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H L Bonkovsky

Publications and source records attributed to H L Bonkovsky.

At least 91 records · Page 5Linked to original sources
PubMed1992

Porphyrogenic properties of the terpenes camphor, pinene, and thujone (with a note on historic implications for absinthe and the illness of Vincent van Gogh).

Camphor, alpha-pinene (the major component of turpentine), and thujone (a constituent in the liqueur called absinthe) produced an increase in porphyrin production in primary cultures of chick embryo liver cells. In the presence of desferrioxamine (an iron chelator which inhibits heme synthesis and thereby mimics the effect of the block associated with acute porphyria), the terpenes enhanced porphyrin accumulation 5- to 20-fold. They also induced synthesis of the rate-controlling enzyme for the pathway, 5-aminolevulinic acid synthase, which was monitored both spectrophotometrically and immunochemically. These effects are shared by well-known porphyrogenic chemicals such as phenobarbital and glutethimide. Camphor and glutethimide alone led to the accumulation of mostly uro- and heptacarboxylporphyrins, whereas alpha-pinene and thujone resulted in lesser accumulations of porphyrins which were predominantly copro- and protoporphyrins. In the presence of desferrioxamine, plus any of the three terpenes, the major product that accumulated was protoporphyrin. The present results indicate that the terpenes tested are porphyrogenic and hazardous to patients with underlying defects in hepatic heme synthesis. There are also implications for the illness of Vincent van Gogh and the once popular, but now banned liqueur, called absinthe.

5-Aminolevulinate Synthetase↗
PubMed1992

Differences in glycogen, lipids, and enzymes in livers from rats flown on COSMOS 2044.

Livers from rats flown aboard COSMOS 2044 were analyzed for protein, carbohydrate (glycogen), and lipids as well as the activities of a number of key enzymes involved in metabolism of these compounds and xenobiotics. The major differences between the flight group and the synchronous control were elevations in microsomal protein, liver glycogen content, tyrosine aminotransferase, and tryptophan oxygenase and reductions in sphingolipids and the rate-limiting enzyme of heme biosynthesis, delta-aminolevulinic acid synthase. These results provide further evidence that spaceflight has pronounced and diverse effects on liver function; however, some of the results with samples from COSMOS 2044 differed notably from those from previous spaceflights. This may be due to conditions of spaceflight and/or the postflight recovery period for COSMOS 2044.

Amino Acids↗
PubMed1991

Synergistic induction of delta-aminolevulinate synthase by glutethimide and iron: relationship to the synergistic induction of heme oxygenase.

Relationships between activities of delta-aminolevulinate synthase and heme oxygenase, respectively the rate-limiting enzymes of heme biosynthesis and degradation, have been studied in chick embryo liver cell cultures following exposure of the cultures to glutethimide and iron, a combination known to produce a synergistic induction of both enzymes. In time-course experiments, synergistic induction of heme oxygenase activity by glutethimide and iron preceded that of delta-aminolevulinate synthase by 4 h. Effects of selective inhibitors of both heme synthesis and degradation have also been studied with respect to effects on delta-aminolevulinate synthase and heme oxygenase activities. The synergistic induction of heme oxygenase by glutethimide and iron appears to be dependent upon cellular heme synthesis because addition of inhibitors of heme biosynthesis, 4,6-dioxoheptanoic acid or N-methyl-mesoporphyrin abolishes this synergistic induction. Exposure of cultures to tin-mesoporphyrin, a potent inhibitor of heme oxygenase, prevented the synergistic induction of delta-aminolevulinate synthase produced by glutethimide and iron, or, when added after induction was already established, promptly halted any further induction. These results suggest that the level of activity of heme oxygenase can reciprocally modulate intracellular heme levels and thus activity of delta-aminolevulinate synthase.

5-Aminolevulinate Synthetase↗
PubMed1991

Immunochemical studies of haem oxygenase. Preparation and characterization of antibodies to chick liver haem oxygenase and their use in detecting and quantifying amounts of haem oxygenase protein.

