RESUMEN
The porphyrias are a group of rare inborn errors of metabolism associated with various clinical presentations and long-term complications, making them relevant differential diagnoses to consider for many clinical specialities, especially hepatologists, gastroenterologists and dermatologists. To diagnose a patient with porphyria requires appropriate biochemical investigations, as clinical features alone are not specific enough. Furthermore, it is important to be aware that abnormalities of porphyrin accumulation and excretion occur in many other disorders that are collectively far more common than the porphyrias. In this review, we provide an overview of porphyria-related tests with their strengths and limitations, give recommendations on requesting and diagnostic approaches in non-expert and expert laboratories for different clinical scenarios and discuss the role of genetic testing in the porphyrias. To diagnose porphyria in a currently symptomatic patient requires analysis of biochemical markers to demonstrate typical patterns of haem precursors in urine, faeces and blood. The use of genomic sequencing in diagnostic pathways for porphyrias requires careful consideration, and the demonstration of increased porphyrin-related markers is necessary prior to genomic testing in symptomatic patients. In the acute porphyrias, genomic testing is presently a useful adjunct for genetic counselling of asymptomatic family members and the most common cutaneous porphyria, porphyria cutanea tarda, is usually a sporadic, non-hereditary disease. Getting a correct and timely porphyria diagnosis is essential for delivering appropriate care and ensuring best patient outcome.
RESUMEN
Congenital erythropoietic porphyria (CEP) is a rare autosomal recessive disease due to the deficient, but not absent, activity of uroporphyrinogen III synthase (UROS), the fourth enzyme in the heme biosynthesis pathway. Biallelic variants in the UROS gene result in decreased UROS enzymatic activity and the accumulation of non-physiologic Type I porphyrins in cells and fluids. Overproduced uroporphyrins in haematopoietic cells are released into the circulation and distributed to tissues, inducing primarily hematologic and dermatologic symptoms. The clinical manifestations vary in severity ranging from non-immune hydrops fetalis in utero to mild dermatologic manifestations in adults. Here, the biochemical, molecular and clinical features of CEP as well as current and new treatment options, including the rescue of UROS enzyme activity by chaperones, are presented.
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Porfiria Eritropoyética , Uroporfirinógeno III Sintetasa , Humanos , Porfiria Eritropoyética/genética , Porfiria Eritropoyética/diagnóstico , Porfiria Eritropoyética/terapia , Uroporfirinógeno III Sintetasa/genética , Uroporfirinógeno III Sintetasa/metabolismo , Uroporfirinas/genéticaRESUMEN
Defects in hydroxymethylbilane synthase (HMBS) can cause acute intermittent porphyria (AIP), an acute neurological disease. Although sequencing-based diagnosis can be definitive, â¼â of clinical HMBS variants are missense variants, and most clinically reported HMBS missense variants are designated as "variants of uncertain significance" (VUSs). Using saturation mutagenesis, en masse selection, and sequencing, we applied a multiplexed validated assay to both the erythroid-specific and ubiquitous isoforms of HMBS, obtaining confident functional impact scores for >84% of all possible amino acid substitutions. The resulting variant effect maps generally agreed with biochemical expectations and provide further evidence that HMBS can function as a monomer. Additionally, the maps implicated specific residues as having roles in active site dynamics, which was further supported by molecular dynamics simulations. Most importantly, these maps can help discriminate pathogenic from benign HMBS variants, proactively providing evidence even for yet-to-be-observed clinical missense variants.
