[Sclerodermatous changes in porphyria cutanea tarda: six cases]. / Lésions sclérodermiformes dans la porphyrie cutanée tardive : six observations.

BACKGROUND: The clinical features of porphyria cutanea tarda (PCT) are usually distinctive and include blistering on sun-exposed areas, fragile skin, hypertrichosis and hyperpigmentation. Sclerodermatous changes are much less common, and may either reveal PCT or else appear later. We carried out a retrospective study of the files of six female patients presenting such lesions. PATIENTS AND METHODS: Six women (age: 45 to 72 years) were referred for sclerodermatous lesions on sun-exposed areas of the upper body. In four patients, these lesions revealed PCT and in the remaining two patients they were indicative of previously treated but relapsing PCT. Four had sclerodermatous skin changes mimicking morphea of the neck and neckline, the top of the back and the face, while one presented more diffuse facial and cervical sclerosis. Associated alopecia was seen in three patients. The last patient presented isolated sclerodermiform alopecia. Associated malar hypertrichosis was seen in five cases and facial hyperpigmentation was noted in three cases. Four exhibited no blisters, cutaneous fragility, milia or photosensitivity. Histological findings were consistent with morphea or scleroderma in all cases. All patients presented abnormal liver tests: cirrhosis was present in four cases (primitive biliary cirrhosis, alcoholic cirrhosis and hepatitis C) and fatty liver in two cases. In four cases, there was excessive alcohol intake. Uroporphyrin levels were above the normal range in all cases. Local corticosteroid therapy associated with phlebotomy and/or low-dose hydroxychloroquine resulted in complete normalisation of porphyrin levels in four patients, with complete resolution of the cutaneous lesions in two patients and partial improvement in the other two. DISCUSSION: Sclerodermatous changes are uncommon in PCT. They are not always late and secondary to the process of healing of blisters but can in fact constitute the first cutaneous symptom of the disease while revealing the underlying liver disease. Even in the absence of blisters, photosensitivity or cutaneous fragility, a diagnosis of PCT must be suspected in a setting of sclerodermatous changes distributed on the neck and face, or the neckline, or scarring alopecia, if associated with abnormal liver tests. Skin biopsy to confirm the diagnosis of scleroderma may delay the diagnosis, which is in fact based on porphyrin level. Normalization of the latter parameter under treatment allows regression of lesions.

Essential role of the AH receptor in the dysfunction of heme metabolism induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin.

The dysfunction of hepatic heme synthesis by 2,3,7,8-tetrachlordibenzo- p-dioxin (TCDD) in mice, enhanced by iron, leads to accumulation of uroporphyrins I and III (uroporphyria) and resembles the human disorder porphyria cutanea tarda (PCT) precipitated by alcohol and estrogenic drugs. Although consequences of TCDD are considered entirely dependent on the aryl hydrocarbon receptor (AHR), this is not proven for uroporphyria. Administration of TCDD (75 microg/kg) caused uroporphyria in susceptible C57BL/6J mice with high-affinity AHR after 5 weeks (>600-fold increase in hepatic uroporphyrins). Transcriptomics showed significant modified gene expressions for intermediary, heme, and iron metabolism as well as for oxidative stress and cell injury. Resistant low-affinity AHR DBA/2 mice (no increase in porphyrins) showed far fewer changes. At this dose of TCDD, persistent up-regulation of some traditional AH battery genes occurred in both strains. Essentiality of AHR was demonstrated with C57BL/6 Ahr knockout mice. Elevation of hepatic uroporphyrins was 964-fold in Ahr (+/+) mice, lower in Ahr (+/-) (60-fold), but undetectable with Ahr (-/-) . Consistent with an oxidative mechanism, iron overload enhanced porphyria as well as general liver injury in Ahr (+/+) and Ahr (+/-) mice but had no interactive effect in Ahr (-/-) . In contrast, when iron-treated mice received, instead of TCDD, the heme precursor 5-aminolevulinic acid (ALA), causing uroporphyia in Ahr (+/+) mice (242-fold rise in uroporphyrins), elevation of uroporphyrins I and III (42-fold) also occurred in Ahr (-/-) mice and was seemingly associated with AHR-independent expression of Cyp1a2. The findings prove that AHR is a key factor in porphyria induced in mice by TCDD. However, in other models of human PCT, participation of AHR may not be an essential requirement.

