Neurosurgical bleeding in platelet storage pool disorder: a case report.

Dense-granule deficiency (DGD) is an inherited platelet disorder due to the absence of dense granules essential for activation of platelets in the event of vascular injury. Decreased platelet dense granules can be detected by electron microscopy, while other tests of hemostasis, including platelet function analyzer (PFA®) closure times, may be normal. The present case report describes a patient with a lifelong history of mucocutaneous bleeding and excessive hemorrhage with resection of vestibular Schwannoma. After hemostasis was obtained the case was aborted and the neurosurgeon noted bleeding resembled as if patient was on an antiplatelet drug. Subsequent hematologic workup revealed a severe platelet function disorder. There is a paucity of literature on management of intracranial neurosurgery in patients with inherited platelet disorders. Patients undergoing major surgical procedures often receive tranexamic acid (TXA), desmopressin, and/or human-leukocyte antigen (HLA)-matched platelet transfusions. We review the clinical management of intracranial tumor surgery, as well as Cyberknife radiosurgery, in our patient with DGD. After diagnosis was known, thoughtful hemostatic planning with empiric platelet transfusions and TXA prevented recurrent bleeding.

Platelet disorders that affect platelet function require high index of suspicion and special laboratory evaluation for diagnosis. We provide a case report of storage pool deficiency with management of bleeding in Schwannoma resection and radiosurgery. This case report adds to the limited literature to guide treatment of platelet function disorder in neurosurgery.

A case of acquired transient bleeding diathesis associated with acquired platelet storage pool deficiency and defective thromboxane A2 production.

Acquired disorders of platelet function are an underdiagnosed cause of bleeding tendency. A 14-year-old girl developed moderate mucocutaneous bleeding two weeks after a Mycoplasma pneumoniae infection successfully treated with clarithromycin. The patient was referred to us 7 months later for laboratory investigation of the persisting bleeding diathesis. The patient's personal and family histories were negative for bleeding disorders. Complete blood count, von Willebrand Factor levels and coagulation tests were normal; platelet aggregation, ATP secretion, δ-granules content and serum thromboxane B2 levels were defective. At follow-up visits, laboratory parameters and the bleeding diathesis progressively normalized within 2 years. The patient's condition is compatible with a diagnosis of acquired Storage Pool Deficiency (SPD), associated with defective thromboxane A2 production. To our knowledge, this is the first case of acquired, transient SPD with spontaneous remission. The pathogenic role of Mycoplasma pneumoniae infection or clarithromycin is possible, albeit uncertain.

Circumcision bleeding rates in patients diagnosed with delta-storage pool deficiency later in life.

BACKGROUND: Male circumcision is a common procedure, generally performed during the newborn period. Bleeding is an uncommon but feared complication of circumcision. Few reports have evaluated circumcision bleeding rates in patients with bleeding disorders. OBJECTIVES: To study circumcision bleeding rates in male subjects who were diagnosed later in life with delta-storage pool deficiency (δ-SPD). METHODS: We retrospectively reviewed the medical records of male subjects (≤18 years of age) who were circumcised without hemostatic prophylaxis prior to δ-SPD diagnosis from 2000 to 2020. Bleeding rates and severity were the main outcomes evaluated. We collected demographic data, bleeding scores using a validated assessment tool, laboratory data, and platelet electron microscopy results. A descriptive analysis was performed. RESULTS: Twenty-five male subjects were included. The median bleeding score at presentation was 3.5 (range: 2-9). The diagnosis was confirmed using platelet electron microscopy. A value of <2 dense granules/platelet was considered abnormal. Circumcision was performed at a median age of 2 days (range: 1 day to 4 months). None of the subjects had intraoperative or postoperative bleeding. With a bleeding rate of zero, we suggest that newborn males with a family history of δ-SPD may be safely circumcised without hemostatic prophylaxis.

Platelet Electron Microscopy: Utilizing LEAN Methodology to Optimize Laboratory Workflow.