Monospecific polyclonal rabbit antibodies to a purified form of haem oxygenase of chick liver, showing sequence similarity to mammalian haem oxygenase-1, were raised and used to study characteristics of the oxygenase. The antibodies inhibited activity of the purified oxygenase, but not other enzyme components (NADPH:cytochrome reductase and biliverdin reductase) of the standard assay mixture of haem oxygenase. In addition, the antibodies inhibited activity of haem oxygenase in microsomes (microsomal fractions) from Cd(2+)-treated chick liver, spleen, testis and brain. Western (immuno-) blots of microsomal proteins of selected organs from chick, rat and man, and homogenates of chick-embryo liver-cell cultures, probed with the antibodies, showed a major protein with a molecular mass of 33-34 kDa and a lower-molecular-mass protein (28-29 kDa) of variable intensity. Studies with trypsin and selected proteinase inhibitors established that the smaller peptide was a proteolytic product of the larger. Treatment of chick-embryo liver-cell cultures with CdCl2, a potent inducer of haem oxygenase, increased the degree of proteinase-mediated cleavage of the 33 kDa protein to the lower-molecular-mass form. These results indicate that, under at least some conditions, such cultures should be homogenized in the presence of trypsin inhibitor to prevent proteolytic degradation of the enzyme and allow maximal expression of haem oxygenase activity. The antibodies also reacted with haem oxygenase from spleen, testis and brain of both chicks and rats, and the spleen of humans. A method for quantifying the amount of haem oxygenase protein was developed with use of slot-blots and laser densitometry; linearity was observed from 0 to 5 ng of haem oxygenase protein per slot, and the method was applied to sonicated cultured chick-embryo liver cells treated with Cd2+ (0.3 mM) or iron plus glutethimide. In both cases, increases in enzyme activity were of similar magnitude to increases in amounts of enzyme protein. Approximate amounts of haem oxygenase protein in microsomes of several organs from intact animals could also be estimated by the use of slot-blot-laser densitometry, and the amounts measured were increased by the addition of purified haem oxygenase to the microsomal preparations. Results of these studies indicated that haem oxygenase-1 could be detected in microsomes from all chick or rat organs studied, including testis and brain.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1991

Cloning, sequencing and expression of cDNA for chick liver haem oxygenase. Comparison of avian and mammalian cDNAs and deduced proteins.

A cDNA from a chick liver library that encodes for haem oxygenase has been cloned and sequenced. Positive clones were identified with monospecific antibodies to the purified enzyme from chick liver and a cDNA of rat haem oxygenase-1. The length of the cDNA is 1258 bases. An open reading frame of 888 bases was identified by comparison of nucleotide and amino acid sequences with those previously identified for haem oxygenase of mammalian or avian origin. The protein corresponding to this fragment of DNA is composed of 296 amino acid residues and has a molecular mass of 33,509 Da, which is similar to that previously estimated for haem oxygenase purified from chick liver. Unequivocal identification of this clone as that complementary to haem oxygenase was provided by (a) comparison of amino acid compositions and partial sequences with those previously established for the purified enzyme, (b) comparison with nucleotide and amino acid sequences for haem oxygenase from rat and human sources and (c) expression in Escherichia coli with production of high levels of mRNA, protein and haem oxygenase activity after exposure of the transfected bacteria to isopropyl beta-D-thiogalactopyranoside (IPTG). Overall, the similarity of chick haem oxygenase to rat and human haem oxygenase (nucleotides 66% and amino acids 62%) is moderately high. The region between proline-129 and alanine-157 is identical in all three enzymes, including histidine-135, which is proposed to play a key role in binding the substrate haem at the active centre of the enzyme. Northern blots also show that treatment of chicks with CdCl2, a potent inducer of haem oxygenase, results in increases in 1.65-1.70 kb mRNA, which hybridizes selectively to the full-length cDNA or to a synthetic 24-base oligonucleotide with sequence identical to that of a portion of the haem oxygenase cDNA. These results suggest that Cd-dependent induction of haem oxygenase is due to increased transcription of the gene or stabilization of its message.

