Paraparesis, hypermanganesaemia, and polycythaemia: a novel presentation of cirrhosis.

Progressive myelopathy is a rare complication of chronic hepatic disease which has never been reported in the paediatric age group. We describe the 11 year course of an adolescent male with hepatic myelopathy caused by cryptogenic micronodular cirrhosis. His condition has been associated with persistent polycythaemia and extraordinary increases of whole blood manganese, with magnetic resonance imaging evidence of manganese deposition within the basal ganglia and other regions of the brain. The patient has developed neither liver failure nor parkinsonism. The pathophysiological bases of this multiorgan system disorder are described.

Biochemical and viral response to consensus interferon (CIFN) therapy in chronic hepatitis C patients: effect of baseline viral concentration. Consensus Interferon Study Group.

OBJECTIVE: The effect of baseline viral concentration on response was assessed as part of a multicenter phase 3 trial evaluating the safety and efficacy of CIFN therapy for chronic HCV infection. METHODS: Patients (n = 472) received either CIFN 9 microg or IFN alpha-2b 3 MU subcutaneously t.i.w. for 24 wk, followed by 24 wk of observation. RESULTS: Efficacy was assessed by the percentage of patients who achieved normal ALT values or undetectable HCV RNA values (using RT-PCR with a sensitivity of 100 copies/ml). There was a clear relationship between baseline viral concentration and either ALT or HCV RNA response; patients with lower titer HCV RNA had better response rates. End-of-treatment HCV RNA responses were better for patients with low viral concentrations treated with CIFN (51%) than for patients treated with IFN a-2b (31%) (p = 0.03). ALT responses in patients with low viral concentrations were 60% for CIFN-treated patients and 27% for IFN alpha-2b-treated patients (p < 0.01) at the end of treatment. Patients with high titer HCV RNA were more likely to have a sustained HCV RNA response after treatment with CIFN 9 microg, compared with those treated with IFN alpha-2b (7% vs 0%, p = 0.03). CONCLUSIONS: Both genotype and baseline viral concentration were independent factors that affected response to interferon.

Utility of hepatitis C virus serotypes in predicting response to treatment of chronic hepatitis C. Consensus Interferon Study Group.

Hepatitis C virus (HCV) genotyping has been shown to predict response to interferon, but is expensive. HCV serotyping is less expensive and simpler, and may be similarly useful. Using data from a large, randomized trial comparing consensus interferon (CIFN) and interferon alfa-2b (IFN alfa-2b) in patients with chronic HCV, we evaluated response rates based on HCV serotypes versus genotypes. Patients included in this analysis received subcutaneous injection of 9 microg CIFN (n = 232) or 3 MU IFN alfa-2b (n = 240) three times weekly for 24 weeks followed by 24 weeks of observation. Serum HCV RNA concentrations were measured regularly during treatment and at the end of both the treatment and post-treatment periods. Response to interferon was similar for HCV antibody types and their corresponding genotypes. The end-of-treatment HCV RNA rate of response (defined as undetectable serum on two consecutive assessments) was 29% for serotype 1 versus 24% for genotype 1 after CIFN; and 14% versus 15%, respectively, after IFN alfa-2b. Independently of treatment, patients infected with serotype or genotype 2 or 3 had a better therapeutic response than those infected with serotype or genotype 1. Similar results were obtained based on HCV antibody typing and genotyping, suggesting the potential of the former for predicting response to interferon.

Diagnostic performance of a receptor-binding radiopharmacokinetic model.

