RESUMEN
Vaccination is crucial in combatting the severe acute respiratory syndrome coronavirus 2 pandemic. The rare complication of thrombocytopenia and thrombotic complications at unusual sites after ChAdOx1 nCov-19 vaccination is caused by platelet-activating antibodies directed against platelet factor 4 (PF4). We present a widely applicable whole-blood standard flow cytometric assay to identify the pathogenic antibodies associated with vaccine-induced immune-mediated thrombotic thrombocytopenia (VITT) after ChAdOx1 nCov-19 vaccination. This assay will enable rapid diagnosis by many laboratories. This trial was registered at www.clinicaltrials.gov as #NCT04370119.
Asunto(s)
Autoanticuerpos/sangre , Vacunas contra la COVID-19/efectos adversos , COVID-19/prevención & control , Citometría de Flujo/métodos , Inmunoglobulina G/sangre , Activación Plaquetaria/inmunología , Factor Plaquetario 4/inmunología , Púrpura Trombocitopénica Idiopática/diagnóstico , Receptores de IgG/inmunología , SARS-CoV-2 , Vacunación/efectos adversos , Especificidad de Anticuerpos , Autoanticuerpos/biosíntesis , Autoanticuerpos/inmunología , Vacunas contra la COVID-19/inmunología , ChAdOx1 nCoV-19 , Heparina/efectos adversos , Heparina/inmunología , Humanos , Técnicas para Inmunoenzimas , Inmunogenicidad Vacunal , Inmunoglobulina G/biosíntesis , Inmunoglobulina G/inmunología , Selectina-P/análisis , Púrpura Trombocitopénica Idiopática/etiología , Púrpura Trombocitopénica Idiopática/inmunologíaAsunto(s)
Enfermedades Autoinmunes , Factor Plaquetario 4 , Trombocitopenia , Trombosis , Humanos , Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/inmunología , Heparina/efectos adversos , Factor Plaquetario 4/inmunología , Trombocitopenia/inmunología , Enfermedad Crónica , Trombosis/inmunologíaRESUMEN
Genetic variants in growth factor-independent 1B (GFI1B), encoding transcription factor GFI1B, are causative of platelet-type bleeding disorder-17. Presently, 53 cases of GFI1B associated inherited thrombocytopenia (IT) have been published, however only three were homozygous. The bleeding- and platelet phenotypes of these patients depend on location and inheritance pattern of the GFI1B variant. We report a novel homozygous GFI1B (Thr174Ile) variant located in the first Zinc finger domain of GFI1B in two sisters of Palestinian ancestry born to consanguineous parents. They experienced severe bleeding tendency at moderately reduced platelet counts. Flow cytometry and immunofluorescent microscopy confirmed the diagnostic features of GFI1B associated IT: a reduced content of alpha granules and aberrant expression of the stem cell marker CD34 on platelets. Transcription factor GFI1B is differentially expressed during hemato- and lymphopoiesis. In addition, to platelet function investigations, we present results of lymphoid subgroup analyses and deformability of red cells measured by ektacytometry.
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Hemorragia/fisiopatología , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Trombocitopenia/fisiopatología , Adulto , Femenino , Homocigoto , Humanos , Persona de Mediana Edad , MutaciónRESUMEN
Patients with inherited thrombocytopenias often require platelet transfusions to raise their platelet count before surgery or other invasive procedures; moreover, subjects with clinically significant spontaneous bleeding may benefit from an enduring improvement of thrombocytopenia. The hypothesis that thrombopoietin-mimetics can increase platelet count in inherited thrombocytopenias is appealing, but evidence is scarce. We conducted a prospective, phase II clinical trial to investigate the efficacy of the oral thrombopoietin-mimetic eltrombopag in different forms of inherited thrombocytopenia. We enrolled 24 patients affected by MYH9-related disease, ANKRD26-related thrombocytopenia, X-linked thrombocytopenia/ Wiskott-Aldrich syndrome, monoallelic Bernard-Soulier syndrome, or ITGB3-related thrombocytopenia. The average pre-treatment platelet count was 40.4 ×109/L. Patients received a 3- to 6-week course of eltrombopag in a dose-escalated manner. Of 23 patients evaluable for response, 11 (47.8%) achieved a major response (platelet count >100 ×109/L), ten (43.5%) had a minor response (platelet count at least twice the baseline value), and two patients (8.7%) did not respond. The average increase of platelet count compared to baseline was 64.5 ×109/L (P<0.001). Four patients with clinically significant spontaneous bleeding entered a program of long-term eltrombopag administration (16 additional weeks): all of them obtained remission of mucosal hemorrhages, with the remission persisting throughout the treatment period. Treatment was globally well tolerated: five patients reported mild adverse events and one patient a moderate adverse event. In conclusion, eltrombopag was safe and effective in increasing platelet count and reducing bleeding symptoms in different forms of inherited thrombocytopenia. Despite these encouraging results, caution is recommended when using thrombopoietinmimetics in inherited thrombocytopenias predisposing to leukemia. ClinicalTrials.gov identifier: NCT02422394.