Utility of plasma fluorometric emission scanning for diagnosis of the first 2 cases reports of variegate porphyria: a very rare type of porphyrias in Thai.

Two Thai women who are siblings presented with a history of recurrent pruritic vesicles on dorsum of both hands and extensor surface of forearms where the sun-exposed areas are. The excoriated vesicles were healed with depressed scars. They had no previous history of intense abdominal pain, seizure, or psychiatric disorder Urinary porphyrins were analyzed by High Performance Liquid Chromatography (HPLC). The level of coproporphyrin III was detected to be higher than the uroporphyrin level. Fluorescence emission scanning of both patients' plasma was performed and demonstrated typical emission peak at 626 nm, that confirmed the diagnosis of variegate porphyria.

Effect of an oral iron chelator or iron-deficient diets on uroporphyria in a murine model of porphyria cutanea tarda.

UNLABELLED: Porphyria cutanea tarda is a liver disease characterized by elevated hepatic iron and excessive production of uroporphyrin (URO). Phlebotomy is an effective treatment that probably acts by reducing hepatic iron. Here we used Hfe(-/-) mice to compare the effects on hepatic URO accumulation of two different methods of hepatic iron depletion: iron chelation using deferiprone (L1) versus iron-deficient diets. Hfe(-/-) mice in a 129S6/SvEvTac background were fed 5-aminolevulinic acid (ALA), which results in hepatic URO accumulation, and increasing doses of L1 in the drinking water. Hepatic URO accumulation was completely prevented at low L1 doses, which partially depleted hepatic nonheme iron. By histological assessment, the decrease in hepatic URO accumulation was associated with greater depletion of iron from hepatocytes than from Kupffer cells. The L1 treatment had no effect on levels of hepatic cytochrome P4501A2 (CYP1A2). L1 also effectively decreased hepatic URO accumulation in C57BL/6 Hfe(-/-) mice treated with ALA and a CYP1A2 inducer. ALA-treated mice maintained on defined iron-deficient diets, rather than chow diets, did not develop uroporphyria, even when the animals were iron-supplemented either directly in the diet or by iron dextran injection. CONCLUSION: The results suggest that dietary factors other than iron are involved in the development of uroporphyria and that a modest depletion of hepatocyte iron by L1 is sufficient to prevent URO accumulation.

High-performance liquid chromatography/electrospray ionization tandem mass spectrometry of hydroxylated uroporphyrin and urochlorin derivatives formed by photochemical oxidation of uroporphyrinogen I.

Hydroxylated uroporphyrin I and urochlorin I derivatives formed by photochemical oxidation of uroporphyrinogen I were separated by high-performance liquid chromatography and fully characterized by electrospray ionization tandem mass spectrometry. The porphyrins and chlorins were identified by analysis of their product ion spectra with each hydroxylated derivative giving a characteristic collision-induced dissociation fragmentation pattern. The porphyrins and chlorins characterized were meso-hydroxyuroporphyrin I, alpha-hydroxypropionic acid uroporphyrin I, beta-hydroxypropionic acid uroporphyrin I, hydroxyacetic acid uroporphyrin I, trans-7-hydroxy-8-spirolactoneurochlorin I, cis-7-hydroxy-8-spirolactoneurochlorin I and trans- and cis-7,8-dihydroxyurochlorins I.

Improvement in HPLC separation of porphyrin isomers and application to biochemical diagnosis of porphyrias.

BACKGROUND: Identification of porphyrias relies on the measurement of different porphyrins in urine, feces and plasma. Separation of porphyrin isomers is essential for the differential diagnosis of some porphyrias. METHOD: Separation of naturally occurring porphyrins was achieved on a Chromolith RP-18 column with fluorimetric detection using a methanol/ammonium acetate gradient mobile phase. Fecal and plasma porphyrins were extracted with acetonitrile and water at different pH values. RESULTS: Eight porphyrins including protoporphyrin eluted within 20 min with good resolution of each of the I and III positional isomer pairs for standards, urine and plasma, and within 50 min for feces. Improvement of the extraction method for fecal and plasmatic porphyrins resulted in high recovery (up to 89%) and reliable quantification of protoporphyrin. CONCLUSIONS: The present RP-HPLC method is specific and efficient for routine analysis of porphyrins in human urine, feces and plasma.

Diagnostic traps in porphyria: case report and literature review.

