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Acute intermittent porphyria (AIP) is a disease affecting the heme biosynthesis pathway caused by mutations of the hydroxymethylbilane synthase (HMBS) gene. AIP is thought to display autosomal dominant inheritance with incomplete penetrance. We evaluated the prevalence, penetrance and heritability of AIP, in families with the disease from the French reference center for porphyria (CFP) (602 overt patients; 1968 relatives) and the general population, using Exome Variant Server (EVS; 12 990 alleles) data. The pathogenicity of the 42 missense variants identified was assessed in silico, and in vitro, by measuring residual HMBS activity of the recombinant protein. The minimal estimated prevalence of AIP in the general population was 1/1299. Thus, 50 000 subjects would be expected to carry the AIP genetic trait in France. Penetrance was estimated at 22.9% in families with AIP, but at only 0.5-1% in the general population. Intrafamily correlation studies showed correlations to be strong overall and modulated by kinship and the area in which the person was living, demonstrating strong influences of genetic and environmental modifiers on inheritance. Null alleles were associated with a more severe phenotype and a higher penetrance than for other mutant alleles. In conclusion, the striking difference in the penetrance of HMBS mutations between the general population and the French AIP families suggests that AIP inheritance does not follow the classical autosomal dominant model, instead of being modulated by strong environmental and genetic factors independent from HMBS. An oligogenic inheritance model with environmental modifiers might better explain AIP penetrance and heritability.

Acute intermittent porphyria (AIP) is a genetic disorder of the synthesis of heme caused by a deficiency in hydroxymethylbilane synthase (HMBS), leading to the overproduction of the porphyrin precursors δ-aminolevulinic acid and porphobilinogen. The aim of this study is to describe the clinical and biological characteristics, the renal pathology, and the cellular mechanisms of chronic kidney disease associated with AIP. A total of 415 patients with HMBS deficiency followed up in the French Porphyria Center were enrolled in 2003 in a population-based study. A follow-up study was conducted in 2013, assessing patients for clinical, biological, and histological parameters. In vitro models were used to determine whether porphyrin precursors promote tubular and endothelial cytotoxicity. Chronic kidney disease occurred in up to 59% of the symptomatic AIP patients, with a decline in the glomerular filtration rate of ~1 ml/min per 1.73 m(2) annually. Proteinuria was absent in the vast majority of the cases. The renal pathology was a chronic tubulointerstitial nephropathy, associated with a fibrous intimal hyperplasia and focal cortical atrophy. Our experimental data provide evidence that porphyrin precursors promote endoplasmic reticulum stress, apoptosis, and epithelial phenotypic changes in proximal tubular cells. In conclusion, the diagnosis of chronic kidney disease associated with AIP should be considered in cases of chronic tubulointerstitial nephropathy and/or focal cortical atrophy with severe proliferative arteriosclerosis.

Background & Aims: Prothrombin induced by vitamin K absence-II (PIVKA-II) is a diagnostic and surveillance marker for HCC mainly used in Asia, and has also been shown to be a predictor of microvascular invasion (MVI), a major prognostic factor in HCC. However, experience with PIVKA-II in Europe remains limited., Methods: In a French cohort, we conducted a case-control study to compare the performances of α-fetoprotein (AFP) and PIVKA-II serum levels for diagnosis of early stage HCC, and we determined the value of PIVKA-II serum and tissue expression in pre-operative detection of MVI. 43 cirrhotic control patients and 85 HCC cases were included, of which 54 (63.5%) had early stage HCC (n=22 very early, n=32 early). PIVKA-II tissue expression was assessed by immunohistochemistry in HCC surgical samples., Results: For the diagnosis of early HCC, PIVKA-II had a sensitivity of 77% and a specificity of 82% at a cut-off of 42 mAU/ml, vs. 61% and 50% for AFP at a cut-off of 5.5 ng/ml (AUC 0.81 vs. 0.58, respectively). A PIVKA-II level >90 mAU/ml was an independent predictor of MVI (HR 3.5; 95% CI 1.08-11.8; p=0.043). High PIVKA-II tissue expression was significantly associated with the presence of MVI (p=0.001). When combining PIVKA-II immunostaining with the PIVKA-II serum level, sensitivity and specificity for the diagnosis of MVI increased from 70% to 87% and 63% to 90%, respectively., Conclusions: PIVKA-II was more efficient than AFP for the diagnosis of early HCC, and could be used as a predictive biomarker of MVI., (Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background: Porphyria cutanea tarda (PCT), the most frequent type of porphyria, results from decreased uroporphyrinogen decarboxylase (UROD) activity. Two forms of PCT have been described: a familial form (fPCT) characterized by the inherited decrease of UROD activity in all tissues and a sporadic form (sPCT) characterized by decreased UROD activity in the liver. Cytochrome P450 CYP1A2 plays a major role in triggering experimental uroporphyria in rodents. It has been suggested that the highly inducible -163A/A genotype of the CYP1A2 gene could confer a heightened risk of PCT in patients., Objectives: To examine the impact of CYP1A2 polymorphisms on the clinical course of PCT., Methods: We performed an extensive CYP1A2 gene analysis in 96 (48 fPCT and 48 sPCT) unrelated French caucasian patients with PCT and in 99 healthy volunteers of similar ethnic origin. Results  We did not observe any difference in CYP1A2 allele distribution, including a novel and rare CYP1A2 c.1063C>T (p.R355W) single nucleotide polymorphism. In addition, we compared the frequency of the -163A highly inducible allele both in patients with symptomatic fPCT (n = 48) and in asymptomatic UROD gene mutations carrier relatives (n=54). This variant was not over-represented in patients with PCT vs. either healthy volunteers or asymptomatic UROD gene mutation carriers., Conclusions: The CYP1A2 genotype does not appear to be a major susceptibility factor in the development of fPCT or sPCT in the French population., (© 2011 The Authors. BJD © 2011 British Association of Dermatologists.)

