Autosomal Dominant Platelet Disorder Research Articles

Clinical and Genetic Characteristics of 30 Patients Misdiagnosed As Having ITP with Non-Muscle Myosin Heavy Chain 9-Related Disease (MYH9-RD): A Retrospective Analysis in China

Introduction : Non-muscle myosin heavy chain 9-related diseases (MYH9-RD) are rare autosomal dominant platelet (PLT) disorders presenting as nonsyndromic forms characterized by macrothrombocytopenia with giant PLTs and leukocyte inclusion bodies or as syndromic forms combining these hematological features with deafness, nephropathy, or cataracts. Owing to its low incidence and similar clinical manifestations, it is easily misdiagnosed as immune thrombocytopenia (ITP), leading to inappropriate treatment. To improve the diagnosis of this rare thrombocytopenic disease and minimize the misdiagnosis rate, we retrospectively analyzed the clinical, treatment, and genetic characteristics of patients with MYH9-RD who were initially misdiagnosed with ITP and compared the differences between pediatric and adult patients. Methods: Patients from various sites in China were enrolled. The inclusion criteria were macrothrombocytopenia on peripheral blood smear, MYH9 gene mutation detected by next-generation sequencing or whole exon sequencing and verified by Sanger sequencing, and absence of other acquired thrombocytopenia diseases. We collected basic information, clinical manifestations, and laboratory test results via medical records and telephone interviews. Results: Of the 30 patients, 17 were male, 13 were female, 14 were children, and 16 were adults. There was no significant difference in the mean PLT count (×10 9/L) at the time of diagnosis between children and adults (50.5 vs. 33.4, P=0.188). However, the median duration of misdiagnosis was significantly shorter in children (0.1 y vs. 9 y, P=0.000) (Table 1). Overall, 5 patients showed no signs of bleeding, while 25 presented with bleeding manifestations: 11 with skin bleeding spots and ecchymosis, 15 with epistaxis, 6 with gingival bleeding, and 3 with increased menstrual volume. Moreover, 21 patients showed no non-hematological manifestations, while 9 presented with non-hematological manifestations, which occurred exclusively in adult patients. Among the latter, 6 patients had nephropathy (2 underwent renal transplantation, and 2 underwent regular dialysis), 5 had hearing impairment (4 underwent high-frequency hearing impairment, and 1 received a cochlear implant due to deafness), and 1 had a congenital cataract. All patients received first- or second-line treatment for ITP. The most common treatment in children was immunoglobulins (12/14), followed by steroids (5/14) and traditional Chinese medicine (2/14). No children received >3 types of treatment, 4 children received >1, and 6 adult patients received >3. The highest number of treatment type for a single patient was up to 7. The median duration of ITP treatment was 1.5 y for adults and 0.1 y for children ( P=0.002). All patients presented heterozygous missense mutations in MYH9; 17/30 were spontaneous mutations, 7/30 were familial mutations (involving 6 paternal and 1 maternal origin), and 6/30 cases had an unknown inherited type. Fourteen cases were located in the N-terminal globular head region, and 16 cases were located in the C-terminal tail domain (Figure 1). Two novel mutations (not recorded in the Global Variome database), p. G1517V and p. K1674Q, were identified. In 2 adult patients with p.R702C mutations, proteinuria occurred in adolescence, followed by chronic renal failure (CRF), end-stage renal failure, and hearing loss. The 3 children with p.R702C mutation were aged <4 years and exhibited no non-hematological symptoms. Three of the 5 adults with p.E1841K mutations had CRF and underwent kidney transplantation. Conclusion s : Compared with pediatric patients, adult patients are more likely to be misdiagnosed with ITP, receive more types of inappropriate treatment for a longer period of time, and have more common non-hematological manifestations. To improve diagnostic accuracy and reduce the misdiagnosis rate for patients with thrombocytopenia, especially those with a poor response to ITP treatments, attention should be paid to mean PLT volume, family history, and genetic screening. Consistent with previous reports, patients with the p.R702C mutation showed early-stage kidney injury and hearing loss. The incidence of spontaneous mutations (56.7%) was significantly higher than previously reported (25-35%). The genetic characteristics of MYH9-RD and genotype-phenotype relationships in China require further clarification.

