δ-aminolevulinic acid synthetase activity. A similar or identical heme fraction appears to mediate stimulation of heme oxygenase, which suggests that the regulation of δ-aminolevulinic acid synthetase and of heme oxygenase in the liver are closely interrelated.

Editorial: Hematin and porphyria.

The discovery that hematin represses induction of δ-aminolevulinic acid synthetase (ALA-S), the rate-limiting enzyme for hepatic porphyrin and heme synthesis,1 is central to recent studies seeking to provide a means of terminating or ameliorating the acute attack of acute intermittent porphyria, porphyria variegata or hepatic coproporphyria. The induction of ALA-S in the acute attack of AIP, acute intermittent porphyria, porphyria variegata or hepatic coproporphyria markedly increases the porphyrin precursors, ALA and porphobilinogen (PBG), in liver, blood plasma and urine. The enzyme is induced because of heme deficiency in the liver secondary to the genetic partial lack of uroporphyrinogen synthetase (UPG-S) . . .

Induction of δ-aminolevulinic acid synthetase in the kidney of chicks treated with porphyrinogenic drugs

1. 1. The administration of allylisopropylacetamide induced an increase in δ-aminolevulinic acid synthetase activity in both kidney and liver of one-day-old chicks. 2. 2. The induction of δ-aminolevulinic acid synthetase in chick kidney was measurable by colorimetric assay and was comparable in activity to the increase observed in the liver. 3. 3. One of the most potent inducers of δ-aminolevulinic acid synthetase in chick liver, 3,5-dicarbethoxy-1,4-dihydrocollidine, did not increase the activity of chick kidney although the activity in liver was higher than that obtained with allylisopropylacetamide. Ferrochelatase activity was decreased in both liver and kidney by dicarbethoxy-1,4-dihyrocollidine.

Studies on the induction of δ-aminolevulinic acid synthetase in mouse liver

Administration of 3,5-diethoxy carbonyl-1,4-dihydrocollidine (DDC) to mice resulted in a striking increase in the level of δ-aminolevulinic acid (ALA) synthetase in liver. Although the enzyme activity was primarily localized in mitochondria and postmicrosomal supernatant fluid, a significant level of activity was also detected in purified nuclei. The time course of induction showed a close parallelism between the bound and free enzyme activities with the former always accounting for a higher percentage of the total activity as compared to the latter. Studies with cycloheximide indicated a half-life of around 3 hr for both the bound and free ALA synthetase. Actinomycin D and hemin prevented enzyme induction when administered along with DDC, but when administered 12 hr after DDC treatment Actinomycin D did not lead to a decay of either the bound or free enzyme activity and hemin inhibited the bound enzyme activity but not the free enzyme level. The molecular sizes of the mitochondrial and cytosolic ALA synthetase(s) were found to be similar on sephadex columns.

The Induction of δ-Aminolevulinic Acid Synthetase in Cultured Liver Cells: THE EFFECTS OF END PRODUCT AND INHIBITORS OF HEME SYNTHESIS

Abstract The regulation of heme biosynthesis has been studied in cultured avian hepatocytes utilizing a specific and sensitive assay for the rate-limiting enzyme in the pathway, δ-aminolevulinic acid synthetase. This enzyme appears to be induced by 2-allyl-2-isopropylacetamide, a drug known to produce experimental porphyria, and by inhibitors of heme synthesis such as lead acetate. However, 2-allyl-2-isopropylacetamide and lead acetate appear to induce δ-aminolevulinic acid synthetase by different mechanisms since inhibitors of protein synthesis at the transcriptional level block induction by 2-allyl-2-isopropylacetamide but not block induction by lead acetate. Furthermore, induction by the two chemicals is synergistic. Addition of the end product of the pathway, heme, and the heme precursor, δ-aminolevulinic acid, prevents induction by 2-allyl-2-isopropylacetamide and lead acetate. These data further support the role of heme as the natural repressor of δ-aminolevulinic acid synthetase. Experiments with inhibitors of protein synthesis are consistent with a translational site for heme repression. A mechanism whereby drugs may induce δ-aminolevulinic acid synthetase by interfering with heme synthesis is demonstrated. The relevance of these observations to the biochemical aberration characterizing the human genetic disease, intermittent acute porphyria, is discussed.