Porphyrias are metabolic disorders of heme biosynthesis, which encompass a broad range of symptoms and signs, neurologic, cutaneous or mixed. Because of lack of specificity and polymorphous clinical picture, porphyrias can mimic either neuropsychiatric, dermatologic, or gastrointestinal diseases. We present the case of a 58 years old man to whom clinical presentation suspicious of Addison's disease (melanoderma, fatigue, weight loss, intermittent abdominal pain) was the disguise of porphyria cutanea tarda. A general background of porphyrias and differential diagnosis with other forms of hepatic porphyria, as well as other causes of hyperpigmentation, are given. The clinician should be aware of the protean manifestations of porphyrias and include them in clinical judgment in various situations.

Experimental hepatic uroporphyria induced by the diphenyl-ether herbicide fomesafen in male DBA/2 mice.

Hepatic uroporphyria can be readily induced by a variety of treatments in mice of the C57BL strains, whereas DBA/2 mice are almost completely resistant. However, feeding of the protoporphyrinogen oxidase-inhibiting herbicide fomesafen (0.25% in the diet for 18 weeks) induced hepatic uroporphyria in male DBA/2N mice (liver porphyrin content up to 150 nmol/g, control animals 1 nmol/g), whereas fomesafen-treated male C57BL/6N mice displayed only a slight elevation of liver porphyrins (approximately 5 nmol/g). The profile of accumulated hepatic porphyrins in fomesafen-treated DBA/2N mice resembled the well-characterised uroporphyria induced by polyhalogenated aromatic hydrocarbons, while histological examination confirmed the presence of uroporphyria-specific cytoplasmic inclusions in the hepatocytes. Uroporphyrinogen decarboxylase activity decreased to about 30% of control values in fomesafen-treated DBA/2N mice; microsomal methoxyresorufin O-dealkylase activity was slightly reduced. The amount of CYP1A1 and CYP1A2 mRNA, as determined by real-time PCR, was not significantly changed; mRNA encoding the housekeeping 5-aminolevulinic acid synthase was elevated 10-fold. Total liver iron was slightly increased. A similar uroporphyria was induced by the herbicide formulation Blazer, containing a structurally related herbicide acifluorfen, when fed to DBA/2N mice at a dose corresponding to 0.25% of acifluorfen in the diet. Since DBA/2 mice are almost completely resistant to all well-characterised porphyrogenic chemicals, the results suggest the possible existence of a yet unknown mechanism of uroporphyria induction, to which the DBA/2 mouse strain is more sensitive than the C57BL strain.

Congenital erythropoietic porphyria (Günther's disease): two cases with very early prenatal manifestation and cystic hygroma.

Congenital erythropoietic porphyria (CEP) or Günther's disease is the rarest form of the porphyrias. The disease is usually diagnosed at birth or during early infancy, but rarely in utero. We describe here the first two cases of very early prenatal expression of CEP with cystic hygroma diagnosed at 14 weeks in the first fetus and at 19 weeks in the second. Both fetuses presented with severe nonimmune hydrops fetalis as early as 19 and 22 weeks, associated with intrauterine growth retardation, hyperechogenic kidneys and bones. Amniotic fluid was dark brown and uro- and coproporphyrin I was dramatically increased. Molecular screening of the CEP gene detected heterozygous C73R mutation in both fetuses, the other parental mutation being as yet unknown.

Measurement of δ-aminolevulinate dehydratase activity.

Vertebrate ALAD is a cytosolic enzyme that catalyzes the second step of porphyrin formation, and it is expressed abundantly in liver and erythroid tissues as well as in other tissues. This unit describes a colorimetric assay for measuring ALAD activity in mammalian blood or tissues and the extent of reactivation of the enzyme in lead-inhibited samples.

Hereditary coproporphyria in Germany: clinical-biochemical studies in 53 patients.