Characterization of Sensorineural Hearing Loss in Patients With MYH9-Related Disease: A Systematic Review.

To review the characteristics and progression of hearing loss in MYH9-related disease (MYH9-RD) patients and present a unique case of bilateral non-simultaneous sudden sensorineural hearing loss (SNHL) in an MYH9-RD patient. MYH9-RD is a rare autosomal dominant platelet disorder. Patients with this disorder have a variable risk of developing SNHL. A comprehensive literature search for scientific articles in PubMed, Scopus, and Web of Science that reported hearing loss outcomes in MYH9-RD patients. Initial search yielded 270 studies. Eight studies with a total of 23 patients met inclusion criteria and were used for data analysis. MYH9-RD patients typically present with progressive bilateral SNHL affecting predominantly the high frequencies. Mean age of hearing loss onset was 17.1 years, progressing to severe-profound SNHL over a mean period of 14.4 years. Seventeen of the 23 patients received cochlear implant (CI) at a mean age of 37.9 years. In comparison, the study patient presented initially with bilateral progressive SNHL as a teenager and developed bilateral non-simultaneous sudden SNHL, first in her right ear at the age of 31 and 7 months later in her left ear at the age of 32. She is now a successful bilateral CI user. This is the first systematic investigation of the relationship between MYH9-RD patients and SNHL. Hearing loss in MYH9-RD patients is generally characterized as progressive SNHL. However, the study patient presented with the unique feature of bilateral non-simultaneous sudden SNHL, potentially expanding the hearing loss sequela associated with this disorder.

Platelet Ultrastructure in Familial Platelet Disorder with Associated Myeloid Malignancy (FPDMM)

The familial platelet disorder with associated myeloid malignancy (FPDMM) is an autosomal dominant platelet disorder, caused by germline RUNX1 mutations, with predisposition to develop hematologic malignancies, especially acute myeloid leukemia. In many of the FPDMM families reported, the platelet defect was a delta-storage-pool disease (d-SPD) which can also be found without leukemia propensity. However, it has not been studied whether the two types of d-SPD have a common nature. Platelet ultrastructure, previously very little studied, may be one of the aspects to be analyzed to solve this question. We analyzed the ultrastructural characteristics of platelets in 5 members of a family with FPDMM. The family included three generations and all affected members had a RUNX1 deletion: chr21:36349450-36572837 (Rio-Machin et al. Nat Commun 2020;11:1044). None of the patients studied had developed leukemia at the time of the platelet study. We compared the results with those of 24 patients with d-SPD non-associated with leukemia and with those of 15 healthy individuals. Platelets were processed by transmission electron microscopy by standard methods. On the electron micrographs, morphometric analysis of the following structures was performed: 1) Platelets: size and shape, 2) Intraplatelet corpuscular structures (dense granules, alpha granules, mitochondria, lipid droplets): size and number (per platelet and per square micrometer of platelet area), 3) Surface-connected canalicular system (SCS): mean area of individual channel sections and mean percentage of the total SCS area with respect to the platelet area, 4) Glycogen masses: total area with respect to the platelet area. The morphological traits of the platelets and organelles measured above were also evaluated and the dense granules were classified into 4 different types depending on the appearance of their solid core (Weiss et al. Br J Haematol 1993;83:282). The dense tubular system and other ultrastructural characteristics were evaluated by morphology only. The main features of the platelet ultrastructure in patients with FPDMM were (Fig 1, Table 1): 1) slight increase in platelet size while preserving the discoidal shape, 2) moderate reduction in the number of dense granules, which showed a reduced proportion of type 1 granules (with the solid core occupying more than 50 % of the granule) and an increased proportion of type 2 and type 3 granules, with a solid core reduced or fragmented respectively, 3) marked increase and dilatation of SCS with some elements filled by a substance of unknown origin, 4) moderate increase in dense tubular system with occasional complex formation. The platelet ultrastructure was similar to that described in the non-associated d-SPD group (Pujol-Moix et al. Haematologica 2000;85:619) although there were some differences (Table 1): in FPDMM platelets the dense granules were less reduced but more dysmorphic, and the SCS, equally dilated, contained a substance that was not observed in the d-SPD platelets. Given that all the findings described belong to the same family, it would be necessary to evaluate the platelet ultrastructure in additional families and extend the study to other characteristics of d-SPD. Only in this way, it would be possible to know to what extent the platelet defect of FPDMM and that of non-associated d-SPD shared a pathogenic mechanism. Disclosures No relevant conflicts of interest to declare.

