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Mikhailova DM, Sudnitsyna J, Kovgan P, Naida L, Kharazova A, Mindukshev I, Gambaryan S. Analysis of Ferric Protoporphyrin IX Effects on Human Platelets: Hematin Is a More Potent Agonist than Hemin. Cells 2025; 14:255. [PMID: 39996728 PMCID: PMC11853094 DOI: 10.3390/cells14040255] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/27/2024] [Revised: 01/27/2025] [Accepted: 02/06/2025] [Indexed: 02/26/2025] Open
Abstract
Hemolysis during severe diseases (malaria, hemorrhagic stroke, sickle cell disease, etc.) and blood transfusion induces the release of free hemoglobin, which degrades to highly reactive and toxic compounds-hemin and hematin. Oxidized heme derivatives induce platelet activation, aggregation, and degranulation, leading to prothrombotic and inflammatory events. In the present study, we showed that hematin is a more potent agonist of platelet activation than hemin, and using several methods, including the original laser diffraction method, flow cytometry, and confocal microscopy, we demonstrated that hematin at low doses induces platelet activation and aggregation without reducing cell viability and affecting calcium efflux. On the contrary, hematin at high concentrations triggered phosphatidylserine exposure, severe loss of platelet viability, and calcium dysregulation, which was not inhibited by cGMP/PKG and cAMP/PKA pathways. Additionally, we showed that albumin could initiate disaggregation processes in hematin-activated platelets.
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Affiliation(s)
- Diana M. Mikhailova
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (J.S.); (P.K.); (L.N.); (I.M.)
- Department of Cytology and Histology, Saint Petersburg State University, 7/9 Universitetskaya Emb., 199034 Saint Petersburg, Russia;
| | - Julia Sudnitsyna
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (J.S.); (P.K.); (L.N.); (I.M.)
| | - Polina Kovgan
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (J.S.); (P.K.); (L.N.); (I.M.)
- Institute of Biomedical Systems and Biotechnologies, Peter the Great Saint Petersburg Polytechnic University, 195251 Saint Petersburg, Russia
| | - Lidia Naida
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (J.S.); (P.K.); (L.N.); (I.M.)
- Institute of Biomedical Systems and Biotechnologies, Peter the Great Saint Petersburg Polytechnic University, 195251 Saint Petersburg, Russia
| | - Alexandra Kharazova
- Department of Cytology and Histology, Saint Petersburg State University, 7/9 Universitetskaya Emb., 199034 Saint Petersburg, Russia;
| | - Igor Mindukshev
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (J.S.); (P.K.); (L.N.); (I.M.)
| | - Stepan Gambaryan
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (J.S.); (P.K.); (L.N.); (I.M.)
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Mikhailova DM, Skverchinskaya E, Sudnitsyna J, Butov KR, Koltsova EM, Mindukshev IV, Gambaryan S. Hematin- and Hemin-Induced Spherization and Hemolysis of Human Erythrocytes Are Independent of Extracellular Calcium Concentration. Cells 2024; 13:554. [PMID: 38534398 PMCID: PMC10969559 DOI: 10.3390/cells13060554] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2024] [Revised: 03/10/2024] [Accepted: 03/15/2024] [Indexed: 03/28/2024] Open
Abstract
Pathologies such as malaria, hemorrhagic stroke, sickle cell disease, and thalassemia are characterized by the release of hemoglobin degradation products from damaged RBCs. Hematin (liganded with OH-) and hemin (liganded with Cl-)-are the oxidized forms of heme with toxic properties due to their hydrophobicity and the presence of redox-active Fe3. In the present study, using the original LaSca-TM laser particle analyzer, flow cytometry, and confocal microscopy, we showed that both hematin and hemin induce dose-dependent RBC spherization and hemolysis with ghost formation. Hematin and hemin at nanomolar concentrations increased [Ca2+]i in RBC; however, spherization and hemolysis occurred in the presence and absence of calcium, indicating that both processes are independent of [Ca2+]i. Both compounds triggered acute phosphatidylserine exposure on the membrane surface, reversible after 60 min of incubation. A comparison of hematin and hemin effects on RBCs revealed that hematin is a more reactive toxic metabolite than hemin towards human RBCs. The toxic effects of heme derivatives were reduced and even reversed in the presence of albumin, indicating the presence in RBCs of the own recovery system against the toxic effects of heme derivatives.
