Genetic variants associated with Hermansky-Pudlak syndrome.

Hermansky-Pudlak syndrome (HPS) is a rare autosomal recessive disorder characterized by defective biogenesis of lysosome-related organelles. Clinical manifestations include a bleeding diathesis due to a platelet delta storage pool deficiency, oculocutaneous albinism, inflammatory bowel disease, neutropenia, and pulmonary fibrosis. Ten genes associated with HPS are identified to date, and each gene encodes a protein subunit of either Biogenesis of Lysosome-related Organelles Complex (BLOC)-1, BLOC-2, BLOC-3, or the Adaptor Protein-3 complex. Several genetic variants and phenotypic heterogeneities are reported in individuals with HPS, who generally exhibit easy bruisability and increased bleeding. Desmopressin, pro-coagulants, or platelet transfusion may be used as prophylaxis or treatment for excessive bleeding in patients with HPS. However, response to desmopressin can be variable. Platelets are effective in preventing or treating bleeding in individuals with HPS, but platelets should be transfused judiciously to limit alloimmunization in patients with HPS who are at risk of developing pulmonary fibrosis and may be potential candidates for lung transplantation. The discovery of new genes associated with HPS in people with excessive bleeding and hypopigmentation of unknown etiology may be facilitated by the use of next-generation sequencing or panel-based genetic testing.

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.

Determination of ATP and ADP Secretion from Human and Mouse Platelets by an HPLC Assay.

BACKGROUND: Secretion of ADP and ATP is an essential prerequisite for platelet aggregation. Impaired nucleotide secretion can cause aggregation defects and increased bleeding risk. Quantitative determination of platelet nucleotide content and exocytosis is thus of importance for the characterization and diagnosis of bleeding phenotypes. For transgenic animal models with hemostatic defects analysis of potential secretion defects is as well imperative. METHODS: Supernatants of washed platelets and platelet-rich plasma were analyzed by HPLC for ADP and ATP concentration. Calibration of the HPLC data was accomplished with an internal standard compensating for loss of analyte, detection sensitivity, and interference of the biomatrix. RESULTS: HPLC analysis of nucleotide secretion was carried out with human and mouse platelets. Detection limits were determined for washed platelet and platelet-rich plasma samples. In the physiological concentration range linearity with respect to the peak area is maintained. CONCLUSION: The method combines reasonable sensitivity with robustness. The internal standard ensures reliable quantification of nucleotide concentrations even in presence of otherwise interfering substances. The low sample consumption renders possible the application to analysis of small samples like in mouse experiments.

Experience With Pre-procedural Hemostatic Medications versus Platelet Transfusion in Patients With Platelet Storage Pool Deficiency.

BackgroundStorage pool deficiency (SPD) is a rare bleeding disorder characterized by reduction in the number of delta granules within platelets, interfering with hemostasis. Current literature lacks well-designed studies from which to draw concrete conclusions regarding pre-procedural management of bleeding complications. Objective: The purpose of this study is to describe bleeding and safety outcomes of SPD patients receiving either pre-procedural platelet transfusions or platelet-sparing regimens. Methods: An exploratory retrospective cohort study was conducted among SPD patients, comparing major bleeding events between those who received platelet transfusion and those who received desmopressin, tranexamic acid, and/or aminocaproic acid within 24 hours prior to procedure. Results: Rates of major bleeding were not found to be higher among patients who received a platelet-sparing regimen [platelet-sparing: 2/25 (8%); platelet transfusion: 2/29 (6.9%); P = .99]. Incidence of non-major bleeding was higher in the platelet transfusion group, but this was not statistically significant [platelet-sparing: 0/25 (0%); platelet transfusion: 3/29 (10.3%); P = .24]. Treatment-related adverse effects were observed following 8 of 54 procedures (14.8%). Conclusion: Use of a platelet-sparing regimen was not associated with a significantly higher incidence of major or non-major bleeding events. Future prospective trials are recommended to compare outcomes between therapies.

Reference Range of Platelet Delta Granules in the Pediatric Age Group: An Ultrastructural Study of Platelet Whole Mount Preparations from Healthy Volunteers.

This study sought to determine delta granule normal ranges for children and to validate methodology for the appropriate diagnosis of delta granule deficiency (storage pool disease) by using the whole-mount technique in electron microscopy. Specimens obtained from 40 healthy volunteers (2 months of age through 21 years old, 21 females and 19 males) were tested. Results showed dense granules/platelet (DG/Plt) ranged from 1.78 to 5.25. The 5th percentile was 1.96 DG/Plt with an overall mean ± SEM 3.07 ± 0.12 DG/Plt. In comparison, a previously published lower cutoff value, 3.68 DG/Plt, was significantly higher than the mean from our volunteers (P < 0.0001). We found no variability in dense granules/platelet based on race or sex and no significant variation by age subgroup. Pending wider studies, the value of 2 DG/Plt is a more appropriate lower limit of normal. In the absence of wider studies (in healthy volunteers and patients), laboratories should consider establishing their own reference ranges.

Serum antibody-negative Goodpasture syndrome with delta granule pool storage deficiency and eosinophilia.

Goodpasture syndrome is a rare, life-threatening autoimmune disease characterized by a triad of rapidly progressive glomerulonephritis, a hemorrhagic pulmonary condition and the presence of anti-glomerular basement membrane (anti-GBM) antibodies. The antibodies initiate destruction of the kidney glomeruli, resulting in a focal necrotizing glomerulitis, which may progress rapidly to renal failure. Autoantibody-mediated damage of alveolar basement membranes leads to diffuse pulmonary hemorrhage, which in some cases may be severe enough to cause respiratory failure. Many clinicians use a variety of assays to detect serum anti-GBM antibodies; however, these tests may be falsely negative in up to 15% of patients with Goodpasture syndrome. Here, we report an unusual case of a 40-year-old man with clinical evidence of Goodpasture syndrome, a negative anti-GBM antibody serum result, eosinophilia and delta granule pool storage deficiency. After a 14-day hospital stay and extensive workup, as well as treatment with antibiotics, steroids and ventilator support for respiratory failure, the patient continued to deteriorate and entered multisystem organ failure. The family decided to withdraw ventilator support, and the patient expired. Immunofluorescence testing for anti-GBM autoantibodies on lung and kidney tissues during an autopsy confirmed the diagnosis of Goodpasture syndrome.