Purpose: Inflammation-induced anemia is frequent among critically ill patients and can be aggravated by true iron deficiency (ID) resulting from blood losses. The serum hepcidin level controls the availability of iron for erythropoiesis, and its determination offers new perspectives for the diagnosis of ID in the presence of inflammation. We conducted a prospective observational study to determine the cutoff value and diagnostic accuracy of hepcidin levels for detecting ID in critically ill anemic patients., Methods: Patients suffering from anemia (hemoglobin <100 g/l) and expected to stay for more than 7 days in intensive care had weekly determinations of hematological and iron parameters, including hepcidin levels (ELISA test). The iron status for each set of measures was determined by the consensus of three experts, blinded to hepcidin values., Results: Of 51 patients (36 male/15 female), 5 had ID at inclusion, while 8 developed ID during their stay. A total of 128 iron profiles were analyzed. Median hepcidin levels were 80.5 (0.05-548.3) and 526.6 (246.7-891.4) microg/l for ID and non-ID profiles, respectively. The onset of ID during the ICU stay led to a progressive decline in hepcidin levels, whereas a persistent inflammatory profile remained associated with high hepcidin concentrations. The optimal threshold for serum hepcidin for ID diagnosis was assessed by building 100 ROC curves using a resampling method and found at 129.5 microg/l [95% CI = (115.5-143.4)]., Conclusions: Hepcidin levels may be suppressed by ID even in case of inflammation. Serum hepcidin of 129.5 microg/l was the most accurate threshold for ID diagnosis in critically ill patients with anemia.

Erythropoietic protoporphyria (EPP) is an inherited disorder of heme biosynthesis that results from a partial deficiency of ferrochelatase (FECH). Recently, we have shown that the inheritance of the common hypomorphic IVS3-48C allele trans to a deleterious mutation reduces FECH activity to below a critical threshold and accounts for the photosensitivity seen in patients. Rare cases of autosomal recessive inheritance have been reported. We studied a cohort of 173 white French EPP families and a group of 360 unrelated healthy subjects from four ethnic groups. The prevalences of the recessive and dominant autosomal forms of EPP are 4% (95% confidence interval 1-8) and 95% (95% confidence interval 91-99), respectively. In 97.9% of dominant cases, an IVS3-48C allele is co-inherited with the deleterious mutation. The frequency of the IVS3-48C allele differs widely in the Japanese (43%), southeast Asian (31%), white French (11%), North African (2.7%), and black West African (<1%) populations. These differences can be related to the prevalence of EPP in these populations and could account for the absence of EPP in black subjects. The phylogenic origin of the IVS3-48C haplotypes strongly suggests that the IVS3-48C allele arose from a single recent mutational event. Estimation of the age of the IVS3-48C allele from haplotype data in white and Asian populations yields an estimated age three to four times younger in the Japanese than in the white population, and this difference may be attributable either to differing demographic histories or to positive selection for the IVS3-48C allele in the Asian population. Finally, by calculating the KA/KS ratio in humans and chimpanzees, we show that the FECH protein sequence is subject to strong negative pressure. Overall, EPP looks like a Mendelian disorder, in which the prevalence of overt disease depends mainly on the frequency of a single common single-nucleotide polymorphism resulting from a unique mutational event that occurred 60,000 years ago.

