Platelet size for distinguishing between inherited thrombocytopenias and immune thrombocytopenia: a multicentric, real life study.

The most frequent forms of inherited thrombocytopenia (IT) are characterized by platelet size abnormalities and it has been suggested that this parameter is useful for their differentiation from immune thrombocytopenia (ITP). Recently, a monocentric study identified cut-off values for mean platelet volume (MPV) and mean platelet diameter (MPD) with good diagnostic accuracy in this respect. To validate these cut-off values in a different and larger case series of patients, we enrolled 130 subjects with ITP and 113 with IT in six different centres. The platelet count and MPV was each measured by the instrument routinely used in each institution. In some centres, platelet count was also measured by optical microscopy. MPD was evaluated centrally by image analysis of peripheral blood films. The previously identified cut-off value for MPV had 91% specificity in distinguishing ITP from inherited macrothrombocytopenias (mono and biallelic Bernard-Soulier, MYH9-related disease), while its sensitivity was greatly variable depending on the instrument used. With an appropriate instrument, specificity was 83%. The diagnostic accuracy of MPD was lower than that obtained with MPV. We concluded that MPV is a useful parameter for differentiating ITP from IT provided that it is measured by appropriate cell counters

Clinical and genetic aspects Bernard-Soulier syndrome: searching for genotype/phenotype correlations

Background. Bernard-Soulier syndrome is a severe bleeding disease due to a defect of GPIb/IX/V, a platelet complex that binds the von Willebrand factor. Due to the rarity of the disease, there are reports only on a few cases preventing any attemptable correlations between genotype and phenotype. In order to reveal any associations, we described the largest case series ever reported, which was evaluated systematically at the same centre. Design and Methods. Thirteen patients with the disease and seven obligate carriers were enrolled. We collect clinical aspects and determined platelet features, including number and size, expression of membrane glycoproteins, and ristocetin induced platelet aggregation. Mutations were identified by directly sequencing of the GP1BA, GP1BB, and GP9 genes and their effect revealed by molecular modelling analyses. Results. Patients had all a moderate thrombocytopenia with giant platelets and a bleeding tendency whose severity varied among individuals. Consistent with an expression levels of GPIbα always lower than 10% of control values, platelet aggregation was absent or severely reduced. Homozygous mutations were identified in the GP1BA, GP1BB and GP9 gene. Six were novel alterations expected to destabilize the conformation of the respective protein. Except for obligate carriers of a GP9 mutation with a reduced GPIb/IX/V expression and defective aggregation, all the others had any obvious anomalies. Conclusions. Regardless of mutations identified, the patients' bleeding diathesis did not correlate with thrombocytopenia, which was always moderate, and platelet GPIbα expression, which was always severely impaired. Obligate carriers had features similar to controls though their GPIb/IX/V expression showed discrepancies. Aware of the limitations of our cohort, we cannot define any correlations. However, further investigations are should be encouraged to better understand the causes of this rare and underestimated disease

Platelet size for distinguishing between inherited thrombocytopenias and immune thrombocytopenia: A multicentric, real life study

The most frequent forms of inherited thrombocytopenia (IT) are characterized by platelet size abnormalities and it has been suggested that this parameter is useful for their differentiation from immune thrombocytopenia (ITP). Recently, a monocentric study identified cut-off values for mean platelet volume (MPV) and mean platelet diameter (MPD) with good diagnostic accuracy in this respect. To validate these cut-off values in a different and larger case series of patients, we enrolled 130 subjects with ITP and 113 with IT in six different centres. The platelet count and MPV was each measured by the instrument routinely used in each institution. In some centres, platelet count was also measured by optical microscopy. MPD was evaluated centrally by image analysis of peripheral blood films. The previously identified cut-off value for MPV had 91% specificity in distinguishing ITP from inherited macrothrombocytopenias (mono and biallelic Bernard-Soulier, MYH9-related disease), while its sensitivity was greatly variable depending on the instrument used. With an appropriate instrument, specificity was 83%. The diagnostic accuracy of MPD was lower than that obtained with MPV. We concluded that MPV is a useful parameter for differentiating ITP from IT provided that it is measured by appropriate cell counters. © 2013 John Wiley & Sons Ltd

β-1 tubulin R307H SNP alters microtubule dynamics and affects severity of a hereditary thrombocytopenia.

