Interruption of agitation of platelet concentrates: a multicenter in vitro study by the BEST Collaborative on the effects of shipping platelets.

BACKGROUND: Transported platelets (PLTs) are not under continuous agitation. The aim of this study was to determine whether PLTs shipped between 24 and 48 hours would be able to maintain a pH(22 degrees C) value of 6.5 at the end of 7 days of storage. STUDY DESIGN AND METHODS: Six laboratories prepared leukoreduced PLTs. PLT pools were divided into low and high PLT concentration with paired shipped (20-43 hr) and unshipped controls. Units were under continuous agitation at 22 +/- 2 degrees C when not being transported. In vitro measures including pH, pO(2), and pCO(2) were determined over 7 days. RESULTS: Ninety-two PLT components from 24 pools were eligible for analysis. One unshipped control and three shipped products failed to maintain a pH(22 degrees C) value of 6.5 through 7 days. In vitro characteristics were maintained slightly better over 7 days of storage in the unshipped control arms. PLT concentration, shipping time, and their interaction were significant determinants of the final pH at the end of storage (p < 0.05). Lactate generation rate increased by 35 +/- 2 (mean +/- SE) micromol per 10(12) PLTs per hour over baseline during shipping (p < 0.0001). After restoration of standard blood banking conditions with agitation, this rate dropped 24 +/- 2 micromol per 10(12) PLTs per hour (p < 0.0001). CONCLUSION: PLTs in plasma shipped for at least 20 to 24 hours maintain a pH(22 degrees C) value of 6.5 for 7 days. A longer shipping time may result in a pH(22 degrees C) value of 6.5. During shipping, glycolysis was up regulated in these PLTs resulting in increased lactic acid production. After restoration of agitation, shipped products down regulated glycolysis. These effects should be accounted for in the development of PLT storage and transportation systems.

A critical comparison of platelet preparation methods.

PURPOSE OF REVIEW: Platelet concentrates may be prepared from whole blood or by plateletpheresis. Currently, the non-evidence-based preponderance of apheresis units in the United States and the 50: 50 ratio in Europe may not optimize the gifts of whole-blood donors or minimize healthcare costs. Post-storage pooled, whole-blood-derived platelets, on the other hand, do not provide the convenience of or an equivalent level of safety as apheresis platelets. RECENT FINDINGS: Some data suggest that different methods of manufacture of whole-blood-derived platelets (platelet-rich plasma or buffy coat intermediate steps) result in differing degrees of platelet activation, which may impact on the quality of stored concentrates. Recent studies have observed superior radiolabel recovery and post-transfusion increments for platelets derived from apheresis compared with platelet-rich plasma whole-blood-derived platelets. A pre-storage pooling system for whole-blood-derived platelets has just been licensed in the USA, and may eventually combine the benefits of apheresis-derived and whole-blood-derived platelets. The advantages of the European method of manufacture of buffy coat whole-blood-derived platelet concentrate have convinced the Canadian Blood Services to abandon platelet-rich-plasma-derived concentrates. SUMMARY: We present a literature-based review of the relative merits of apheresis-derived and whole-blood-derived platelets. Additional studies are needed in order to define the optimal proportion of the platelet supply from apheresis collections and the choice of whole-blood-derived production method for US blood providers.

In vitro pH effects on in vivo recovery and survival of platelets: an analysis by the BEST Collaborative.

BACKGROUND: The pH environment of stored platelet (PLT) products is recognized as an important factor and is generally used as a key surrogate measure of PLT viability. It is the only in vitro measurement that has been translated into industry standards and regulatory rules or specifications for storage of PLT products. The objective of this study was to evaluate the effect of in vitro pH on the in vivo recovery and survival of autologous PLT products. STUDY DESIGN AND METHODS: Data from individual autologous radiolabeled PLT kinetic studies were solicited from independent laboratories. PLTs stored for at least 5 days in 100 percent autologous plasma with a pH(22 degrees C) of at least 6.2 were analyzed. Data were fit to a mixed-effects regression model with fixed effects of pH(22 degrees C), time of storage, and preparation method-storage bag combination. RESULTS: Eight research laboratories reported 476 individual recovery and survival results with associated pH before labeling from a variety of autologous, radiolabeled PLT kinetic studies from September 1999 to March 2005. These results are from 254 individual subjects who donated a total of 386 PLT units, with up to nine collections per subject reported. The effect of pH on either PLT recovery (p = 0.86) or survival (p = 0.55) was not significant. Time of storage and the method-bag combination both had significant effects on these outcomes (p < 0.0001). CONCLUSION: These data suggest that there is no relationship between in vitro pH at a pH(22 degrees C) of at least 6.2 and in vivo PLT viability as measured by radiolabeled recovery and survival of autologous PLTs.