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Affiliation(s)
- Diana M. Mikhailova
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (E.S.); (J.S.); (I.V.M.)
- Department of Cytology and Histology, Saint Petersburg State University, 7/9 Universitetskaya Emb., 199034 Saint Petersburg, Russia
| | - Elisaveta Skverchinskaya
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (E.S.); (J.S.); (I.V.M.)
| | - Julia Sudnitsyna
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (E.S.); (J.S.); (I.V.M.)
| | - Kirill R. Butov
- Department of Molecular Biology and Medical Biotechnology, Pirogov Russian National Research Medical University, 117997 Moscow, Russia;
- Dmitry Rogachev National Medical Research Center of Pediatric Hematology, Oncology and Immunology, 117997 Moscow, Russia;
| | - Ekaterina M. Koltsova
- Dmitry Rogachev National Medical Research Center of Pediatric Hematology, Oncology and Immunology, 117997 Moscow, Russia;
- Center for Theoretical Problems of Physicochemical Pharmacology, Russian Academy of Sciences, 30 Srednyaya Kalitnikovskaya st., 109029 Moscow, Russia
| | - Igor V. Mindukshev
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (E.S.); (J.S.); (I.V.M.)
| | - Stepan Gambaryan
- Sechenov Institute of Evolutionary Physiology and Biochemistry, Russian Academy of Sciences, 44 Thorez Ave., 194223 Saint Petersburg, Russia; (D.M.M.); (E.S.); (J.S.); (I.V.M.)
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Lin J, Liu J, Wang A, Si Z. A case report of acute intermittent porphyria leading to severe disability. Front Neurol 2024; 14:1334743. [PMID: 38274883 PMCID: PMC10808997 DOI: 10.3389/fneur.2023.1334743] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2023] [Accepted: 12/19/2023] [Indexed: 01/27/2024] Open
Abstract
Acute intermittent porphyria (AIP) is a rare inherited metabolic disorder resulting from increased production of porphyrins and their precursors, δ-aminolevulinic acid (ALA) and porphobilinogen (PBG), due to deficiencies in the enzymatic activity of the heme synthesis pathway. The disease is typically characterized by a triad of abdominal pain, neurologic impairment symptoms, and psychiatric abnormalities. However, only a small percentage of patients present with this classic triad of symptoms. Our female patient, aged 23, was admitted to the hospital with a 4-year history of abnormal mood episodes and weakness in the limbs for over 1 week. She had a previous medical history of intestinal obstruction. After admission, a cranial MRI revealed reversible posterior leukoencephalopathy imaging manifestations, and the patient exhibited weakness of the extremities, respiratory failure, seizures, and severely reduced serum sodium concentration. The diagnosis of AIP was ultimately confirmed by a positive urine PBG-sunlight test and analysis of HMBS gene variants. The absence of typical triadic signs in acute attacks of AIP can make early recognition of the disease challenging. We present a case with multiple typical clinical manifestations of AIP in the hope of aiding clinicians in fully recognizing acute intermittent porphyria.