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Hidroximetilbilano Sintasa , Porfiria Intermitente Aguda , Humanos , Hidroximetilbilano Sintasa/química , Hidroximetilbilano Sintasa/genética , Hidroximetilbilano Sintasa/metabolismo , Mutación Missense/genética , Porfiria Intermitente Aguda/diagnóstico , Porfiria Intermitente Aguda/genética , Sustitución de Aminoácidos , Simulación de Dinámica MolecularRESUMEN
Defects in hydroxymethylbilane synthase (HMBS) can cause Acute Intermittent Porphyria (AIP), an acute neurological disease. Although sequencing-based diagnosis can be definitive, ~â of clinical HMBS variants are missense variants, and most clinically-reported HMBS missense variants are designated as "variants of uncertain significance" (VUS). Using saturation mutagenesis, en masse selection, and sequencing, we applied a multiplexed validated assay to both the erythroid-specific and ubiquitous isoforms of HMBS, obtaining confident functional impact scores for >84% of all possible amino-acid substitutions. The resulting variant effect maps generally agreed with biochemical expectation. However, the maps showed variants at the dimerization interface to be unexpectedly well tolerated, and suggested residue roles in active site dynamics that were supported by molecular dynamics simulations. Most importantly, these HMBS variant effect maps can help discriminate pathogenic from benign variants, proactively providing evidence even for yet-to-be-observed clinical missense variants.
RESUMEN
Acute intermittent porphyria (AIP) is a rare disease caused by a deficiency of hydroxymethylbilane synthase (HMBS), the third enzyme of the heme-synthesis pathway. Decreased enzymatic activity in the liver induces an overproduction of heme-precursors and acute neurological attacks. We report a 36-years-old female with AIP with a long-term history of severe, disabling, recurrent attacks, who underwent curative liver transplantation. Tissue samples from the explant were obtained for transcriptome analysis. Whole RNA was extracted and 16 gene-transcripts were selected and investigated by quantitative polymerase chain reaction. These included nine genes encoding enzymes that consecutively catalyze heme-synthesis and catabolism in the liver (ALAS1; ALAD; HMBS; UROS; UROD; CPOX; PPOX; FECH; HMOX1). Additionally, we studied genes related to inflammation (IL6; TNF) insulin signaling (PGC-1α; IGF-1; FOXO-1) and tryptophan metabolism (TDO2; IDO). Transcripts of eight house-keeping genes were co-measured for normalization. All transcripts were also measured in five control samples from healthy living liver donors. The transcriptome of the controls showed important differences between the various genes, with the first two genes of the heme-synthesis pathway, ALAS1 and ALAD showing strikingly high mRNA levels compared to the consecutive HMBS gene. Transcripts of several genes significantly differed in the AIP liver compared to controls. Transcripts of HMOX1 and UROS were increased in the AIP liver whereas transcripts of UROD; CPOX, PPOX, and TDO2 were decreased. ALAS1 expression was not increased, possibly due to hemin administered to the patient before transplantation. These results highlight several transcriptomic changes related to heme homeostasis in AIP.
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Patients with major forms of acute hepatic porphyria present acute neurological attacks with overproduction of porphobilinogen (PBG) and δ-aminolevulinic acid (ALA). Even if ALA is considered the most likely agent inducing the acute symptoms, the mechanism of its accumulation has not been experimentally demonstrated. In the most frequent form, acute intermittent porphyria (AIP), inherited gene mutations induce a deficiency in PBG deaminase; thus, accumulation of the substrate PBG is biochemically obligated but not that of ALA. A similar scenario is observed in other forms of acute hepatic porphyria (i.e., porphyria variegate, VP) in which PBG deaminase is inhibited by metabolic intermediates. Here, we have investigated the molecular basis of δ-aminolevulinate accumulation using in vitro fluxomics monitored by NMR spectroscopy and other biophysical techniques. Our results show that porphobilinogen, the natural product of δ-aminolevulinate deaminase, effectively inhibits its anabolic enzyme at abnormally low concentrations. Structurally, this high affinity can be explained by the interactions that porphobilinogen generates with the active site, most of them shared with the substrate. Enzymatically, our flux analysis of an altered heme pathway demonstrates that a minimum accumulation of porphobilinogen will immediately trigger the accumulation of δ-aminolevulinate, a long-lasting observation in patients suffering from acute porphyrias.
