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Acute intermittent porphyria (AIP) is a rare metabolic disease caused by mutations within the hydroxymethylbilane synthase gene. Previous studies have reported increased levels of plasma total homocysteine (tHcy) in symptomatic AIP patients. In this study, we present long-term data for tHcy and related parameters for an AIP patient cohort (n = 37) in different clinical disease-states. In total, 25 patients (68%) presented with hyperhomocysteinemia (HHcy; tHcy > 15 μmol/L) during the observation period. HHcy was more frequent in AIP patients with recurrent disease receiving heme arginate, than in nonrecurrent (median tHcy: 21.6 μmol/L; range: 10-129 vs median tHcy: 14.5 μmol/L; range 6-77). Long-term serial analyses showed a high within-person tHcy variation, especially among the recurrent patients (coefficient of variation: 16.4%-78.8%). HHcy was frequently associated with low blood concentrations of pyridoxal-5'-phosphate and folate, while cobalamin concentration and the allele distribution of the methylene-tetrahydrofolate-reductase gene were normal. Strikingly, 6 out of the 9 recurrent patients who were later included in a regime of givosiran, a small-interfering RNA that effectively reduced recurrent attacks, showed further increased tHcy (median tHcy in 9 patients: 105 μmol/L; range 16-212). Screening of amino acids in plasma by liquid-chromatography showed co-increased levels of methionine (median 71 μmol/L; range 23-616; normal <40), suggestive of acquired deficiency of cystathionine-β-synthase. The kynunerine/tryptophan ratio in plasma was, however, normal, indicating a regular metabolism of tryptophan by heme-dependent enzymes. In conclusion, even if HHcy was observed in AIP patients receiving heme arginate, givosiran induced an aggravation of the dysregulation, causing a co-increase of tHcy and methionine resembling classic homocystinuria., (© 2021 SSIEM.)

Givosiran is a novel approach to treat patients with acute intermittent porphyrias (AIP) by silencing of ∂-ALA-synthase 1, the first enzyme of heme biosynthesis in the liver. We included two patients in the Envision study who responded clinically well to this treatment. However, in both patients, therapy had to be discontinued because of severe adverse effects: One patient (A) developed local injection reactions which continued to spread all over her body with increasing number of injections and eventually caused a severe systemic allergic reaction. Patient B was hospitalized because of a fulminant pancreatitis. Searching for possible causes, we also measured the patients plasma homocysteine (Hcy) levels in fluoride-containing collection tubes: by LC-MS/MS unexpectedly, plasma Hcy levels were 100 and 200 in patient A and between 100 and 400 μmol/l in patient B. Searching for germline mutations in 10 genes that are relevant for homocysteine metabolism only revealed hetero- and homozygous polymorphisms in the MTHFR gene. Alternatively, an acquired inhibition of cystathionine-beta-synthase which is important for homocysteine metabolism could explain the plasma homocysteine increase. This enzyme is heme-dependent: when we gave heme arginate to our patients, Hcy levels rapidly dropped. Hence, we conclude that inhibition of ∂-ALA-synthase 1 by givosiran causes a drop of free heme in the hepatocyte and therefore the excessive increase of plasma homocysteine. Hyperhomocysteinemia may contribute to the adverse effects seen in givosiran-treated patients which may be due to protein-N-homocysteinylation.