OBJECTIVES: To describe the biochemical and clinical features in hereditary coproporphyria (HCP). DESIGN AND METHOD: Within the last 20 years, we investigated 53 patients (male:female = 1:2.5; age = 8-86 years) suffering from HCP. We describe the characteristic levels of urine, and fecal porphyrins and their precursors in hereditary coproporphyria and present the clinical features. Especially, we measured the coproporphyrin isomers I and III. RESULTS AND CONCLUSION: The group of hereditary coproporphyria patients exhibited a significantly higher (p<0.0001) excretion of urinary porphyrin precursors, delta-aminolevulinic acid (median = 84 micromol/24 h) and porphobilinogen (median = 39 micromol/24 h), as compared to controls (delta-aminolevulinic acid: 22 micromol/24 h, porphobilinogen: 3 micromol/24 h; median, n = 20). The median of coproporphyrin in urine (1315 nmol/24 h) and feces (1855 nmol/g) were enhanced 12- and 168-fold, as compared to healthy subjects (urinary coproporphyrin: 106 nmol/24 h, fecal coproporphyrin: 11 nmol/g; median, n = 20). During therapy on one female patient, with IV application of heme arginate, a considerable decline of porphyrin precursors and porphyrin excretion was observed. The examination of urinary and fecal coproporphyrin isomers I and III revealed an excessive elevation of the coproporphyrin isomer III of 87% in urine and 94% in feces, respectively (normal: urinary isomer III = 69-83% and fecal isomer III = 25-40%). In feces the increase of isomer III caused an inversion of the physiologic coproporphyrin isomer III:I ratio that could be recognized in all various stages in hereditary coproporphyria and in children. Acute attacks of hereditary coproporphyria are accompanied by an acute polysymptomatic clinical syndrome, and this is associated with high levels of urinary porphyrin precursors. On review of our patients, the highest percentage had abdominal pain (89%), followed by neurologic (33%), psychiatric (28%), cardiovascular (25%), and skin symptoms (14%).

Arsenite alters heme synthesis in long-term cultures of adult rat hepatocytes.

Arsenite (As[III]) effects on the intermediate steps of heme biosynthesis were studied in adult rat hepatocytes seeded on a feeder layer of 3T3 cells (3T3-hepatocytes) and maintained for 2 weeks with culture medium non-supplemented or supplemented with 150 microM 5-aminolevulinic acid (ALA). The activities of the intracellular enzymes porphobilinogen deaminase (PBG-D), uroporphyrinogen III synthase (UROIII-S), and uroporphyrinogen III decarboxylase (URO-D), and the intermediary uroporphyrins (URO), coproporphyrins (COPRO) and protoporphyrin IX (PROTO) were determined in these cultures. The 3T3-hepatocytes maintained the activities of PBG-D, UROIII-S and URO-D during 2 weeks and ALA addition to the culture medium increased PBG-D (2-3-fold) and UROIII-S (50%) activities and porphyrin production, which accumulated as PROTO. Exposure to 3.9 microM As(III) inhibited UROIII-S activity (down to 34%), and PBG-D and URO-D activities to a lower extent; these effects were magnified by ALA supplementation. As(III) also produced an intracellular accumulation and a decreased excretion of PROTO, and a 31% reduction of the COPRO/URO ratio in the culture medium. Additionally, As(III) caused cytoplasmic vacuolization and lipid accumulation. Our results show that As(III) exposure selectively inhibits several intermediary enzymes of heme metabolism and affects the intra- and extracellular content of porphyrins and their ratio in the culture medium. They also confirm that 3T3-hepatocytes are a suitable in vitro model to study hepatic heme metabolism and its alterations by hepatotoxic chemicals.

Congenital erythropoietic porphyria.

Congenital erythropoietic porphyria is a rare autosomal-recessive disorder of the porphyrin metabolism caused by the homozygous defect of uroporphyrinogen III cosynthase. High amounts of uroporphyrin I accumulate in all cells and tissues, reflected by an increased erythrocyte porphyrin concentration and excretion of high porphyrin amounts in urine and feces. Dermal deposits of uroporphyrin frequently induce a dramatic phototoxic oxygen-dependent skin damage with extensive ulcerations and mutilations. Splenomegaly and hemolytic anemia are typical internal symptoms. Skeletal changes such as osteolysis and calcifications are frequent. To date 130 cases of congenital erythropoietic porphyria have been published and are summarized here. Splenectomy, erythrocyte transfusions, and bone marrow transplantation have shown some beneficial effect. The best therapy is the avoidance of sunlight. In the two patients with congenital erythropoietic porphyria described here, oral administration of the oxygen quenchers ascorbic acid and alpha-tocopherol resulted in an improvement in the reduced hemoglobin and erythrocyte concentrations.

A modified spectrophotometric assay for porphobilinogen deaminase: its application in the detection of both carriers and patients with acute intermittent porphyria.