UNLABELLED: The aim of this study was to determine which measurement obtained from a radiopharmacokinetic model of a receptor-binding radiotracer provides the highest diagnostic performance for the detection of diffuse hepatocellular disease. METHODS: Twenty-seven healthy subjects and 46 patients with diffuse hepatocellular disease were studied with the receptor-binding radiopharmaceutical, 99mTc-galactosyl-neoglycoalbumin. A radiopharmacokinetic model was used to produce estimates of receptor concentration [R]o, the scaled forward-binding rate constant Kb, hepatic plasma volume, Vh, extrahepatic plasma volume, Ve and hepatic plasma flow, F. Receiver operating characteristic analysis of each model estimate was conducted. RESULTS: Receptor concentration [R]o and the metrics [R]o/tbw and kb[R]o[R]o/tbw provided the best discrimination between healthy and diseased liver. The forward-binding rate constant kb and the metrics F/Ve and Vh/tbw provided no discrimination. CONCLUSION: Based on simplicity and higher measurement precision, [R]o was selected as the most accurate index of hepatic function.

Evaluation of hepatocellular function by way of receptor-mediated uptake of a technetium-99m-labeled asialoglycoprotein analog.

UNLABELLED: We have developed a quantitative functional imaging study of the liver using a radiolabeled asialoglycoprotein analog, Tc-galactosyl-neoglycoalbumin. Heart and liver time-activity data can be transformed by automated kinetic analysis into asialoglycoprotein hepatocyte receptor concentration. Twenty-eight healthy controls, 46 patients with noncholestatic chronic liver injury and 11 patients with primary biliary cirrhosis were studied. Liver function was also assessed by Pugh modified-Child-Turcotte criteria, 14C-aminopyrine breath test and indocyanine green clearance (24 patients). RESULTS: (a) In normal controls with a Child-Turcotte criteria score of 5, receptor concentration ranged from 0.63 to 1.19 mumol/L, with a mean 0.83 +/- 2 S.D. 0.06 mumol/L, which was significantly higher (p < 0.001) than that of the patient group (mean receptor concentration = 0.44 +/- 2 S.D. 0.04 mumol/L). In cirrhotic patients with Child-Turcotte criteria score of 5, the mean receptor concentration was 0.60 +/- 2 S.D. 0.07 mumol/L, which was significantly lower than controls (p < 0.01). In end-stage cirrhosis (Child-Turcotte criteria score 11 to 15), a group in which patients died or required orthotopic liver transplantation within 1 yr, the mean receptor concentration was 0.35 +/- 2 S.D. to 0.07 mumol/L. The sensitivity and specificity for receptor concentration in relation to liver disease, with values above 0.65 mumol/L being normal, were 0.96 and 0.88, respectively. Receptor concentration correlated well with Child-Turcotte criteria score (r = 0.78, p = < 0.001), with aminopyrine breath test (r = 0.75, p = < 0.001) and with indocyanine green clearance (r = 0.88, p = < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Uroporphyrinogen decarboxylases from human erythrocytes: purification, complete separation and partial characterization of two isoenzymes.

1. Two distinct molecular forms of uroporphyrinogen decarboxylase have been completely separated and highly purified from human erythrocytes. 2. Each protein, with molecular masses of about 52-54 kDa and 35 kDa, are apparently composed of a single polypeptide chain. 3. They may form a functional decarboxylating complex for heme biosynthesis.

Free radical mechanism of oxidation of uroporphyrinogen in the presence of ferrous iron.

