Standard Platelet Research Articles

Pathogen reduction with amotosalen/UVA reduces platelet refractoriness in a dog platelet transfusion model.

Pathogen reduction of donor platelets with amotosalen/UVA has been shown to effectively inactivate pathogens and also contaminating white blood cells (WBCs). We wanted to determine whether WBC inactivation could also decrease alloimmune refractoriness to donor platelets. Platelets were prepared from a donor dog's whole blood, and the platelets were either transfused without modification [standard (STD) platelets] or treated with amotosalen/UVA under conditions modelling the amotosalen/UVA Blood System for human platelets (APR) using either 4 or 3J/cm2 of UVA exposure. Platelets were transfused weekly from a single donor dog for 8weeks or until the recipient dog became refractory to their donor's platelets. Antibody samples were drawn weekly and tested against the donor dog's platelets and WBCs (CD8 and B cells). Only 1/7 (14%) dogs that received STD platelets accepted 8weeks of donor transfusions. Following APR 4J/cm2 donor transfusions, 3/9 (33%) recipients accepted their donor's transfusions, but only one recipient remained antibody negative. Following APR 3J/cm2 donor transfusions, the same dose as used for human platelet transfusions, 7/10 (70%) recipients accepted their donor's transfusions, but only two remained antibody negative. As a very high percentage of recipient dogs (70%) accepted APR 3J/cm2 donor transfusions, these data suggest that preventing alloimmune platelet refractoriness may be another benefit of pathogen reduction using amotosalen/UVA.

Use of a high platelet-to-RBC ratio of 2:1 is more effective in correcting trauma-induced coagulopathy than a ratio of 1:1 in a rat multiple trauma transfusion model

BackgroundPlatelet dysfunction importantly contributes to trauma-induced coagulopathy (TIC). Our aim was to examine the impact of transfusing platelets (PLTs) in a 2:1 PLT-to-red blood cell (RBC) ratio versus the standard 1:1 ratio on transfusion requirements, correction of TIC, and organ damage in a rat multiple trauma transfusion model.MethodsMechanically ventilated male Sprague Dawley rats were traumatized by crush injury to the small intestine and liver and a fracture of the femur, followed by exsanguination until a mean arterial pressure (MAP) of 40 mmHg. Animals were randomly assigned to receive resuscitation in a high PLT dose (PLT to plasma to RBC in a ratio of 2:1:1) or a standard PLT dose (ratio of 1:1:1) until a MAP of 60 mmHg was reached (n = 8 per group). Blood samples were taken for biochemical and thromboelastometry (ROTEM) assessment. Organs were harvested for histopathology.Outcome measures were transfusion requirements needed to reach a pretargeted MAP, as well as ROTEM correction and organ failure.ResultsTrauma resulted in coagulopathy as assessed by deranged ROTEM results. Mortality rate was 19%, with all deaths occurring in the standard dose group. The severity of hypovolemic shock as assessed by lactate and base excess was not different in both groups. The volume of transfusion needed to reach the MAP target was lower in the high PLT dose group compared to the standard dose, albeit not statistically significant (p = 0.054). Transfusion with a high PLT dose resulted in significant stronger clot firmness compared to the standard dose at all time points following trauma, while platelet counts were similar. Organ failure as assessed by biochemical analysis and histopathology was not different between groups, nor were there any thromboembolic events recorded.ConclusionsResuscitation with a high (2:1) PLT-to-RBC ratio was more effective compared to standard (1:1) PLT-to-RBC ratio in treating TIC, with a trend towards reduced transfusion volumes. Also, high PLT dose did not aggravate organ damage. Transfusion strategies using higher PLT dose regiments might be a feasible treatment option in hemorrhaging trauma patients for the correction of TIC.

Impact of cold storage on platelets treated with Intercept pathogen inactivation.

BACKGROUNDPathogen inactivation and cold or cryopreservation of platelets (PLTs) both significantly affect PLT function. It is not known how PLTs function when both are combined.STUDY DESIGN AND METHODSStandard PLT concentrates (PCs) were compared to pathogen‐inactivated PCs treated with amotosalen photochemical treatment (AS‐PCT) when stored at room (RT, 22°C), cold (4°C, n = 6), or cryopreservation (−80°C, n = 8) temperatures. The impact of alternative storage methods on both arms was studied in flow cytometry, light transmittance aggregometry, and hemostasis in collagen‐coated microfluidic flow chambers.RESULTSPlatelet aggregation of cold‐stored AS‐PCT PLTs was 44% ± 11% compared to 57% ± 14% for cold‐stored standard PLTs and 58% ± 21% for RT‐stored AS‐PCT PLTs. Integrin activation of cold‐stored AS‐PCT PLTs was 53% ± 9% compared to 77% ± 6% for cold‐stored standard PLTs and 69% ± 13% for RT‐stored AS‐PCT PLTs. Coagulation of cold‐stored AS‐PCT PLTs started faster under flow (836 ± 140 sec) compared to cold‐stored standard PLTs (960 ± 192 sec) and RT‐stored AS‐PCT PLTs (1134 ± 220 sec). Fibrin formation rate under flow was also highest for cold‐stored AS‐PCT PLTs. This was in line with thrombin generation in static conditions because cold‐stored AS‐PCT PLTs generated 297 ± 47 nmol/L thrombin compared to 159 ± 33 nmol/L for cold‐stored standard PLTs and 83 ± 25 nmol/L for RT‐stored AS‐PCT PLTs. So despite decreased PLT activation and aggregation, cold storage of AS‐PCT PLTs promoted coagulation. PLT aggregation of cryopreserved AS‐PCT PLTs (23% ± 10%) was not significantly different from cryopreserved standard PLTs (25% ± 8%).CONCLUSIONThis study shows that cold storage of AS‐PCT PLTs further affects PLT activation and aggregation but promotes (pro)coagulation. Increased procoagulation was not observed after cryopreservation.