Background: 5-Aminolevulinic acid dehydratase (ALAD) porphyria (ADP) is an ultrarare autosomal recessive disease, with only eight documented cases, all of whom were males. Although classified as an acute hepatic porphyria (AHP), induction of the rate limiting hepatic enzyme 5-aminolevulinic acid synthase-1 (ALAS1) has not been demonstrated, and the marrow may also contribute excess 5-aminolevulinic acid (ALA). Two patients have died and reported follow up for the others is limited, so the natural history of this disease is poorly understood and treatment experience limited., Methods: We report new molecular findings and update the clinical course and treatment of the sixth reported ADP patient, now 31 years old and the only known case in the Americas, and review published data regarding genotype-phenotype correlation and treatment., Results: Circulating hepatic 5-aminolevulinic acid synthase-1 (ALAS1) mRNA was elevated in this case, as in other AHPs. Gain of function mutation of erythroid specific ALAS2 - an X-linked modifying gene in some other porphyrias - was not found. Seven reported ADP cases had compound heterozygous ALAD mutations resulting in very low residual ALAD activity and symptoms early in life or adolescence. One adult with a germline ALAD mutant allele developed ADP in association with a clonal myeloproliferative disorder, polycythemia vera., Conclusions: Elevation in circulating hepatic ALAS1 and response to treatment with hemin indicate that the liver is an important source of excess ALA in ADP, although the marrow may also contribute. Intravenous hemin was effective in most reported cases for treatment and prevention of acute attacks of neurological symptoms., Competing Interests: Declaration of Competing Interest AP Lahiji and VM Sadagopa Ramanujam declare no conflicts. A Chan and A Simon are employees of Alnylam Pharmaceuticals. KE Anderson and RJ Desnick consult for Alnylam Pharmaceuticals, Recordati Rare Diseases and Mitsubishi Tanabe Pharma America. All authors had access to the data and a role in writing the manuscript., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background and Aims: The acute porphyrias are characterized by defects in heme synthesis, particularly in the liver. In some affected patients, there occurs a critical deficiency in a regulatory heme pool within hepatocytes that leads to up-regulation of 5-aminolevulinic acid [ALA] synthase-1, which is the first and normally rate-controlling enzyme in the pathway. In earlier work, we described defects in mitochondrial functions in cultured skin fibroblasts from patients with acute intermittent porphyria [AIP]. Others described defects in livers of murine models of AIP. Here, we explored mitochondrial energetics in peripheral blood mononuclear cells [PBMCs] and platelets in persons with AIP and hereditary coproporphyria [HCP]. Our hypotheses were that there are deficits in bioenergetic capacity in acute porphyrias and that subjects with more severe acute porphyria have more pronounced reductions in mitochondrial oxygen consumption rates [OCR]., Methods: We studied 17 subjects with acute hepatic porphyrias, 14 with classical AIP, one with severe AIP due to homozygous deficiency of hydroxymethylbilane synthase [HMBS], 2 with HCP, and 5 non-porphyric controls. We collected peripheral blood, isolated PBMCs, which we assayed either immediately or after frozen storage [80C] for up to 14 days. Using Seahorse XF-24-3, we measured OCR in the presence of glucose + pyruvate under basal condition, and after additions of oligomycin, carbonylcyanide p-trifluoromethoxyphenylhydrazone [FCCP], and antimycin+rotenone., Results: Most subjects [13/17, 76%] were female. Subjects with moderate/severe symptoms associated with acute porphyria had significantly lower basal and maximal-OCR than those with no/mild symptoms who were the same as controls. We observed significant inverse correlation between urinary porphobilinogen [PBG] excretion and OCR. The subject with homozygous AIP had a much lower-OCR than his asymptomatic parents., Summary/conclusions: Results support the hypothesis that active acute hepatic porphyria is characterized by a deficiency in mitochondrial function that is detectable in PBMCs, suggesting that limitations in electron transport and ATP production exist in such individuals., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background and Aims: Acute intermittent porphyria (AIP) results from a partial deficiency of porphobilinogen deaminase (PBGD). Symptomatic AIP patients, most of whom are women, experience acute attacks characterized by severe abdominal pain and abrupt increases in blood pressure. Here, we characterized the reactivity of mesenteric arteries from male and female AIP mice with ~30% of normal PBGD activity and wild type C57BL/6 mice., Methods: An acute porphyric attack was induced in AIP mice by treatment with phenobarbital. Vascular responses to K + , phenylephrine (PE), acetylcholine (ACh), and hemin were determined (Wire Multi Myograph)., Results: Maximal contraction to PE was increased in arteries from male and female AIP mice (p < .05) during an induced attack of acute porphyria. Female AIP arteries had increased sensitivity to PE (p < .05) even after nitric oxide (NO) blockade with N ω -nitro-L-arginine methyl ester (L-NAME) (p < .05). Maximal relaxation to ACh was similar in males and females with lower sensitivity in female AIP arteries (p < .05). Hemin induced greater relaxation in AIP arteries in both males and females (p < .05)., Summary/conclusions: Sex differences in this AIP mouse model include a pro-contractile response in females. These alterations may contribute to the increased blood pressure during an acute attack and provide a novel mechanism of action whereby heme ameliorates the attacks., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Acute intermittent porphyria (AIP) is an inborn error of heme biosynthesis due to the deficiency of hydroxymethylbilane synthase (HMBS) activity. Human AIP heterozygotes have episodic acute neurovisceral attacks that typically start after puberty, whereas patients with homozygous dominant AIP (HD-AIP) have early-onset chronic neurological impairment, including ataxia and psychomotor retardation. To investigate the dramatically different manifestations, knock-in mice with human HD-AIP missense mutations c.500G>A (p.Arg167Glu) or c.518&#95;519GC>AG (p.Arg173Glu), designated R167Q or R173Q mice, respectively, were generated and compared with the previously established T1/T2 mice with ~30% residual HMBS activity and the heterozygous AIP phenotype. Homozygous R173Q mice were embryonic lethal, while R167Q homozygous mice (R167Q+/+) had ~5% of normal HMBS activity, constitutively elevated plasma and urinary 5-aminolevulinic acid (ALA) and porphobilinogen (PBG), profound early-onset ataxia, delayed motor development and markedly impaired rotarod performance. Central nervous system (CNS) histology was grossly intact, but CNS myelination was delayed and overall myelin volume was decreased. Heme concentrations in liver and brain were similar to those of T1/T2 mice. Notably, ALA and PBG concentrations in the cerebral spinal fluid and CNS regions were markedly elevated in R167Q+/+ mice compared with T1/T2 mice. When the T1/T2 mice were administered phenobarbital, ALA and PBG markedly accumulated in their liver and plasma, but not in the CNS, indicating that ALA and PBG do not readily cross the blood-brain barrier. Taken together, these studies suggest that the severe HD-AIP neurological phenotype results from decreased myelination and the accumulation of locally produced neurotoxic porphyrin precursors within the CNS., (© The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: Acute intermittent porphyria (AIP) is caused by diminished activity of porphobilinogen deaminase (PBGD). The purpose of this study was to validate and compare two assays for PBGD activity. The clinical sensitivity of the PBGD activity assays in AIP diagnosis was also evaluated., Methods: This study included 74 subjects from 18 Taiwanese families including symptomatic patients with AIP, asymptomatic carriers, and healthy family members. The specific mutations in AIP patients were identified by DNA sequencing. PBGD activity was measured in erythrocytes by quantifying formation of coproporphyrin or uroporphyrin by the enzyme using porphobilinogen (PBG) as a substrate and fluorimetry for detection., Results: The calibration curves obtained with pure coproporphyrin or uroporphyrin were linear with correlation coefficients >0.99 in the range of 0-200nM for coproporphyrin and 0-150nM for uroporphyrin. The coefficients of variation for within-run and between-day imprecision were <9.8% for both assays. The three groups of subjects were used to establish the best cut-off of PBGD activity for identifying symptomatic AIP patients by using area under receiver operating characteristic curve analysis. The symptomatic AIP patients and asymptomatic carriers had significantly lower PBGD activity compared with the healthy family members (all p<.001)., Conclusion: Two different PBGD activity assays were validated. The best cut-off for coproporphyrin was derived as 46.4nmol/h/mL RBC with corresponding sensitivity of 100% and specificity of 100% and the best cut-off for uroporphyrin was derived as 43.7nkat/L RBC with corresponding sensitivity of 100% and specificity of 97.4%., (Copyright © 2018 Elsevier B.V. All rights reserved.)

This study aimed to examine whether acute intermittent porphyria (AIP) is associated with systemic inflammation and whether the inflammation correlates with disease activity. A case-control study with 50 AIP cases and age-, sex- and place of residence-matched controls was performed. Plasma cytokines, insulin and C-peptide were analysed after an overnight fast using multiplex assay. Long pentraxin-3 (PTX3) and complement activation products (C3bc and TCC) were analysed using enzyme-linked immunosorbent assay (ELISA). Urine porphobilinogen ratio (U-PBG, µmol/mmol creatinine), haematological and biochemical tests were performed using routine methods. Questionnaires were used to register AIP symptoms, medication and other diseases. All 27 cytokines, chemokines and growth factors investigated were increased significantly in symptomatic AIP cases compared with controls (P < 0·0004). Hierarchical cluster analyses revealed a cluster with high visfatin levels and several highly expressed cytokines including interleukin (IL)-17, suggesting a T helper type 17 (Th17) inflammatory response in a group of AIP cases. C3bc (P = 0·002) and serum immunoglobulin (Ig)G levels (P = 0·03) were increased significantly in cases with AIP. The U-PBG ratio correlated positively with PTX3 (r = 0·38, P = 0·006), and with terminal complement complex (TCC) levels (r = 0·33, P = 0·02). PTX3 was a significant predictor of the biochemical disease activity marker U-PBG in AIP cases after adjustment for potential confounders in multiple linear regression analyses (P = 0·032). Prealbumin, C-peptide, insulin and kidney function were all decreased in the symptomatic AIP cases, but not in the asymptomatic cases. These results indicate that AIP is associated with systemic inflammation. Decreased C-peptide levels in symptomatic AIP cases indicate that reduced insulin release is associated with enhanced disease activity and reduced kidney function., (© 2016 British Society for Immunology.)