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Porfiria Intermitente Aguda , Porfirias Hepáticas , Humanos , Porfiria Intermitente Aguda/genética , Porfiria Intermitente Aguda/metabolismo , Porfobilinógeno , Hidroximetilbilano Sintasa/genética , Hidroximetilbilano Sintasa/metabolismo , Porfirias Hepáticas/genéticaRESUMEN
Acute intermittent porphyria (AIP) is an inherited rare hepatic disorder due to mutations within the hydroxymethylbilane gene. AIP patients with active disease overproduce aminolevulinic acid (ALA) and porphobilinogen (PBG) in the liver which are exported inducing severe neurological attacks. Different hepatic metabolic abnormalities have been described to be associated with this condition. The goal of this research was to explore the metabolome of symptomatic AIP patients by state-of-the art liquid chromatography-tandem mass spectrometry (LC-MS/MS). A case versus control study including 18 symptomatic AIP patients and 33 healthy controls was performed. Plasmatic levels of 51 metabolites and 16 ratios belonging to four metabolic pathways were determined. The results showed that the AIP patients presented significant changes in the two main areas of the metabolome under study: (a) the tryptophan/kynurenine pathway with an increase of tryptophan in plasma together with increase of the kynurenine/tryptophan ratio; and (b) changes in the tricarboxylic acid cycle (TCA) including increase of succinic acid and decrease of the fumaric acid/succinic acid ratio. We performed a complementary in vitro study adding ALA to hepatocytes media that showed some of the effects on the TCA cycle were parallel to those observed in vivo. Our study confirms in plasma previous results obtained in urine showing that AIP patients present a moderate increase of the kynurenine/tryptophan ratio possibly associated with inflammation. In addition, it also reports changes in the mitochondrial TCA cycle that, despite requiring further research, could be associated with an energy misbalance due to sustained overproduction of heme-precursors in the liver.
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Porfiria Intermitente Aguda , Ácido Aminolevulínico/orina , Cromatografía Liquida , Humanos , Quinurenina , Metabolómica , Ácido Succínico , Espectrometría de Masas en Tándem , TriptófanoRESUMEN
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-ß-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.
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Acetilgalactosamina/análogos & derivados , Arginina/deficiencia , Hemo/deficiencia , Hiperhomocisteinemia/etiología , Porfiria Intermitente Aguda/tratamiento farmacológico , Pirrolidinas/uso terapéutico , Acetilgalactosamina/efectos adversos , Acetilgalactosamina/uso terapéutico , Adulto , Arginina/uso terapéutico , Cistationina betasintasa/genética , Femenino , Ácido Fólico/sangre , Hemo/uso terapéutico , Homeostasis , Homocisteína/metabolismo , Homocistinuria/complicaciones , Humanos , Hidroximetilbilano Sintasa/sangre , Hidroximetilbilano Sintasa/genética , Masculino , Metionina/sangre , Persona de Mediana Edad , Porfiria Intermitente Aguda/sangre , Porfiria Intermitente Aguda/complicaciones , Porfiria Intermitente Aguda/genética , Fosfato de Piridoxal/sangre , Pirrolidinas/efectos adversos , Adulto JovenRESUMEN
BACKGROUND: Amphetamine urine drug screening by immunoassay is prone to cross-react with other compounds leading to false positive results. Tetracaine is a local anesthetic drug used in the clinical setting as an ointment during urinary catheterization. In our laboratory, tetracaine is often detected by gas chromatography-mass spectrometry in the urine of patients admitted in the emergency department with false positive amphetamine results. The objectives of this study were to investigate if there was cross-reactivity to tetracaine in an amphetamine immunoassay and to retrospectively evaluate the potential contribution of tetracaine to false positive amphetamine results. METHODS: An interference study was conducted using negative urine samples spiked with increasing concentrations of tetracaine hydrochloride and analyzed with the CEDIA Amphetamine/Ecstasy immunoassay. Retrospectively, urine samples of patients which yielded positive amphetamine immunoassay results and were analyzed by gas chromatography-mass spectrometry were reviewed (n = 417). The presence of tetracaine and/or other drugs by gas chromatography-mass spectrometry were gathered. RESULTS: Tetracaine caused false positive amphetamine results by immunoassay (cut-off 1000 µg/L) with a concentration of above 40 mg/L. Retrospective analysis of all positive amphetamine immunoassay samples showed that in 45 out of the 417 (10.8%) urine samples no amphetamine-like derivative was identified by gas chromatography - mass spectrometry. In 37 out of 45 (82.2%) of these false positive cases tetracaine was detected, of whom 59.5% (22/37) had an estimated tetracaine concentration of ≥40 mg/L. CONCLUSIONS: This study confirmed the interference of tetracaine in the CEDIA Amphetamine/Ecstasy immunoassay and that tetracaine may have contributed to around 80% of the false positive amphetamine cases in the urine samples of patients admitted to the emergency department at our institution.