Reevaluation of the resting time period when preparing whole blood-derived platelet concentrates with the platelet-rich plasma method.

BACKGROUND: The resting of platelet (PLT) pellets during the preparation of whole blood-derived PLT concentrates (PCs) is considered an essential step. A reevaluation of the rest period was conducted because preparation and storage conditions have been modified during the past 20 years. STUDY DESIGN AND METHODS: A two-site in vitro study (Study 1) was conducted with rest times of 0 to 5 minutes, 1 hour, and 4 hours (n = 31-33 per rest period). Leukoreduced PCs were stored for 5 days. A second study (Study 2) measuring in vivo viability was conducted at a third site (14 paired studies). PCs were prepared from 2 units of whole blood on the same day to simultaneously compare viability following storage and radioisotopic labeling. RESULTS: In Study 1, comparable in vitro parameters and swirling were observed with the three rest periods. The mean (+/-1 SD) values after storage for the extent of shape change and hypotonic stress parameters were 0 to 5 minutes, 16.7 +/- 7.2 and 66.0 +/- 15.7%; 1 hour, 19.1 +/- 6.9 and 69.3 +/- 17.9%; and 4 hours, 17.6 +/- 5.5 and 64.1 +/- 11.3%. In Study 2, the in vivo recovery was 49.9 +/- 15.3 and 50.9 +/- 20.2% with 0- to 5-minute and 1-hour rest periods, respectively. The corresponding survival time was 111.2 +/- 50.7 and 114.9 +/- 43.8 hours. CONCLUSION: These studies indicate comparable in vitro and in vivo viability properties with 0- to 5-minute and 1-hour rest periods and at 4 hours (in vitro only).

Platelet dose consistency and its effect on the number of platelet transfusions for support of thrombocytopenia: an analysis of the SPRINT trial of platelets photochemically treated with amotosalen HCl and ultraviolet A light.

BACKGROUND: The SPRINT trial examined efficacy and safety of photochemically treated (PCT) platelets (PLTs). PCT PLTs were equivalent to untreated (control) PLTs for prevention of bleeding. Transfused PLT dose and corrected count increments (CIs), however, were lower and transfusion intervals were shorter for PCT PLTs, resulting in more PCT than control transfusions. PLT dose was analyzed to determine the impact of the number of PLTs transfused on transfusion requirements. STUDY DESIGN AND METHODS: Transfusion response was compared for patients with all doses of >or=3.0 x 10(11) and the complementary subset of patients with any dose of fewer than 3.0 x 10(11). Analyses included comparison of bleeding, number of PLT and red blood cell (RBC) transfusions, transfusion intervals, and CIs between PCT and control groups within each PLT dose subset. RESULTS: Mean PLT dose per transfusion in the PCT group was lower than in the control group (3.7 x 10(11) vs. 4.0 x 10(11); p<0.001). More PCT patients received PLT doses of fewer than 3.0 x 10(11) (n=190) than control patients (n=118; p<0.01). Comparisons of patients receiving comparable PLT doses showed no significant differences between PCT and control groups for bleeding or number of PLT or RBC transfusions; however, transfusion intervals and CIs were significantly better for the control group. CONCLUSIONS: When patients were supported with comparable doses of PCT or conventional PLTs, the mean number of PLT transfusions was similar. Lower CIs and shorter transfusion intervals for PCT PLTs suggest that some PLT injury may occur during PCT. This injury does not result in a detectable increase in bleeding, however.

The influence of various hematology analyzers on component platelet counts.