Background: Acute intermittent porphyria (AIP) is an autosomal inherited disease caused by deficiency of the third enzyme in the heme biosynthetic pathway, porphobilinogen deaminase. The clinical course of the disease is characterized by acute attacks, most often with abdominal pain.The aim of our study was to investigate selected markers of oxidative and carbonyl stress in plasma and saliva in patients with AIP and to find out whether saliva could be used for monitoring the disease progression. Saliva is an attractive biological fluid for determination of biochemical markers in various pathological conditions. The advantage is that saliva can be collected non-invasively, and the examination needs only a small volume of saliva., Methods: Blood and total non-stimulated saliva were collected from 16 patients with AIP in remission, and from 20 healthy individuals. Markers of oxidative and carbonyl stress - advanced glycation end products (AGEs), and thiobarbituric acid reacting substances (TBARS) were determined by spectrofluorometric methods, advanced oxidation protein products (AOPPs), total antioxidant capacity (TAC) and ferric reducing ability of plasma/saliva (FRAP/FRAS) were investigated by spectrophotometric methods in the above mentioned groups., Results: Advanced glycation end products and thiobarbituric acid reacting substances in plasma and saliva were significantly higher in patients with AIP in comparison to the control group (p < 0.001) and (p < 0.05). Advanced oxidation protein products in AIP if compared to the control group did not show statistical significance (p > 0.05), but the levels in the saliva were significantly lower (p < 0.001). The concentrations of markers of antioxidant status of plasma and saliva were significantly lower in AIP if compared to the control group (p < 0.001)., Conclusions: According to the best of our knowledge this is the first study to demonstrate increased concentrations of markers of oxidative and carbonyl stress and decreased antioxidant status of plasma and saliva in patients with AIP. Moreover, the study suggests that the saliva might be a promising fluid to study relevant biomarkers in a wide array of human biomedical conditions.Key words: acute intermittent porphyria - biomarkers - oxidative and carbonyl stress - plasma and saliva.

The aims of the present study were to explore the expression pattern of haem biosynthesis enzymes in circulating cells of patients affected by two types of porphyria (acute intermittent, AIP, and variegate porphyria, VP), together with the antioxidant enzyme pattern in AIP in order to identify a possible situation of oxidative stress. Sixteen and twelve patients affected by AIP and VP, respectively, were analysed with the same numbers of healthy matched controls. Erythrocytes, neutrophils and peripheral blood mononuclear cells (PBMCs) were purified from blood, and RNA and proteins were extracted for quantitative real time PCR (qRT-PCR) and Western-blot analysis, respectively. Porhobilinogen deaminase (PBGD) and protoporphyrinogen oxidase (PPOX) gene and protein expression was analysed. Antioxidant enzyme activity and gene expression were additionally determined in blood cells, together with protein carbonyl content in plasma. PBMCs isolated from AIP patients presented low mRNA levels of PBGD when compared to controls, while PBMCs isolated from VP patients presented a decrease in PPOX mRNA. PPOX protein content was higher in AIP patients and lower in VP patients, compared to healthy controls. Regarding antioxidant enzymes, PBMCs and erythrocyte superoxide dismutase (SOD) presented statistically significant higher activity in AIP patients compared to controls, while catalase activity tended to be lower in these patients. No differences were observed regarding antioxidant gene expression in white blood cells. Circulating cells in AIP and VP patients present altered expression of haem biosynthetic enzymes, which could be useful for the differential diagnosis of these two types of porphyria in certain difficult cases. AIP patients present a condition of potential oxidative stress similar to VP patients, evidenced by the post-transcriptional activation of SOD and possible catalase impairment., Competing Interests: The authors have declared that no competing interests exist.

Porphobilinogen deaminase (PBGD) haploinsufficiency (acute intermittent porphyria, AIP) is characterized by neurovisceral attacks when hepatic heme synthesis is activated by endogenous or environmental factors including fasting. While the molecular mechanisms underlying the nutritional regulation of hepatic heme synthesis have been described, glucose homeostasis during fasting is poorly understood in porphyria. Our study aimed to analyse glucose homeostasis and hepatic carbohydrate metabolism during fasting in PBGD-deficient mice. To determine the contribution of hepatic PBGD deficiency to carbohydrate metabolism, AIP mice injected with a PBGD-liver gene delivery vector were included. After a 14 h fasting period, serum and liver metabolomics analyses showed that wild-type mice stimulated hepatic glycogen degradation to maintain glucose homeostasis while AIP livers activated gluconeogenesis and ketogenesis due to their inability to use stored glycogen. The serum of fasted AIP mice showed increased concentrations of insulin and reduced glucagon levels. Specific over-expression of the PBGD protein in the liver tended to normalize circulating insulin and glucagon levels, stimulated hepatic glycogen catabolism and blocked ketone body production. Reduced glucose uptake was observed in the primary somatosensorial brain cortex of fasted AIP mice, which could be reversed by PBGD-liver gene delivery. In conclusion, AIP mice showed a different response to fasting as measured by altered carbohydrate metabolism in the liver and modified glucose consumption in the brain cortex. Glucose homeostasis in fasted AIP mice was efficiently normalized after restoration of PBGD gene expression in the liver., (© The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Acute intermittent porphyria is the most common acute porphyria caused by a decrease in hepatic porphobilinogen deaminase activity, resulting in an accumulation of delta-aminolevulinic acid and porphobilinogen. This disease shows nonspecific signs and symptoms that can be confused with other diseases, thereby making the diagnosis difficult. We report a case of acute intermittent porphyria, reviewing clinical and laboratory aspects, highlighting the hematological and biochemical parameters during and after the crisis. A female patient, aged 28 years, suffered two crises, both presenting gastrointestinal disorders. The second presented neuropsychiatric symptoms. The analysis of hematological and biochemical parameters during the second crisis showed anemia, leukocytosis, hyponatremia, mild hypokalemia, uremia and elevated C-reactive protein. The initial treatment included glucose infusion, a diet rich in carbohydrates and interruption of porphyrinogenic drugs. Subsequently, treatment was maintained with oral contraceptive use. According to the observed data, signs and symptoms of gastrointestinal, neurological and psychiatric disorders, associated with laboratory results presented in this paper can be applied to screen acute porphyria, contributing to early diagnosis.

AIP is an acute liver disorder caused by a deficiency of porphobilinogen deaminase (PBGD) characterized by neuroabdominal symptoms. It is an autosomal dominant disease. However, homozygous dominant AIP (HD-AIP) have been described. In some cases erythrodontia was observed. CEP is an autosomal recessive disease produced by mutations in the uroporphyrinogen III synthase gene (UROS), characterized by severe cutaneous lesions and erythrodontia. The aim of the work was to establish the differential diagnosis of porphyria in a patient with abdominal pain, neurological attacks, skin symptoms and erythrodontia. The PBGD activity was reduced 50% and the genetic analysis indicated the presence of two genetic variants in the PBGD gene, p.G111R and p.E258G, a new genetic variant, revealing a case of heteroallelic HD-AIP. The patient, first diagnosed as a carrier of a dual porphyria: AIP / CEP based on the excretion profile of porphyrins, precursors and her clinical symptoms, would be an atypical case of human HD-AIP. These results would also suggest the presence of a phenocopy of the CEP, induced by an endogenous or exogenous factor. Our findings highlight the importance of genetic studies for a proper diagnosis of porphyria, prevention of its manifestation and its treatment.

Serum/plasma concentrations of 5-aminolaevulinic acid (ALA) and porphobilinogen (PBG) are elevated in patients with acute hepatic porphyrias, especially during acute attacks. Current assays require lengthy sample pre-treatment and derivatization steps. We report here a rapid, sensitive and specific hydrophilic interaction liquid chromatography-tandem mass spectrometry method for the direct and simultaneous quantitation of ALA and PBG in serum or plasma following simple protein precipitation with acetonitrile and centrifugation prior to injection. ALA and PBG were detected using selected reaction monitoring mode, following positive atmospheric pressure chemical ionization. Calibration was linear from 0.05 to 50 µmol/L for ALA and PBG. For both analytes, imprecision (relative standard deviation) was <13% and accuracy (percentage nominal concentrations) was between 92 and 107%. The method was successfully applied to the measurement of ALA and PBG in serum or plasma samples for the screening, biochemical diagnosis and treatment monitoring of patients with acute hepatic porphyrias., (Copyright © 2012 John Wiley & Sons, Ltd.)