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Affiliation(s)
- Jie Lin
- Department of Neurology, Shandong Provincial Qianfoshan Hospital, Shandong University, Shandong Institute of Neuroimmunology, Shandong Key Laboratory of Rheumatic Disease and Translational Medicine, Shandong, China
| | - Jinzhi Liu
- Department of Neurology, The First Affiliated Hospital of Shandong First Medical University & Shandong Provincial Qianfoshan Hospital, Shandong Institute of Neuroimmunology, Shandong Key Laboratory of Rheumatic Disease and Translational Medicine, Shandong, China
| | - Aihua Wang
- Department of Neurology, The First Affiliated Hospital of Shandong First Medical University & Shandong Provincial Qianfoshan Hospital, Shandong Institute of Neuroimmunology, Shandong Key Laboratory of Rheumatic Disease and Translational Medicine, Shandong, China
| | - Zhihua Si
- Department of Neurology, The First Affiliated Hospital of Shandong First Medical University & Shandong Provincial Qianfoshan Hospital, Shandong Institute of Neuroimmunology, Shandong Key Laboratory of Rheumatic Disease and Translational Medicine, Shandong, China
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Brito Avô L, Pereira L, Oliveira A, Ferreira F, Filipe P, Coelho Rodrigues I, Couto E, Ferreira F, Airosa Pardal A, Morgado P, Moreira S. Portuguese Consensus on Acute Porphyrias: Diagnosis, Treatment, Monitoring and Patient Referral. ACTA MEDICA PORT 2023; 36:753-764. [PMID: 37924314 DOI: 10.20344/amp.20323] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2023] [Accepted: 09/11/2023] [Indexed: 11/06/2023]
Abstract
Acute porphyrias are a group of rare genetic metabolic disorders, caused by a defect in one of the enzymes involved in the heme biosynthesis, which results in an abnormally high accumulation of toxic intermediates. Acute porphyrias are characterized by potentially life-threatening attacks and, for some patients, by chronic manifestations that negatively impact daily functioning and quality of life. Clinical manifestations include a nonspecific set of gastrointestinal, neuropsychiatric, and/or cutaneous symptoms. Effective diagnostic methods are widely available, but due to their clinical heterogeneity and non-specificity, many years often elapse from symptom onset to diagnosis of acute porphyrias, delaying the treatment and increasing morbidity. Therefore, increased awareness of acute porphyrias among healthcare professionals is paramount to reducing disease burden. Treatment of acute porphyrias is centered on eliminating the potential precipitants, symptomatic treatment, and suppressing the hepatic heme pathway, through the administration of hemin or givosiran. Moreover, properly monitoring patients with acute porphyrias and their relatives is fundamental to preventing acute attacks, hospitalization, and long-term complications. Considering this, a multidisciplinary panel elaborated a consensus paper, aiming to provide guidance for an efficient and timely diagnosis of acute porphyrias, and evidence-based recommendations for treating and monitoring patients and their families in Portugal. To this end, all authors exhaustively reviewed and discussed the current scientific evidence on acute porphyrias available in the literature, between November 2022 and May 2023.
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Affiliation(s)
- Luís Brito Avô
- Serviço de Medicina Interna. Hospital CUF Tejo. Lisboa; Unidade de Doenças Raras. Hospital CUF Tejo. Lisboa; Nucleo de Estudos de Doenças Raras da Sociedade Portuguesa de Medicina Interna. Lisboa. Portugal
| | - Luísa Pereira
- Nucleo de Estudos de Doenças Raras da Sociedade Portuguesa de Medicina Interna. Lisboa; Unidade de Cuidados Paliativos. Hospital CUF Tejo. Lisboa. Portugal
| | - Anabela Oliveira
- Serviço de Medicina I. Centro de Referência de Doenças Hereditárias do Metabolismo. Centro Hospitalar Universitário Lisboa Norte. Lisboa. Portugal
| | - Filipa Ferreira
- Unidade de Rastreio Neonatal, Metabolismo e Genética. Departamento de Genética Humana. Instituto Nacional de Saúde Doutor Ricardo Jorge. Porto. Portugal
| | - Paulo Filipe
- Unidade de Investigação de Dermatologia. Instituto de Medicina Molecular João Lobo Antunes. Lisboa. Portugal
| | - Inês Coelho Rodrigues
- Serviço de Gastrenterologia. Centro Hospitalar e Universitário de Lisboa Norte. Lisboa. Portugal
| | - Eduarda Couto
- Departamento de Medicina Interna. Serviço de Hematologia Clínica. Centro Hospitalar Póvoa de Varzim - Vila do Conde. Póvoa de Varzim. Portugal
| | - Fátima Ferreira
- Serviço de Hematologia Clínica. Centro Hospitalar e Universitário de São João. Porto. Portugal
| | - André Airosa Pardal