A spectrophotometric method for porphobilinogen deaminase assay in erythrocytes is described. This test is determinant for the definite diagnosis of acute intermittent porphyria. In the method described, delta-aminolevulinic acid is used as substrate. Mercaptoethanol and zinc ions are introduced to maintain delta-aminolevulinic acid dehydratase in optimal conditions and to guarantee the in vitro production of porphobilinogen. An incubation temperature of 45 degrees C leads to the production of uroporphyrins, which are measured spectrophotometrically at 405 nm, giving reproducible results. The assay can be performed easily in any clinical laboratory and is valuable for detecting both patients and carriers of acute intermittent porphyria.

Resonance Raman study of sirohydrochlorin and siroheme in sulfite reductases from sulfate reducing bacteria.

Soret-excited resonance Raman (RR) spectra are reported for the sirohemes in the oxidized and Cr11(EDTA)-reduced forms of both desulforubidin from D. baculatus (DSR) and the low molecular weight sulfite reductase from D. vulgaris (1SIR) and for sirohydrochlorin in the oxidized form of desulfoviridin from D. gigas (DSV). Several patterns in the RR spectra of these enzymes can be utilized as signatures for the siroheme/sirohydrochlorin moiety. The active site for DSR and 1SIR consists of a siroheme exchange-coupled to a [4Fe-4S]2+ cluster. Upon addition of Cr11(EDTA), the active center of DSR and 1SIR undergoes a one-electron and two-electron reduction, respectively. The RR spectra of DSR suggest that the siroheme iron is high spin and 5-coordinate in the oxidized enzyme and probably remains high spin and 5-coordinate upon reduction. The iron in the siroheme of oxidized 1SIR changes from a low spin and probably 6-coordinate configuration to a high spin, 5-coordinate complex upon two-electron reduction of the active site. Close similarities between the RR spectral features of the two-electron-reduced assimilatory sulfite reductases from E. coli and from D. vulgaris (1SIR) are discussed.

The role of iron in experimental porphyria and porphyria cutanea tarda.

Porphyria cutanea tarda (PCT) and experimental porphyria are characterized by a decreased activity of the enzyme uroporphyrinogen decarboxylase, and accumulation of uroporphyrins and heptacarboxylporphyrins in the liver. Iron (Fe) plays an important role in PCT and experimental porphyria. Biochemically and electron microscopically, we examined the relationship between Fe and porphyrins in liver tissue of C57BL/10 mice made porphyric by administration of iron dextran as Imferon (IMF), and in liver biopsies of patients with symptomatic PCT. Accumulation of uroporphyrins and heptacarboxylporphyrins, and an increased amount of Fe were observed in livers of mice treated with IMF and in liver biopsies of patients with PCT. In mice treated with IMF, the activity of uroporphyrinogen decarboxylase was decreased. Both in livers of mice treated with IMF and in livers of patients with PCT, needle-like structures, representing uroporphyrin crystals, were observed by electron microscopy. Uroporphyrin crystals and Fe (as ferritin) were observed in the same hepatocyte. Moreover, there was a striking morphological correlation between uroporphyrin crystals and ferritin-Fe, suggesting a role for (ferritin-)Fe in the pathogenesis of porphyria.

Preparation, high-performance liquid chromatographic separation and characterization of hexacarboxylic porphyrinogens.

A simple method for the preparation and reversed-phase high-performance liquid chromatographic separation of hexacarboxylic porphyrinogen isomers is described. Uroporphyrin I or III was partially decarboxylated in 0.5 M hydrochloric acid at 150 degrees C. Unreacted uroporphyrin and the hepta-, hexa- and pentacarboxylic porphyrins formed were esterified and then group-separated by thin-layer chromatography. After hydrolysis, the porphyrins were reduced to the corresponding porphyrinogens with 3% (w/w) sodium amalgam. The hexacarboxylic porphyrinogens were separated on an ODS-Hypersil column by elution with acetonitrile-methanol-1 M ammonium acetate, pH 5.16 (8:12:80, v/v/v) as mobile phase. Isomers were identified by high-performance liquid chromatography of the characteristic mixture of two pentacarboxylic porphyrins formed after partial decarboxylation of individual isomers. Except for the two type I isomers, resolution of the hexacarboxylic porphyrinogens was superior to that of the corresponding porphyrins.