BACKGROUND: Platelet electron microscopy (PEM) is the gold standard methodology for diagnosing storage pool disorder, defined as a paucity of delta granules, alpha granules, or both. PEM literature is limited with few published resources and without well-developed interlaboratory standardization for the preparation and examination of platelet samples. METHODS: Whole mount (WM) dense body (DB) counts for 300 pediatric cases were reviewed to determine whether counting fewer platelets could yield the same results. For 6 cases, DB average was determined on the day of WM preparation and on 2 consecutive days. Both WM and thin section (TS) preparations were examined for all cases. RESULTS: Employing LEAN methodology, an algorithm was developed to markedly decrease the number of platelets counted and still ensure accurate results. WMs decay with time; a statistically significant difference in DB counts was noted between day 0 and day 1 (p < .1). Twelve of 300 cases required both WM and TS preparations for a complete diagnosis. CONCLUSION: It is possible to maintain accuracy and decrease 100 platelet DB counts by >75%. WMs must be counted on the day they are prepared to avoid false paucity of DB secondary to sample decay. An accurate evaluation of platelet morphology requires both the WM and TS techniques.

Weak correlation of bleeding scores to platelet electron microscopy: A retrospective chart review of pediatric patients with delta-storage pool disorder.

BACKGROUND: Delta granule storage pool deficiency (δ-SPD) is a rare platelet disorder in which a deficiency of platelet granules leads to poor aggregation, resulting in varying clinical bleeding phenotypes. Children with δ-SPD have variable laboratory results, making the proper diagnosis and evaluation controversial. OBJECTIVES: To describe the demographic and laboratory trends of this population and to assess the value of electron microscopy in diagnostic evaluation and its correlation to bleeding symptoms. METHODS: We performed a retrospective review of 109 pediatric patients diagnosed with δ-SPD. We collected demographic information and bleeding scores using a validated bleeding assessment tool. A descriptive and exploratory analysis was performed. RESULTS: The majority of patients were female, with an average age at diagnosis of 11.61 years. Females were diagnosed at a significantly older age presenting most often with menorrhagia, while males presented most commonly with epistaxis. The majority showed normal lumiaggregometry, the mean platelet electron microscopy (PEM) value was 2.37, and the mean bleeding score was 6. Bleeding assessment tool and PEM had a significantly weak correlation. CONCLUSIONS: Patients with more dense granules per platelet had higher bleeding scores than those with fewer dense granules per platelet. The current body of evidence does not favor the use of PEM in routine clinical practice, and results are difficult to interpret. In patients with severe mucocutaneous bleeding symptoms and normal platelet aggregation studies, consideration should be given to an alternative diagnosis and further evaluation is warranted.

Variation in Platelet Delta Granules Over Time in Young Women Undergoing Evaluation for Heavy Menstrual Bleeding.

Delta-granule platelet storage pool deficiency (δ-PSPD) is a qualitative platelet function defect associated with variable bleeding phenotypes. Platelet electron microscopy (EM) is commonly utilized to evaluate for δ-PSPD, but intrapatient variability in platelet δ-granule numbers by EM is currently unknown. Fifteen young women aged 11 to 17 years presenting to a young women's hematology clinic for the evaluation of heavy menstrual bleeding underwent platelet EM testing at their initial hematology clinic visit and at 1 and 3 months later. Platelet aggregation of platelet-rich plasma by light transmission was also performed on all patients at their initial visit. Eight patients had average δ-granules per platelet consistently ≥2. Three patients were found to have average δ-granules per platelet <2 on initial testing, 2 of which reverted to ≥2 on subsequent testing. When initial average δ-granules per platelet was ≥2, initial repeat testing remained so in 83% (95% confidence interval [CI], 52%-98%) of cases and subsequent repeat testing remained so in 75% (95% CI, 43%-95%) of the cases. Platelet aggregation testing was abnormal in 53% of patients, and there was no apparent correlation between platelet EM findings and platelet aggregation testing. In this small group of young women presenting for the evaluation of bleeding symptoms, we found that almost half of the patients had substantial variability in platelet EM results. Given other identified limitations in platelet EM testing, and the intrapatient variability identified in this study, providers should use caution in utilizing EM in isolation to diagnose δ-PSPD.

Combined alpha-delta platelet storage pool deficiency is associated with mutations in GFI1B.

Combined alpha-delta platelet storage pool deficiency is characterized by the absence or reduction in the number of both alpha granules and dense bodies. This disorder can have variable severity as well as a variable inheritance pattern. We describe two patients from unrelated families with combined alpha-delta storage pool deficiency due to mutations in GFI1B, a zinc finger protein known to act as a transcriptional repressor of various genes. We demonstrate that this disease is associated with either a heterozygous mutation (de novo or familial) abrogating the binding of the zinc fingers with the promoter of its target genes, or by hypomorphic biallelic mutations in GFI1B leading to autosomal recessive inheritance.