Comparative Analysis of Plateletpheresis Using Different Cell Separators Fenwal Amicus, Fresenius COM.TEC and MCS Plus

Objective: This objective of the study was to compare the three commonly used apheresis instruments available for Plateletpheresis, i.e., MCS Plus, COM.TEC and Amicus in terms of their Pre and Post donor CBC variables, instrument efficacy and product variables. Methodology: Donors undergoing Plateletpheresis are categorized into three groups. Sixty donors were selected according to the selection criteria of donor for Plateletpheresis by AABB. Later the procedure was performed on MCS Plus, Amicus and COM.TEC. Twenty donors were processed with each instrument. Results: The study revealed that there is no significant difference in pre and post count of donors on CBC in all the instruments. It is observed that the blood volume processed in order to have the standard platelet yield of ≥3. 3 × 1011 is higher in the COM.TEC as compared to Amicus and MCS plus (p 3.3 × 1011. But in COM.TEC, 100% of the products have a platelet count of >3.3 × 1011. Products collected from Amicus and COM.TEC is leucodepleted. On the contrary, the products by MCS Plus are not leucodepleted. The collection efficiency is significantly low in MCS Plus (47 ± 13.6) when compares to Amicus (64 ± 7.9) and COM.TEC (59 ± 10.5). However, the collection rate is significantly higher with Amicus (0.07 ± 0.007) followed by COM.TEC (0.06 ± 0.006) and MCS plus (0.04 ± 0.004). Conclusion: The study concludes that all kits for Plateletpheresis are very efficient for platelet collection. However, both Amicus and COM. TEC is better than MCS Plus, as they give leucodepleted platelet concentrates. Amicus supersedes all as the target platelet yield is achieved more rapidly as compared to COM.TEC and MCS plus.

The Usefulness of Improved, Newly Developed Polyolefin Container of Platelet, PO-100, with Higher Oxygen Permeability and Content

The demand for platelet concentrate (PC) has been growing because of the increase of more intensive chemotherapy and hematopoietic stem cell transplantation, and highly invasive surgical operations of several specific areas (i.e. cardiovascular surgery, etc.), which has brought about considerable impact on its decent availability. Furthermore, the short storage period for room temperature stored PC, four days in Japan, five days in worldwide and seven days in the U.S. if they meet several requirements by FDA for the control of the risk of bacterial contamination, has put many PCs become outdated. In response to these requirements, measures have been taken to maximize platelet use, including development of polyolefin containers for platelets that elongate storage time and formulations of higher units.Here, we have developed a polyolefin container, PO-100, which has a 40%/m2 higher oxygen permeability than PO-80, a former, and standard platelet storage bags across the world. Furthermore, PO-100 has two types, one is PO-100/1000 with 1000mL capacity for 4-5 x 1011 platelets content and the other is PO-100/1500 with 1500mL capacity for over 6 x 1011 platelets content.In this study, we evaluated the influence of storage of higher content of platelet in PO-100 on the condition of platelet in the bag. The IRB of Nagasaki University approved this study. After informed consent was obtained from healthy volunteers, single-donor apheresis was performed. Then, platelets were pooled, separated equally into two types of bags, PO-100/1000 (n=6) or PO-100/1500 (n=6), and stored in plasma for up to day 7. We monitored various parameters, which related the quality of PC, in bags over the storage period.The result showed that the in vitro characteristics of platelets; platelet counts, mean platelet volume (MPV), and adenosine diphosphate (ADP)/collagen combined-induced platelet aggregation did not significantly change through 7 days storage period both in PO-100/1000 and PO-100/1500. Furthermore, MPV (fL) (8.63±0.35 vs 8.85±0.58, respectively, p=0.463) and platelet aggregation (%) (81.58±4.45 vs 84.67±1.21, respectively, p=0.118) were equivalent between these two conditions.Although pH gradually declined, it was maintained around 7.0 at day 7 without significant difference in both conditions (7.00±0.10 vs 6.92±0.08, respectively, p=0.063). On the other hand, lactate concentration elevated gradually, but there was no significant difference between two bags at day 7 (133.07±23.70 vs 104.52±15.56, respectively, p=0.097) and the pO2 declined by 30 to 40% from day 0 through day 1 in both conditions, suggesting aerobic metabolism was well maintained with efficient O2 consumption and less lactate accumulation in PO-110 bags regardless of its capacity.Intriguingly, our previous investigation demonstrated that pH and ADP-induced platelet aggregation in PO-80 bags decline steeply, notably after day 3, and values at day 7 were 6.67±0.24 and 56.67±11.34%, respectively, which suggested that higher oxygen permeability might contribute to keep platelets in better condition in PO-100 than PO-80.In conclusion, the present study demonstrated that PO-100 had significantly high utility as a noble container for PC. It was considered that better maintenance of the pH of the medium in PC within the appropriate range through improved higher oxygen permeability and efficient aerobic metabolism resulted in platelets in a better functional state even after 7 days storage. Furthermore, the results in this study might open the possibility of practical application of higher unit products, so that the supply chain of PCs could be conducted favorably. DisclosuresNagai:Kawasumi Laboratories Inc.: Research Funding; Kaneka Corporation: Research Funding; Ono Pharmaceutical Co.Ltd.: Consultancy. Yakushiji:Kawasumi Laboratories Inc.: Employment. Kino:Kawasumi Laboratories Inc.: Employment. Tokunaga:Kawasumi Laboratories Inc.: Employment. Yamaoka:Kawasumi Laboratories Inc.: Employment.