Porphyria cutanea tarda (PCT), a disorder characterized by a photosensitive dermatosis and hepatic siderosis, is caused by a decreased activity of the hepatic enzyme uroporphyrinogen decarboxylase (UROD). Two forms of PCT have been described: a familial one (fPCT) with an inherited decrease of UROD activity in all tissues and a sporadic one (sPCT) with a decreased UROD activity restricted to the liver. Iron overload and acquired factors including hepatic viral infections, alcohol, drugs contribute to the expression of PCT. In 65 French sPCT patients and 108 controls we have evaluated the respective role of iron and HCV status, the hemochromatosis (HFE) gene mutations frequencies (H63D. S65C, C282Y), and in a case control study we searched for an association between sPCT and the human transferrin receptor-1 (TFRC1) gene whose product is thought to be in functional association with the HFE protein: three single nucleotide polymorphisms (SNPs) previously characterized and 2 novel ones were studied. The iron-related parameters and transaminases were higher in sPCT patients than those of non-porphyric controls. Of the sPCT patients studied, 28% were HCV positive. In the HFE gene, 17% of sPCT patients carried C282Y mutation compared to 4% in controls, no significant differences were found with H63D and S65C variants. Compound heterozygous genotypes, C282Y/H63D or C282Y/S65C, were not significantly different in sPCT and control groups. Independently from HFE gene mutations, an association was found between the IVS4+198 T allele in the TFRC1 gene and sPCT patients. Analysis of HFE genotypes indicated that C282Y (but not H63D nor S65C) is a susceptibility factor for the development of sPCT in West European continental patients. However, analysis of TFRC1 genotypes suggest that sPCT should be considered as a multifactorial disorder in which other intracellular iron metabolism genes could be involved.

Erythropoietic protoporphyria (EPP) is an inherited disorder of heme biosynthesis caused by a partial deficiency of ferrochelatase (FECH, EC 4.99.1.1). EPP is transmitted as an autosomal dominant disorder with an incomplete penetrance. Using haplotype segregation analysis, we have identified an intronic single nucleotide polymorphism (SNP), IVS3-48T/C, that modulates the use of a constitutive aberrant acceptor splice site. The aberrantly spliced mRNA is degraded by a nonsense-mediated decay mechanism (NMD), producing a decreased steady-state level of mRNA and the additional FECH enzyme deficiency necessary for EPP phenotypic expression.

Erythropoietic protoporphyria (EPP; MIM 177000) is an inherited disorder caused by partial deficiency of ferrochelatase (FECH), the last enzyme in the heme biosynthetic pathway. In EPP patients, the FECH deficiency causes accumulation of free protoporphyrin in the erythron, associated with a painful skin photosensitivity. In rare cases, the massive accumulation of protoporphyrin in hepatocytes may lead to a rapidly progressive liver failure. The mode of inheritance in EPP is complex and can be either autosomal dominant with low clinical penetrance, as it is in most cases, or autosomal recessive. To acquire an in-depth knowledge of the genetic basis of EPP, we conducted a systematic mutation analysis of the FECH gene, following a procedure that combines the exon-by-exon denaturing-gradient-gel-electrophoresis screening of the FECH genomic DNA and direct sequencing. Twenty different mutations, 15 of which are newly described here, have been characterized in 26 of 29 EPP patients of Swiss and French origin. All the EPP patients, including those with liver complications, were heterozygous for the mutations identified in the FECH gene. The deleterious effect of all missense mutations has been assessed by bacterial expression of the respective FECH cDNAs generated by site-directed mutagenesis. Mutations leading to a null allele were a common feature among three EPP pedigrees with liver complications. Our systematic molecular study has resulted in a significant enlargement of the mutation repertoire in the FECH gene and has shed new light on the hereditary behavior of EPP.

Objectives: Acute intermittent porphyria (AIP) is an autosomal dominant disorder resulting from a 50% deficiency in porphobilinogen deaminase (PBG deaminase). The true prevalence in the general population of mutations in the PBG deaminase gene capable of causing AIP is unknown. However, it is important to identify asymptomatic carriers of AIP mutations because all are at risk to have an acute attack., Design: We measured erythrocyte PBG deaminase from 3350 healthy blood donors. When a clear cut deficiency (< mean minus 2.5 SD) was found, the PBG deaminase gene was analysed by molecular biology technics., Subjects: Four subjects with PBG deaminase deficiency were identified. Two had mutations in the PBG deaminase gene which are known to cause AIP., Conclusion: We conclude that, in France, the mutations of the PBG deaminase gene show a high prevalence in the healthy population. If only these two confirmed latent cases are used for the calculation, in France the minimal prevalence of the AIP gene is 1:1675.

Acute intermittent porphyria (AIP) is an autosomal dominant disorder characterized by alterations of the gene encoding porphobilinogen deaminase (PBGD: EC 4.3.1.8), the third enzyme of the heme biosynthetic pathway. The molecular heterogeneity of the mutations causing AlP has been demonstrated with a reported predominance of single base substitutions resulting in amino acid changes. The molecular basis of AIP in four French patients was investigated using denaturing gradient gel electrophoresis followed by direct sequencing. We describe four different novel mutations that affected exon 12 (a frameshift and an exon skipping), exon 4 (a stop codon) and exon 15 (a frameshift inducing a stop codon). This study further documents the molecular heterogeneity of mutations in the PBGD gene in the French Caucasian population and reports types of mutations relatively uncommon in AIP.