Objective: To measure serum concentrations of progesterone, estradiol and 5α- and 5β-reduced progesterone metabolites in the follicular and luteal phases of the menstrual cycle in women with latent acute intermittent porphyria and manifest acute intermittent porphyria in comparison with healthy control women., Design: A descriptive study with repeated measurements during a complete, ovulatory menstrual cycle., Setting: University hospital out-patient clinic., Population: Thirty-two women with DNA-diagnosed acute intermittent porphyria and 20 healthy control women., Methods: Blood samples for serum progesterone, estradiol, allopregnanolone and pregnanolone were drawn on predefined menstrual cycle days, twice in the follicular phase and three times in the luteal phase. Serum levels of estradiol and progesterone were analysed with commercial kits. Allopregnanolone and pregnanolone levels were analysed with radioimmunoassay following diethylether extraction and celite column chromatography., Main Outcome Measures: Changes in serum levels of progesterone, estradiol, allopregnanolone and pregnanolone throughout the menstrual cycle., Results: Women with acute intermittent porphyria displayed lower serum concentrations of allopregnanolone in comparison with healthy control women, the difference being most prominent in the luteal phase (p < 0.001). Levels of pregnanolone did not differ significantly between groups. No significant difference was found between women with latent acute intermittent porphyria and manifest acute intermittent porphyria., Conclusions: Decreased levels of the 5α-reduced progesterone metabolite allopregnanolone were found in the menstrual cycle of women with acute intermittent porphyria. This has not been reported previously and could indicate a reduced 5α-reductase type 1 capacity in the ovary and liver among these women., (© 2012 The Authors Acta Obstetricia et Gynecologica Scandinavica© 2012 Nordic Federation of Societies of Obstetrics and Gynecology.)

A review of the literature about the toxic effects of cadmium on the human body. We describe a patient with clinical and biochemical signs of an attack of acute porphyria imitated or severe lead poisoning. In the patient's blood was revealed a 3-fold, compared to the allowable rate, increase of cadmium in the normal lead content. We discuss the etiologic role of cadmium and the possible pathogenetic mechanisms of this pathological condition.

This work deals with the study of how porphyrinogenic drugs modeling acute porphyrias interfere with the status of carbohydrate-regulating hormones in relation to key glucose enzymes and to porphyria, considering that glucose modulates the development of the disease. Female Wistar rats were treated with 2-allyl-2-isopropylacetamide (AIA) and 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) using different doses of AIA (100, 250 and 500mg/kg body weight) and a single dose of DDC (50mg DDC/kg body weight). Rats were sacrificed 16h after AIA/DDC administration. In the group treated with the highest dose of AIA (group H), hepatic 5-aminolevulinic acid synthase (ALA-S) increased more than 300%, phosphoenolpyruvate carboxykinase (PEPCK) and glycogen phosphorylase (GP) activities were 43% and 46% lower than the controls, respectively, plasmatic insulin levels exceeded normal values by 617%, and plasmatic glucocorticoids (GC) decreased 20%. GC results are related to a decrease in corticosterone (CORT) adrenal production (33%) and a significant reduction in its metabolization by UDP-glucuronosyltransferase (UGT) (62%). Adrenocorticotropic hormone (ACTH) stimulated adrenal production 3-fold and drugs did not alter this process. Thus, porphyria-inducing drugs AIA and DDC dramatically altered the status of hormones that regulate carbohydrate metabolism increasing insulin levels and reducing GC production, metabolization and plasmatic levels. In this acute porphyria model, gluconeogenic and glycogenolytic blockages caused by PEPCK and GP depressed activities, respectively, would be mainly a consequence of the negative regulatory action of insulin on these enzymes. GC could also contribute to PEPCK blockage both because they were depressed by the treatment and because they are positive effectors on PEPCK. These disturbances in carbohydrates and their regulation, through ALA-S de-repression, would enhance the porphyria state promoted by the drugs on heme synthesis and destruction. This might be the mechanism underlying the "glucose effect" observed in hepatic porphyrias. The statistical correlation study performed showed association between all the variables studied and reinforce these conclusions., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Accurate determinations of 5-aminolevulinic acid (ALA) and porphobilinogen (PBG) in physiologic fluids are required for the diagnosis and therapeutic monitoring of acute porphyrias. Current colorimetric methods are insensitive and over-estimate ALA and PBG due to poor specificity, while LC-MS/MS methods increase sensitivity, but have limited matrices. An LC-MS/MS method was developed to simultaneously determine ALA and PBG concentrations in fluids or tissues which were solid phase extracted, butanol derivatized, and quantitated by selective reaction monitoring using (13)C(5), (15)N-ALA and 2,4-(13)C(2)-PBG internal standards. ALA was separated from interfering compounds on a reverse phase C8-column. For ALA and PBG, the matrix effects (87.3-105%) and process efficiencies (77.6-97.8% and 37.2-41.6%, respectively) were acceptable in plasma and urine matrices. The assay was highly sensitive for ALA and PBG (LLOQ=0.05 μM with 25 μL urine or 100 μL plasma), and required ∼4 h from extraction to results. ALA and PBG accuracy ranged from 88.2 to 110% (n=10); intra- and inter-assay coefficients of variations were <10% for urine and plasma. In clinical applications, patients with mutation-confirmed acute porphyrias had normal to slightly increased urinary ALA and PBG levels when asymptomatic, and high levels during acute attacks, which decreased with hemin therapy. In AIP mice, baseline ALA and PBG levels in urine, plasma, and liver were increased after phenobarbital induction 28-/63-, 42-/266-, and 13-/316-fold, respectively. This LC-MS/MS method is rapid, specific, highly sensitive, accurate, and simultaneously measures ALA and PBG in urine, plasma, and tissues permitting porphyria clinical diagnoses, therapeutic monitoring, and research., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Background: Acute intermittent porphyria (AIP) is an autosomal dominant disorder of the haem biosynthesis resulting from a partial deficiency of hydroxymethylbilane synthase (HMBS) with incomplete penetrance. By conventional means, it is able to identify asymptomatic mutation carrier by molecular diagnosis, but one cannot reliably predict an acute porphyric attack. The presence of fluorescent red cells (fluorocytes) in AIP is probably under-recognized since AIP is a hepatic porphyria and not associated with photosensitivity., Methods: We used an automatic image acquisition platform to detect the circulating fluorocytes at 700 nm emission in a diabetic AIP patient during acute attack. We screened the patient and her family members for the mutation on HMBS, urine porphobilinogen and circulating fluorocytes., Results: The patient was heterozygous for a disease-causing mutation on HMBS and several bright circulating fluorocytes were detected. We showed evidence that protoporphyrin contributed to the erythrocyte auto-fluorescence. Interestingly, asymptomatic mutation carriers with increased urine porphobilinogen did not have circulating fluorocytes. All mutation-negative family members revealed no circulating fluorocytes., Conclusion: Sudden decrease in plasma glucose concentration might invoke acute attack of AIP and appearance of circulatory fluorocytes. Potential of detecting fluorocytes as screening test or for predicting an acute attack of AIP in diabetes is worth investigating., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Background: Acute intermittent and variegate porphyria are an autosomal dominant hereditary diseases caused by the deficient activity of porphobilinogen deaminase in the haem biosynthesis. Acute intermittent porphyria (AIP) in 11 patients (8 women and 3 men) and variegate porphyria (VP) in one patient were diagnosed and long-term treated during 15-22 years. Eleven patients had in acute attack abdominal pain, they were agitated and restless and suffered from insomnia. Besides they had various neurological signs. Examination of kidney function during remission showed hypertension and tubulointerstitial impairment of the kidneys in 10 patients (hyposthenuria and impairment of tubular excretory phase in isotopic renography). Deficiency of serum erythropoietin in 4 patients, significant deficiency of plasma and erythrocyte vitamin B6, significant hyperoxalaemia and hyperoxaluria in all patients were found. Direct relationship between plasma oxalic acid and effect of pyridoxal-5-phosphate (PLP), (effect of PLP was in indirect relationship with the concentration of erythrocyte vitamin B6), in AIP patients was found. Deficiency of vitamin B6 was probably a cause of hyperoxalaemia and hyperoxaluria in those patients. The effective therapy was repeated i.v. administration of haem-arginate during acute attacks (4-5 days). Besides during remission the patients were treated by pyridoxine (40-60 mg/day), by glucose, sodium chloride and phenothiazines. All patients showed significant improvement and had regular ambulatory check-up every three months. Currently, they are in clinical and laboratory remission.