Differential proteomics study of platelets in asymptomatic constitutional macrothrombocytopenia: altered levels of cytoskeletal proteins.

Harris platelet syndrome (HPS), also known as asymptomatic constitutional macrothrombocytopenia (ACMT), is an autosomal dominant platelet disorder characterized by mild-to-severe thrombocytopenia and giant platelets with normal platelet aggregation and absence of bleeding symptoms. We have attempted a comparative proteomics study for profiling of platelet proteins in healthy vs. pathological states to discover characteristic protein expression changes in macrothrombocytes and decipher the factors responsible for the functionally active yet morphologically distinct platelets. We have used 2-D gel-based protein separation techniques coupled with MALDI-ToF/ToF-based mass spectrometric identification and characterization of the proteins to investigate the differential proteome profiling of platelet proteins isolated from the peripheral blood samples of patients and normal volunteers. Our study revealed altered levels of actin-binding proteins such as myosin light chain, coactosin-like protein, actin-related protein 2/3 complex, and transgelin2 that hint toward the cytoskeletal changes necessary to maintain the structural and functional integrity of macrothrombocytes. We have also observed over expressed levels of peroxiredoxin2 that signifies the prevailing oxidative stress in these cells. Additionally, altered levels of protein disulfide isomerase and transthyretin provide insights into the measures adapted by the macrothrombocytes to maintain their normal functional activity. This first proteomics study of platelets from ACMT may provide an understanding of the structural stability and normal functioning of these platelets in spite of their large size.

Mutation spectrum and genotype-phenotype correlations in a large French cohort of MYH9-Related Disorders.

MYH9-Related Disorders are a group of rare autosomal dominant platelet disorders presenting as nonsyndromic forms characterized by macrothrombocytopenia with giant platelets and leukocyte inclusion bodies or as syndromic forms combining these hematological features with deafness and/or nephropathy and/or cataracts. They are caused by mutations in the MYH9 gene encoding the nonmuscle myosin heavy chain II-A (NMMHC-IIA). Until now, at least 49 MYH9 mutations have been reported in isolated cases or small series but only rarely in large series. We report the results of an 8-year study of a large cohort of 109 patients from 37 sporadic cases and 39 unrelated families. We have identified 43 genetic variants, 21 of which are novel to our patients. A majority, 33 (76.7%), were missense mutations and six exons were preferentially targeted, as previously published. The other alterations were three deletions of one nucleotide, one larger deletion of 21 nucleotides, and one duplication. For the first time, a substitution T>A was found in the donor splice site of intron 40 (c.5765+2T>A). Seven patients, four from the same family, had two genetic variants. The analysis of the genotype-phenotype relationships enabled us to improve the knowledge of this heterogeneous but important rare disease.

Syndrome MYH9 et anémie hémolytique auto-immune : une association fortuite ?

IntroductionThe MYH9 syndrome is a group of rare autosomal dominant platelet disorders associating in most of the cases a macrothrombocytopenia and characteristic leukocyte inclusions. Clinical features may include renal, visual, or hearing impairment. The bleeding tendency is usually moderate. Case reportWe report a 28-year-old-man, with an auto-immune haemolytic anaemia associated with a MYH9 syndrome. ConclusionTo our knowledge, this is the first report of such an association.