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Anfetamina/orina , Inmunoensayo , Tetracaína/orina , Reacciones Cruzadas , Reacciones Falso Positivas , Cromatografía de Gases y Espectrometría de Masas , Humanos , Pomadas , Estudios Retrospectivos , Cateterismo UrinarioRESUMEN
Introduction: 6-Monoacetylmorphine (6-MAM) is a specific metabolite of heroin. Thus, the presence of 6-MAM in urine is a definitive indication of heroin intake. The possibility of having an immunoassay procedure to measure 6-MAM would be a diagnosis tool to discriminate, among opiates-positive, those patients who have consumed heroin and those who have not.Methods: EMIT® II Plus 6-Acetylmorphine Assay was used to measure 6-MAM in urine. The positive opiate screening results were confirmed at the Toxicology laboratory of our hospital by GC-MS.Results: This study includes 63 urine samples from subjects admitted to emergency department with suspicion of opiate consumption. Specificity was evaluated in the two groups of samples studied. In the first group all samples which resulted negative by opiate immunoassay (n = 33) were negative for 6-MAM immunoassay test. Thus, the specificity obtained for 6-MAM immunoassay in this group was 100%. Regarding the second specificity study, performed in positive samples by opiate immunoassay which were negative to 6 MAM by GC-MS, the specificity decreased down to 75%. In the study of sensitivity all samples confirmed as positive to 6-MAM by confirmatory method (GC-MS) resulted positive by the screening method, thus sensitivity obtained was 100%.Discussion: In this study no FN for 6-MAM was observed and therefore the new Emit® II Plus 6- Acetylmorphine Assay procedure has a high NPV, thus a negative result with 6-MAM immunoassay practically excludes heroine consume. The positive results to 6-MAM by immunoassay should be confirmed by a more analytically specific method, such as GCMS.
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Dependencia de Heroína/diagnóstico , Inmunoensayo , Derivados de la Morfina/orina , Detección de Abuso de Sustancias , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Automatización de Laboratorios , Biomarcadores/orina , Femenino , Cromatografía de Gases y Espectrometría de Masas , Dependencia de Heroína/orina , Humanos , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Reproducibilidad de los Resultados , Urinálisis , Adulto JovenRESUMEN
BACKGROUND: Hepatitis C virus (HCV) is a risk factor for porphyria cutanea tarda (PCT), a rare disease originating in the liver characterised by overproduction of porphyrins. Although hepatitis C infection is highly prevalent among patients with porphyria, only a minority of hepatitis C patients develop PCT. AIMS: To explore the presence of porphyrin abnormalities in a cohort of asymptomatic hepatitis C-infected patients and the impact of anti-viral therapy. METHODS: Eighty-four consecutive patients with HCV infection treated with direct-acting antivirals after 1 January 2018 were longitudinally evaluated for the presence of porphyrin abnormalities. Those patients with biochemical abnormalities at baseline were additionally evaluated at follow-up. Porphyrins in urine were screened by fluorometry and isomer separation was performed by liquid chromatography. RESULTS: In five patients, all of them asymptomatic, porphyrin profile abnormalities were detected: three presented significant increased urinary porphyrins with a typical PCT profile, and two showed normal levels of urinary porphyrins, but abnormal porphyria-like profiles. Urine evaluation after hepatitis C cure showed complete normalisation of the urinary porphyrins in all patients, confirming the biochemical cure of the disease. CONCLUSIONS: We document the existence of rare cases of hepatitis C-infected patients with significant uroporphyrinuria in the absence of dermatological manifestations. Anti-viral therapy normalises the biochemical disorder, preventing patients from presenting PCT associated complications.