Objective: Vitamin B(6) (VB(6)) is a water-soluble vitamin, which is important for the normal functioning of multiple organ systems. It is metabolized to the active molecule pyridoxal-5-phosphate (PLP). Oxalic acid (OA) is thought to be a uremic toxin that participates in the pathogenesis of the uremic syndrome. The objectives of this study were as follows: (1) to evaluate the plasma and erythrocyte VB(6) (effect of PLP; effect of PLP was in indirect relationship with the concentration of erythrocyte VB(6)), and plasma and urinary OA in marathon runners, in patients with acute intermittent porphyria (AIP) and variegate porphyria, and in patients with stage 1 chronic kidney disease (CKD), chronic glomerulonephritis and nephrotic syndrome (CGNS); (2) to examine the influence of water diuresis in healthy subjects, and the influence of sodium diuresis (high sodium intake) and an intravenous administration of furosemide on the urinary excretion of VB(6) and OA in CKD stage 3-4 patients; and (3) to evaluate the influence of erythropoietin treatment on erythrocyte VB(6) (effect of PLP) in hemodialysis (HD) patients, and the influence of continuous ambulatory peritoneal dialysis (CAPD) therapy on plasma VB(6) and OA and their peritoneal clearance and transfer., Design and Setting: This study was conducted at the Nephrological Clinic of L. Pasteur Faculty Hospital and of Medical School of P. J. Safarik University. A combination of 29 marathon runners, 15 patients with CG and NS, 11 patients with AIP, 1 patient with variegate porphyria, 15 healthy subjects, 27 CKD stage 3-4 patients, 30 HD, and 27 CAPD patients were used in the study., Results: After a marathon run, plasma and erythrocyte VB(6) significantly decreased and plasma OA increased. Plasma (15.5 +/- 3.8 nmol/L) and erythrocyte VB(6) (effect of PLP: 42.1% +/- 7.5%) were decreased and plasma OA (9.8 +/- 2.3 micromol/L) was significantly elevated in patients with CGNS and stage 1 CKD. In patients with AIP, deficiency of plasma (24.3 +/- 5.2 nmol/L) and erythrocyte VB(6) (effect of PLP: 46.2% +/- 7.0%) and hyperoxalemia (9.39 +/- 2.5 micromol/L) were present. The urinary excretion of VB(6) and of OA during maximal water diuresis and after intravenous administration of furosemide increased significantly (P < .01), but was not affected by the high intake of NaCl (P > .05). Erythropoietin treatment in HD patients led to the erythrocyte VB(6) deficiency. This finding is an indirect evidence that erythrocyte VB(6) is consumed by the hemoglobin synthesis much more during EPO treatment. In CAPD patients, plasma value of VB(6) (127.3 +/- 66.9 micromol/L) was in the normal range and plasma OA (23.6 +/- 7.4 micromol/L) was significantly elevated. Mean value of peritoneal clearance of VB(6) was 8.8% and of OA was 76.9% of urea clearance., Conclusion: Our study indicates that deficiency of VB(6) led to hyperoxalemia and hyperoxaluria in patients with CKD. Deficiency of VB(6) in CKD stage 4-5 patients potentiates the uremic hyperoxalemia and hyperoxaluria., (Copyright 2010 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.)

Objective: Acute intermittent porphyria (AIP) is caused by a deficiency of hydroxymethylbilane synthase. Clinical manifestations are abdominal pain and neurovisceral symptoms, accompanied by overproduction of heme-precursors in the liver, which frequently remains long-lasting in AIP patients. We tested the hypothesis that this condition may be associated with alterations of hepatic proteins known to be either increased or decreased in serum according to diverse pathological conditions including malnutrition, inflammation or liver disease., Design: Serum proteins were analyzed in 26 biochemically active AIP patients that were classified according to the EPI (European Porphyria Initiative) guidelines as follows: (i) patients who presented a single acute attack having remained so far free of clinical symptoms; (ii) patients who present recurrent attacks or chronic symptoms associated with exacerbations of AIP., Results: Most of the serum proteins were within normal limits, however insulin-like growth factor 1 (IGF-1) was decreased in 53.8% of AIP patients (z-score = -2.86 +/- 0.37) and transthyretin (prealbumin) was found significantly decreased in 38.5% of them. The IGF-1 z-score was lower in group B versus group A patients (-2.66 vs. -1.43; P = 0.024). The coincident decrease of both IGF-1 and transthyretin was associated with worsening of the clinical condition., Conclusions: This first study in humans suggests that the clinical expression AIP is associated with a state of under-nutrition and/or with hepatic inflammation due to the sustained accumulation of heme-precursors. We propose the use of both IGF-1 and transthyretin as biomarkers of disease morbidity/severity for the clinical follow-up of AIP patients.

Background: Acute intermittent porphyria (AIP) is a metabolic disease affecting hepatic heme biosynthesis. The clinical course in overt disease is characterized by acute attacks of neurovisceral symptoms. Treatment is based on symptomatic relief together with carbohydrate loading and in more severe attacks heme therapy. During an acute attack the heme precursors porphobilinogen (PBG) and 5-aminolevulinic acid (ALA) are produced in high amounts by the liver and are found in high concentrations in plasma and urine. These metabolites represent the acute phase reactants confirming an ongoing attack and are used to evaluate therapeutic measures. The aim of this study was to measure PBG and ALA in plasma and urine during an acute attack and to match the biochemical pattern with the clinical and therapeutical course., Methods: Three consecutive AIP patients were included during four acute attacks. Plasma PBG and ALA were measured by a LC-MS method and in urine by ion-exchange chromatography. The patients received symptomatic and glucose treatment at admission to hospital, and four days later, if necessary, heme therapy., Results: In the three attacks that required heme therapy, plasma PBG concentrations had further increased after admission (p=0.01). In the patient that did not require heme therapy, plasma PBG had decreased after admission., Conclusions: Biochemical monitoring of an acute attack was more accurately reflected by plasma PBG than plasma ALA or urinary PBG and ALA. Glucose administration, in contrast to heme therapy, was not sufficient to achieve clinical and biochemical remission in the more serious attacks.