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Hidrazinas , Trombocitopenia , Benzoatos/efectos adversos , Humanos , Hidrazinas/efectos adversos , Estudios Prospectivos , Pirazoles , Trombocitopenia/tratamiento farmacológicoRESUMEN
Major surgery is associated with an increased risk of venous thromboembolism (VTE), thus the application of mechanical or pharmacologic prophylaxis is recommended. The incidence of VTE in patients with inherited platelet disorders (IPD) undergoing surgical procedures is unknown and no information on the current use and safety of thromboprophylaxis, particularly of low-molecular-weight-heparin in these patients is available. Here we explored the approach to thromboprophylaxis and thrombotic outcomes in IPD patients undergoing surgery at VTE-risk participating in the multicenter SPATA study. We evaluated 210 surgical procedures carried out in 155 patients with well-defined forms of IPD (VTE-risk: 31% high, 28.6% intermediate, 25.2% low, 15.2% very low). The use of thromboprophylaxis was low (23.3% of procedures), with higher prevalence in orthopedic and gynecological surgeries, and was related to VTE-risk. The most frequently employed thromboprophylaxis was mechanical and appeared to be effective, as no patients developed thrombosis, including patients belonging to the highest VTE-risk classes. Low-molecular-weight-heparin use was low (10.5%) and it did not influence the incidence of post-surgical bleeding or of antihemorrhagic prohemostatic interventions use. Two thromboembolic events were registered, both occurring after high VTE-risk procedures in patients who did not receive thromboprophylaxis (4.7%). Our findings suggest that VTE incidence is low in patients with IPD undergoing surgery at VTE-risk and that it is predicted by the Caprini score. Mechanical thromboprophylaxis may be of benefit in patients with IPD undergoing invasive procedures at VTE-risk and low-molecular-weight-heparin should be considered for major surgery.
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Trombosis , Tromboembolia Venosa , Anticoagulantes , Fibrinolíticos/uso terapéutico , Heparina de Bajo-Peso-Molecular/uso terapéutico , Humanos , Complicaciones Posoperatorias/epidemiología , Complicaciones Posoperatorias/etiología , Complicaciones Posoperatorias/prevención & control , Trombosis/epidemiología , Trombosis/etiología , Trombosis/prevención & control , Tromboembolia Venosa/epidemiología , Tromboembolia Venosa/etiología , Tromboembolia Venosa/prevención & controlRESUMEN
MYH9-related disease (MYH9-RD) is a rare, autosomal dominant disorder caused by mutations in MYH9, the gene encoding the actin-activated motor protein non-muscle myosin IIA (NMIIA). MYH9-RD patients suffer from bleeding syndromes, progressive kidney disease, deafness, and/or cataracts, but the impact of MYH9 mutations on other NMIIA-expressing tissues remains unknown. In human red blood cells (RBCs), NMIIA assembles into bipolar filaments and binds to actin filaments (F-actin) in the spectrin-F-actin membrane skeleton to control RBC biconcave disk shape and deformability. Here, we tested the effects of MYH9 mutations in different NMIIA domains (motor, coiled-coil rod, or non-helical tail) on RBC NMIIA function. We found that MYH9-RD does not cause clinically significant anemia and that patient RBCs have normal osmotic deformability as well as normal membrane skeleton composition and micron-scale distribution. However, analysis of complete blood count data and peripheral blood smears revealed reduced hemoglobin content and elongated shapes, respectively, of MYH9-RD RBCs. Patients with mutations in the NMIIA motor domain had the highest numbers of elongated RBCs. Patients with mutations in the motor domain also had elevated association of NMIIA with F-actin at the RBC membrane. Our findings support a central role for motor domain activity in NMIIA regulation of RBC shape and define a new sub-clinical phenotype of MYH9-RD.