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Antivirales/uso terapéutico , Hepatitis C/tratamiento farmacológico , Porfiria Cutánea Tardía/virología , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Porfirinas/orina , Factores de RiesgoRESUMEN
With the advent of precision and genomic medicine, a critical issue is whether a disease gene variant is pathogenic or benign. Such is the case for the three autosomal dominant acute hepatic porphyrias (AHPs), including acute intermittent porphyria, hereditary coproporphyria, and variegate porphyria, each resulting from the half-normal enzymatic activities of hydroxymethylbilane synthase, coproporphyrinogen oxidase, and protoporphyrinogen oxidase, respectively. To date, there is no public database that documents the likely pathogenicity of variants causing the porphyrias, and more specifically, the AHPs with biochemically and clinically verified information. Therefore, an international collaborative with the European Porphyria Network and the National Institutes of Health/National Center for Advancing Translational Sciences/National Institute of Diabetes and Digestive and Kidney Diseases (NIH/NCATS/NIDDK)-sponsored Porphyrias Consortium of porphyria diagnostic experts is establishing an online database that will collate biochemical and clinical evidence verifying the pathogenicity of the published and newly identified variants in the AHP-causing genes. The overall goal of the International Porphyria Molecular Diagnostic Collaborative is to determine the pathogenic and benign variants for all eight porphyrias. Here we describe the overall objectives and the initial efforts to validate pathogenic and benign variants in the respective heme biosynthetic genes causing the AHPs.
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Porfirias/genética , Porfirias/fisiopatología , Virulencia/genética , Curaduría de Datos/métodos , Bases de Datos Factuales , Femenino , Humanos , Masculino , Patología Molecular , Porfobilinógeno Sintasa/deficiencia , Porfobilinógeno Sintasa/genética , Porfiria Intermitente Aguda/genética , Porfiria Intermitente Aguda/fisiopatología , Porfirias Hepáticas/genética , Porfirias Hepáticas/fisiopatología , Estados UnidosRESUMEN
Porphyria cutanea tarda (PCT) arises from a deficiency of uroporphyrinogen decarboxylase (UROD) in the liver. Several exogenous risk factors are associated with the acquired form of the disease. In Southern Europe, PCT is strongly linked to hepatitis C virus (HCV) infection to the point that a high prevalence of viral infection in some geographic areas generated an increase of PCT cases as a complication. In spite of the association, PCT is a rare complication of HCV infection, thus suggesting the existence of susceptibility factors operating in only some patients. Investigation of liver specimens of PCT patients showed iron accumulation, which albeit moderate, was higher in comparison with HCV-infected patients without PCT. Measurements of hepcidin in serum of HCV-infected patients with and without PCT and calculation of hepcidin/ferritin ratio were compatible with the hypothesis that HCV induced inadequate response of hepcidin to iron accumulation. Administration of direct-acting antivirals (DAA) to HCV-infected patients with active PCT showed that eradication of the virus was followed by resolution of PCT and rapid disappearance of urinary porphyrins. This suggests a direct participation of the virus in the oxidative mechanism leading to UROD inhibition. If clinical evolution of HCV- PCT-patients is placed within a time-frame, rapid PCT resolution by DAA is in striking contrast with a long-delay (in most cases of decades) between viral infection and appearance of overt porphyria. This could be explained if HCV infection (a): enhanced an oxidative environment in the vicinity of UROD and (b): facilitated iron accumulation through hepdicin down-regulation. Thus, only when iron accumulation reached a threshold, inhibition of UROD attained a critical level. However, the enigma is why only a minority of HCV-infected patients develop PCT. If additional risk factors (i.e. alcohol abuse) are not concurring, it should be concluded that modifier genes or epigenetic mechanisms related to iron homeostasis, facilitate iron progressive accumulation in only a minority susceptible patients.