Human porphyria cutanea tarda is an unusual consequence of common hepatic disorders such as alcoholic liver disease. Hepatic iron plays a key role in the expression of the metabolic lesions, i.e., defective hepatic decarboxylation of porphyrinogens, catalyzed by uroporphyrinogen decarboxylase. This prompted the present study to determine the in vitro effects of iron on the uroporphyrinogen substrate in the absence and presence of atmospheric oxygen. We observed that (i) unless oxygen is the limiting reactant, autoxidation of ferrous iron and iron-catalyzed oxidation of uroporphyrinogen occurred soon after initiating the reaction at pH 7.4 and 30 degrees C in buffers which are non- or poor chelators of iron; (ii) the rates of uroporphyrinogen oxidation were proportional to the initial concentration of ferrous ion; (iii) about 70% of the oxidations of uroporphyrinogen were accountable due to a free-radical chain reaction pathway involving superoxide radical and hence inhibitable by superoxide dismutase; (iv) uroporphyrinogen could be further oxidized to completion by the hydroxyl radical since the reaction was partially inhibited by both mannitol and catalase which prevent hydroxyl radical production; (v) the oxidizing effects of ferric ion on uroporphyrinogen were none or negligible as compared to those of ferrous ion. Ferric was reduced to ferrous ion in the presence of dithiothreitol. When the ferrous ion thus formed was reoxidized in the presence of atmospheric oxygen, minor but definite oxidations of both uroporphyrinogen and dithiothreitol were observed. The oxidations of Fe2+ and uroporphyrinogen could be blocked by 1,10-phenanthroline, a ferrous iron chelator. The data suggest that ferrous is the reactive form of iron that may contribute to pathogenic development of the disease by irreversibly oxidizing the porphyrinogen substrates to nonmetabolizable porphyrins, which accumulate in porphyric liver.

Defective human erythrocyte uroporphyrinogen decarboxylase in familial porphyria cutanea tarda: the metabolic lesion or the result of endogenous porphyrinemia?

We have demonstrated that oral charcoal therapy is as effective as therapeutic phlebotomy in reducing porphyrinemia in porphyria cutanea tarda. The effects of immediate and sustained reduction of porphyrinemia on the catalytic properties of partially purified (approximately 200-fold) preparations of red cell uroporphyrinogen decarboxylase of a patient with familial porphyria cutanea tarda were studied. All populations of the patient's red cells exhibited defective enzyme activity, and the apparent Michaelis constants (Km) determined with penta-, hepta-, and octa-carboxylic I porphyrinogen substrates were approximately 3-4 times higher as compared to the normal controls. Mixing experiments (normal and defective enzyme), and preincubation of the normal enzyme with porphyric plasma prior to purification, yielded data supporting the concept that the catalytic defects of red cell uroporphyrinogen decarboxylase in familial porphyria cutanea tarda are independent of interactions between circulating endogenous porphyrins and the enzyme.

Evidence for two uroporphyrinogen decarboxylase isoenzymes in human erythrocytes.

In animals and plants, uroporphyrinogen decarboxylase catalyzes the stepwise decarboxylations of uroporphyrinogen, the precursor of heme and chlorophyll. To better understand its metabolic roles, we characterized the enzyme purified to electrophoretic homogeneity (about 11,000-fold) from human erythrocytes by a novel uroporphyrin-sepharose affinity chromatographic method. Native polyacrylamide disc gel electrophoresis of the purified enzyme preparation showed two bands detected by staining either for protein or with uroporphyrin-I. Each individual protein eluted from the gel when subjected to re-electrophoresis on SDS-polyacrylamide gel, appeared as a single protein band with molecular masses of approximately 54,000 and approximately 35,000 daltons respectively. Both proteins were able to catalyze all four decarboxylation steps, though the ratios of enzyme activity using octa-, hepta-, hexa- to pentacarboxylic porphyrinogen substrates were distinctly different. Also, their kinetic analysis with heptacarboxylic porphyrinogen-I substrate provided distinctly different apparent Michaelis constants. This provides the first evidence that decarboxylations of uroporphyrinogen to coproporphyrinogen are catalyzed by two isoenzymes.

Selective in vivo tumor localization of uroporphyrin isomer I in mouse mammary carcinoma: superiority over other porphyrins in a comparative study.