- Serviço de Hematologia Clínica. Centro Hospitalar Universitário de São João. Porto. Portugal
| | - Pedro Morgado
- Instituto de Investigação em Ciências da Vida e Saúde. Escola de Medicina. Universidade do Minho. Braga; Laboratório Associado do Governo Português ICVS/3B's. Braga/Guimarães; Serviço de Psiquiatria. Hospital de Braga. Braga. Portugal
| | - Sónia Moreira
- Nucleo de Estudos de Doenças Raras da Sociedade Portuguesa de Medicina Interna. Lisboa; Serviço de Medicina Interna. Centro de Referência de Doenças Hereditárias do Metabolismo. Centro Hospitalar e Universitário de Coimbra. Coimbra; Faculdade de Medicina. Universidade de Coimbra. Coimbra. . Portugal
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Zübarioğlu T, Kıykım E, Aktuğlu-Zeybek Ç, Aktuglu Zeybek C. An Overview of Acute Hepatic Porphyrias: Clinical Implications, Diagnostic Approaches, and Management Strategies. Turk Arch Pediatr 2023; 58:3-9. [PMID: 36598205 PMCID: PMC9885782 DOI: 10.5152/turkarchpediatr.2022.22301] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/03/2023]
Abstract
Porphyrias are inborn errors of heme biosynthesis pathway that result in neurovisceral and/ or cutaneous manifestations which occur with episodic attacks, usually accompanied by a multisystemic involvement. Acute hepatic porphyrias include acute intermittent porphyria, variegate porphyria, hereditary coproporphyria, and aminolevulinic acid dehydratase deficiency porphyria. Acute hepatic porphyrias may present with symptoms of an affected central, peripheral, and autonomic nervous system and are generally diagnosed in time of an acute neurovisceral attack. In children, clinical picture is more complicated and presents with neurological findings predominantly. First-line investigation should be the urinary porphobilinogen and aminolevulinic acid performance when acute hepatic porphyria is clinically suspected. Comprehensive testing including urine porphyrin separation, fluorescence scanning of diluted plasma at neutral pH, evaluation of fecal porphyrins, and measurement of erythrocyte porphobilinogen deaminase activity is indicated for confirmation or exclusion of the porphyria and define the type of acute hepatic porphyrias. The main aim of the treatment is to decrease aminolevulinic acid, porphobilinogen, and porphyrins by reducing hepatic ALAS1 activity. The first measure should always be the avoidance of any porphyrinogenic drugs. Hemin therapy should not be delayed in the treatment of a severe acute attack. Gonadotropin-releasing hormone analogs and prophylactic hemin protocols can be used for selected cases with more than 4 attacks per year. Givosiran is a promising treatment option for severe cases.
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Recent Insights into the Pathogenesis of Acute Porphyria Attacks and Increasing Hepatic PBGD as an Etiological Treatment. LIFE (BASEL, SWITZERLAND) 2022; 12:life12111858. [PMID: 36430993 PMCID: PMC9694773 DOI: 10.3390/life12111858] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 10/20/2022] [Revised: 11/03/2022] [Accepted: 11/09/2022] [Indexed: 11/16/2022]
Abstract
Rare diseases, especially monogenic diseases, which usually affect a single target protein, have attracted growing interest in drug research by encouraging pharmaceutical companies to design and develop therapeutic products to be tested in the clinical arena. Acute intermittent porphyria (AIP) is one of these rare diseases. AIP is characterized by haploinsufficiency in the third enzyme of the heme biosynthesis pathway. Identification of the liver as the target organ and a detailed molecular characterization have enabled the development and approval of several therapies to manage this disease, such as glucose infusions, heme replenishment, and, more recently, an siRNA strategy that aims to down-regulate the key limiting enzyme of heme synthesis. Given the involvement of hepatic hemoproteins in essential metabolic functions, important questions regarding energy supply, antioxidant and detoxifying responses, and glucose homeostasis remain to be elucidated. This review reports recent insights into the pathogenesis of acute attacks and provides an update on emerging treatments aimed at increasing the activity of the deficient enzyme in the liver and restoring the physiological regulation of the pathway. While further studies are needed to optimize gene therapy vectors or large-scale production of liver-targeted PBGD proteins, effective protection of PBGD mRNA against the acute attacks has already been successfully confirmed in mice and large animals, and mRNA transfer technology is being tested in several clinical trials for metabolic diseases.
