Donald P. Tschudy

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

Abstract 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.

Succinylacetone, a potent inhibitor of heme biosynthesis: Effect on cell growth, heme content and δ-aminolevulinic acid dehydratase activity of malignant murine erythroleukemia cells

Abstract 4,6-Dioxoheptanoic acid (succinylacetone, SA) was examined with regard to its ability to a) inhibit the second enzyme of the heme pathway, δ-aminolevulinic acid (ALA) dehydratase, b) lower the heme concentration, and c) inhibit cell growth of murine erythroleukemia (MEL) cells in culture. SA profoundly inhibited ALA dehydratase in broken cell preparations at concentrations as low as 10−7 M. The stimulation of hemoglobin production by DMSO and butyrate in MEL cells was inhibited by the addition of SA to the cell medium. When 1 mM SA was added to the medium, there was a profound inhibition of ALA dehydratase activity, and the heme concentration of cells declined progressively with each cell division. Cell growth was markedly inhibited after two cell divisions.

Acute Intermittent Porphyria: Clinical and Selected Research Aspects

Acute intermittent porphyria is an inborn error of metabolism characterized by the excretion of excess porphyrin precursors (porphobilinogen and usually delta-aminolevulinic acid) in the urine, and by sporadic attacks of neurologic dysfunction. The disease is complex, involving variable patterns of autonomic and peripheral neuropathy as well as the central nervous system manifestations. There may be alterations in carbohydrate, lipid, water, and electrolyte metabolism in addition to clinically inapparent endocrine abnormalities. The fundamental defect is thought to be a 50% decrease of uroporphyrinogen I synthetase, the third enzyme of the heme biosynthetic pathway. This is associated with a marked increase of hepatic delta-aminolevulinic acid synthetase, the first and rate controlling enzyme of the pathway. The measurement of uroporphyrinogen I synthetase in erythrocytes now provides an enzyme diagnostic test for the disease. Two therapeutic approaches that may prove to reverse the fundamental disease process, at least in some patients, involve [1] a high carbohydrate intake, and [2] intravenous administration of hematin. The latter, only recently introduced, is now being investigated.

FACTORS AFFECTING THE EXCRETION OF PORPHYRIN PRECURSORS BY PATIENTS WITH ACUTE INTERMITTENT PORPHYRIA. I. THE EFFECT OF DIET.

Abstract Daily fluctuation of porphyrin-precursor excretion by asymptomatic patients with acute intermittent porphyria (AIP) was markedly diminished by a regimen which included a balanced constant diet adequate in content of protein, fat and carbohydrate to maintain weight. Acute reduction in daily food intake, such that caloric intakes decreased by 60 to 80 per cent, was associated with a rise in the excretion of both δ-aminolevulinic acid (ALA) and porphobilinogen (PBG). Return to the adequate diet reversed this effect. A rise in porphyrin-precursor excretion followed the isocaloric substitution of fat for protein alone or for protein together with carbohydrate. ALA and PBG excretion decreased following the addition of carbohydrate to the diet. Elevation of porphyrin-precursor excretion was accompanied by negative nitrogen balance in the studies where this was measured. With 1 exception the patients remained asymptomatic. It is concluded that the effect of diet in causing changes in porphyrin-precursor excretion by patients with AIP is related to protein and carbohydrate content rather than total caloric content per se.

Abnormal electrolyte and water metabolism in acute intermittent porphyria. The transient inappropriate secretion of antidiuretic hormone.

Abstract Two patients with symptomatic acute intermittent porphyria transiently exhibited a syndrome of hyponatremia, water retention and urinary loss of electrolytes. It is suggested that this was due to inappropriate secretion of antidiuretic hormone, rendering them incapable of excreting water loads normally which, in turn, induced the observed derangements in sodium metabolism, and may have contributed to urinary loss of calcium and magnesium as well. In one patient, hypomagnesemia and hypocalcemia developed, with concomitant tetany. Excessive loss of magnesium and calcium, coupled with inadequate intake, presumably was responsible for the low serum values.

Hematin therapy for acute porphyria.