Twenty-eight porphyrins were evaluated for tumor localization as delineated by fluorescence using a transplantable KHJJ mammary carcinoma in BALB/c mice as the tumor model. Five of the 28 porphyrins were found to localize and of these, one, i.e., uroporphyrin I (UROP I), showed a higher tumor:skin ratio than any of the others; moreover, as no measurable UROP I was present in the gut, the tumor:intestinal porphyrin ratio under the conditions of assay was infinity. Because hematoporphyrin derivative (HPD), a complex mixture of porphyrins has been studied extensively as a tumor localizer, we compared HPD with UROP I at differing doses (2-40 mg/kg) and at different times (3-96 h) following i.v. administration. Dose response curves showed tissue levels of porphyrin to plateau out at doses above 20 mg/kg. Peak tumor HPD and UROP I levels attained 6-18 h after i.v. administration (40 mg porphyrin/kg) were comparable, but tumor retention of HPD over the ensuing 96 h was higher. The ratio of UROP I in tumor compared to skin was significantly greater throughout the period of observation. At all times, no UROP I was detectable in gastrointestinal mucosa. At differing doses (10-40 mg/kg), the tumor:skin ratio for HPD ranged from 1.47-1.85, and for UROP I from 6.06-12.33. As a function of time (6-72 h), the tumor:skin ratios respectively were 1.03-2.38, and 11.9 to infinity. At all times, the tumor:colon mucosa ratio at different doses for HPD approached 1 and for UROP I was infinity. We conclude that the greater specificity of tumor uptake by UROP I and its lack of retention by gut mucosa warrants further study to determine its potential clinical application as a diagnostic marker, particularly for early mucosal cancer, and in photoradiation therapy.

In vitro studies of the mechanism of inhibition of rat liver uroporphyrinogen decarboxylase activity by ferrous iron under anaerobic conditions.

Human porphyria cutanea tarda (PCT) is an unusual consequence of common hepatic disorders such as alcoholic liver disease and iron overload, where hepatic iron plays a key role in the expression of the metabolic lesion, i.e., defective hepatic decarboxylation of porphyrinogens. In this investigation, kinetic studies on a partially purified rat liver uroporphyrinogen decarboxylase have been conducted under controlled conditions to determine how iron perturbs porphyrinogen decarboxylation in vitro. The enzyme, assayed strictly under anaerobic conditions in the dark, was inhibited progressively by ferrous iron. Approximately 0.45 mM ferrous ammonium sulfate was required to observe about 50% inhibition of enzyme activity measured with uroporphyrinogen I as substrate. We showed that (a) all the steps of enzymatic decarboxylation (octa-, hepta-, hexa-, and pentacarboxylic porphyrinogen of isomer I series) were inhibited by ferrous iron. The inhibition was competitive with respect to uroporphyrinogen I and III substrates; (b) the cations, e.g., Fe3+ and Mg2+, had no effect, whereas sulfhydryl group specific cations and compounds such as Hg2+, Zn2+, p-mercuribenzoate, and 5,5'-dithiobis(2-nitrobenzoate) all inhibited the enzyme; (c) the enzyme could be protected from inhibition by Fe2+ and p-mercuribenzoate by preincubation with pentacarboxylic porphyrinogen, a natural substrate and competitive inhibitor. These data suggest for the first time a direct interaction of ferrous iron with cysteinyl residue(s) located at the active site(s) of the enzyme.

Biochemical diagnosis and monitoring therapeutic modulation of disease activity in an unusual case of congenital erythropoietic porphyria.

We describe the methodology used for quantifying and characterizing porphyrins in various tissues and in excreta, in the diagnosis and monitoring of the therapeutic modulation of biochemical disease activity in a 53-year-old white man who has a rare form of familial porphyria cutanea tarda with bone marrow rather than hepatic expression of the disease. Liquid-chromatographic and thin-layer chromatographic analyses of the patients's urine and skin showed predominantly heptacarboxylic porphyrin and uroporphyrin, whereas his stool and bile contained isocoproporphyrin and coproporphyrin as the major products. The data reflect defective uroporphyrinogen decarboxylation. Both analytical methods gave quantitatively similar results for urinary and fecal porphyrins. A triple-lumen perfusion study of samples procured both at the ampulla of Vater and 15 cm downstream provided data for porphyrins excreted in the bile and their reabsorption in the small intestine. We evaluated: suppression by hypertransfusion of bone marrow overproduction of porphyrins and reduction of enteral absorption of porphyrins by orally administered charcoal (Acta Char) and cholestyramine.