Budget impact of implementing platelet pathogen reduction into the Italian blood transfusion system.

Despite improvements in blood donor selection and screening procedures, transfusion recipients can still develop complications related to infections by known and emerging pathogens. Pathogen reduction technologies (PRT) have been developed to reduce such risks. The present study, developed whithin a wider health technology assessment (HTA) process, was undertaken to estimate the costs of the continuing increase in the use of platelet PRT in Italy. A multidisciplinary team was established to perform the HTA and conduct a budget impact analysis. Quantitative data on platelet use were derived from the 2015 national blood transfusion report and from the Italian Platelets Transfusion Assessment Study (IPTAS). The current national fee of 60 Euro per platelet PRT procedure was used to quantify the costs to the Italian National Health Service (INHS). The analysis adopts a 3-year time-frame. In order to identify the impact on budget we compared a scenario representing an increased use of PRT platelets over time with a control scenario in which standard platelets are used. Progressive implementation of PRT for 20%, 40% and 66% of annual adult platelet doses could generate an increase in annual costs for the INHS amounting to approximately 7, 14 and 23 million Euros, respectively. Use of kits and devices suitable for the treatment of multiple adult platelet doses in one PRT procedure could lower costs. In order to fully evaluate the societal perspective of implementing platelet PRT, the increase in costs must be balanced against the expected benefits (prevention of transfusion-transmissible infections, white cell inactivation, extension of platelet storage, discontinuation of pathogen detection testing). Further studies based on actual numbers of platelet transfusion complications and their societal cost at a local level are needed to see the full cost to benefit ratio of platelet PRT implementation in Italy, and to promote equal treatment for all citizens.

A Useful Device to Measure Kinetics of Platelet Contraction.

Platelet contraction provides a minimally invasive source for physiologic information. In this article, we describe a device that directly measures the kinetics of platelet contraction. Whole blood is injected between acrylic plates and an adherent clot forms. The bottom plate is fixed, and the top plate is attached to a wire cantilever. Platelet contraction drives deflection of the wire cantilever which is captured by a camera. Force generated by the clot with time is derived using beam equations. Force derivations were verified using a microelectromechanical (MEMS) force sensor. Kinetics of clot contraction were defined, including maximum contraction force (FMAX), lift-off time (TLIFTOFF), and contraction rate (CR). Metrics were compared with optical aggregometry and thromboelastography. FMAX correlates with optical aggregometry maximal amplitude with a Spearman's rho of 0.7904 and p = 0.0195 and thromboelastography maximal amplitude with a Spearman's rho of 0.8857 and p = 0.0188. Lift-off time correlates with optical aggregometry lag time with a Spearman's rho of 0.9048 and p = 0.002. This preliminary study demonstrates the repeatability of a useful platelet contraction device and its correlation with thromboelastography and optical aggregometry, the gold standard platelet function test.

Population Pharmacokinetic/Pharmacodynamic Analyses of Avatrombopag in Patients With Chronic Liver Disease and Optimal Dose Adjustment Guide With Concomitantly Administered CYP3A and CYP2C9 Inhibitors.

Avatrombopag, a c-Mpl agonist, has been developed to provide an alternative therapy to standard platelet transfusion care for the treatment of thrombocytopenia. The main objectives of this article were to describe the pharmacokinetics (PK) of avatrombopag, to characterize the pharmacokinetic/pharmacodynamic (PK/PD) relationship between plasma avatrombopag concentrations and platelet count, and to identify potential intrinsic and extrinsic factors affecting PK or PK/PD in patients with chronic liver disease (CLD). Platelet count following avatrombopag administration with and without concomitant medication was further simulated using the final population PK/PD model to explore potential dose adjustments. Avatrombopag PK was described by a 1-compartment model with combined first- and zero-order absorption and linear elimination. The relationship between the plasma avatrombopag concentrations and platelet count was well described by a 6-compartment life-span model with a linear drug effect. The final PK and PK/PD models included statistically significant but not clinically relevant effects of body weight and CLD on apparent volume distribution and East Asian ethnicity, albumin, and thrombopoietin level on the slope parameter in the PK/PD relationship. PK/PD simulations showed comparable elevation in platelet count with and without concomitant cytochrome P450 (CYP) 3A and CYP2C9 inhibitors for the dosing regimens of 40 and 60 mg for 5 days, with predictions of <10% of CLD patients exceeding platelet count >200 × 109 /L. Dose adjustment is therefore not necessary with concomitant use of CYP3A and CYP2C9 interacting drugs considering the limited treatment duration (ie, 5 days) and lack of significant safety concerns in CLD patients.