Single nucleotide polymorphisms (SNPs) in platelet-associated genes partly explain inherent variability in platelet counts. Patients with monoallelic Bernard Soulier syndrome due to the Bolzano mutation (GPIBA A156V) have variable platelet counts despite a common mutation for unknown reasons.We investigated the effect of the most common SNP (R307H) in the hematopoietic-specific tubulin isotype β-1 in these Bernard Soulier patients and potential microtubule-based mechanisms of worsened thrombocytopenia.Ninety-four monoallelic Bolzano mutation patients were evaluated for the R307H β-1 SNP and had platelet counts measured by three methods; the Q43P SNP was also evaluated. To investigate possible mechanisms underlying this association, we used molecular modeling of β-1 tubulin with and without the R307H SNP. We transfected SNP or non-SNP β-1 tubulin into MCF-7 and CMK cell lines and measured microtubule regrowth after nocodazole-induced depolymerization.We found that patients with at least one R307H SNP allele had significantly worse thrombocytopenia; manual platelet counting revealed a median platelet count of 124 in non-SNP patients and 76 in SNP patients (both ×10(9)  L(-1) ; P < 0.01). The Q43P SNP had no significant association with platelet count. Molecular modeling suggested a structural relationship between the R307H SNP and microtubule stability via alterations in the M-loop of β tubulin; in vitro microtubule recovery assays revealed that cells transfected with R307H SNP β-1 had significantly impaired microtubule recovery.Our data show that the R307H SNP is significantly associated with the degree of thrombocytopenia in congenital and acquired platelet disorders, and may affect platelets by altering microtubule behavior

Platelet size for distinguishing between inherited thrombocytopenias and immune thrombocytopenia: a multicentric, real life study

The most frequent forms of inherited thrombocytopenia (IT) are characterized by platelet size abnormalities and it has been suggested that this parameter is useful for their differentiation from immune thrombocytopenia (ITP). Recently, a monocentric study identified cut-off values for mean platelet volume (MPV) and mean platelet diameter (MPD) with good diagnostic accuracy in this respect. To validate these cut-off values in a different and larger case series of patients, we enrolled 130 subjects with ITP and 113 with IT in six different centres. The platelet count and MPV was each measured by the instrument routinely used in each institution. In some centres, platelet count was also measured by optical microscopy. MPD was evaluated centrally by image analysis of peripheral blood films. The previously identified cut-off value for MPV had 91% specificity in distinguishing ITP from inherited macrothrombocytopenias (mono and biallelic Bernard-Soulier, MYH9-related disease), while its sensitivity was greatly variable depending on the instrument used. With an appropriate instrument, specificity was 83%. The diagnostic accuracy of MPD was lower than that obtained with MPV. We concluded that MPV is a useful parameter for differentiating ITP from IT provided that it is measured by appropriate cell counters

Clinical and laboratory features of 103 patients from 42 Italian families with inherited thrombocytopenia derived from the monoallelic Ala156Val mutation of GPIb alpha (Bolzano mutation)