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Actinas , Membrana Eritrocítica , Eritrocitos Anormales , Pérdida Auditiva Sensorineural , Mutación , Cadenas Pesadas de Miosina , Trombocitopenia/congénito , Actinas/genética , Actinas/metabolismo , Membrana Eritrocítica/genética , Membrana Eritrocítica/metabolismo , Membrana Eritrocítica/patología , Eritrocitos Anormales/metabolismo , Eritrocitos Anormales/patología , Femenino , Pérdida Auditiva Sensorineural/genética , Pérdida Auditiva Sensorineural/metabolismo , Pérdida Auditiva Sensorineural/patología , Humanos , Masculino , Cadenas Pesadas de Miosina/genética , Cadenas Pesadas de Miosina/metabolismo , Trombocitopenia/genética , Trombocitopenia/metabolismo , Trombocitopenia/patologíaRESUMEN
MYH9-related disease (MYH9-RD) is a rare autosomal-dominant disorder caused by mutations in the gene for nonmuscle myosin heavy chain IIA (NMMHC-IIA). MYH9-RD is characterized by a considerable variability in clinical evolution: patients present at birth with only thrombocytopenia, but some of them subsequently develop sensorineural deafness, cataract, and/or nephropathy often leading to end-stage renal disease (ESRD). We searched for genotype-phenotype correlations in the largest series of consecutive MYH9-RD patients collected so far (255 cases from 121 families). Association of genotypes with noncongenital features was assessed by a generalized linear regression model. The analysis defined disease evolution associated to seven different MYH9 genotypes that are responsible for 85% of MYH9-RD cases. Mutations hitting residue R702 demonstrated a complete penetrance for early-onset ESRD and deafness. The p.D1424H substitution associated with high risk of developing all the noncongenital manifestations of disease. Mutations hitting a distinct hydrophobic seam in the NMMHC-IIA head domain or substitutions at R1165 associated with high risk of deafness but low risk of nephropathy or cataract. Patients with p.E1841K, p.D1424N, and C-terminal deletions had low risk of noncongenital defects. These findings are essential to patients' clinical management and genetic counseling and are discussed in view of molecular pathogenesis of MYH9-RD.
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Catarata/genética , Estudios de Asociación Genética , Pérdida Auditiva Sensorineural/genética , Proteínas Motoras Moleculares/genética , Cadenas Pesadas de Miosina/genética , Trombocitopenia/congénito , Adulto , Edad de Inicio , Sustitución de Aminoácidos , Femenino , Genotipo , Pérdida Auditiva Sensorineural/complicaciones , Pérdida Auditiva Sensorineural/diagnóstico , Humanos , Italia , Modelos Lineales , Masculino , Mutación , Fenotipo , Factores de Riesgo , Trombocitopenia/complicaciones , Trombocitopenia/diagnóstico , Trombocitopenia/genéticaAsunto(s)
Benzoatos/uso terapéutico , Pérdida de Sangre Quirúrgica/prevención & control , Hidrazinas/uso terapéutico , Cadenas Pesadas de Miosina/genética , Hemorragia Posoperatoria/prevención & control , Pirazoles/uso terapéutico , Trombocitopenia/cirugía , Femenino , Humanos , Masculino , Trombocitopenia/genética , Resultado del TratamientoRESUMEN
BACKGROUND: The transcription factor GATA1 is an essential regulator of erythroid cell gene expression and maturation and is also relevant for platelet biogenesis. GATA1-related thrombocytopenia (GATA1-RT) is a rare X-linked inherited platelet disorder (IPD) characterized by macrothrombocytopenia and dyserythropoiesis. Enlarged platelet size, reduced platelet granularity, and noticeable red blood cell anisopoikilocytosis are characteristic but unspecific morphological findings in GATA1-RT. OBJECTIVES: To expand the investigation of platelet phenotype of patients with GATA1-RT by light- and immunofluorescence microscopy on a blood smear. METHODS: We assessed blood smears by light- and immunofluorescence microscopy after May-Grünwald Giemsa staining using a set of 13 primary antibodies against markers belonging to different platelet structures. Antibody binding was visualized by fluorescently labeled secondary antibodies. RESULTS: We investigated 12 individuals with genetically confirmed GATA1-RT from 8 unrelated families. While confirming the already known characteristic of platelet morphology (platelet macrocytosis and reduced expression of markers for α-granules), we also found aggregates of nonmuscular myosin heavy chain II A (NMMIIA) in the erythrocytes in all individuals (1-3 aggregates/cell, 1-3 µm diameter). By systematically reanalyzing blood smears from a cohort of patients with 19 different forms of IPD, we found similar NMMIIA aggregates in the red blood cells only in subjects with GFI1B-related thrombocytopenia (GFI1B-RT), the other major IPD featured by dyserythropoiesis. CONCLUSION: Aggregates of NMMIIA in the erythrocytes associate with GATA1-RT and GFI1B-RT and can facilitate their diagnosis on blood smears. This previously unreported finding might represent a novel marker of dyserythropoiesis assessable in peripheral blood.