Clinical and Laboratory Findings in Patients with δ-Storage Pool Disease: A Case Series.

Platelet δ-storage pool disease (δ-SPD) is a platelet function disorder characterized by a reduction in the number or content of dense granules. Reports on δ-SPD are mostly limited to case presentations. We aimed to retrospectively describe a series of patients with δ-SPD to better characterize the disease. We studied 16 patients with congenital or acquired δ-SPD. Lumiaggregometry, α- and δ-granules content, platelet ultrastructure, αIIbß3 integrin, and glycoprotein Ib (GPIb) activation were assessed. Most of the patients generally demonstrate mild to moderate bleeding diathesis. Platelet aggregation studies showed moderate abnormalities with variable profiles, while all the individuals had almost complete absence of adenosine triphosphate release. Mepacrine capture, CD63 expression, and study of dense granules by electron microscopy enabled to distinguish different subtypes of δ-SPD with quantitative or qualitative defect. Surprisingly, significantly decreased GPIb expression levels after platelet activation with thrombin receptor activating peptide 50 µM were found, suggesting that GPIb-impaired mobilization may represent an additional feature of the disorder. In conclusion, δ-SPD represents a complex disorder with various clinical and biological aspects, requiring a great deal of expertise to be properly diagnosed.

Storage pool diseases illuminate platelet dense granule biogenesis.

Platelet dense granules (DGs) are membrane bound compartments that store polyphosphate and small molecules such as ADP, ATP, Ca2+, and serotonin. The release of DG contents plays a central role in platelet aggregation to form a hemostatic plug. Accordingly, congenital deficiencies in the biogenesis of platelet DGs underlie human genetic disorders that cause storage pool disease and manifest with prolonged bleeding. DGs belong to a family of lysosome-related organelles, which also includes melanosomes, the compartments where the melanin pigments are synthesized. These organelles share several characteristics including an acidic lumen and, at least in part, the molecular machinery involved in their biogenesis. As a result, many genes affect both DG and melanosome biogenesis and the corresponding patients present not only with bleeding but also with oculocutaneous albinism. The identification and characterization of such genes has been instrumental in dissecting the pathways responsible for organelle biogenesis. Because the study of melanosome biogenesis has advanced more rapidly, this knowledge has been extrapolated to explain how DGs are produced. However, some progress has recently been made in studying platelet DG biogenesis directly in megakaryocytes and megakaryocytoid cells. DGs originate from an endosomal intermediate compartment, the multivesicular body. Maturation and differentiation into a DG begins when newly synthesized DG-specific proteins are delivered from early/recycling endosomal compartments. The machinery that orchestrates this vesicular trafficking is composed of a combination of both ubiquitous and cell type-specific proteins. Here, we review the current knowledge on DG biogenesis. In particular, we focus on the individual human and murine genes encoding the molecular machinery involved in this process and how their deficiencies result in disease.

Usefulness of Flow Cytometric Mepacrine Uptake/Release Combined with CD63 Assay in Diagnosis of Patients with Suspected Platelet Dense Granule Disorder.

Dense granule disorder is one of the most common platelet abnormalities, resulting from dense granule deficiency or secretion defect. This study was aimed to evaluate the clinical usefulness of the flow cytometric combination of mepacrine uptake/release assay and CD63 expression detection in the management of patients with suspected dense granule disorder. Over a period of 5 years, patients with abnormal platelet aggregation and/or reduced adenosine triphosphate (ATP) secretion suggestive of dense granule disorder were consecutively enrolled. The flow cytometric assays were systematically performed to further investigate dense granule functionality. Among the 26 included patients, 18 cases showed impaired mepacrine uptake/release and reduced CD63 expression on activated platelets, consistent with δ-storage pool deficiency (SPD). Another seven patients showed decrease in mepacrine release and CD63 expression but mepacrine uptake was normal, indicating secretion defect rather than δ-SPD. Unfortunately, ATP secretion could not be measured in 7 out of the 26 patients due to insufficient sample and/or severe thrombocytopenia. This test combination provides a rapid and effective method to detect the heterogeneous abnormalities of platelet dense granule by distinguishing between storage and release defects. This combination is particularly advantageous for severely thrombocytopenic patients and pediatric patients in which only minimal sample is required.