BACKGROUND: Bernard-Soulier syndrome is a very rare form of inherited thrombocytopenia that derives from mutations in GPIb\u3b1, GPIb\u3b2, or GPIX and is typically inherited as a recessive disease. However, some years ago it was shown that the monoallelic c.515C>T transition in the GPIBA gene (Bolzano mutation) was responsible for macrothrombocytopenia in a few Italian patients. DESIGN AND METHODS: Over the past 10 years, we have searched for the Bolzano mutation in all subjects referred to our institutions because of an autosomal, dominant form of thrombocytopenia of unknown origin. RESULTS: We identified 42 new Italian families (103 cases) with a thrombocytopenia induced by monoallelic Bolzano mutation. Analyses of the geographic origin of affected pedigrees and haplotypes indicated that this mutation originated in southern Italy. Although the clinical expression was variable, patients with this mutation typically had a mild form of Bernard-Soulier syndrome with mild thrombocytopenia and bleeding tendency. The most indicative laboratory findings were enlarged platelets and reduced GPIb/IX/V platelet expression; in vitro platelet aggregation was normal in nearly all of the cases. CONCLUSIONS: Our study indicates that monoallelic Bolzano mutation is the most frequent cause of inherited thrombocytopenia in Italy, affecting 20% of patients recruited at our institutions during the last 10 years. Because many people from southern Italy have emigrated during the last century, this mutation may have spread to other countries

Clinical and laboratory features of 103 patients from 42 italian families with inherited thrombocytopenia derived from the monoallelic Ala156Val mutation of gpibα (Bolzano mutation)

Background. Bernard-Soulier syndrome is a very rare form of inherited thrombocytopenia that derives from mutations in GPIb alpha, GPIb beta, or GPIX and is typically inherited as a recessive disease. However, some years ago it has been shown that the monoallelic c.515C>T transition in the GPIBA gene (Bolzano mutation) was responsible for macrothrombocytopenia in a few Italian patients. Design and Methods. In the past 10 years, we have searched for Bolzano mutation in all subjects referred to our institutions for an autosomal, dominant form of thrombocytopenia of unknown origin. Results. We identified 42 new Italian families (103 cases) with a thrombocytopenia induced by monoallelic Bolzano mutation. Analyses of the geographic origin of affected pedigrees and haplotypes indicated that this mutation originated in southern Italy. Although the clinical expressivity was variable, patients with this mutation typically had a mild form of Bernard-Soulier syndrome with a mild thrombocytopenia and bleeding tendency. The most indicative laboratory findings were an enlarged platelet size and reduced GPIb/IX/V platelet expression, although in vitro platelet aggregation was normal in nearly all of the cases. Conclusions. Our study indicates that monoallelic Bolzano mutation is the most frequent cause of inherited thrombocytopenia in Italy, affecting 20% of patients recruited at our institutions during the last 10 years. Because many people from southern Italy have emigrated during the last century, this mutation may have spread to other countries

Clinical and laboratory features of 103 patients from 42 Italian families with inherited thrombocytopenia derived from the monoallelic Ala156Val mutation of GPIb{alpha} (Bolzano mutation).

Background. Bernard-Soulier syndrome is a very rare form of inherited thrombocytopenia that derives from mutations in GPIb alpha, GPIb beta, or GPIX and is typically inherited as a recessive disease. However, some years ago it has been shown that the monoallelic c.515C>T transition in the GPIBA gene (Bolzano mutation) was responsible for macrothrombocytopenia in a few Italian patients. Design and Methods. In the past 10 years, we have searched for Bolzano mutation in all subjects referred to our institutions for an autosomal, dominant form of thrombocytopenia of unknown origin. Results. We identified 42 new Italian families (103 cases) with a thrombocytopenia induced by monoallelic Bolzano mutation. Analyses of the geographic origin of affected pedigrees and haplotypes indicated that this mutation originated in southern Italy. Although the clinical expressivity was variable, patients with this mutation typically had a mild form of Bernard-Soulier syndrome with a mild thrombocytopenia and bleeding tendency. The most indicative laboratory findings were an enlarged platelet size and reduced GPIb/IX/V platelet expression, although in vitro platelet aggregation was normal in nearly all of the cases. Conclusions. Our study indicates that monoallelic Bolzano mutation is the most frequent cause of inherited thrombocytopenia in Italy, affecting 20% of patients recruited at our institutions during the last 10 years. Because many people from southern Italy have emigrated during the last century, this mutation may have spread to other countries