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Anemia , Factor de Transcripción GATA1 , Miosina Tipo IIA no Muscular , Proteínas Proto-Oncogénicas , Proteínas Represoras , Trombocitopenia , Humanos , Plaquetas/metabolismo , Eritrocitos , Factor de Transcripción GATA1/genética , Factor de Transcripción GATA1/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genéticaRESUMEN
BACKGROUND: Inherited platelet disorders (IPDs) are rare diseases characterized by reduced blood platelet counts and/or impaired platelet function. Recognizing IPDs is advisable but often challenging. The diagnostic tools include clinical evaluation, platelet function tests, and molecular analyses. Demonstration of a pathogenic genetic variant confirms IPDs. We established a method to assess the platelet phenotype on blood smears using immunofluorescence microscopy as a diagnostic tool for IPDs. OBJECTIVES: The aim of the present study was to validate immunofluorescence microscopy as a screening tool for IPDs in comparison with genetic screening. METHODS: We performed a blinded comparison between the diagnosis made using immunofluorescence microscopy on blood smears and genetic findings in a cohort of 43 families affected with 20 different genetically confirmed IPDs. In total, 76% of the cases had inherited thrombocytopenia. RESULTS: Immunofluorescence correctly predicted the underlying IPD in the vast majority of patients with 1 of 9 IPDs for which the typical morphologic pattern is known. Thirty of the 43 enrolled families (70%) were affected by 1 of these 9 IPDs. For the other 11 forms of IPD, we describe alterations of platelet structure in 9 disorders and normal findings in 2 disorders. CONCLUSION: Immunofluorescence microscopy on blood smears is an effective screening tool for 9 forms of IPD, which include the most frequent forms of inherited thrombocytopenia. Using this approach, typical changes in the phenotype may also be identified for other rare IPDs.
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Trastornos de las Plaquetas Sanguíneas , Trombocitopenia , Humanos , Trastornos de las Plaquetas Sanguíneas/diagnóstico , Trastornos de las Plaquetas Sanguíneas/genética , Plaquetas/patología , Trombocitopenia/patología , Pruebas de Función Plaquetaria , Técnica del Anticuerpo FluorescenteRESUMEN
BACKGROUND: Germline mutations in RUNX1 can cause a familial platelet disorder that may lead to acute myeloid leukemia, an autosomal dominant disorder characterized by moderate thrombocytopenia, platelet dysfunction, and a high risk of developing acute myeloid leukemia or myelodysplastic syndrome. Discerning the pathogenicity of novel RUNX1 variants is critical for patient management. OBJECTIVES: To extend the characterization of RUNX1 variants and evaluate their effects by transcriptome analysis. METHODS: Three unrelated patients with long-standing thrombocytopenia carrying heterozygous RUNX1 variants were included: P1, who is a subject with recent development of myelodysplastic syndrome, with c.802 C>T[p.Gln268∗] de novo; P2 with c.586A>G[p.Thr196Ala], a variant that segregates with thrombocytopenia and myeloid neoplasia in the family; and P3 with c.476A>G[p.Asn159Ser], which did not segregate with thrombocytopenia or neoplasia. Baseline platelet evaluations were performed. Ultrapure platelets were prepared for platelet transcriptome analysis. RESULTS: In P1 and P2, but not in P3, transcriptome analysis confirmed aberrant expression of genes recognized as RUNX1 targets. Data allowed grouping patients by distinct gene expression profiles, which were partitioned with clinical parameters. Functional studies and platelet mRNA expression identified alterations in the actin cytoskeleton, downregulation of GFI1B, defective GPVI downstream signaling, and reduction of alpha granule proteins, such as thrombospondin-1, as features likely implicated in thrombocytopenia and platelet dysfunction. CONCLUSION: Platelet phenotype, familial segregation, and platelet transcriptomics support the pathogenicity of RUNX1 variants p.Gln268∗ and p.Thr196Ala, but not p.Asn159Ser. This study is an additional proof of concept that platelet RNA analysis could be a tool to help classify pathogenic RUNX1 variants and identify novel RUNX1 targets.