A reciprocal relationship between the induction of δ-aminolevulinic acid synthetase and drug metabolism produced by m-dichlorobenzene

The daily administration of large doses of m-dichlorobenzene (m-DCB) causes experimental hepatic porphyria in rats by induction of δ-aminolevulinic acid synthetase (ALA synthetase). However, smaller doses of m-DCB produce a biphasic stimulation of both urinary coproporphyrin excretion and liver ALA synthetase. The decline of ALA synthetase and urinary coproporphyrin despite continued daily dosage is associated with an increase in activity of the liver drug-metabolizing systems and decreasing serum m-DCB levels. The most probable explanation for the self-limiting action of m-DCB is stimulation by the drug of its own metabolism.

Chronic hepatic porphyria in chronic aggressive hepatitis.

An acquired (“symptomatic”) chronic hepatic porphyria (CHP) is described, which develops secondarily following damage to the liver tissue and is characterized by the absence of clinical symptoms. It can be differentiated biochemically from latent porphyria cutanea tarda (PCT) by a lower total porphyrin excretion (0.18–0.64 mg/l) and a relatively increased excretion of corproporphyrin (C) (18–55 %) in urine. In liver biopsies from 4 patients with chronic aggressive hepatitis (CAH) and 2 patients with fibrosis of the liver, uro-(U) and heptacarboxylic porphyrin (VII) were found to have accumulated in the liver, just as in PCT. The elevated excretion of porphyrins with the urine involved all components, especially U, VII, and C. Independently of an inversion of the C/U ratio, VII increases in accordance with the rule of the biochemical PCT index: VII/C increases (U+VII)/C by 18–37 %. U was found to predominate in 3 patients with CAH; in fibrosis, however, larger amounts of C were present in the urine, although the liver stored U and VII. On the basis of the relative distributions of the individual urinary porphyrins, two types of CHP are distinguished: Type A with C/U>1 (in fibrosis), and type B, in which C/U<1 (in CAH). Apparently, enhanced renal excretion of U in type B of CHP is indicative of an active hepatic process. The concentrations of U and VII in the liver are greater in type B than in type A. CHP can remain stable, or undergo remission: type A can develop into type B; PCT can arise from type B with an increase of U plus VII. The key to the biochemical pathogenesis is probably to be found in the induction ofδ-aminolevulinic acid synthetase and, in addition, a reduced decarboxylation of heptacarboxylic porphyrinogen.

The regulation of porphyrin-heme biosynthesis: The effects of certain steroids and other porphyrin-inducing agents in different in vitro systems

The effects of certain steroids, sedormid, and a barbiturate on the biosynthesis of porphyrins were studied in several in vitr systems using either avian erythrocytes, rat liver homogenates, or bovine liver acetone powder as the porphyrin-synthesizing system. No direct stimulatory or inhibitory effect was observed under the conditions employed. The present results thus show that in vitro systems lacking (avian erythrocytes) or having limited capacity (livers of adult animals) for the induction of δ-aminolevulinic acid synthetase exhibit little or no response to the in vivo porphyria-inducing agents studied. In contrast, in vitro systems having the ability of ALAS induction (bone marrow cells, embryonic liver cells) show increased porphyrin formation upon treatment with these same chemical agents. The present results are in accord with the current concept that the effect of these substances on porphyrinogenesis is indirect apparently by way of the induction of the key enzyme of the porphyrin biosynthetic pathway, δ-aminolevulinic acid synthetase. Impairment of terminal cellular oxidation by the inhibition of NADH oxidase activity, resulting in lower cellular ATP levels, may be a common, primary factor involved in inducing the biosynthesis of ALAS by the chemical agents employed.