Amino Acid Sequence↗
PubMed1991

Iron and the liver.

Iron is essential for life, but iron overload is toxic and potentially fatal. The liver is a major site of iron storage and is particularly susceptible to injury from iron overload, especially when (as in primary hemochromatosis) the iron accumulates in hepatocytes. Iron can be taken up by the liver in several forms and by several pathways including: (1) receptor-mediated endocytosis of diferric or monoferric transferrin or ferritin, (2) reduction and carrier-facilitated internalization of iron from transferrin without internalization of the protein moiety of transferrin, (3) electrogenic uptake of low molecular weight, non-protein bound forms of iron, and (4) uptake of heme from heme-albumin, heme-hemopexin, or hemoglobin-haptoglobin complexes. Normally, pathway 2 is probably the major one for uptake of iron by hepatocytes. Iron is stored in the liver in the cores of ferritin shells and as hemosiderin, an insoluble product derived from iron-rich ferritin. Iron in hepatocytes stimulates translation of ferritin mRNA and represses transcription of DNA for transferrin and transferrin receptors. The major pathologic effects of chronic hepatic iron overload are: (1) fibrosis and cirrhosis, (2) porphyria cutanea tarda, and (3) hepatocellular carcinoma. Although precise pathogenetic mechanisms remain unknown, iron probably produces these and other toxic effects by increasing oxidative stress and lysosomal lability. Vigorous efforts at diagnosis and treatment of iron overload are essential since the pathologic effects of iron are totally preventable by early vigorous iron removal and prevention of iron re-accumulation.

Ascorbic Acid↗
PubMed1991

Intravenous heme-albumin in acute intermittent porphyria: evidence for repletion of hepatic hemoproteins and regulatory heme pools.

The purpose of this study was to assess effects of heme administered intravenously, complexed to human serum albumin, on activities of the hepatic hemoproteins, cytochrome(s) P-450, and tryptophan pyrrolase, and on the size of the heme pool that regulates activity of 5-aminolevulinate synthase. Effects were compared in six normal women and four women with acute intermittent porphyria. All porphyric subjects over-excreted heme precursors and had histories of acute neurovisceral porphyric attacks. All subjects were placed on a constant daily diet that included at least 3 g carbohydrate/kg body weight and sufficient total intake to provide 1.4 times the estimated resting energy expenditure. Urinary excretions of 5-aminolevulinate, porphobilinogen, porphyrins, and metabolites of tryptophan were measured daily before, during, and after infusions of heme-albumin. In the porphyric subjects, intravenous heme [4 mg (6.1 mumol)/kg body weight (BWt) with equimolar albumin], given daily for 4 days, markedly reduced overexcretion of 5-aminolevulinate, porphobilinogen, and porphyrins, indicating repletion of the regulatory heme pool. The heme infusions also decreased mean urinary excretion of 5-hydroxyindoleacetic acid from 4.9 to 2.9 mg/g creatinine per day, suggesting increased activity of hepatic tryptophan pyrrolase, the rate-controlling enzyme for metabolism of tryptophan to products not in the serotonin-5-hydroxyindoleacetic acid pathway. Heme-albumin infusions were without detectable effects on excretions of heme precursors or tryptophan metabolites in normal subjects. In contrast, in both normals and porphyrics, heme-albumin infusions significantly increased rates of antipyrine metabolism (by 159% and 330%, respectively), suggesting increased activities of cytochrome(s) P-450 were produced by the infusions. The infusions were well tolerated; no subject developed thrombophlebitis or bleeding. We conclude that such infusions are safe and effective in repleting deficient heme pools and hemoproteins in patients with acute porphyria, and that activities of cytochrome(s) P-450 in normal subjects may also be increased by heme administration. The therapeutic effect of heme in acute porphyria probably relates to its ability to decrease overproduction of precursors of heme or serotonin, as the result of its increasing critical cellular heme pools.