Porphyrin precursors and porphyrins were measured in three patients with recurrent attacks of acute intermittent porphyria and end-stage renal disease (ESRD): two patients on hemodialysis and one on peritoneal dialysis. Plasma porphobilinogen (PBG) and porphyrins were considerably increased in the three patients. In a previous study, the mean urinary and plasma PBG/ALA ratio in biochemically active AIP patients with conserved renal function was 2.0 (normal 0.3) and plasma porphyrin levels were normal (< 10 nmol/L). In this study we show that the progression to ESRD was paralleled by an increase in urinary and plasma PBG/ALA ratio reaching levels above 6 and higher. Plasma porphyrin increased to levels above 1000 nmol/L causing cutaneous lesions resembling porphyria cutanea tarda. The porphyrin precursors were readily filtered by dialysis membranes but not the porphyrins. The development of kidney failure was a devastating complication in these AIP patients with chronic active disease, leading to unavoidable deterioration of peripheral veins, progression of peripheral neuropathy, dialysis treatment and secondary cutaneous lesions. The clinical course could not be reversed by medical treatment in any of the cases. Today, combined liver and kidney transplantation should be considered as a therapeutic option.

The objective of this study was to update the clinical issues of acute intermittent porphyria (AIP), as they have not been in focus for years, and to be aware of potentially associated disorders and social consequences. A total of 356 gene carriers of AIP from northern Sweden participated in this retrospective population-based study. Eight mutations were found with a predominance of W198X (89%). Clinical manifestations of AIP (manifest AIP) were identified in 42%, 65% were women. Women were more severely stricken by AIP attacks concerning number and duration, hospital admission and early onset. Men reporting most attacks were > 40 years of age. In addition to traditional symptoms during attacks, fatigue was commonly described. Chronic AIP symptoms and disability pension due to AIP were reported in about 20% of subjects. Precipitating factors were reported with evident sex differences. Half of the gene carriers who were on medications used drugs considered not safe (in 1999), mainly antihypertensive drugs. Smoking was associated with high AIP attack frequency. Elevated levels of ALT, bile acids, creatinine, U-ALA and U-PBG and decreased levels of creatinine clearance were associated with manifest AIP. The same was true for hypertension and myalgia in the legs. Hepatoma was strikingly overrepresented. The high prevalence of manifest AIP in this study could be explained by a mutation-dependent penetrance. Our results emphasize the importance of early diagnosis, counselling and treatment of attacks, screening and treatment of associated disorders.

Acute intermittent porphyria (AIP) is a rare metabolic disorder characterized by mutations of the porphobilinogen deaminase gene. Clinical manifestations of AIP are caused by the neurotoxic effects of increased porphyrin precursors, although the underlying pathophysiology of porphyric neuropathy remains unclear. To further investigate the neurotoxic effect of porphyrins, excitability measurements (stimulus-response, threshold electrotonus, current-threshold relationship and recovery cycle) of peripheral motor axons were undertaken in 20 AIP subjects combined with the results of genetic screening, biochemical and conventional nerve conduction studies. Compared with controls, excitability measurements from five latent AIP patients were normal, while 13 patients who experienced acute porphyric episodes without clinical neuropathy (AIPWN) showed clear differences in their responses to hyperpolarizing currents (e.g. reduced hyperpolarizing I/V slope, P < 0.01). Subsequent mathematical simulation using a model of human axons indicated that this change could be modelled by a reduction in the hyperpolarization-activated, cyclic nucleotide-dependent current (I(H)). In contrast, in one patient tested during an acute neuropathic episode, axons of high threshold with reduced superexcitability, consistent with membrane depolarization and reminiscent of ischemic changes. It is proposed that porphyrin neurotoxicity causes a subclinical reduction in I(H) in AIPWN axons, whereas porphyric neuropathy may develop when reduced activity of the Na(+)/K(+) pump results in membrane depolarization.

Acute intermittent porphyria is a rare disorder of heme biosynthesis with a clinical course characterized by exacerbations of neurologic symptoms. Drugs may precipitate these exacerbations; however, little is known about the safety of most drugs used to treat the disease. We describe a patient with acute intermittent porphyria who developed bone marrow failure and was successfully treated with cyclosporine. This agent has been reported as being unsafe for patients with acute intermittent porphyria based on an in vitro model. However, cyclosporine was used in this patient based on two published case reports of successful cyclosporine therapy in patients with acute intermittent porphyria undergoing kidney transplantation. Our patient tolerated cyclosporine well, and her blood counts demonstrated improvement. To our knowledge, this is the first case report of cyclosporine used for treatment of bone marrow failure in a patient with acute intermittent porphyria. Until more data are available, clinicians should consider immunosuppressive therapy as a safe option for treating certain patients with acute intermittent porphyria.

The acute or inducible hepatic porphyrias comprise four inherited disorders of heme biosynthesis. They usually remain asymptomatic for most of the lifespan of individuals who inherit the specific enzyme deficiencies but may cause life-threatening attacks of neurovisceral symptoms. Failure to consider the diagnosis frequently delays effective treatment, and inappropriate diagnostic tests and/or mistaken interpretation of results may lead to misdiagnosis and inappropriate treatment. The four disorders are ALA dehydratase deficiency porphyria, acute intermittent porphyria, hereditary coproporphyria, and variegate porphyria. Other conditions that clinically and biochemically may mimic acute porphyria include lead poisoning and hereditary tyrosinemia type I. The diagnosis of one of these acute porphyric syndromes should be considered in many patients with otherwise unexplained abdominal pain, severe constipation, systemic arterial hypertension, or other characteristic symptoms. Critical to the rapid diagnosis of the three most common of these disorders is demonstration of markedly increased urinary porphobilinogen (PBG) in a single-void urine specimen. The treatment of choice for all but mild attacks of the acute porphyrias is intravenous hemin therapy, which should be started as soon as possible. Intravenous glucose alone is recommended only for mild attacks (no weakness or hyponatremia) or until hemin is available.