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Trastornos de las Plaquetas Sanguíneas , Leucemia Mieloide Aguda , Síndromes Mielodisplásicos , Trombocitopenia , Humanos , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Mutación de Línea Germinal , Trastornos de las Plaquetas Sanguíneas/complicaciones , Trombocitopenia/genética , Trombocitopenia/complicaciones , Leucemia Mieloide Aguda/genética , Perfilación de la Expresión Génica , Células Germinativas/metabolismo , MutaciónAsunto(s)
Hematopoyesis Extramedular/genética , Mediastino/patología , Proteínas Motoras Moleculares/genética , Cadenas Pesadas de Miosina/genética , Trombocitopenia/genética , Trombocitopenia/patología , Biopsia , Médula Ósea/patología , Femenino , Humanos , Mediastino/diagnóstico por imagen , Persona de Mediana Edad , Tomografía Computarizada por Tomografía de Emisión de Positrones , Radiografía Torácica , Trombocitopenia/diagnósticoRESUMEN
MYH9-related disease patients with mutations in the contractile protein nonmuscle myosin heavy chain IIA display, among others, macrothrombocytopenia and a mild-to-moderate bleeding tendency. In this study, we used three mouse lines, each with one point mutation in the Myh9 gene at positions 702, 1424, or 1841, to investigate mechanisms underlying the increased bleeding risk. Agonist-induced activation of Myh9 mutant platelets was comparable to controls. However, myosin light chain phosphorylation after activation was reduced in mutant platelets, which displayed altered biophysical characteristics and generated lower adhesion, interaction, and traction forces. Treatment with tranexamic acid restored clot retraction in the presence of tPA and reduced bleeding. We verified our findings from the mutant mice with platelets from patients with the respective mutation. These data suggest that reduced platelet forces lead to an increased bleeding tendency in patients with MYH9-related disease, and treatment with tranexamic acid can improve the hemostatic function.
RESUMEN
The GATA1 transcription factor is essential for normal erythropoiesis and megakaryocytic differentiation. Germline GATA1 pathogenic variants in the N-terminal zinc finger (N-ZF) are typically associated with X-linked thrombocytopenia, platelet dysfunction, and dyserythropoietic anemia. A few variants in the C-terminal ZF (C-ZF) domain are described with normal platelet count but altered platelet function as the main characteristic. Independently performed molecular genetic analysis identified a novel hemizygous variant (c.865C>T, p.H289Y) in the C-ZF region of GATA1 in a German patient and in a Spanish patient. We characterized the bleeding and platelet phenotype of these patients and compared these findings with the parameters of two German siblings carrying the likely pathogenic variant p.D218N in the GATA1 N-ZF domain. The main difference was profound thrombocytopenia in the brothers carrying the p.D218N variant compared to a normal platelet count in patients carrying the p.H289Y variant; only the Spanish patient occasionally developed mild thrombocytopenia. A functional platelet defect affecting αIIbß3 integrin activation and α-granule secretion was present in all patients. Additionally, mild anemia, anisocytosis, and poikilocytosis were observed in the patients with the C-ZF variant. Our data support the concept that GATA1 variants located in the different ZF regions can lead to clinically diverse manifestations.