A potential biochemical explanation for the genesis of porphyria cutanea tarda. Studies on the inherent biochemical defect in highly purified human erythrocyte uroporphyrinogen decarboxylase and its amplification by iron.

Familial porphyria cutanea tarda (PCT) is a photocutaneous disease in which subnormal activity of uroporphyrinogen decarboxylase is observed both in the liver and red cells. Hepatic iron plays a key role in the genesis of overt biochemical and clinical PCT. In this report, we have studied the properties of 10 000-fold purified erythrocyte uroporphyrinogen decarboxylase preparations from two familial PCT patients and a non-porphyric control subject. The apparent Michaelis constants (Km), determined by using uroporphyrinogen III substrate, were approx. 3.2-times higher for the enzyme from the diseased subjects (Km = approximately 1.0 microM) as compared to the normal (Km = 0.3 microM). Though both abnormal and normal enzymes were inhibited progressively with increasing concentrations of iron, the enzymes from diseased subjects exhibited greater susceptibility e.g. 0.1 mM Fe2+ inhibited the former about 50% and the latter about 20%. These observations suggest that the inherent biochemical defect in PCT is the reduced enzyme-substrate affinity and the intrinsic abnormal conformation renders the PCT enzyme particularly susceptible to inhibition by iron.

The interaction of tumour-localizing porphyrins with collagen, elastin, gelatin, fibrin and fibrinogen.

We have already reported in Balb C mouse transplantable mammary carcinoma, that uroporphyrin I and III are superior as tumour localizers when compared to hematoporphyrin derivative and a derivative thereof, photofrin II. This study compares the binding of porphyrins to proteins which may be found in tumour cells or stroma to investigate whether there is a common binding determinant. Coproporphyrin III and deuteroporphyrin IX which are non-tumour localizing porphyrins, were also part of the comparative study. The interaction of these porphyrins with acid soluble collagen and acid insoluble collagen, elastin, and fibrin was evaluated, and the binding of uroporphyrin isomers I and III and deuteroporphyrin IX to gelatin and fibrinogen, was also determined. The results suggest that collagen, especially the acid soluble form, and gelatin preferentially bind the four porphyrins which localize in mammary carcinoma tissue. The well reported observations that malignant epithelial cells, including breast cancer, produce collagen and contain a rate-limiting enzyme in collagen biosynthesis would support the notion that de novo synthesis of this protein may in part govern the tumour uptake and retention of porphyrins. Elastin, fibrinogen and fibrin showed non-discriminant binding to the porphyrins under study.

Hematologic and hepatic manifestations of the cutaneous porphyrias.