Background: Measurement of 5-aminolevulinic acid (ALA) in serum is potentially useful in acute porphyrias, lead poisoning and hereditary tyrosinemia. Because levels of ALA in serum are about 100 times less than in urine, a highly sensitive method is required. We describe a simple and sensitive fluorometric method that does not require HPLC., Methods: ALA is separated from serum using a cation-exchange resin (DOWEX 50WX8-400), followed by addition of acetylacetone and formaldehyde to produce a fluorescent ALA derivative by the Hantzsch reaction. The fluorescence was measured at an excitation wavelength of 370 nm, and the emission peak was near 463 nm. Results were compared with measurements of ALA by a published HPLC method on the same samples., Results: The fluorometric method was linear for ALA concentrations up to 500 microg/l with a detection limit of about 8.7 microg/l. Within-run variations (N=8) at 25 and 100 microg/l ALA were 3.7% and 3.1%, respectively, and day-to-day variations (N=10) for the same levels were 7.2% and 6.1%, respectively. The regression equation for this method in reference to the HPLC method (Y=0.99X+10.34 for N=34, r=0.98, S(y/x)=36.1) had a slope of near unity and an insignificant y-intercept, although values less than 50 microg/l were generally slightly higher by the fluorometric method than by HPLC. The reference range for serum ALA was 0-79.4 microg/l (35.2+/-22.1, mean+/-S.D., N=42), with a distribution skewed to the left because levels in eight subjects were below the detection limit., Conclusions: This simple, fluorometric method for serum ALA correlated well with the results of the HPLC method, and may be suitable for assessing biochemical expression of acute porphyrias and response to treatment especially when HPLC is not available.

A 7-year-old boy demonstrating hepatosplenomegaly, mild anaemia, mild mental retardation, yellow-brown teeth and dark red urine had excessively elevated levels of urinary delta-aminolevulinic acid, porphobilinogen and uroporphyrin. Furthermore hepta-, hexa-, penta- and copro(I)porphyrins were highly increased in urine. This pattern of porphyrin precursor and metabolite excretion is characteristic of acute intermittent porphyria. The decreased copro(III)/copro(I+III) ratio, normally not found in acute intermittent porphyria, is discussed. The porphobilinogen deaminase activity in red cells was decreased to 2-4%. Mutation analysis revealed a novel homozygous L81P mutation in exon 6 of the porphobilinogen deaminase gene. The father and mother, shown to be gene carriers of the same mutation, are asymptomatic and have normal urinary porphyrin precursor and metabolite excretion.

Background: Case reports display similarities between multiple sclerosis and acute intermittent porphyria (AIP). This study examines whether patients with AIP in general demonstrate white-matter lesions on brain magnetic resonance imaging (MRI) and/or abnormalities in plasma and/or cerebrospinal fluid (CSF) when examined outside attacks. We looked particularly for the presence of oligoclonal bands (OB) of immunoglobulin (Ig) in liquor., Methods: Eight AIP gene carriers without previous episodes of porphyria, mean age 42.8 years (range 30-60), and 8 AIP gene carriers with previous episodes of porphyria, mean age 42.8 years (range 33-62), were examined with brain MRI, venous blood samples and lumbar punctures., Results: Two male AIP gene carriers with previous episodes of porphyria, 58 and 35 years of age, had multiple white-matter, high-signal lesions on T(2)- weighted MRI sequences. Two AIP gene carriers without previous episodes of porphyria, 1 male and 1 female, had less than 5 such lesions. No OB were seen in the CSF in any patient, but 1 carrier had an increased level of protein in the CSF. Seven of 16 subjects (44%) had increased levels of HbA1c (>6.0), suggesting protracted hyperglycemia, and 3 further subjects had borderline levels (5.9)., Conclusion: T(2)-weighted MRI sequences demonstrated multiple white-matter, high-signal lesions in 4 out of 16 AIP gene carriers (25%). No carrier demonstrated OB of Ig in CSF, making it unlikely that demyelinating lesions play a pivotal role in the pathogenesis of CNS symptoms in AIP. Only 1 AIP gene carrier had an increased level of protein in CSF; this contrasts with studies during acute attacks of porphyria. Seven subjects (44%) had abnormally high levels of HbA1c, in spite of the fact that no patient had a previous diagnosis of diabetes mellitus., (Copyright 2004 S. Karger AG, Basel)

Background: Patients with the acute porphyrias may develop renal failure and autonomic dysfunction. Renal damage and sympathetic failure may both cause erythropoietin (EPO) deficiency. In this study, we have investigated serum erythropoietin levels and autonomic function in patients with acute porphyria in clinical remission., Methods: Serum erythropoietin levels and the corresponding haemoglobin (Hb) were assayed in 31 patients with acute porphyria and were compared to 15 type 1 diabetic patients with autonomic neuropathy, 23 patients with iron-deficiency anaemia and 18 healthy individuals., Results: 9 out of 31 porphyric patients showed a normochromic normocytic anaemia with normal ferritin levels. Three patients had borderline-raised serum creatinine levels, and one of them was anaemic. Autonomic function was investigated in seven patients, six of them being anaemic, and the results were normal. Patients with iron-deficiency anaemia showed the expected increase in serum erythropoietin levels in response to a decreasing haemoglobin (r=-0.86, p<0.001). Patients with porphyria had inappropriately low serum erythropoietin levels for the degree of anaemia compared to iron-deficiency patients (p<0.001) although there was still a significant increase in serum erythropoietin with decreasing haemoglobin levels (r=-0.46, p=0.01). In contrast, diabetic autonomic neuropathy patients demonstrated a significant decrease in serum erythropoietin with decreasing Hb levels (r=+0.53, p=0.05)., Conclusions: Patients with acute porphyria may have inappropriately low levels of EPO. In contrast to the diabetic patients, this does not appear to be due to autonomic neuropathy but it may reflect mild renal tubular impairment.

Background: Acute intermittent porphyria is a hereditary error of porphyrin metabolism in which the main metabolic defect is caused by a decrease in porphobilinogen deaminase activity. Previous work has demonstrated a higher prevalence of acute intermittent porphyria in the psychiatric patient population than in the general population. The goal of this study was evaluate 300 psychiatric patients and 150 control subjects to detect acute intermittent porphyria by measurement of porphobilinogen (PBG) deaminase activity in blood., Methods: Screening for porphobilinogen deaminase activity was carried out by fluorometric measurement of porphyrins synthesized during 1 h in blood and the measurement of delta-aminolevulinic acid and porphobilinogen in urine., Results: We found two psychiatric patients, one male and one female, with decreased porphobilinogen deaminase activity. When the families of these patients were studied, one brother was found to have an abnormality. Among controls, a woman was found to have the abnormality and her father was found to have typical features of the disease., Conclusions: These results indicate a prevalence of porphyria in Mexican psychiatric patients similar to controls, and that measurement of PBG deaminase activity is a good tool for defining acute intermittent porphyria carriers.

A 44-year-old woman with acute intermittent porphyria (AIP) was admitted to Kudanzaka Hospital because of abdominal pain. A cholecystectomy was performed in another hospital without improvement. On admission, her transaminases were elevated to greater than 1,000 mU/ml. After an intravenous drip of mainly glucose, her transaminases returned to normal. Her acute attacks occurred during stress, and she died of respiratory failure after repetitive acute episodes. AIP should be included in a list of the differential diagnosis of gastrointestinal diseases, neurosis, and hysteria. This is the first case of AIP accompanied by transient marked elevation of transaminases during an acute attack.