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Hepacivirus/patogenicidad , Hepatitis C/complicaciones , Porfiria Cutánea Tardía/virología , Animales , Europa (Continente) , Hepatitis C/epidemiología , Hepcidinas/sangre , Homeostasis , Humanos , Hierro/metabolismo , Hígado/patología , Hígado/virología , Ratones , Porfiria Cutánea Tardía/epidemiología , Prevalencia , Factores de RiesgoRESUMEN
BACKGORUND & AIMS: Mushroom poisoning with Amanita phalloides or similar species can lead to liver failure with 10-30% mortality rates. We aimed at defining the prognostic value of urinary amatoxin quantification in patients with hepatotoxic mushroom poisoning. METHODS: Data from 32 patients with hepatotoxic mushroom poisoning (Hospital Clínic Barcelona, 2002-16) in whom urinary amatoxins were determined (ELISA) were retrospectively reviewed. Correlations between urinary amatoxin and collected baseline variables with outcomes including hepatotoxicity (ALT>1000 U/L), severe acute liver injury (ALI, prothrombin <50%), acute liver failure (ALF, ALI and encephalopathy), transplantation/death and hospital length-of-stay, were evaluated. RESULTS: 12/32 patients developed increased aminotransferase activity. Among the 13/32 amatoxin negative patients, 1 developed ALI and 12/13 no hepatotoxicity. Among the 19/32 amatoxin positive patients, 8/19 (42%) developed hepatotoxicity, including 5 who progressed to severe ALI, of whom 3 developed ALF (2 deaths, 1 transplantation). Urinary amatoxin and prothrombin were independent predictors of hepatotoxicity, ALT peak values (along with age) and hospital length-of-stay. In positive amatoxins patients, urinary concentrations > 55 ng/ml (or a baseline prothrombin ≤ 83%), were associated to hepatotoxicity (presented by 8/9 patients with ALT>1000 U/L). Among 5 patients with urinary amatoxin ≥ 70 ng/ml, 4 developed severe ALI. CONCLUSIONS: In patients with hepatotoxic mushroom poisoning, a negative urinary amatoxin quantification within 72h of intake ruled out the risk of hepatotoxicity in 92% of patients, whereas positive urinary amatoxins were associated with hepatotoxicity and severe ALI. Concentrations >55 ng/ml and ≥ 70 ng/ml were predictive of hepatotoxicity and severe ALI, respectively.
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Amanitinas/orina , Fallo Hepático Agudo/inducido químicamente , Intoxicación por Setas/diagnóstico , Intoxicación por Setas/orina , Adolescente , Adulto , Anciano , Niño , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Fallo Hepático Agudo/diagnóstico , Modelos Logísticos , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Estudios Retrospectivos , España , Adulto JovenRESUMEN
BACKGROUND: Acute intermittent porphyria (AIP) is a rare metabolic disorder due to a deficiency of porphobilinogen deaminase, the third enzyme of the heme biosynthetic pathway. This low enzymatic activity may predispose to the appearance of acute neurological attacks. Seminal studies suggested that AIP was associated with changes in tryptophan homeostasis with inconclusive results. Therefore, the aim of this study was to analyze the urinary metabolome of AIP patients focusing on tryptophan metabolism using state-of-the-art technology. METHODS: This was a case-control study including a group of 25 AIP patients with active biochemical disease and increased excretion of heme-precursors and 25 healthy controls. Tryptophan and related compounds and metabolites including: large neutral amino acids (LNAAs), serotonin, kynurenine, kynurenic acid and anthranilic acid were quantified in urine by liquid chromatography tandem-mass spectrometry (LC-MS/MS). Twenty-nine biological markers (including metabolic ratios and absolute concentrations) were compared between patients and controls. RESULTS: Significant differences were found in the tryptophan-kynurenine metabolic pathway. Compared to controls, AIP patients showed: (a) increased urinary excretion of kynurenine and anthranilic acid (P<0.005); (b): elevation of the kynurenine/tryptophan ratio (P<0.001) and (c): decrease of the kynurenic acid/kynurenine ratio (P=0.001). In contrast, no differences were found in the serotonin metabolic pathway independently of the markers and ratios used. CONCLUSIONS: The results of the study demonstrate that there is an imbalance in the kynurenine metabolic pathway in AIP patients, with an increase of the kynurenine/tryptophan ratio in urine and a reduction of the kynurenic acid/kynurenine ratio. The modified ratios suggest induction of indoleamine 2,3-deoxygenase and decreased activity of kynurenine aminotransferase in the liver. The results confirm that LC-MS/MS is useful for the characterization of the urinary metabolome of hepatic porphyrias.