Syndromic mental retardation with thrombocytopenia due to 21q22.11q22.12 deletion: Report of three patients

During the last few years, an increasing number of microdeletion/microduplication syndromes have been delineated. This rapid evolution is mainly due to the availability of microarray technology as a routine diagnostic tool. Microdeletions of the 21q22.11q22.12 region encompassing the RUNX1 gene have been reported in nine patients presenting with syndromic thrombocytopenia and mental retardation. RUNX1 gene is responsible for an autosomal dominant platelet disorder with predisposition to acute myelogenous leukemia. We report on three novel patients with an overlapping "de novo" interstitial deletion involving the band 21q22 characterized by array-CGH. All our patients presented with severe developmental delay, dysmorphic features, behavioral problems, and thrombocytopenia. Comparing the clinical features of our patients with the overlapping ones already reported two potential phenotypes related to 21q22 microdeletion including RUNX1 were highlighted: thrombocytopenia with +/- mild dysmorphic features and syndromic thrombocytopenia with growth and developmental delay.

The May-Hegglin anomaly in a kidney transplant recipient.

We report a 37-year-old renal transplanted patient with thrombocytopaenia. He had May–Hegglin anomaly (MHA) inherited from his father, who died due to myocardial infarction subsequent to two renal transplantations. At the age of 15 years, proteinuria was found, but he was not biopsied. In 1982, he underwent splenectomy because of trauma. In 1989, when he was 17, renal function was found to be decreased. Five years later, he started haemodialysis and at 24 years he underwent live related kidney transplantation. Serum creatinine was always found stable around 1.9 mg/dl on cyclosporine, azathioprine and steroids. Automated blood count reported a number of platelets around 8000/μl with a mean volume of 20.3 fl (normal range 7.2– 11.1 fl). However, blood cell counters have a defined volume window for platelet counting and large macrothrombocytes are classified as red blood cells, meaning that platelet number is underestimated (Figure 1). The true value of platelets after count with the microscope was around 30 000/μl. He was completely asymptomatic, without any bleeding tendency. MHA is a rare autosomal dominant platelet disorder, characterized by macrothrombocytopaenia and granulocyte inclusion bodies. The mutation involves the MYH9 gene, localized in chromosome 22q, which encodes a heavy chain of non-muscle myosin IIA. MYH9-related hereditary macrothrombocytopaenia disorders include: MHA, Ebstein, Fechtner and Sebastian syndromes, which are characterized by nephritis, hearing loss and cataract [1]. In 1992, Nel et al. [2] reported two cases of MHA incidentally discovered in a patient and his brother during investigation for end-stage renal failure and workup for renal transplantation. Alhindawi and Al-Jbour [3] described an association between Epstein syndrome and chronic nephropathy. Recently, Singh et al. [4] suggested that MYH9 gene alterations are associated with a greater risk of focal segmental glomerulosclerosis and hypertensive nephrosclerosis in African Americans.

Mutations inMYH9Exons 1, 16, 26, and 30 Are Infrequently Found in Japanese Patients with Nonsyndromic Deafness

Mutations in MYH9 result in the autosomal dominant giant platelet disorders with leukocyte inclusion bodies with varying degrees of Alport manifestations, including nephritis, deafness, and cataracts. A specific MYH9 mutation in exon 16, R705H, causes nonsyndromic deafness DFNA17. We searched for mutations in MYH9 exons 1, 16, 26, and 30 in a total of 157 Japanese patients with nonsyndromic deafness without known cause of hearing loss, but no mutations were found. We conclude that mutations in MYH9 are infrequently found in patients with nonsyndromic deafness and suggest that MYH9 mutations infrequently cause isolated sensorineural hearing loss. Thus, MYH9 may not currently be a good candidate gene for efficient screening of genetic causes in nonsyndromic deafness.