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Anemia Diseritropoyética Congénita , Factor de Transcripción GATA1 , Enfermedades Genéticas Ligadas al Cromosoma X , Variación Genética , Trombocitopenia , Dedos de Zinc , Humanos , Masculino , Factor de Transcripción GATA1/genética , Factor de Transcripción GATA1/metabolismo , Integrinas/metabolismo , Fenotipo , Trombocitopenia/genética , Dedos de Zinc/genética , Enfermedades Genéticas Ligadas al Cromosoma X/genética , Anemia Diseritropoyética Congénita/genética , Plaquetas/patologíaRESUMEN
Src-related thrombocytopenia (SRC-RT) is a rare autosomal dominant, inherited platelet disorder resulting from the p.E527K heterozygous germline gain-of-function variant of Src. To date, genetic diagnosis of the disease has only been reported in 7 patients from 3 unrelated families. The clinical features ranged from isolated thrombocytopenia to complex syndromic manifestations characterized by thrombocytopenia, bleeding, myelofibrosis, splenomegaly, and bone disease. We report a new 3-generation kindred with the Src p.E527K variant. Patients presented with rather variable platelet counts (38-139 × 109/L), mildly impaired platelet function, >15% immature platelet fraction, and with a significant proportion of large-giant platelets. Four adults from the family were diagnosed with immune thrombocytopenia (ITP) and underwent splenectomy, achieving sustained platelet counts >75 × 109/L for several years; increases in platelet counts were also observed after corticosteroid therapy. Four of 7 Src p.E527K variant carriers showed immune defects and recurrent infections. In addition, a range of neurological symptoms, from specific language impairment to epilepsy, was seen in some family members. Patient platelets exhibited constitutive Src, Bruton tyrosine kinase, and phospholipase Cγ2 activation, and after stimulating CD19 cells by crosslinking surface immunoglobulin M, phosphorylated extracellular signal-regulated kinase (ERK) was significantly increased in B cells from individuals carrying the Src p.E527K substitution. In summary, in addition to causing impaired platelet production, SRC-RT may associate immune dysregulation and increased platelet consumption. In families in whom several members are responsive to ITP-directed therapies, an underlying Src p.E527K variant should be excluded.
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Púrpura Trombocitopénica Idiopática , Trombocitopenia , Adulto , Plaquetas , Humanos , Megacariocitos , Púrpura Trombocitopénica Idiopática/diagnóstico , Púrpura Trombocitopénica Idiopática/genética , Trombocitopenia/genética , TrombopoyesisRESUMEN
Platelet disorders comprise heterogeneous diseases featured by reduced platelet counts and/or impaired platelet function causing variable bleeding symptoms. Despite their bleeding diathesis, patients with platelet disorders can develop transient or permanent prothrombotic conditions that necessitate prophylactic or therapeutic anticoagulation. Anticoagulation in patients with platelet disorders is a matter of concern because the bleeding risk could add to the hemorrhagic risk related to the platelet defect. This review provides an overview on the evidence on anticoagulation in patients with acquired and inherited thrombocytopenia and/or platelet dysfunction. We summarize tools to evaluate and balance bleeding- and thrombotic risks and describe a practical approach on how to manage these patients if they have an indication for prophylactic or therapeutic anticoagulation.
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Anticoagulantes/uso terapéutico , Trastornos de las Plaquetas Sanguíneas/tratamiento farmacológico , Trastornos Hemorrágicos/tratamiento farmacológico , Trombocitopenia/metabolismo , Anticoagulantes/farmacología , HumanosRESUMEN
Inherited platelet disorders (IPDs) are a group of rare conditions featured by reduced circulating platelets and/or impaired platelet function causing variable bleeding tendency. Additional hematological or non hematological features, which can be congenital or acquired, distinctively mark the clinical picture of a subgroup of patients. Recognizing an IPD is challenging, and diagnostic delay or mistakes are frequent. Despite the increasing availability of next-generation sequencing, a careful phenotyping of suspected patients-concerning the general clinical features, platelet morphology, and function-is still demanded. The cornerstones of IPD diagnosis are clinical evaluation, laboratory characterization, and genetic testing. Achieving a diagnosis of IPD is desirable for several reasons, including the possibility of tailored therapeutic strategies and individual follow-up programs. However, detailed investigations can also open complex scenarios raising ethical issues in case of IPDs predisposing to hematological malignancies. This review offers an overview of IPD diagnostic workup, from the interview with the proband to the molecular confirmation of the suspected disorder. The main implications of an IPD diagnosis are also discussed.