Pathogen-reduced platelets for the prevention of bleeding.

Blood for transfusion is collected from donors and then processed and stored as bags of different blood components. One of these components is platelets. Platelets are cells which help the body form clots and prevent bleeding. As for all transfusions, there are risks related to giving platelets to patients, including a small risk of transfusion-transmitted infections. A number of methods are used to minimise the risk of transfusion-transmitted infections, including careful selection of donors and rigorous donor testing. One new method of preventing infection is pathogen reduction by which, through a process of adding chemicals to the donated platelets and exposing them to a wavelength of ultraviolet light, the number of infecting organisms can be reduced. The aim of this review was to assess whether these specially treated, pathogen-reduced platelets, work as effectively as normal platelets when transfused. Specifically, can they stop or prevent bleeding as well as standard platelets, do they produce the same increase in platelet count and does their use affect further transfusion requirements? Also, this review assessed whether pathogen-reduced platelets are as safe as normal platelets - for example, are they associated with any difference in the rate of death following transfusion and are there any side effects associated with the use of these products? Ten randomised controlled trials, involving 1422 patients, were included assessing two different pathogen-reduced platelet transfusion products, Intercept® and Mirasol® platelets, as compared with standard platelets. Nine trials assessed Intercept® platelets and one trial Mirasol® platelets. The 10 trials had different designs with varying methods of outcome assessment and durations of follow-up. Bleeding was assessed as 'any bleeding', 'clinically significant' and 'severe' at short (up to 48 hours) or long (more than seven days) follow-up periods. There was no difference in 'clinically significant' or 'severe bleeding', mortality, transfusion reactions or adverse events between pathogen-reduced and standard platelets. However, there were significantly poorer responses following the transfusion of pathogen-reduced platelets with a requirement for more platelet transfusions. Due to the limitations of the trial sizes and their designs, there is not enough evidence available to be sure that pathogen-reduced platelets work as effectively as standard platelets to prevent bleeding.

Clinical effectiveness of platelets in additive solution treated with two commercial pathogen-reduction technologies.

Two noninferiority, randomized, controlled trials were conducted in parallel comparing the safety and efficacy of platelets treated with Intercept or Mirasol pathogen-reduction technologies versus standard platelets. The primary endpoint was the percentage of hematology patients who developed World Health Organization Grade 2 or greater bleeding. A noninferiority margin of 11% was chosen based on expected Grade 2 or greater bleeding in 20% of controls. The study was closed for financial restrictions before reaching the planned sample size of 828 patients, and an intention-to-treat analysis was conducted on 424 evaluable patients. In the Intercept trial (113 treated vs. 115 control patients), the absolute risk difference in Grade 2 or greater bleeding was 6.1%, with an upper one-sided 97.5% confidence limit of 19.2%. The absolute risk difference in the Mirasol trial (99 treated vs. 97 control patients) was 4.1%, and the upper one-sided 97.5% confidence limit was 18.4%. Neither absolute risk difference was statistically significant. In both trials, posttransfusion platelet count increments were significantly lower in treated versus control patients. Mean blood component use in treated patients versus controls was 54% higher (95% confidence interval, 36%-74%; Intercept) and 34% higher (95% confidence interval, 16%-54%; Mirasol) for platelets and 23% higher (95% confidence interval, 8%-39%; Intercept) and 32% higher (95% confidence interval, 10%-57%; Mirasol) for red blood cells. Unexpected reactions and adverse events were not reported. Mortality did not differ significantly between treated and control patients. Although conclusions on noninferiority could not be drawn due to low statistical power, the study provides additional information on the safety and efficacy of pathogen-reduced platelets treated with two commercial pathogen-reduction technologies.

HLA‐stripped platelets: preclinical developments and clinical outlooks

Antibodies to foreign HLA class I can form in patients exposed to allogeneic cells, for example during blood transfusion. During platelet transfusions, these anti‐HLA class I antibodies can bind to transfused platelets and mediate their destruction, causing platelet refractoriness. Providing HLA‐matched platelets is a therapeutic option for immunized patients. However, sometimes HLA‐matched donors cannot be found or are too few, leading to suboptimal treatment and risks of bleedings. An alternative is to strip standard platelets of HLA class I using a brief incubation at low pH. Such HLA‐stripping results in a reduction in HLA class I expression at the surface, followed by loss of reactivity to HLA class I‐specific antibodies. HLA‐stripped platelets therefore have therapeutic potential in platelet refractory patients. In this article, we summarize our recent work on HLA stripping of platelets and discuss how the HLA‐stripping process affects platelet function and interactions with cells of the immune system. We also review earlier clinical case studies in which HLA‐stripped platelets have been used In patients, and discuss concerns and considerations that should be taken into account when testing their efficacy in platelet refractory patients in clinical trials.