Acute Disease↗
PubMed1991

A randomized, controlled trial of treatment of alcoholic hepatitis with parenteral nutrition and oxandrolone. I. Short-term effects on liver function.

The present studies were designed to provide careful measures of effects of oxandrolone, an anabolic steroid, intravenous nutritional supplementation, and the combination of these two treatments on liver functions, metabolic balances, nitrogen metabolism, and nutritional status in patients with moderate to severe alcoholic hepatitis. Of 43 patients originally recruited, 39 (19 men, 20 women) with typical clinical and laboratory features of alcoholic hepatitis (11 Child's-Pugh class B; 28 class C) were admitted to a metabolic unit and completed a 35-day three-phase protocol. Phase I was a 10-day baseline period of observation, during which routine and special quantitative tests of liver function (galactose and antipyrine metabolism), a 7-day elemental balance study, and a 15N, 13C-leucine metabolism study were done. Phase II was a 21-day treatment period during which patients were randomly assigned to receive one of four regimens: 1) standard therapy, consisting of abstinence, a balanced, nutritionally adequate diet, and multivitamins; 2) oxandrolone (20 mg orally four times a day) plus standard therapy; 3) nutritional supplementation, consisting of 2 L daily of 3.5% crystalline amino acids (in 5% dextrose), given by peripheral vein; or 4) a combination of oxandrolone and nutritional supplementation, along with standard therapy. Metabolic balances were repeated during phase II. Phase III was 2 or 3 days posttreatment, during which special studies of liver functions and volumes and leucine metabolism were repeated. All patients who completed phase I of study and were randomly allocated to one of the four treatment groups completed the subsequent two phases. Overall, with time, patients showed highly significant improvements in most clinical and laboratory features. For most standard laboratory tests (e.g., serum albumin, transferrin, prothrombin time) improvements were more marked in patients treated with nutritional supplementation and/or oxandrolone than in those given standard therapy alone. Liver volumes fell in all treatment groups, with greater improvement in those treated with nutritional supplementation. Improvements in galactose and antipyrine metabolism rates were significant only in those treated with nutritional supplementation or oxandrolone. Effects of treatments on metabolic balances, nitrogen metabolism, and measures of nutrition are described in this issue in a companion paper. We conclude that the addition of nutritional supplementation and oxandrolone to standard therapy of moderately severe or severe alcoholic hepatitis is well tolerated, and leads to more rapid improvement in the laboratory parameters measured.

Adult↗
PubMed1991

A randomized, controlled trial of treatment of alcoholic hepatitis with parenteral nutrition and oxandrolone. II. Short-term effects on nitrogen metabolism, metabolic balance, and nutrition.