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Haverkamp T, Bronisch O, Knösel T, Mogler C, Weichert W, Stauch T, Schmid C, Rummeny C, Beykirch MK, Petrides PE. Heterogeneous molecular behavior in liver tumors (HCC and CCA) of two patients with acute intermittent porphyria. J Cancer Res Clin Oncol 2022; 149:2647-2655. [PMID: 36245063 DOI: 10.1007/s00432-022-04384-5] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/02/2022] [Accepted: 09/26/2022] [Indexed: 11/25/2022]
Abstract
INTRODUCTION Acute intermittent porphyria (AIP) is a very rare (orphan) metabolic disorder of porphyrin biosynthesis which is characterized by elevated plasma and urine levels of 5-aminolevulinic acid (5-ALA) and porphobilinogen (PBG). Patients with this disorder which is caused by a germline mutation of the hydroxymethylbilan-synthase (HMBS)-gene have a high risk of primary liver cancer which may be determined by disease activity. The exact mechanism of carcinogenesis of this rare tumor is unknown, however. MATERIALS AND METHODS We analyzed paraffin-embedded formalin-fixed liver tumor and normal liver specimens of two female AIP patients treated at the Munich EPNET center. One patient had developed hepatocellular carcinoma (HCC), the other intrahepatic cholangiocarcinoma (CCA). Since biallelic inactivation of HMBS had been observed in one study, we used Sanger and next-generation sequencing with a 8 gene porphyria panel plus 6 potential modifier loci to search for mutations in DNA extractions. RESULTS In the patient with the HCC, we found a second inactivating mutation in the HMBS gene in the tumor but not in the adjacent normal liver tissue. No mutation could be found in the liver tissues of the patient with CCA, however. CONCLUSIONS Biallelic inactivation of HMBS or protoporphyrinogen-oxidase (PPOX), another enzyme of porphyrin biosynthesis, has been observed in patients with acute porphyrias and liver tumors. We could confirm this in our patient with HCC with a mutation in HMBS but not in the one with CCA. Since 5-ALA can be converted into carcinogenic substances such as 4,5-dioxovaleric acid (DOVA) or 3,6-dihydropyrazine-2,5-dipropanoic acid (= cyclic dimerization product of 5-ALA), local production of these metabolites in hepatic areas with complete loss of HMBS activity may contribute to liver carcinogenesis.
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Affiliation(s)
- Thomas Haverkamp
- Molecular Genetics Laboratory, MVZ Dr.Eberhard, Brauhausstr.4, 44137, Dortmund, Germany
| | - Olivia Bronisch
- Hematology Oncology Center, EPNET Clinical Center Munich, Ludwig Maximilians University (LMU) Munich, Zweibrückenstr.2, 80331, Munich, Germany
| | - Thomas Knösel
- Institute of Pathology, Ludwig Maximilians University Munich (LMU), Thalkirchner Str.36, 80337, Munich, Germany
| | - Carolin Mogler
- Institute of Pathology, Klinikum Rechts Der Isar (RDI), Technical University of Munich, Trogerstr.36, 80337, Munich, Germany
| | - Wilko Weichert
- Institute of Pathology, Klinikum Rechts Der Isar (RDI), Technical University of Munich, Trogerstr.36, 80337, Munich, Germany
| | - Thomas Stauch
- EPNET-Porphyria Specialist Laboratory MVZ PD Dr, Volkmann Kriegsstraße 99, 76133, Karlsruhe, Germany
| | - Claudia Schmid
- Institute of Radiology Dachau, Frühlingstr.33-34, 85221, Dachau, Germany
| | - Claudia Rummeny
- Institute of Radiology Munich East, Wasserburger Landstr.274-276, 81827, Munich, Germany
| | - Maria K Beykirch
- Hematology Oncology Center, EPNET Clinical Center Munich, Ludwig Maximilians University (LMU) Munich, Zweibrückenstr.2, 80331, Munich, Germany
| | - Petro E Petrides
- Hematology Oncology Center, EPNET Clinical Center Munich, Ludwig Maximilians University (LMU) Munich, Zweibrückenstr.2, 80331, Munich, Germany.
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