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Metaboloma/fisiología , Metabolómica/métodos , Porfiria Intermitente Aguda/orina , Triptófano/metabolismo , Triptófano/orina , Adulto , Biomarcadores/metabolismo , Biomarcadores/orina , Estudios de Cohortes , Femenino , Humanos , Quinurenina/metabolismo , Masculino , Persona de Mediana Edad , Porfiria Intermitente Aguda/metabolismo , Serotonina/metabolismo , Adulto JovenRESUMEN
BACKGROUND: Human immunodeficiency virus (HIV) infection has been reported to be a risk factor for porphyria cutanea tarda (PCT). PURPOSE: To assess the strength of the association between HIV infection and PCT in a large hospital setting. METHODS: All patients (N = 210) diagnosed of PCT between 1990 and 2014 were retrospectively investigated for HIV infection and co-precipitating factors. High-performance liquid chromatography was used to assess the appearance of pre-PCT urinary porphyrin abnormalities among a group (N = 22) of HIV-infected patients without PCT using the urine of patients co-infected with hepatitis C virus (HCV) without PCT for comparison. RESULTS: Twenty-six HIV-infected patients (19 males and seven females) were diagnosed of PCT. During the same interval of time, ~8000 different patients infected with HIV were attended in the hospital of infectious diseases department. Examination of risk factors showed that 25 out of 26 of the PCT patients with HIV were co-infected with HCV. No chromatographic abnormalities were found in the urine of non-PCT-HIV-infected patients, whereas 39% co-infected patients showed urinary porphyrin abnormalities. CONCLUSIONS: In our large hospital series, the appearance of PCT among HIV-infected patients is low (<1%) and most present co-infection with HCV. Therefore, in most HIV-infected patients with PCT, hepatitis C and not HIV may induce uroporphyrinogen decarboxylase deficiency.
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Infecciones por VIH , VIH-1 , Porfiria Cutánea Tardía , Porfirinas/orina , Adulto , Femenino , Infecciones por VIH/complicaciones , Infecciones por VIH/orina , Humanos , Masculino , Persona de Mediana Edad , Porfiria Cutánea Tardía/etiología , Porfiria Cutánea Tardía/orinaRESUMEN
We document the presence of a previously unknown species in the urine of patients with acute intermittent porphyria (AIP). The compound was fully characterised by liquid chromatography tandem mass spectrometry. Interpretation of both full spectrum acquisition and product ion spectra acquired in positive and negative ionisation modes by quadrupole time of flight MS allowed for the identification of a condensation product arising from porphobilinogen (PBG, increased in the urine of AIP patients) and indolyl-3-acryloylglycine (IAG, derived from indolylacrylic acid and present in human urine). The structure was unequivocally confirmed through comparison between the selected reaction monitoring chromatograms obtained from the urinary species and the condensation product qualitatively synthesised in the laboratory. Owing to the large amounts of both PBG and IAG in urine of AIP patients, the possible ex vivo formation of PBG-IAG in urine samples was evaluated. The product was spontaneously formed at room temperature, at 4 °C and even during storage at -20 °C when spiking a control sample with PBG. A positive correlation was found between PBG and PBG-IAG in samples collected from AIP patients. However, no correlation was found between PBG-IAG and IAG. Purified PBG-IAG did not form the characteristic chromogen after application of p-dimethylaminobenzaldehyde in HCl, thus suggesting that the current techniques used to measure PBG in urine of AIP patients based on Ehlrich's reaction do not detect this newly characterised PBG-IAG fraction.