Platelet-Rich Plasma Powder: A New Preparation Method for the Standardization of Growth Factor Concentrations

Background: Platelet-rich plasma (PRP) is widely used in sports medicine. Available PRP preparations differ in white blood cell, platelet, and growth factor concentrations, making standardized research and clinical application challenging. Purpose: To characterize a newly standardized procedure for pooled PRP that provides defined growth factor concentrations. Study Design: Controlled laboratory study. Methods: A standardized growth factor preparation (lyophilized PRP powder) was prepared using 12 pooled platelet concentrates (PCs) derived from different donors via apheresis. Blood samples and commercially available PRP (SmartPrep-2) served as controls (n = 5). Baseline blood counts were analyzed. Additionally, single PCs (n = 5) were produced by standard platelet apheresis. The concentrations of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), platelet-derived growth factor AB (PDGF-AB), transforming growth factor β1 (TGF-β1), insulin-like growth factor 1 (IGF-1), interleukin (IL)–1α, IL-1β, and IL-1 receptor agonist (IL-1RA) were analyzed by enzyme-linked immunosorbent assay, and statistical analyses were performed using descriptive statistics, mean differences, 95% CIs, and P values (analysis of variance). Results: All growth factor preparation methods showed elevated concentrations of the growth factors VEGF, bFGF, PDGF-AB, and TGF-β1 compared with those of whole blood. Large interindividual differences were found in VEGF and bFGF concentrations. Respective values (mean ± SD in pg/mL) for whole blood, SmartPrep-2, PC, and PRP powder were as follows: VEGF (574 ± 147, 528 ± 233, 1087 ± 535, and 1722), bFGF (198 ± 164, 410 ± 259, 151 ± 99, and 542), PDGF-AB (2394 ± 451, 17,846 ± 3087, 18,461 ± 4455, and 23,023), and TGF-β1 (14,356 ± 4527, 77,533 ± 13,918, 68,582 ± 7388, and 87,495). IGF-1 was found in SmartPrep-2 (1539 ± 348 pg/mL). For PC (2266 ± 485 pg/mL), IGF-1 was measured at the same levels of whole blood (2317 ± 711 pg/mL) but was not detectable in PRP powder. IL-1α was detectable in whole blood (111 ± 35 pg/mL) and SmartPrep-2 (119 ± 44 pg/mL). Conclusion: Problems with PRP such as absent standardization, lack of consistency among studies, and black box dosage could be solved by using characterized PRP powder made by pooling and lyophilizing multiple PCs. The new PRP powder opens up new possibilities for PRP research as well as for the treatment of patients. Clinical Relevance: The preparation of pooled PRP by means of lyophilization may allow physicians to apply a defined amount of growth factors by using a defined amount of PRP powder. Moreover, PRP powder as a dry substance with no need for centrifugation could become ubiquitously available, thus saving time and staff resources in clinical practice. However, before transferring the results of this basic science study to clinical application, regulatory issues have to be cleared.

Management of Anticoagulation in Patients with Hematologic Malignancy Undergoing Autologous Hematopoietic Cell Transplantation

▪Introduction: Venous thromboembolism (VTE) is a significant cause of morbidity in patients with hematologic malignancy who undergo hematopoietic cell transplantation (HCT); however, clinically significant bleeding is not uncommon in this setting and anticoagulation is often contraindicated during prolonged periods of severe thrombocytopenia. This study aims to determine the relative risks of continuing versus temporarily withholding therapeutic anticoagulation during periods of chemotherapy-induced thrombocytopenia in patients who undergo autologous HCT.Methods: Adult patients with hematologic malignancies who underwent first autologous HCT at our institution between 2006 and 2015 were selected. Among those, patients with VTE (as identified by ICD9 code and confirmed by chart review) that occurred in the year preceding transplant were selected as the study population. Patients were allocated into two cohorts at the onset of thrombocytopenia based on whether they continued anticoagulation using a high platelet transfusion threshold (50k), or whether they temporarily withheld anticoagulation using a standard platelet transfusion threshold (10k) and restarted treatment upon platelet engraftment. Patient characteristics and VTE risk factors were captured, depicted as number (percentage) and median (interquartile range), and compared using Fisher's exact and rank sum tests.Primary outcomes included rate of VTE extension/recurrence (assessed by appropriate imaging modality) and major bleeding (defined as grade 3 or 4 bleeding by WHO criteria) by 30 days. Secondary outcomes included clinical bleeding (grade 2-4), overall mortality, median number of platelet and red blood cell (RBC) transfusion, median time to neutrophil and platelet engraftment. Pre-defined exploratory subgroup analysis was performed. Logistic regression, Cox regression, and linear regression models without adjustment were generated for outcomes where a P value of < 0.05 was considered significant (Stata 14.1).Results: Among 1,631 patients who underwent first autologous HCT between 2006 and 2015, 204 patients developed VTE (12.5%) in the year preceding transplant. The median duration of thrombocytopenia, defined as the period during which the platelet < 50k, was 12 days. The median follow-up period was 359 days and only 4% of patients were lost to follow up prior to 30 days. In addition to routine clinical assessment for VTE and bleeding, repeat imaging surveillance was done in 55% of patients prior to discharge from the transplant service. Except for the timing of prior VTE occurrence, there were no significant differences in baseline characteristics between the cohort that continued anticoagulation (N=132) and the cohort that discontinued anticoagulation (N=72) (Table 1).There were no significant differences in the rate of VTE extension/recurrence or major bleeding between the treatment groups by 30 days (Table 2). The rates of VTE in both groups were low (1 new pulmonary embolism and 2 new catheter-associated thromboses). There was 1 fatal spontaneous retroperitoneal bleeding (grade 4) in the cohort that continued anticoagulation. The number of platelet transfusions was significantly higher in the patients who continued anticoagulation. Further comparison of the 2 cohorts by different subgroups did not reveal significant interactions (Figure 1, all interaction P values > 0.300).Conclusions: In patients undergoing HCT for hematologic malignancy who also have VTE, continuation of anticoagulation (compared to temporary cessation) during chemotherapy-induced thrombocytopenia is associated with increased platelet utilization but no significant difference in the rate of VTE extension/recurrence. The low rate of recurrent thrombosis among patients who discontinued anticoagulation during pre-engraftment thrombocytopenia suggests that temporary interruption of anticoagulation may be the better option for selected patients who face this difficult clinical situation. [Display omitted] [Display omitted] [Display omitted] DisclosuresGopal:Seattle Genetics: Research Funding. Garcia:Daiichi Sankyo: Consultancy, Research Funding; Pfizer: Consultancy; Boehringer Ingelheim: Consultancy; BMS: Consultancy; Janssen: Consultancy, Research Funding.