This chapter has dealt with five photocutaneous forms of human porphyria. The forms are a diverse group of disorders with many different hematologic, hepatologic, and neurologic manifestations. In essence, most photocutaneous porphyrias occurring in childhood will relate to congenital erythropoietic porphyria or protoporphyria. The nature of the skin lesions and a study of the heme precursor profile in red cells, plasma, urine, and feces should easily distinguish these two conditions. CEP is a disease wherein photomutilation is a dominant concern and aggressive new approaches of therapy also have been discussed. In protoporphyria, the dermatologic problem is less severe and the dermatologist should be aware that a subset of patients could develop active liver disease that may lead to fatal cirrhosis. Novel approaches of therapy have been briefly alluded to. With regard to postpubertal photocutaneous porphyria, the classic porphyria cutanea tarda syndrome is associated with liver disease, usually alcoholic with siderosis, and the treatment by phlebotomy to reduce hepatic iron is highly effective. The potential danger of liver carcinoma has been discussed. In subsets of porphyria cutanea tarda, this can be an endemic disease relating to environmental factors, ie, ingestion of polyhalogenated hydrocarbons. The biochemical diagnosis can be attained by fairly straight-forward solvent extraction analyses of urine and feces, showing the dominance of uroporphyrin excretion in the urine and coproporphyrin in the feces. Chromatographic techniques in plasma, bile, and feces reveal a PCT-specific porphyrin: isocoproporphyrin. Rare subtypes with hematologic manifestations, ie, hepatoerythropoietic porphyria and CEP, indicate the wide spectra of disorders that might be associated with a spontaneous deficiency of uroporphyrinogen decarboxylase activity. These latter syndromes are, however, rare. Two hereditary hepatic porphyrias, ie, autosomal dominantly inherited VP and HCP, have been briefly discussed. The hepatic lesion is metabolic, not morphologic, and its expression by the liver relates to its adaptive response to induction of microsomal hemoproteins by a variety of exogeneous and endogeneous compounds, eg, drugs and hormones. Photocutaneous lesions of HCP and VP are identical to PCT, the latter having no neurologic sequelae. In the former two, however, exposure of persons to drugs, such as the hydantoins and barbiturates, can lead to potentially fatal acute porphyric attacks.(ABSTRACT TRUNCATED AT 400 WORDS)

Reduced substrate affinity for human erythrocyte uroporphyrinogen decarboxylase constitutes the inherent biochemical defect in porphyria cutanea tarda.

The pathogenesis of human porphyria cutanea tarda (PCT) is associated with an intrinsic abnormality of the uroporphyrinogen decarboxylase enzyme. To characterize this, we studied the kinetic properties of the red cell enzyme procured from patients with various forms of PCT and non-porphyric controls. The enzyme activity (units/mg hemoglobin) in the red cell hemolysate was close to normal in sporadic PCT but about 75% diminished in the familial PCT. The Michaelis constants (Km) of 200-fold purified red cell enzyme preparations, determined by using pentacarboxylic porphyrinogen I and uroporphyrinogen I as substrates, were more than 3.8-4.0 times higher, and the maximum velocity (Vmax) was about 70% diminished in familial PCT, whereas the Km was about 1.7-1.9 times higher and the Vmax was more or less normal for sporadic PCT. These observations suggest for the first time that the primary lesion in familial PCT is a genetically determined kinetic abnormality of uroporphyrinogen decarboxylase which appears to be different from the sporadic form of the disease.

Dual mechanism of inhibition of rat liver uroporphyrinogen decarboxylase activity by ferrous iron: its potential role in the genesis of porphyria cutanea tarda.

Hepatic iron overload amplifies the uroporphyrinogen decarboxylase enzyme defect in human porphyria cutanea tarda. To understand its mechanism, we studied the effects of iron on the enzyme activity from rat liver cytosol. Enzyme activity was inhibited about 50% by 0.10 mM Fe2+ or by 0.16 mM Zn2+ directly regardless of whether the cations were added immediately, or were first preincubated for 2 h at 37 degrees C in the absence or presence of oxygen. Cysteine (6.7 mM) protected the enzyme from inhibition by Fe2+ under strictly anaerobic preincubation conditions; cysteine also protected enzyme inhibition by Zn2+ even in the presence of oxygen. Under aerobic conditions, cysteine enhanced the inhibition by Fe2+ to about 70%. This additional 20% inhibition was reversed by vitamin E, an antioxidant. The results suggest dual inhibitory effects of iron (a) by direct interaction of Fe2+, as well as Zn2+, with the essential sulfhydryl group(s) of the enzyme and (b), indirectly, due to generation of free radicals in the presence of oxygen and an electron donor such as cysteine. These radicals might interact directly with the enzyme and/or oxidize the porphyrinogen substrates to nonmetabolizable porphyrins, which accumulate in porphyric patients.