Assessment and validation of a defined fluid restriction protocol in the use of subcutaneous desmopressin for children with inherited bleeding disorders.

INTRODUCTION: Despite the availability of subcutaneous desmopressin (1-deamino-8-d-arginine vasopressin, SC-DDAVP) as a haemostatic agent for children with mild bleeding disorders, few publications specifically address the safety or efficacy of this mode of administration. AIM: Our aim was to assess whether a defined fluid restriction protocol was effective in preventing hyponatremia in children receiving perioperative SC-DDAVP, and to document adequate biological and clinical response in this setting. METHODS: We retrospectively analysed a cohort of children with mild bleeding disorders prescribed SC-DDAVP over a 5-year period following institution of a 'two-thirds maintenance' fluid restriction protocol. RESULTS: Sixty-nine patients received SC-DDAVP following this protocol, including 15 with mild haemophilia A, 49 with von Willebrand disease (VWD) and five with platelet storage pool disorder. In patients who underwent formal preoperative assessment a complete or partial response was observed in 28/29 with type 1 VWD and 14/15 with mild haemophilia A. Perioperative SC-DDAVP provided excellent haemostasis in all patients, with no requirement for factor concentrate or blood products. Mild asymptomatic hyponatremia was detected in seven children who received multiple doses of DDAVP (lowest sodium 129 mmol L(-1) ); however, adherence to the prescribed fluid restriction protocol was questionable in six of these cases. Symptomatic hyponatremia was not observed. CONCLUSION: Subcutaneous desmopressin was well-tolerated, with no serious side-effects observed, and good biological responses in preoperative trials. A two-thirds maintenance fluid regimen was effective at preventing symptomatic hyponatremia in our cohort, and is now the standard protocol for fluid restriction post-DDAVP administration in our centre.

Light Transmission Aggregometry Does Not Correlate With the Severity of δ-Granule Platelet Storage Pool Deficiency.

Delta-granule platelet storage pool deficiency (δ-PSPD) is a poorly studied bleeding diathesis resulting from either decreased granule content or decreased average number of platelet δ-granules. Light transmission aggregometry (LTA) is commonly used to evaluate for δ-PSPD and platelet electron microscopy (EM) is used to confirm the diagnosis. Currently, little data exist examining the relationship between the likelihood of abnormal platelet aggregation findings, severity of δ-granule deficiency on platelet EM, and severity of bleeding symptoms in patients with δ-PSPD. Patients diagnosed with δ-PSPD by platelet EM who also underwent LTA testing were identified at a single institution for correlation between severity of bleeding, average number of platelet δ-granules, and number of agonist abnormalities on LTA. No statistically significant association was identified between the average number of δ-granules per platelet and likelihood of an abnormal LTA. LTA abnormalities were quite varied and only 50% diagnosed with δ-PSPD on EM had abnormal aggregation testing. Also, no correlation was seen between the number of clinical bleeding symptoms, number of average δ-granules per platelet, and the number of LTA agonist abnormalities. Our findings highlight the difficulties inherent in the laboratory evaluation of platelet function.

Platelets: still a therapeutical target for haemostatic disorders.

Platelets are cytoplasmatic fragments from bone marrow megakaryocytes present in blood. In this work, we review the basis of platelet mechanisms, their participation in syndromes and in arterial thrombosis, and their potential as a target for designing new antithrombotic agents. The option of new biotechnological sources is also explored.

Molecular determinants of platelet delta storage pool deficiencies: an update.

Delta storage pool deficiency (δ-SPD) is a rare heterogeneous group of platelet disorders characterized by a reduction in the number or content of dense granules. δ-SPD causes a mild to moderate bleeding diathesis characterized mainly by mucocutaneous bleeding. Currently, no specific treatment is available and the therapeutic approach is based on prevention of excessive bleeding. However, during the last few years, important insights into the pathophysiology of δ-SPD have been achieved using mouse models and dense granule deficiency-associated congenital diseases, such as Hermansky-Pudlak syndrome and Chediak-Higashi syndrome. It thus appears that δ-SPD represents a genetically heterogeneous group of intracellular vesicle biogenesis and/or trafficking disorders. This review summarizes recent data regarding the molecular mechanisms together with clinical features of the different types of δ-SPD. Although the molecular basis of isolated inherited δ-SPD remains currently unknown, next-generation sequencing strategies should enable researchers to identify the causative genes. Identification of those genes should contribute to our understanding of the pathophysiology, represent useful tools for genetic diagnosis, and eventually lead to new specific therapeutic approaches.