Effects of various antiplatelet drugs on ex vivo platelet activation induced by equine herpesvirus type 1.

OBJECTIVE To evaluate the effects of treatment of horses with standard platelet inhibitors on ex vivo inhibition of platelet activation by equine herpesvirus type I (EHV-I). ANIMALS II healthy adult horses. PROCEDURES In a double-blinded, placebo-controlled crossover study, horses were treated orally for 5 days with theophylline (5 mg/kg, q 12 h), pentoxifylline (10 mg/kg, q 12 h), clopidogrel bisulfate (4 mg/kg, q 24 h), acetylsalicylic acid (20 mg/kg, q 24 h), or placebo. Horses received all treatments, each separated by a 3-week washout period. Platelet-rich plasma was prepared from citrated blood samples obtained before each treatment session and 4 hours after each final drug dose. Platelets were exposed to 2 EHV-I strains (at I plaque forming units/cell) or positive (thrombin-convulxin) and negative control substances for 10 minutes, then platelet activation was assessed by determining the percentages of P-selectin-positive platelets and platelet-derived microparticles (PDMPs; small events positive for annexin V) with flow cytometry. Platelet aggregation in response to 10μM ADP was also assessed. RESULTS No significant differences in median percentages of P-selectin-positive platelets and PDMPs in EHV-I-exposed platelets were identified between measurement points (before and after treatment) for all drugs, nor were differences identified among drugs at each measurement point. Only clopidogrel significantly inhibited platelet aggregation in response to ADP in platelet-rich plasma samples obtained after that treatment session. CONCLUSIONS AND CLINICAL RELEVANCE Treatment of horses with standard platelet inhibitors had no effect on EHV-I-induced platelet α-granule exteriorization or microvesiculation and release of PDMPs ex vivo, suggesting these drugs will not prevent platelet activation induced directly by EHV-I in vivo.

Practice patterns in the diagnosis of inherited platelet disorders within a single institution.

: The diagnosis of inherited platelet disorders (IPDs) is challenging with variable diagnostic practices existing between institutions. To determine patterns and utility of diagnostic testing practices for IPDs within a single institution, a retrospective cohort study was performed. Records of 50 patients (50% women), median age 32 years (1 day to 81 years) were analyzed. In total, 28 (53%) had a positive International Society of Thrombosis and Hemostasis Bleeding Assessment Tool score. Test-ordering patterns were highly variable. All patients had platelet morphology analysis by light microscopy. In total, 42 (84%) underwent light transmission aggregometry, 43 (86%) platelet function analyzer, 37 (74%) platelet electron microscopy, 25 (50%) flow cytometry, and 15 (30%) genetic testing. Platelet function analyzer and light transmission aggregometry were always used as first-order tests, followed by platelet transmission electron microscopy and flow cytometry (81 and 84%, respectively). Genetic testing was obtained up front in five cases (33% of orders), mostly in patients with syndromic thrombocytopenia or in the setting of a known genetic disorder. Test-ordering practices did not adhere to published algorithms. Even within a single institution, great heterogeneity exists in the testing approach to IPDs. Although, a large proportion of cases were studied with platelet transmission electron microscopy and flow cytometry, standard platelet assays established the diagnosis in a great majority. Standardization of testing practices, first beginning at the institutional level is a much needed step forward.

Dynamic platelet function on von Willebrand factor is different in preterm neonates and full‐term neonates: changes in neonatal platelet function

Background Very low birth weight (VLBW) preterm neonates have an increased risk of hemorrhage-related morbidity and mortality as compared with their full-term counterparts. It is unclear whether platelet function differs between preterm and full-term neonates. This is partly because of the large volumes of blood required to perform standard platelet function tests, and the difficulty in obtaining such samples in neonates. Objectives This study was designed to characterize platelet behavior in neonates with a physiologic flow-based assay that quantifies platelet function in microliter volumes of blood under arterial shear. Methods Blood from VLBW preterm neonates of ≤ 32 weeks' gestation (n = 15) and full-term neonates (n = 13) was perfused under arterial shear over surface-immobilized von Willebrand factor (VWF). Platelet behavior was recorded by digital-image microscopy and analyzed. Surface expression of platelet glycoprotein (GP) Ibα and GPIIIa of VLBW preterm and full-term neonates was also measured. Results VLBW preterm neonates had increased numbers of platelets interacting with VWF, and increased GPIbα expression on the platelet surface. Despite the increased numbers of VWF interactions as reflected by flow-driven platelet translocation along the protein surface, no significant differences were observed in the numbers of platelets that adhered in a stationary fashion to VWF. Platelets from VLBW preterm neonates and those from full-term neonates behaved differently on VWF. Conclusions These differences in platelet function may contribute to the higher incidence of bleeding observed in VLBW preterm neonatal populations, or may represent a compensatory mechanism to counteract this risk of bleeding.