Objectives: To differentiate the porphyrias by clinical and biochemical methods., Design and Methods: We describe levels of blood, urine, and fecal porphyrins and their precursors in the porphyrias and present an algorithm for their biochemical differentiation. Diagnoses were established using clinical and biochemical data. Porphyrin analyses were performed by high performance liquid chromatography., Results and Conclusions: Plasma and urine porphyrin patterns were useful for diagnosis of porphyria cutanea tarda, but not the acute porphyrias. Erythropoietic protoporphyria was confirmed by erythrocyte protoporphyrin assay and erythrocyte fluorescence. Acute intermittent porphyria was diagnosed by increases in urine delta-aminolevulinic acid and porphobilinogen and confirmed by reduced erythrocyte porphobilinogen deaminase activity and normal or near-normal stool porphyrins. Variegate porphyria and hereditary coproporphyria were diagnosed by their characteristic stool porphyrin patterns. This appears to be the most convenient diagnostic approach until molecular abnormalities become more extensively defined and more widely available.

5-Aminolevulinic acid dehydratase (ALAD) activity in two patients with compound heterozygous 5-aminolevulinic acid dehydratase deficiency porphyria was studied over the last 20 years. The patients' enzyme activity was <10% from 1977 to 1997. An acute crisis in each patient was successfully treated by infusion of glucose and heme arginate. After this therapy both urinary 5-aminolevulinic acid (ALA) and total porphyrins were diminished to 65% in patient B. In patient H, ALA was decreased to 80%, and total porphyrins were reduced to 15% after treatment with heme arginate and glucose. The patients remained free of symptoms after this therapy. Family studies of patient B showed cross-reactive immunological material (CRIM), in which the maternal mutation is CRIM(+), whereas the paternal mutation is CRIM(-). Incubation of erythrocyte lysates with ALA decreased porphyrin formation, whereas incubation with porphobilinogen produced porphyrin concentrations within reference values in both patients, confirming that ALAD activity is rate-limiting in these cells.

In order to elucidate the question of free radical involvement in acute porphyric crisis, antioxidants were administered to two acute intermittent porphyria patients with long-standing recurrent attacks. Clinical condition and urinary excretion of porphyrins and porphyrin precursors were monitored before, during and after an eight week therapy with daily doses of vitamin E, beta-carotene, ascorbic acid, selenium, vitamin Q, acetylcysteine, mannitol and carnitine. Blood cell trace element profiles were followed. The administration of the compound antioxidant formula was found not to further impair the clinical or biochemical conditions of the patients but the incidence of the recurrent crises or the severity of the symptoms were not positively affected. Aberrant blood cell trace element profiles with increased granulocyte manganese were normalized during treatment, on cessation of the therapy again resuming the abnormal pretreatment patterns, which may suggest an origin in oxidative stress. No correlation was observed between the concentration of granulocyte manganese and the excretion of 5-aminolaevulinic acid. Indications for participation of this porphyrin precursor in a radical generating process leading to generalized mitochondrial superoxide dismutase induction, as conceivably signalled by increased intracellular manganese, were thus not obtained. The failure to note a clinical response to antioxidant therapy may be due to factors dependent upon dosage of, or interaction between, the antioxidant compounds given, or on restricted bioavailability of the antioxidants at critical anatomical sites, and does not per se invalidate the model of acute porphyria as a hyperoxidative condition.

Acute intermittent porphyria (AIP) is an autosomally dominant inherited metabolic disorders caused by decreased activity of porphobilinogen deaminase, the third enzyme in the human heme biosynthetic pathway. We report here the first mutations in the human porphobilinogen deaminase gene in seven unrelated patients from the Czech and Slovak Republics with acute intermittent porphyria. We used denaturing gradient gel electrophoresis to screen all 15 exons and exon/intron boundaries of the porphobilinogen deaminase gene. Polymerase chain reaction products of abnormal migration patterns were subjected to direct sequencing to identify the causative mutations. Thus we revealed four novel mutations and three which have been previously described. Of the four novel mutations, two were mis-sense (G24S, V267M), one was a single base insertion (158insA) that produced a stop codon 12 codons downstream, and one was a single base substitution in intron 12 (771 + 1) resulting in a splicing defect. The three previously detected mutations were mis-sense mutations (R26C, R26H, G111R). These results suggest a high allelic heterogeneity in Czech and Slovak patients.

Three patients from Moroccan kindreds with acute intermittent porphyria (AIP) are described. The propositus of family A originating from Mrirt, Morocco, is living in Embrun, France. This 26 year-old woman who experienced an acute attack with visceral manifestations presented an elevated urinary level of 5-ALA and PBG, and a half-normal activity in porphobilinogen deaminase (PBG-D) in red blood cells (RBC). The family's survey was carried out by measuring the PBG-D activity in RBC (normal values = 125 +/- 40 U). Three of the 16 subjects tested, beside the propositus, were found to be asymptomatic carriers (PBG-D < 70 U). The two patients of family B, originating from Tetouan in the Rif area, were living in Bastia, Corsica. The two brothers, respectively 37 and 39 years old, had a long history (6 years) of neuropsychiatric manifestations before the AIP diagnosis was evidenced by elevated urinary level of 5-ALA and PBG, and showed a partial deficiency, approximately, 50%, of PBG-D activity in RBC. The youngest patient also presented a peripheral neuropathy and recently died after surgery from an unknown reason at the age of 45.

Fluorescence spectra of plasma porphyrin were measured in 6 patients with the acute intermittent porphyria (AIP), 30 patients with variegate porphyria (PV), 2 patients with hereditary coproporphyria, 2 patients with porphyria cutanea tarda, and in 6 patients with erythropoietic protoporphyria (EPP). It was found that the excitation and emission wavelengths at which maximum fluorescence is seen may help to diagnose and differentiate PV and EPP. In patients with AIP spectrum characteristic for porphyria of such a type was noted in all patients during the attack of disease and in only 33% of patients in remission. Fluorescence spectrum was normal in asymptomatic family members. In variegate porphyria spectrum with a characteristic maximum of fluorescence was noted in all patients during an attack and remission, and 62% of the asymptomatic family members.

Acute intermittent porphyria (AIP) is a hereditary disease characterized biochemically by a defect in the heme pathway enzyme porphobilinogen deaminase. There is wide variability in the neurologic clinical expression of AIP, and the disorder remains latent in most gene carriers. The natural history of the disease and results in a porphyric rat model suggest a significant relationship between tryptophan metabolites and clinical expression of the disease. In the present study, we examined urine and blood tryptophan metabolite levels in AIP women before, during and after acute attacks and treatment by heme arginate. Heme arginate treatment promptly decreased total tryptophan levels (from 69 +/- 9, to 44 +/- 5, mean +/- SEM, mumole/l, p < 0.001), serotonin blood levels (from 629 +/- 103, to 356 +/- 80, nmole/l, p < 0.01) and the urinary excretion of 5-HIAA (from 3.9 +/- 0.6, to 2.2 +/- 0.4, mumole/mmole creatinine, p < 0.01). The plasma level of melatonin was found much lower than the normal control level at night (86.2 +/- 70.3, vs the normal range, 409 +/- 78.9, pmole/l +/- SEM) and day time (38.8 +/- 22.0, vs 75 +/- 13.7). Heme arginate treatment did not influence melatonin levels. Our results support the involvement of abnormal tryptophan metabolism in the pathophysiology of AIP acute attacks. Low melatonin plasma levels in porphyric women suggest that the defect of the pineal hormone may be responsible for the recurrent aspect of porphyric attacks. A desynchronization of biological rhythms in AIP patients may increase the inducibility of hepatic ALA synthase to environmental risk factors and, specially, to sex steroid hormones.