1. A therapeutic trial of intravenous hematin is presented. Eleven cases of AIP and one of VP who did not improve with conventional treatment (high carbohydrate intake) received this new agent. 2. Urinary ALA, PBG and, when possible, uroporphyrin and coproporphyrin were used to monitor the chemical response to the treatment. Objective clinical parameters of hypertension and tachycardia were followed when present in addition to subjective estimates of acute porphyric symptomatology (abdominal pain, backache, extremity pain and paresthesias, weakness, depression, etc.). 3. At a dosage of approximately 3 mg/kg, diminution of urinary ALA and PBG excretion was achieved in every patients. Hypertension and tachycardia improved in those instances where they were observed in association with the attack. Also, subjective improvements in the clinical status of the patients were observed frequently. 4. Hematin appears to be a promising therapeutic agent for the treatment of acute attack forms of porphyria.

Acute intermittent porphyria: New morphologic and biochemical findings

Abstract This paper presents the results of postmortem morphologic and biochemical studies on a patient with acute intermittent porphyria (AIP); there was evidence suggesting the syndrome of inappropriate antidiuretic hormone (ADH) release. Examination of the brain revealed injury to the median eminence and bilateral loss of neurones of the supraoptic and paraventricular nuclei. This represents the first correlation of a specific hypothalamic lesion with the syndrome of inappropriate ADH release. The activity of hepatic δ-aminolevulinic acid synthetase (ALA synthetase), a mitochondrial enzyme which normally is rate-limiting for porphyrin biosynthesis, was increased sevenfold above normal. The high level of this enzyme explains the increased porphyrin precursor excretions seen in AIP. Three general mechanisms, including an operator constitutive mutation, have been discussed as possible explanations of the induction of hepatic ALA synthetase.

A simple micro method for the direct determination of δ-amino[14Clevulinic acid production in murine spleen and liver homogenates

Abstract 1. 1. A simple and sensitive radiochemical procedure for the estimation of the activity of δ-aminolevulinate synthetase (succinyl-CoA: glycine succinyltransferase) in homogenates of normal murine spleen and liver is described. 2. 2. Homogenates of spleen or liver are incubated with α- keto [ 14 C] glutarate or [14C]succinate respectively, and the [14-amino14Clevulinic acid generated in the reaction is selectively adsorbed onto a Dowex 50 column run at pH 3.9. Since labeled α-amino acids and unreacted radioactive substrate are not reatained by the column, contamination of α-aminolevulinic acid by these substances does. δ- Amino- [ 14 C] levulinic can be recovered in yields of 97–100%. 3. 3. Maximal δ- amino[ 14 C]levulinic acid production was obtainedin spleen homogenates in the presence of α- keto[ 14 C]glutarate , glycine, EDTA, and pyridoxal phosphate. δ- Amino[ 14 C]leculinic acid utilization was minimal at homogenate concentrations of 2.5% or less. Varying degrees of inhibition of δ-amino14Clevulinic acid production were seen with malate, malonate, arsenite, ATP, CoA, and NAD+. 4. 4. [14]Succinate, glycine, EDTA, and pyridoxal phosphate produced the highest levels of δ- amino[ 14 C]levulinic acid in liver homogenates. δ- Amino[ 14 C]levulinic acid utilization was minimal at homogenate concentrations of 1% or less. Varying degrees of inhibition of δ- amino[ 14 ]levulinic acid production were seen with malate, malonate, arsenite, ATP, and CoA. 5. 5. The sensitivity of the procedure will allow the measurement of δ-amino-levulinate synthetase activity in normal spleen and liver and will permit more detailed investigation of mechanisms controlling heme synthesis. Smaller amounts of tissues can be used than have been previously employed by colorimetric methods. The method can be easily adapted to preparations of mitochondria and blood cells.

Coordinate synthesis of heme and apoenzyme in the formation of tryptophan pyrrolase.

Reciprocal control mechanisms between hemoprotein and 8-aminolevulinic acid synthetase take part in coordinate synthesis of the heme and apoenzyme moieties of tryptophan pyrrolase. Stimulation of heme biosynthesis increases tryptophan pyrrolase, whereas enhancement of heme binding by apotryptophan pyrrolase secondarily increases the formation of δ-aminolevulinic acid synthetase, the rate-limiting enzyme in heme formation. Tryptophan-mediated induction of δ-aminolevulinic acid synthetase suggests that heme participates in repression of that enzyme

Abnormalities of growth hormone regulation in acute intermittent porphyria

Abstract Administration of glucose to patients with acute intermittent porphyria often produces a paradoxical rise in the level of circulating growth hormone rather than the decline seen in normals. In other patients with the disease, the rate and magnitude of the decline may be less than that of normals. These findings suggest that abnormalities of hypothalamic function occur frequently in acute intermittent porphyria and are in keeping with previously demonstrated abnormalities in the regulation of antidiuretic hormone in this disease.