Hematology analyzers designed to count platelets in samples of whole blood are used to enumerate the total number of platelets in components prepared for transfusion. This report addresses the issue of variability in platelet counts obtained with different models of hematology analyzers. The influence of a common calibration procedure, involving one level of porcine platelets, on the extent of variability was also evaluated. Identical sets of samples of simulated and apheresis-derived human platelets were counted by multiple laboratories in 3 separate studies. In the first 2 exercises, 7 samples of both porcine platelets and modified goat erythrocytes with targeted platelets counts from 0.2 to 4.0 x 10(12)/L were counted without prior dilution. In both exercises, the samples were counted multiple times after routine calibration using instructions provided by the manufacturers of the various hematology analyzers used. In the second exercise, the samples were recounted after the hematology analyzers were recalibrated with a common calibrant consisting of porcine platelets at a targeted concentration of 0.5 x 10(12)/L. In the first and second exercises, 20 and 18 hematology analyzers were used, respectively. In the third exercise, 6 samples prepared from a single unit of apheresis platelets with targeted counts from 0.2 to 1.64 x 10(12)/L were shipped by an overnight courier and counted in triplicate on the day of arrival. Eleven hematology analyzers were used. The influence of recalibration was evaluated statistically by using the 95% prediction interval for the mean of a future set of observations. The platelet counts measured with a specific type of hematology analyzer provided the data to calculate the 95% prediction interval. With routine calibration, a wide variability in platelet counts was observed with all levels of both simulated and apheresis-derived human platelets. For example, with porcine platelets at a targeted level of 0.4 x 10 (12)/L, the platelet counts ranged from 0.31 to 0.47 x 10(12)/L. Recalibration reduced the extent of variability observed with all levels of simulated and apheresis-derived human platelets by increasing the observed platelet counts determined with a subset of hematology analyzers that produced platelet counts in the lower portion of the range. With recalibration, the mean platelet counts obtained with most hematology analyzers, especially with samples having targeted platelet levels no greater than 1.0 x 10(12)/L, were within or near the 95% prediction interval determined with the instruments that provided the highest platelet counts with routine calibration. With recalibration, the reproducibility of the platelet counts was considered to be good for all hematology analyzers with all levels of simulated and apheresis-derived human platelets for most of the instruments. The coefficient of variance did not exceed 6%, with most of the values ranging from 1% to 3%. This study therefore found that the platelet counts of platelet concentrates can be markedly influenced by the type of hematology analyzer used. A common calibration procedure designed specifically for the range of platelet counts in platelet products may be beneficial considering that many different hematology analyzers are being used to count platelets.

Improving technology for collecting platelets by apheresis: five-year experience in one blood center.

Over the past decade, newly introduced methods for apheresis platelet collection have led to increased collection yields. This has resulted in "splitting," which allows transfusion of 2 patients from 1 high-yield collection. Although many small studies exist, no large studies have described the impact of methodological changes on routine blood center collections. We constructed a database containing selected parameters from 45,224 apheresis collections spanning July 1997 to April 2002, using Gambro BCT Spectra (Lakewood, CO), Fenwal CS-3000+ (Baxter Healthcare Corp, Fenwal Division, Deerfield, IL), and Baxter Amicus instruments. A Baker 9110+ hematology analyzer (Bio Chem Immunosystems, Inc., Allentown, PA) was used for platelet counting. Monthly average collection yields, distribution yields (product platelet contents after splitting), and split rates (the fraction of donations which may be split) were determined. The monthly mean collection yield and split rate correlated very closely. Both rose throughout the study period. The split rate climbed from 25% to 70% by study end. However, mean monthly distribution yields decreased by 7% because split and unsplit platelet yields both rose as split rates rose. Overcollections with the Amicus correlated with underestimation of donors' true preprocedure platelet counts during machine programming. Undercollections occurred in donors with low counts and, with-single needle Amicus, microcytic platelet collection. These results may assist in the optimization of an apheresis program. Increased collection yields correlated with cell separator type, dual-needle access, donor platelet count >250 x 10(9)/L, programming with true preprocedure platelet counts and capacity for triple product preparation from collection yields exceeding 2-bag storage capacity.

White particulate matter: report of the ad hoc industry review group.

BACKGROUND: In January 2003, blood center personnel in the American Red Cross, Southern Region in Atlanta, noticed whitish particulate material (WPM) that had not been observed previously in several units of red blood cells (RBCs). An expert panel was formed to evaluate studies of the material and make appropriate recommendations STUDY DESIGN AND METHODS: The expert panel reviewed information provided by several investigations and organizations. This included: background information, and experiences relating to WPM; WPM composition; factors promoting WPM formation; risk of WPM (if any) to patients; and recommendations to prevent future occurrences. RESULTS: WPM is derived from blood. No data suggest that external contamination or collection set components contribute to WPM development. A major constituent of WPM is platelets (PLTs). WPM is most commonly observed in RBCs that have been subjected to a hard spin without PLT separation. WPM is rarely, if ever, observed in RBCs that have been subjected to leukoreduction. CONCLUSIONS: (1) WPM is not new, can be prevented, and can be removed. (2) WPM contains PLTs, white blood cells, fibrin, and cellular debris. (3) Changes in blood handling are not necessary. (4) WPM may be more frequent when higher g forces are used in component preparation. (5) Enhanced visual inspection of blood components need not be continued. (6) It appears that WPM may not form in RBC collected using automated devices. (7) WPM did not pose a risk to patients but should be avoided.