Historical hematology: May–Hegglin anomaly

May-Hegglin anomaly is a rare autosomal dominant platelet disorder characterized by thrombocytopenia, giant platelets, and unique leukocyte inclusion bodies. This disorder was first described by May, a German physician, in 1909, and was subsequently described by a Swiss physician, Hegglin, in 1945. The pathogenesis of the disorder had been unknown until recently, when mutations in the gene encoding for nonmuscle myosin heavy chain IIA (MYH9) were identified. Unique cytoplasmic inclusion bodies are aggregates of nonmuscle myosin heavy chain IIA, and are only present in granulocytes. It is not yet known why inclusion bodies are not present in platelets, monocytes, and lymphocytes, or how giant platelets are formed. Interestingly, MYH9 is also found to be responsible for several related disorders with macrothrombocytopenia and leukocytes inclusion, including Sebastian, Fechtner, and Epstein syndromes, which feature deafness, nephritis, and/or cataract. Current interest is centered upon the mechanisms by which a single mutation causes a variety of phenotypes.

Le syndrome MYH9 : à propos d'une nouvelle observation et de la mise en évidence d'une nouvelle mutation du gène MYH9

Introduction Familial macrothrombocytopenias are a group of rare autosomal dominant platelet disorders including many syndromes in particular the May-Hegglin anomaly. They are characterized by thrombocytopenia with giant platelets and in some cases neutrophilic inclusions in peripheral blood granulocytes. Recently these different clinical entities have been demonstrated to be linked to mutations in the same gene, MYH9. Case report We report in a young African woman presenting as a May-Hegglin anomaly a new mutation of the MYH9 gene. In regard of this case we present a brief review of the MYH9 syndrome. Conclusion The MYH9 syndrome includes now several clinical entities who share some common clinical and biological characteristics such as a thrombocytopenia with giant platelets, presence or absence of other manifestations including Dohle like bodies, nephritis, sensineural hearing loss, cataract. We report a new case in which a new mutation of the MYH9 gene was evidenced.

Localization of a lysosomal enzyme in platelets from patients with the White platelet syndrome

The White platelet syndrome (WPS) is an autosomal dominant platelet disorder. Platelet structural abnormalities include the presence of Golgi complexes in up to 13% of their cells, frequently accompanied by centrioles, deficient numbers or absent alpha granules in 30–40% of their platelets and masses of dense tubular system (DTS) channels often forming areas of cytoplasmic sequestration. The degradation of cytoplasm and organelles in the sequestered areas suggested the possibility that hydrolytic enzymes remained in the DTS and were being transferred to sequestration vacuoles. The present study has used ultrastructural cytochemistry to localize the sites of a lysosomal enzyme, aryl sulfatase, in normal and WPS platelets. Enzyme reaction product, lead sulfide, was localized to lysosomal organelles in normal platelets, and only in rare examples appeared in the DTS. Aryl sulfatase activity was found in the Golgi complexes, a few lysosomes, much of the DTS and areas of cytoplasmic sequestration in WPS platelets. The findings indicate that aryl sulfatase, and, most likely, proteins destined for alpha granules in WPS platelets are not completely transferred from the endoplasmic reticulum to Golgi complexes, then to Golgi vesicles and finally to lysosomal and alpha granules in the parent cell before the platelets are delivered to the circulation.