Patients with moderately severe or severe alcoholic hepatitis, described in a companion paper in this issue, had serial studies of energy and protein metabolism and elemental balances before and during treatment for 21 days with one of four randomly assigned regimens: 1) standard therapy, consisting of abstinence, a balanced, nutritionally adequate diet, and multivitamins; 2) oxandrolone (20 mg orally four times a day) plus standard therapy; 3) nutritional supplementation, consisting of 2 liters of 3.5% crystalline amino acids in 5% dextrose given by peripheral vein (PPN) plus standard therapy; and 4) a combination of the other three treatments. Dietary and intravenous intakes and weights were recorded daily, and weekly averages were calculated. Anthropometric measurements and blood studies were done weekly; blood studies included white blood cell counts and differentials, serum prealbumin, transferrin, and total protein and plasma aminograms. Four-days complete balance studies and measures of 15N,1-13C-leucine metabolism also were performed at baseline and after the treatment period. Major findings were as follows: a) Intakes of total calories and protein were significantly higher in PPN-treated than in other groups. b) All patients had positive elemental balances, both at baseline and at the end of the treatment period. However, those treated with PPN (with or without oxandrolone) had higher positive balances of nitrogen, potassium, phosphorus, and magnesium, indicating improvement in lean body mass. c) Anthropometric measurements showed no significant changes, but measures of the visceral protein compartment (serum prealbumin, transferrin, total protein, total lymphocyte count) improved significantly with time. For most of these variables, increases were significantly greater in those treated with PPN with or without oxandrolone than in the other groups. However, for prealbumin, the increase was greatest in the oxandrolone-treated group d) PPN treatment produced dramatic increases in levels of branched-chain amino acids and improvement in the ratio of plasma branched chain to aromatic amino acids. Other treatments had no effect on plasma aminograms. e) Metabolism of 15N,1-13C-leucine was normal and was not affected significantly by treatment. Therapy with PPN and/or oxandrolone was tolerated well. We conclude that PPN has favorable effects on energy and protein metabolism in florid alcoholic hepatitis; oxandrolone has lesser effects, although it may exert some additional action and particularly increases serum prealbumin levels. The results support the use of nutritional supplementation in therapy of moderately severe or severe alcoholic hepatitis.

Adult↗
PubMed1990

Purification and characterization of heme oxygenase from chick liver. Comparison of the avian and mammalian enzymes.

A major inducible form of heme oxygenase (EC 1.14.99.3) was purified from liver microsomes of chicks pretreated with cadmium chloride. The purification involved solubilization of microsomes with Emulgen 913 and sodium cholate, followed by DEAE-Sephacel, carboxymethyl-cellulose (CM-52) and hydroxyapatite chromatography, and FPLC through Superose 6 and 12 columns operating in series. The final product gave a single band on silver-stained SDS/polyacrylamide gels (Mr = 33,000). Optimal conditions for measurement of activity of solubilized heme oxygenase were studied. In a reconstituted system containing purified heme oxygenase, NADPH-cytochrome reductase, biliverdin reductase and NADPH, the Km for free heme was 3.8 +/- 0.5 microM; for heme in the presence of bovine serum albumin (5 mol heme/3 mol albumin) the Km was 5.0 +/- 0.8 microM; and the Km for NADPH was 6.1 +/- 0.4 microM (all values mean +/- SD, n = 3). Oxygen concentration as low as 15 microM, with saturating concentrations of heme and NADPH, did not affect the reaction rate, indicating that the supply of oxygen is not involved in the physiological regulation of activity of the enzyme. The pH optimum of the reaction was 7.4; at 37 degrees C, the apparent Vmax was 580 +/- 44 nmol biliverdin.(mg protein)-1.min-1 and the molecular activity was 19.2 min-1. Biliverdin IXa was the sole biliverdin isomer formed. In the presence of purified biliverdin reductase, biliverdin was converted quantitatively to bilirubin. Addition of catalase to the reconstituted system decreased the breakdown of heme to non-biliverdin products and led to nearly stoichiometric conversion of heme to biliverdin. Activity of the enzyme in the reconstituted system was inhibited by metalloporphyrins in the following order of decreasing potency: tin mesoporphyrin greater than tin protoporphyrin greater than zinc protoporphyrin greater than manganese protoporphyrin greater than cobalt protoporphyrin. Protoporphyrin (3.3 or 6.6 microM) (and several other porphyrins) and metallic ions (100 microM) alone had little if any inhibitory effect, except for Hg2+ which inhibited by 67% at 10 microM and totally at 15 microM. Following partial cleavage, fragments of the purified enzyme were sequenced. Comparison of sequences to those derived from cDNA sequences for the major inducible rat and human heme oxygenase showed 69% and 76% similarities, respectively. The histidine residue at position 132 of rat heme oxygenase-1 and the residues (Lys128-Arg136) flanking His132 were conserved in all three enzymes, as well as in the corresponding portion of a fourth less highly similar rat enzyme, heme oxygenase-2.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1990

Prevalence and prediction of osteopenia in chronic liver disease.