Platelet Transmission Electron Microscopy and Flow Cytometry in the Diagnosis of Congenital/Hereditary Qualitative or Quantitative Platelet Disorders

BackgroundCongenital/hereditary qualitative platelet disorders (CQPD) can be classified by platelet count (eg, normal or decreased) and platelet size as well as by functional defect. Based on mean platelet volume (MPV), congenital thrombocytopenias (CT) are classified as macro-thrombocytopenias (eg, Bernard-Soulier syndrome [BSS], MYH-9 [myosin heavy chain 9, non-muscle]-associated thrombocytopenias, gray platelet syndrome [GPS], etc.); microthrombocytopenias (eg, Wiskott-Aldrich syndrome and variants); and those with normal platelet size. CQPDs with normal platelet count include Glanzmann thrombasthenia (GT), dense granule storage pool disorders (DG-SPD) including Hermansky-Pudlak syndrome (HPS), platelet secretion disorders and many others. Diagnosis has relied on platelet counting and sizing, blood smear light microscopy, platelet aggregation (agg) and platelet function analyzer (PFA-100) studies. Platelet transmission electron microscopy (PTEM) and flow cytometry can provide supplementary information and molecular analysis is also evolving. The objective of this study was to determine the roles of platelet transmission electron microscopy (PTEM) and flow cytometry in the diagnosis of CQPD and CT.MethodsIn this retrospective cohort study, after IRB approval, the electronic medical record system was queried for patients (pt) with a confirmed CQPD or CT seen at Mayo Clinic between 2000 and 2015. A detailed chart review was undertaken.Results54 pt (64% female) met our study criteria; median age was 32 years (range 1 day to 81 years). 18/54 (33%) pt had macrothrombocytopenia: BSS 4 (7%); MYH-9 10 (19%); gray platelet syndrome 4 (7%). 36/54 (66%) had normal MPV: Glanzmann thrombasthenia 6 (11%); storage pool disorders including DG-SPD 10 (19%) and HPS 2 (3%), mild alpha granule deficiency 3 (6%) and York platelet syndrome 4 (7%); platelet secretion disorders 4 (7%); ANKRD26 mutation 3 (6%); congenital amegakaryocytic thrombocytopenia 1 (2%); GATA-1 mutation 1 (2%); RUNX-1 mutation 1 (2%); and Jacobsen syndrome 1 (2%). 44 pt (81%) had a positive bleeding history: epistaxis 52%, cutaneous bleeding 57%, gastrointestinal bleeding 21% and menorrhagia 54%; 10 pt (19%) had a negative bleeding history. Results of standard and esoteric platelet testing are summarized in Table 1.Diagnosis in the 10 pt with glycoprotein deficiency (BSS or GT) was established by agg; flow cytometry was confirmatory but not necessary. Of the 23 pt with SPD, 5 had a negative bleeding history; 4 pt had normal agg and PFA-100 and were diagnosed exclusively by PTEM. In this group, the abnormalities on platelet aggregation and PFA were variable and PTEM allowed for a better definition of the specific abnormalities. For MYH-9 patients, PTEM confirmed leukocyte inclusions that had already been identified on light microscopy. PTEM was the only diagnostic modality able to identify the abnormality in York platelet syndrome. In 5/54 (9%) genetic testing was necessary for diagnosis (GATA-1, ANKRD26 and RUNX-1)ConclusionIn this cohort, standard platelet assays established the diagnosis in the large majority of CQPD and CT. PTEM was essential for confirmation in DG-SPD, mild alpha granule deficiency and York platelet syndrome and useful in combination with molecular testing to establish a diagnosis in selected cases. Guidance on selection of patients for such specialized testing requires further study.Table 1Results of standard and esoteric platelet testing for 54 patients with congenital qualitative platelet disorders and/or thrombocytopeniaDisordern (%)Platelet aggregationPFAPTEMFlow cytometryConfirmation by geneticsDoneAbnormalDoneAbnormalDoneAbnormalDoneAbnormalGlanzmann thrombasthenia6 (11)665510330Bernard-Soulier syndrome & variants4 (7)444430440MYH-9 related disorders10 (19)728355425Dense granule deficiency and HPS12 (22)1281281212400Alpha granule deficiency*7 (13)747677441York platelet syndrome4 (7)101144110TXA2 or PG synthesis/receptor defect4 (7)444330300CATM1 (2)0-0-0-0-1ANKRD263 (6)333233333GATA-11 (2)0-1111111RUNX-11 (2)10110-0-1Jacobsen syndrome1 (2)101111101MYH-9: myosin heavy chain 9, non-muscle (MHY-9) associated thrombocytopenia, HPS: Hermansky-Pudlak Syndrome, TXA2: thromboxane A2, PG: prostaglandin CATM: congenital amegakaryocytic thrombocytopenia. * Includes gray platelet syndrome DisclosuresPardanani:Stemline: Research Funding.