A ‘touch’ of the White platelet syndrome

Investigations into structural defects in platelets from a large family with the White platelet syndrome (WPS) separated the members into three groups. The first group of 22 members was the subject of our first report (White JG, Key NS, King RA, Vercellotti GM. The white platelet syndrome: A new autosomal dominant platelet disorder. Platelets 2004;15:173–184). A third group of 13 members had no abnormalities of platelet ultrastructure. The second group of 17 members, the focus of the present study, had a ‘touch’ of the WPS. Platelet counts, mean platelet volumes (MPVs) and platelet responses to aggregating agents were normal in ‘touch’ patients in contrast to platelets of those with the full WPS in whom these parameters were abnormal. Up to 13% of the full WPS platelets contained large, fully developed Golgi complexes, up to seven in number, extruding innumerable vesicles from the trans-Golgi face and filling the cytoplasm of many platelets. Many Golgi complexes had centrioles associated with them. ‘Touch’ platelets had one or two Golgi complexes of intermediate size in 3–5% of their platelets. Golgi vesicles were uncommon and centrioles absent. Gray platelets and hypogranular cells were infrequent in patients with a ‘touch’ of the WPS, whereas up to 44% of the platelets from those with the WPS were gray or hypogranular. Elements of the dense tubular system were prominent in full WPS platelets, together with their formation into areas of cytoplasmic sequestration and autodigestion. These features were absent in ‘touch’ platelets. As commonly observed in full WPS platelets, mitochondria were larger and more numerous than alpha granules in some ‘touch’ cells. Both ‘touch’ and full WPS platelets frequently contained giant and rod-shaped granules. Dense bodies, however, were normal in size and number in ‘touch’ platelets, and half normal size in full WPS platelets. The separation of ultrastructural abnormalities in the two varieties of the WPS suggests that genetic defects involve more than a single gene and the genes are variable in their penetrance. Genetic studies to determine if this is the case are currently in progress.

Genotype–phenotype correlation in MYH9‐related thrombocytopenia

Mutation of the non-muscle myosin heavy chain type II-A results in MYH9-related hereditary macrothrombocytopenia (HMTC), including four autosomal dominant platelet disorders: May-Hegglin anomaly (MHA), Sebastian (SBS), Fechtner (FS) and Epstein (EPS) syndrome. Denaturing high-performance liquid chromatography (DHPLC) was optimised for rapid screening of the seven exons harbouring all but one of the previously reported mutations of MYH9. Individuals from 13 families with phenotypes suggestive of MYH9-related HMTC were screened for mutations by DHPLC followed by direct sequencing of samples with aberrant column retention time. Mutations were identified in all 13 families. Six distinct missense heterozygous mutations were found in 10 families, including six families with MHA or SBS (E1841K, D1424N), three families with FS (R702H, R1165C, and D1424Y), and one family with EPS (S96L). A truncating mutation (R1933X) was found in three MHA families. A review of all published mutations suggests that mutation in the C-terminal coiled coil region or truncation of the tailpiece is associated with haematological-only phenotype, while mutation of the head ATPase domain frequently is associated with nephropathy and/or hearing loss. Mutations of other regions have intermediate expression of non-haematological characteristics. Further study is required to confirm these associations and understand the molecular basis for this genotype-phenotype relationship.

Perioperative management of a patient with May-Hegglin anomaly requiring craniotomy

May-Hegglin anomaly (MHA) is a rare type of autosomal dominant platelet disorder associated with mutations in the gene encoding nonmuscle myosin heavy chain 9 (MYH9). It is characterized by the presence of large platelets, leukocyte inclusions, and thrombocytopenia. The bleeding tendency is usually mild, but severe hemorrhages have been reported. This is the first reported case of a patient with MHA who underwent craniotomy for intractable seizure disorder of temporal lobe origin. Patients who have thrombocytopenia have a higher likelihood of developing intraoperative or postoperative intracranial hematoma and bleeding complications. The patient was administered desmopressin (DDAVP) prior to the neurosurgical procedure and had no complications. With this approach, the use of platelet concentrates could be avoided. We discuss the role of DDAVP in MHA and related platelet disorders and review the current literature.

Sticky Platelet Syndrome (SPS) as a Cause of Unexplained Thrombosis.