Further studies on the relationship of the stimulatory effects of phenobarbital and 3,4-benzpyrene on hepatic heme synthesis to their effects on hepatic microsomal drug oxidations

The administration of either phenobarbital or 3,4-benzpyrene to rats resulted in the rapid and marked induction of δ-aminolevulinic acid synthetase (EC 2.3.1.13), the proposed initial and rate-limiting enzyme in the hepatic heme biosynthetic pathway. Enhanced formation of δ-aminolevulinic acid was followed sequentially by an enhancement of the liver's capacity to synthesize microsomal heme in vivo, increases in the content of cytochrome P-450 and protoheme in hepatic microsomes and stimulation of certain hepatic microsomal drug oxidations. Changes in the hepatic microsomal levels of cytochrome P-450 paralleled, in part, changes in the activity of hepatic δ-aminolevulinic acid synthetase and in the capacity of the liver to synthesize microsomal heme in vivo, suggesting that the rate of hepatic heme synthesis may control the rate of synthesis of hepatic microsomal cytochome P-450. Increases in the hepatic microsomal content of cytochrome b 5, however, followed a different time course from that observed from either cytochrome P-450 or protoheme. The simultaneous administration of maximum stimulatory doses of phenobarbital and 3,4-benzpyrene did not result in an additive stimulation of δ-aminolevulinic acid synthetase activity, indicating that phenobarbital and 3,4-benzpyrene induce δ-aminolevulinic acid synthetase by the same or closely related mechanisms. However, the stimulatory effects of these agents on cytochrome P-450 and on the N-demethylation of 3-methyl-4-monomethylaminoazobenzene were additive, suggesting that differences may exist in the mechanism by which phenobarbital and 3,4-benzpyrene induce hepatic microsomal cytochrome P-450 and enhance certain hepatic microsomal drug oxidations.

The role of heme synthesis during the induction of hepatic microsomal cytochrome P-450 and drug metabolism produced by benzpyrene

δ-Aminolevulinic acid synthetase, the initial and rate limiting step in hepatic heme synthesis, is induced by both benzpyrene and phenobarbital. Induction of this enzyme by benzpyrene results in the stimulation of glycine-2- 14C incorporation into hepatic microsomal heme in vivo and in the induction of cytochrome P-450 and N-demethylase activity. 3-Amino-1,2,4-triazole, an inhibitor of the second step in hepatic heme synthesis, prevents the stimulation of hepatic heme synthesis and the induction of P-450 and N-demethylase activity. It is suggested that induction of δ-aminolevulinic acid synthetase leading to increases in hepatic heme synthesis may be the mechanism by which benzpyrene induces cytochrome P-450 and certain hepatic microsomal oxidations.

Induction of δ-aminolevulinic Acid Synthetase in Lead Poisoning

Induction of δ-aminolevulinic Acid Synthetase in Lead Poisoning

The effect of carbohydrate feeding on the induction of δ-aminolevulinic acid synthetase

Allylisopropylacetamide (AIA), when administered to rats, induces high levels of hepatic mitochondrial δ-aminolevulinic acid synthetase (ALA synthetase). The activity of this enzyme can be quantitated in porphyric rat liver, since the freshly prepared mitochondria from porphyric rat liver are permeable to succinyl-coenzyme A and do not have high deacylase activity. Administration of carbohydrate to rats markedly inhibits the induction of ALA synthetase by AIA. Since ALA synthetase is thought to be the rate controlling enzyme in hepatic porphyrin biosynthesis, diet is a regulatory factor in hepatic porphyrin synthesis in porphyria. The previously observed effects of diet on porphyrin precursor excretion in both experimental and human acute intermittent porphyria are best explained by the effect of diet on the hepatic ALA synthetase level. The application of the theory of “catabolite repression” to the present findings is discussed.