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Trastornos de las Plaquetas Sanguíneas , Diagnóstico Tardío , Trastornos de las Plaquetas Sanguíneas/diagnóstico , Trastornos de las Plaquetas Sanguíneas/genética , Plaquetas , Pruebas Genéticas , Secuenciación de Nucleótidos de Alto Rendimiento , HumanosRESUMEN
BACKGROUND: The classic Bernard-Soulier syndrome (BSS) is a rare inherited thrombocytopenia (IT) associated with severe thrombocytopenia, giant platelets, and bleeding tendency caused by homozygous or compound heterozygous variants in GP1BA, GP1BB, or GP9. Monoallelic BSS (mBSS) associated with mild asymptomatic macrothrombocytopenia caused by heterozygous variants in GP1BA or GP1BB may be a frequent cause of mild IT. OBJECTIVE: We aimed to examine the frequency of mBSS in a consecutive cohort of patients with IT and to characterize the geno- and phenotype of mBSS probands and their family members. Additionally, we set out to examine if thrombopoietin (TPO) levels differ in mBSS patients. PATIENTS/METHODS: We screened 106 patients suspected of IT using whole exome- or whole genome sequencing and performed co-segregation analyses of mBSS families. All probands and family members were phenotypically characterized. Founder mutation analysis was carried out by certifying that the probands were unrelated and the region around the variant was shared by all patients. TPO was measured by solid phase sandwich ELISA. RESULTS: We diagnosed 14 patients (13%) with mBSS associated with heterozygous variants in GP1BA and GP1BB. Six unrelated probands carried a heterozygous variant in GP1BA (c.58T>G, p.Cys20Gly) and shared a 2.0 Mb region on chromosome 17, confirming that it is a founder variant. No discrepancy of TPO levels between mBSS patients and wild-type family members (P > .05) were identified. CONCLUSION: We conclude that the most frequent form of IT in Denmark is mBSS caused by the Copenhagen founder variant.
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Síndrome de Bernard-Soulier , Trombocitopenia , Síndrome de Bernard-Soulier/diagnóstico , Síndrome de Bernard-Soulier/genética , Dinamarca , Homocigoto , Humanos , Linaje , Complejo GPIb-IX de Glicoproteína Plaquetaria/genética , Trombocitopenia/diagnóstico , Trombocitopenia/genéticaRESUMEN
Thrombocytopenic disorders have been treated with the Thrombopoietin-receptor agonist Eltrombopag. Patients with the same apparent form of thrombocytopenia may respond differently to the treatment. We describe a miniaturized bone marrow tissue model that provides a screening bioreactor for personalized, pre-treatment response prediction to Eltrombopag for individual patients. Using silk fibroin, a 3D bone marrow niche was developed that reproduces platelet biogenesis. Hematopoietic progenitors were isolated from a small amount of peripheral blood of patients with mutations in ANKRD26 and MYH9 genes, who had previously received Eltrombopag. The ex vivo response was strongly correlated with the in vivo platelet response. Induced Pluripotent Stem Cells (iPSCs) from one patient with mutated MYH9 differentiated into functional megakaryocytes that responded to Eltrombopag. Combining patient-derived cells and iPSCs with the 3D bone marrow model technology allows having a reproducible system for studying drug mechanisms and for individualized, pre-treatment selection of effective therapy in Inherited Thrombocytopenias.
Platelets are tiny cell fragments essential for blood to clot. They are created and released into the bloodstream by megakaryocytes, giant cells that live in the bone marrow. In certain genetic diseases, such as Inherited Thrombocytopenia, the bone marrow fails to produce enough platelets: this leaves patients extremely susceptible to bruising, bleeding, and poor clotting after an injury or surgery. Certain patients with Inherited Thrombocytopenia respond well to treatments designed to boost platelet production, but others do not. Why these differences exist could be investigated by designing new test systems that recreate the form and function of bone marrow in the laboratory. However, it is challenging to build the complex and poorly understood bone marrow environment outside of the body. Here, Di Buduo et al. have developed an artificial three-dimensional miniature organ bioreactor system that recreates the key features of bone marrow. In this system, megakaryocytes were grown from patient blood samples, and hooked up to a tissue scaffold made of silk. The cells were able to grow as if they were in their normal environment, and they could shed platelets into an artificial bloodstream. After treating megakaryocytes with drugs to stimulate platelet production, Di Buduo et al. found that the number of platelets recovered from the bioreactor could accurately predict which patients would respond to these drugs in the clinic. This new test system enables researchers to predict how a patient will respond to treatment, and to tailor therapy options to each individual. This technology could also be used to test new drugs for Inherited Thrombocytopenias and other blood-related diseases; if scaled-up, it could also, one day, generate large quantities of lab-grown blood cells for transfusion.