Background: Sticky platelet syndrome (SPS) was first described in 1983 by Holliday. The results of recent studies suggest SPS to be a common cause of arterial and venous thrombosis. With respect to otherwise unexplained venous thrombosis, the prevalence of SPS approximates that of activated protein C resistance (APC-R). It has been found that SPS accounted for about 21% of otherwise unexplained arterial events (acute myocardial infarction, cerebrovascular thrombosis, transient cerebral ischemic attacks, retinal thrombosis, and peripheral arterial thrombosis) and accounted for about 13,2% of otherwise unexplained venous events (deep vein thrombosis, with or without pulmonary embolus).SPS is an autosomal dominant platelet disorder associated with arterial and venous thromboembolic events, frequently recurrent under oral anticoagulant therapy. It is characterized by hyperaggregability of platelets in platelet-rich plasma with adenosine diphosphate (ADP) and epinephrine (type I), epinephrine alone (type II), or ADP alone (type III). Combination of SPS with other congenital thrombophilic defects have been described. Low-dose aspirin treatment (80–100 mg) ameliorates the clinical symptoms and normalizes hyperaggregability. The precise etiology of this defect is at present not known, but receptors on the platelet surface may be involved.Patients and methods: We examined fifty patients with otherwise unexplained arterial or venous thrombosis. SPS evaluations were performed according to the method of Mammen. The equipment used was PACKS-4 aggregometer.Results: SPS was detected in eight patients. Clinically SPS resulted as an arterial thrombosis in five patients (2 coronary thrombosis, 2 cerebral ischemic attacks, 1 peripheral arterial thrombosis), two individuals showed deep venous thrombosis (DVT) and one patient passed recurrent abortions. SPS was classified as type I in 2 cases, 5 patients had type II and only 1 patient was diagnosed as type III, respectively (4 of them showed a glycoprotein IIIa abnormality).Conclusion: Because treatment with heparin or warfarin will not alleviate the thrombotic tendency of SPS, but simple aspirin therapy almost always will correct the defect and protect the individual from second event, it is particularly important to define the presence of this defect.

Nonmuscle Myosin Heavy Chain IIA Mutations Define a Spectrum of Autosomal Dominant Macrothrombocytopenias: May-Hegglin Anomaly and Fechtner, Sebastian, Epstein, and Alport-Like Syndromes

May-Hegglin anomaly (MHA) and Fechtner (FTNS) and Sebastian (SBS) syndromes are autosomal dominant platelet disorders that share macrothrombocytopenia and characteristic leukocyte inclusions. FTNS has the additional clinical features of nephritis, deafness, and cataracts. Previously, mutations in the nonmuscle myosin heavy chain 9 gene (MYH9), which encodes nonmuscle myosin heavy chain IIA (MYHIIA), were identified in all three disorders. The spectrum of mutations and the genotype-phenotype and structure-function relationships in a large cohort of affected individuals (n=27) has now been examined. Moreover, it is demonstrated that MYH9 mutations also result in two other FTNS-like macrothrombocytopenia syndromes: Epstein syndrome (EPS) and Alport syndrome with macrothrombocytopenia (APSM). In all five disorders, MYH9 mutations were identified in 20/27 (74%) affected individuals. Four mutations, R702C, D1424N, E1841K, and R1933X, were most frequent. R702C and R702H mutations were only associated with FTNS, EPS, or APSM, thus defining a region of MYHIIA critical in the combined pathogenesis of macrothrombocytopenia, nephritis, and deafness. The E1841K, D1424N, and R1933X coiled-coil domain mutations were common to both MHA and FTNS. Haplotype analysis using three novel microsatellite markers revealed that three E1841K carriers--one with MHA and two with FTNS--shared a common haplotype around the MYH9 gene, suggesting a common ancestor. The two new globular-head mutations, K371N and R702H, as well as the recently identified MYH9 mutation, R705H, which results in DFNA17, were modeled on the basis of X-ray crystallographic data. Altogether, our data suggest that MHA, SBS, FTNS, EPS, and APSM comprise a phenotypic spectrum of disorders, all caused by MYH9 mutations. On the basis of our genetic analyses, the name "MYHIIA syndrome" is proposed to encompass all of these disorders.