To obtain information on the prevalence and clinical and laboratory correlates of osteopenia in patients with chronic liver disease, we measured bone densities and 30 selected laboratory variables in 133 subjects (70 men, 63 women) with liver disease. Thirty-two had alcoholic liver disease, 18 had primary biliary cirrhosis, 16 had primary sclerosing cholangitis, 48 had other forms of cirrhosis (cryptogenic, posthepatic) and 19 had chronic hepatitis or fibrosis without cirrhosis. Bone densities of the lumbar spine and three sites of the proximal femur (neck, Ward's triangle, greater trochanter) were estimated by dual-photon absorptiometry. Bone densities at all sites were significantly correlated to one another (r = 0.4 to 0.9; 95% confidence intervals = 0.24-0.54 to 0.81-0.90; p less than 0.0001 for all). Compared with an age- and gender-matched reference group, patients with liver disease had highly significant decreases in bone densities (greater than 2 standard deviations below control values; p less than 0.0008 at all sites). Decreases were particularly marked (24% to 42%) at Ward's triangle, the site of the femoral neck particularly prone to fracture. The prevalence of decreased bone densities ranged from 10% to 56%, depending on the site studied and the nature of the liver disease. Among 30 laboratory variables studied, there were significant (p less than 0.05) correlations with bone densities at more than one site for urinary creatinine (r = 0.21, 0.25), urinary calcium (r = -0.18, -0.23), serum total alkaline phosphatase (r = -0.18, -0.27) and the liver-1 isozyme of serum alkaline phosphatase (r = -0.19, -0.26).(ABSTRACT TRUNCATED AT 250 WORDS)

Bone Density↗
PubMed1990

Usefulness and limitations of laboratory and hepatic imaging studies in iron-storage disease.

Liver biopsy with measurement of hepatic iron concentration is the most certain procedure for evaluation of iron-storage disease, although use of computed tomography and magnetic resonance imaging procedures recently have been proposed as alternative, noninvasive methods for estimating the degree of iron overload. The results of these imaging procedures were compared with those of other noninvasive techniques and liver biopsies in 48 patients. Final diagnoses, based on synthesis of clinical and laboratory data, included (a) primary hemochromatosis (n = 25; 19 homozygous, 6 heterozygous); (b) secondary hemochromatosis (n = 7); (c) alcoholic liver disease (n = 11); (d) chronic active hepatitis (n = 3); and (e) other (n = 2). Serum ferritin and computed tomography or magnetic resonance scanning had 100% sensitivity in detecting hepatic iron overload more than fivefold above the upper limit of normal (greater than 10.7 mumol Fe/100 mg dry liver) but did not detect lesser degrees of iron overload reliably, including those found in 6 of 13 patients with untreated homozygous primary hemochromatosis and 3 of 7 with secondary hemochromatosis. Computed tomography and magnetic resonance imaging were more specific than ferritin (64% and 92% vs. 21%) in the detection of iron excess, more than five times the upper limit of normal. Among magnetic resonance imaging measures, the ratio of the second echo signal intensities of liver to paraspinous muscle was the most sensitive and most specific for detection of this degree of iron overload. The degree of correlation between hepatic iron concentration and results of noninvasive laboratory or imaging studies were insufficient to permit prediction of hepatic iron content by noninvasive studies alone. It is concluded that computed tomography or magnetic resonance scanning as currently usually used is not cost-effective in routine evaluation of iron overload, although these imaging procedures may play a role in patients in whom liver biopsy is contraindicated. Because of their low cost and ready availability, serum ferritin and transferrin saturation tests remain the preferred screening studies for iron overload. Liver biopsy with quantitative iron measurement remains the study of choice for the definitive diagnosis of hemochromatosis.

Biopsy↗
PubMed1990

Biochemistry and pharmacology of S-adenosyl-L-methionine and rationale for its use in liver disease.