In vitro Quality of Platelets with Low Plasma Carryover Treated with Ultraviolet C Light for Pathogen Inactivation

Background: The THERAFLEX UV-Platelets system uses shortwave ultraviolet C light (UVC, 254 nm) to inactivate pathogens in platelet components. Plasma carryover influences pathogen inactivation and platelet quality following treatment. The plasma carryover in the standard platelets produced by our institution are below the intended specification (<30%). Methods: A pool and split study was carried out comparing untreated and UVC-treated platelets with <30% plasma carryover (n = 10 pairs). This data was compared to components that met specifications (>30% plasma). The platelets were tested over storage for in vitro quality. Results: Platelet metabolism was accelerated following UVC treatment, as demonstrated by increased glucose consumption and lactate production. UVC treatment caused increased externalization of phosphatidylserine on platelets and microparticles, activation of the GPIIb/IIIa receptor (PAC-1 binding), and reduced hypotonic shock response. Platelet function, as measured with thrombelastogram, was not affected by UVC treatment. Components with <30% plasma were similar to those meeting specification with the exception of enhanced glycolytic metabolism. Conclusion: This in vitro analysis demonstrates that treatment of platelets with <30% plasma carryover with the THERAFLEX UV-Platelets system affects some aspects of platelet metabolism and activation, although in vitro platelet function was not negatively impacted. This study also provides evidence that the treatment specifications of plasma carryover could be extended to below 30%.

Optimal- vs. standard-antiplatelet therapy on platelet function and long-term clinical outcomes in patients with high on-treatment platelet reactivity: 2-year outcomes of the multicentre, randomized Optimal-antiPlatelet Therapy (OPT) trial

Optimal strategy for the management of high on-treatment platelet reactivity (HPR), an independent predictor of ischaemic events, is not yet clear. The aim of this study was to explore the safety and efficacy of personalized antiplatelet therapy based on platelet function testing in patients with HPR. A total of 840 patients with HPR who underwent coronary stenting for acute coronary syndrome (ACS) were randomly assigned in a 1:2 ratio to receive standard dual-antiplatelet therapy ( n = 280) or platelet function-guided optimal-antiplatelet therapy ( n = 560). In the optimal group, patients initially received clopidogrel 150 mg/day on top of aspirin and switched to triple antiplatelet therapy with additional cilostazol if repeat platelet function assay after 3 days indicated a sustained HPR. The primary endpoint was the composite of all-cause death, myocardial infarction, clinically driven target vessel revascularization (TVR), or stroke. Optimal therapy resulted in significantly lower 30-day HPR rate (30.1 vs. 44.3%, P < 0.001). At 1-year follow up, optimal therapy was associated with significantly lower incidence of primary endpoint events (2.7 vs. 6.8%, P = 0.006) compared with standard therapy, mainly due to the reduction of TVR. These beneficial effects were sustained at 2 years (4.5 vs. 9.2%, P = 0.006), with similar bleeding event rates between groups (7.3 vs. 6.5%, P = 0.565). Optimal-antiplatelet therapy based on platelet function testing is effective in attenuating HPR and improving long-term outcomes for ACS patients with no increase in bleeding events.

Riboflavin and amotosalen photochemical treatments of platelet concentrates reduce thrombus formation kinetics in vitro.

Photochemical treatment (PCT) of platelet concentrates using photosensitizers and ultraviolet light illumination reduces the proliferation potential of pathogens by damaging biomolecules. The impact of riboflavin (RF-PRT)- and amotosalen (AS-PCT)-based pathogen inactivation on platelets was studied using microfluidic flow chambers on immobilized collagen using standard platelet concentrates prepared from buffy coats in additive solution. Flow cytometry, metabolic parameters and light transmission aggregometry with thrombin-related peptide, collagen and ristocetin were determined concurrently. Both PCTs significantly decreased the platelet surface coverage kinetics in flow chambers over the course of the 7-day study. Platelet aggregation was affected following RF-PRT in response to all agonists, while AS-PCT mainly impacted low-dose ristocetin agglutination. RF-PRT induces premature platelet activation because integrin αII b β3 was spontaneously activated, and α-degranulation, phosphatidylserine/-ethanolamine exposure and anaerobic metabolism significantly increased following treatment, which was not the case for AS-PCT. On the other hand, AS-PCT significantly diminished thrombus growth onto von Willebrand factor under shear flow. This defect was caused by fewer integrin αII b β3 interactions, not by defective GPIbα-VWF binding as shown by adhesion experiments in the presence of tirofiban. Moreover, integrin αII b β3 activation was also affected following the activation of platelets via GPVI-FcγRIIa or PAR1. Finally, amotosalen illumination as such is sufficient to induce platelet damage, with no additional measurable effect of the chemical adsorption step. Gamma irradiation caused no significant difference compared to controls on any time-point or for any parameter. Both PCTs significantly reduce thrombus formation rate but by different biochemical mechanisms.