Platelets from patients with the Quebec platelet disorder contain and secrete abnormal amounts of urokinase-type plasminogen activator

The Quebec platelet disorder (QPD) is an autosomal dominant platelet disorder associated with delayed bleeding and alpha-granule protein degradation. The degradation of alpha-granule, but not plasma, fibrinogen in patients with the QPD led to the investigation of their platelets for a protease defect. Unlike normal platelets, QPD platelets contained large amounts of fibrinolytic serine proteases that had properties of plasminogen activators. Western blot analysis, zymography, and immunodepletion experiments indicated this was because QPD platelets contained large amounts of urokinase-type plasminogen activator (u-PA) within a secretory compartment. u-PA antigen was not increased in all QPD plasmas, whereas it was increased more than 100-fold in QPD platelets (P <.00009), which contained increased u-PA messenger RNA. Although QPD platelets contained 2-fold more plasminogen activator inhibitor 1 (PAI-1) (P <.0008) and 100-fold greater u-PA-PAI-1 complexes (P <.0002) than normal platelets, they contained excess u-PA activity, predominantly in the form of two chain (tcu-PA), which required additional PAI-1 for full inhibition. There was associated proteolysis of plasminogen in QPD platelets, to forms that comigrated with plasmin. When similar amounts of tcu-PA were incubated with normal platelet secretory proteins, many alpha-granule proteins were proteolyzed to forms that resembled degraded QPD platelet proteins. These data implicate u-PA in the pathogenesis of alpha-granule protein degradation in the QPD. Although patients with the QPD have normal to increased u-PA levels in their plasma, without evidence of systemic fibrinogenolysis, their increased platelet u-PA could contribute to bleeding by accelerating fibrinolysis within the hemostatic plug. QPD is the only inherited bleeding disorder in humans known to be associated with increased u-PA.

Mutations in the NMMHC-A gene cause autosomal dominant macrothrombocytopenia with leukocyte inclusions (May-Hegglin anomaly/Sebastian syndrome)

AbstractMacrothrombocytopenia with leukocyte inclusions is a rare autosomal dominant platelet disorder characterized by a triad of giant platelets, thrombocytopenia, and characteristic Döhle body-like leukocyte inclusions. A previous study mapped a locus for the disease on chromosome 22q12.3-q13.2 by genome-wide linkage analysis. In addition, the complete DNA sequence of human chromosome 22 allowed a positional candidate approach, and results here indicate that the gene encoding nonmuscle myosin heavy chain-A, NMMHC-A, is mutated in this disorder. Mutations were found in 6 of 7 Japanese families studied: 3 missense mutations, a nonsense mutation, and a one-base deletion resulting in a premature termination. Immunofluorescence studies revealed that NMMHC-A distribution in neutrophils appeared to mimic the inclusion bodies. These results provide evidence for the involvement of abnormal NMMHC-A in the formation of leukocyte inclusions and also in platelet morphogenesis.

The Gene for May-Hegglin Anomaly Localizes to a

The May-Hegglin anomaly (MHA) is an autosomal dominant platelet disorder of unknown etiology. It is characterized by thrombocytopenia, giant platelets, and leukocyte inclusion bodies, and affected heterozygotes are predisposed to bleeding episodes. The MHA gene has recently been localized, by means of linkage analysis, to a 13.6-cM region on chromosome 22, and the complete chromosome 22 sequence has been reported. We recently performed a genome scan for the MHA gene in 29 members of a large, multigenerational Italian family, and we now confirm that the MHA locus is on chromosome 22q12. 3-13.1. The maximal two-point LOD score of 4.50 was achieved with the use of marker D22S283, at a recombination fraction of.05. Haplotype analysis narrowed the MHA critical region to 6.6 cM between markers D22S683 and D22S1177. It is of note that the chromosome 22 sequence allowed all markers to be ordered correctly, identified all the candidate genes and predicted genes, and specifically determined the physical size of the MHA region to be 0. 7 Mb. These results significantly narrow the region in which the MHA gene is located, and they represent the first use of chromosome 22 data to positionally clone a disease gene.