The major biological functions of S-adenosyl-L-methionine (SAMe) include methylation of various molecules (transmethylation) and synthesis of cysteine (trans-sulphuration). A stable double salt of SAMe has been found to be effective in intrahepatic cholestasis. The mechanism of its therapeutic effect is not fully understood but presumably involves methylation of phospholipids. Methylation of plasma membrane lipids may affect membrane fluidity and viscosity, which modulate the activities of a number of membrane-associated enzymes, for example, the activity of enzymes involved in Na+/Ca++ exchange (e.g. sarcolemmal vesicles), Na+/K+ adenosine triphosphatase (ATPase) [e.g. hepatocyte plasma membranes], and Na+/H+ exchange (e.g. plasma membranes of colonic cells). Recently, patients with cirrhosis were shown to have an acquired metabolic block in the hepatic conversion of methionine to SAMe. These patients, when administered conventional elemental diets, develop abnormally low plasma concentrations of cysteine and choline, 2 nonessential nutrients present in low concentrations in most elemental diets. These low concentrations probably reflect systemic deficiencies attributable to reduced endogenous syntheses of cysteine and choline caused by limited availability of hepatic SAMe. Such cirrhotic patients are often in negative nitrogen balance and have abnormal hepatic functions, which are corrected by cysteine and choline supplements. Noncirrhotic patients on parenteral elemental diets also become deficient in cysteine and choline. Consequently, these patients may require SAMe as an essential nutrient to normalise their overall hepatic transmethylation and trans-sulphuration activities.

Animals↗
PubMed1990

Four-hour measurement of urinary iron excretion after deferoxine treatment: a rapid, simple method for study of iron excretion.

In a group of 56 patients being evaluated for iron storage disease, all of whom underwent liver biopsy with quantitative measurement of liver iron concentration, urinary excretion of iron was measured following intramuscular injection of deferoxamine (10 mg/kg body weight). Urine was collected in two portions, 0-4 h and 4-24 h after deferoxamine administration. Iron excretion in the first 4 h was closely correlated to that over the entire 24-h period (r = 0.88, p less than 0.0001, 95% confidence interval = 0.80-0.93), indicating that the 4-h deferoxamine urinary excretion test can be adapted readily to an outpatient clinical setting, helping to insure complete and accurate urine collections. Correlation of 0- to 4-h urinary iron excretion and quantitative liver iron (r = 0.335), although higher than for the full 24 h (r = 0.192), was not sufficient to accurately predict hepatic iron concentration in the individual patient.

Deferoxamine↗
PubMed1989

Heme catabolism in cultured hepatocytes: evidence that heme oxygenase is the predominant pathway and that a proportion of synthesized heme is converted rapidly to biliverdin.

Heme oxygenase has been considered to be involved in the predominant pathway of heme degradation in vivo. However, alternative pathways involving cytochrome P-450 reductase, and lipid peroxidation, have previously been demonstrated in vitro, and studies with cultured rat hepatocytes were interpreted to show a majority of endogenous hepatic heme breakdown by non-heme oxygenase pathways. To clarify the pathway of heme breakdown in hepatocytes and the role of heme oxygenase in this process, cultured hepatocytes were pre-labelled with 5-[5-14C]aminolevulinate [( 14C]ALA). Radioactivity in heme, carbon monoxide, and bile pigments was measured for 8-24 h after the removal of [14C]ALA. In cultured chick embryo hepatocytes, which lack biliverdin reductase, the rate of production of biliverdin IXa was closely similar to the rate of catabolism of exogenous heme and radioactivity in carbon monoxide and biliverdin IXa was similar to the loss of radioactivity from endogenous heme. These results support the conclusion that heme breakdown occurred predominantly, if not solely, by heme oxygenase. Also, no evidence of non-heme oxygenase pathways was found in the presence of tin protoporphyrin, an inhibitor of heme oxygenase or mephenytoin, an inducer of both cytochrome P-450 and heme oxygenase. Similarly, in untreated cultured rat hepatocytes, radioactivity in carbon monoxide corresponded with loss of radioactivity in endogenous heme. In other experiments with chick hepatocyte cultures, rates of heme synthesis and breakdown were measured, and data were fitted to various models of hepatic heme metabolism. The results observed were consistent only with models in which an appreciable fraction (control cells, 17%, mephenytoin treated cells, 41%) of the newly synthesized heme was degraded rapidly to biliverdin.