Down-regulation of the RUNX1-target gene NR4A3 contributes to hematopoiesis deregulation in familial platelet disorder/acute myelogenous leukemia.

RUNX1 encodes a DNA-binding α subunit of the core-binding factor, a heterodimeric transcription factor. RUNX1 is a master regulatory gene in hematopoiesis and its disruption is one of the most common aberrations in acute leukemia. Inactivating or dominant-negative mutations in the RUNX1 gene have been also identified in pedigrees of familial platelet disorders with a variable propensity to develop acute myeloid leukemia (FPD/AML). We performed analysis of hematopoiesis from 2 FPD/AML pedigrees with 2 distinct RUNX1 germline mutations, that is, the R139X in a pedigree without AML and the R174Q mutation in a pedigree with AML. Both mutations induced a marked increase in the clonogenic potential of immature CD34(+)CD38(-) progenitors, with some self-renewal capacities observed only for R174Q mutation. This increased proliferation correlated with reduction in the expression of NR4A3, a gene previously implicated in leukemia development. We demonstrated that NR4A3 was a direct target of RUNX1 and that restoration of NR4A3 expression partially reduced the clonogenic potential of patient progenitors. We propose that the down-regulation of NR4A3 in RUNX1-mutated hematopoietic progenitors leads to an increase in the pool of cells susceptible to be hit by secondary leukemic genetic events.

Menstrual bleeding patterns and prevalence of bleeding disorders in a multidisciplinary adolescent haematology clinic.

Heavy menstrual bleeding (HMB) is a frequent complaint in adolescence. Although HMB is often caused by immaturity of the hypothalamic-pituitary-ovarian axis, bleeding disorders are another common yet often unidentified cause. The aim of this study was to examine the bleeding patterns and prevalence of inherited bleeding disorders among females referred for HMB to a multidisciplinary adolescent haematology clinic. We retrospectively reviewed the first 105 patients (ages 8-18 years) referred to this specialty clinic from February 2009 to December 2011. Using menstrual bleeding questionnaires and medical records, data were extracted regarding demographics, bleeding patterns, frequency and types of bleeding disorders identified, and prescribed interventions. Sixty-two per cent of patients were diagnosed with a bleeding disorder, including platelet storage pool deficiency (36%), von Willebrand's disease (9%), other platelet function defect (8%), Ehlers-Danlos syndrome (7%) and combined bleeding disorders (2%). Comparison of the bleeding profiles for females with and without a bleeding disorder revealed only three factors that were significantly different, including the reported regularity of patients' periods (P = 0.02), description of period flow (P = 0.04) and number of days of each period that the bleeding was described as 'heavy' (P = 0.007). Bleeding disorders are prevalent in adolescent females presenting to a specialty clinic. Specifically, a relatively high proportion of adolescents were diagnosed with platelet storage pool deficiency. In our small population, menstrual bleeding profiles, as examined by a standardized questionnaire, could not identify females with an underlying bleeding disorder, demonstrating the important role of haemostasis testing in the evaluation of adolescents with HMB.

The Medich giant platelet syndrome: two new cases.

Hypogranular platelet disorders in human subjects are relatively rare. They include the gray platelet syndrome, αδ storage pool deficiency, the Hermansky-Pudlak syndrome, and the white platelet syndrome. Perhaps the rarest of them all is the Medich giant platelet disorder. No additional cases of this condition have been reported since description of the first case in 2004. This study describes two children with thrombocytopenia and giant, hypogranular platelets found shortly after birth. Electron microscopic study of their platelets revealed sheets of membrane wrapped into tubes resembling scrolls. The scroll-like structures were open at both ends and often filled with glycogen particles. The abnormal structures are identical to those found in the initial case. As a result, the disorder can now be referred to as the Medich giant platelet syndrome.