Animals↗
PubMed1989

Mechanism of iron potentiation of hepatic uroporphyria: studies in cultured chick embryo liver cells.

Effects of iron were studied in cultured chick embryo liver cells to help elucidate the effect of hepatic iron in the human disease porphyria cutanea tarda and in toxic porphyria caused by chemicals. These cultures have proven useful because (a) phenobarbital and phenobarbital-like drugs induce a common form(s) of cytochrome P-450 (P-450-phenobarbital) in these cultures; (b) 20-methylcholanthrene and certain other polycyclic hydrocarbons induce a different form(s) (P-450-methylchol-anthrene), and (c) uroporphyria can be produced rapidly by exposure to suitable chemicals. In these cultures, treatment with iron alone did not produce porphyrin accumulation, and treatment with iron + 5-aminolevulinate caused accumulation of protoporphyrin, as did treatment with 5-aminolevulinate alone. However, treatment with phenobarbital-like drugs and iron, the latter at a concentration as low as 0.2 microM, led to accumulation of uro- and heptacarboxylporphyrins. Potentiation of uroporphyrin accumulation by iron began before there was a detectable synergistic increase in activity of 5-aminolevulinate synthase, the rate-controlling enzyme of heme synthesis. In contrast, treatment of cultures with 20-methylcholanthrene, in the presence or absence of iron, did not result in uroporphyrin accumulation or an increase in the activity of 5-aminolevulinate synthase. Uroporphyrinogen decarboxylase activity was unchanged by drug and iron treatments. Inhibitors of P-450-phenobarbital, SKF525A and piperonyl butoxide, as well as cadmium and cycloheximide prevented the porphyrin accumulation produced by glutethimide + iron, even though, except with cycloheximide, these substances further increased 5-aminolevulinate synthase activity. In vitro, uroporphyrin was oxidized autocatalytically by iron. In intact hepatocytes, even low concentrations of iron (0.2 to 20 microM), in the presence of a form of cytochrome P-450 induced by phenobarbital-like chemicals, produces uroporphyria primarily by enhancing uroporphyrinogen oxidation, not by inhibition of the decarboxylase. Induction of 5-aminolevulinate synthase amplifies the porphyrin overproduction.

5-Aminolevulinate Synthetase↗
PubMed1989

The porphyrias.

The porphyrias are metabolic disorders in which there are excessive accumulation and excretion of porphyrins and porphyrin precursors. Each of the porphyrias has a specific enzyme defect in the pathway of heme biosynthesis that explains the pattern of biochemical abnormalities that occur. However, some patients have the enzyme defect but do not have clinical or biochemical manifestations, indicating that other factors (e.g., demand for increased heme biosynthesis) are also important in causing disease expression. The major clinical manifestations are neurologic dysfunction and photosensitivity. The precise cause of the neurologic dysfunction has not been defined, but the likely possibilities are overproduction of delta-aminolevulinic acid, which may act as a neurotoxin, or a deficiency of heme (or both). The cutaneous lesions in the porphyrias are due to the photo-sensitizing and other effects of porphyrins that are deposited in the skin or are circulating in dermal blood vessels. Therapy is directed to modify the biochemical abnormalities. Most importantly, intravenous administration of hematin is used in the treatment of acute attacks of neurologic dysfunction. Prevention remains a cornerstone in management of patients with porphyria, and those with gene defects should be counseled regarding factors that precipitate acute attacks.

Heme↗