Therapeutic efficacy and safety of platelets treated with a photochemical process for pathogen inactivation: the SPRINT Trial.

We report a transfusion trial of platelets photochemically treated for pathogen inactivation using the synthetic psoralen amotosalen HCl. Patients with thrombocytopenia were randomly assigned to receive either photochemically treated (PCT) or conventional (control) platelets for up to 28 days. The primary end point was the proportion of patients with World Health Organization (WHO) grade 2 bleeding during the period of platelet support. A total of 645 patients (318 PCT and 327 control) were evaluated. The primary end point, the incidence of grade 2 bleeding (58.5% PCT versus 57.5% control), and the secondary end point, the incidence of grade 3 or 4 bleeding (4.1% PCT versus 6.1% control), were equivalent between the 2 groups (P =.001 by noninferiority). The mean 1-hour posttransfusion platelet corrected count increment (CCI) (11.1 x 10(3) PCT versus 16.0 x 10(3) control), average number of days to next platelet transfusion (1.9 PCT versus 2.4 control), and number of platelet transfusions (8.4 PCT versus 6.2 control) were different (P <.001). Transfusion reactions were fewer following PCT platelets (3.0% PCT versus 4.4% control; P =.02). The incidence of grade 2 bleeding was equivalent for PCT and conventional platelets, although posttransfusion platelet count increments and days to next transfusion were decreased for PCT compared with conventional platelets.

Ocular rosacea in childhood.

PURPOSE: To describe the clinical characteristics and treatment response of ocular rosacea in the pediatric population. DESIGN: Retrospective case series. METHODS: The clinic charts of consecutive pediatric cases of ocular rosacea were evaluated over a 34-month period. Minimal diagnostic inclusion criteria were the presence of posterior eyelid inflammation including meibomian gland inspissation and lid margin telangiectasis, in conjunction with conjunctival injection or episcleritis. RESULTS: Six patients ranged from 3 to 12 years of age at presentation. All shared a long history of ocular irritation and photophobia. Five patients (83%) were female and had bilateral involvement. Eyelid telangiectases and meibomian gland disease were present in all cases. Three patients (50%) had sterile corneal ulcers. Only two patients (33%) had cutaneous involvement at the time of diagnosis. All patients experienced significant improvement with a combination of oral antibiotics (doxycycline or erythromycin), with or without topical antibiotics (erythromycin or bacitracin) or topical steroids (fluorometholone). CONCLUSION: Ocular rosacea in children may be misdiagnosed as viral or bacterial infections. Unlike in adults, associated cutaneous changes are uncommon. Most disease is bilateral, although involvement may be asymmetric. Response to conventional treatment is excellent, although long-term treatment may be necessary to prevent relapses.

Warfarin-induced skin necrosis and venous limb gangrene in the setting of heparin-induced thrombocytopenia.

BACKGROUND: Heparin-induced thrombocytopenia (HIT) is a common, often catastrophic, syndrome that produces the most hypercoagulable of states. Emerging therapeutic strategies use alternative anticoagulants; warfarin's place is being reexamined. Early in the course of warfarin therapy, there may be net procoagulant effects because of the inhibition of protein C. With HIT, it has been suggested that unopposed warfarin can precipitate venous limb gangrene. There are also reports of warfarin-induced skin necrosis. We seek to confirm and increase awareness of the risks of warfarin with HIT. METHODS: We describe 6 patients with HIT seen at 3 medical centers in whom frank or impending venous limb gangrene, central skin necrosis, or both were temporally related to warfarin initiation. RESULTS: At warfarin initiation, 5 patients had recognized HIT and 1 had it recognized later. Complications emerged after 2 to 7 days, and consisted of warfarin-induced skin necrosis (n = 5) and venous limb gangrene (n = 2); 1 patient had both. This emerged with unopposed warfarin in 4 patients and as a direct thrombin inhibitor was being withdrawn in 2. All had supratherapeutic international normalized ratios. One patient required leg and breast amputations, and another one died. CONCLUSIONS: Because of the early effects on protein C, warfarin can precipitate venous limb gangrene and/or skin necrosis in the extreme hypercoagulable milieu of HIT. With HIT, unopposed warfarin should be avoided and caution is needed during transition from a direct thrombin inhibitor. Warfarin should be initiated at modest doses in patients with HIT after platelet recovery. Implications extend to warfarin initiation with other thrombotic diatheses.

Preliminary validation of a new standard of efficacy for stored platelets.

BACKGROUND: Platelet preparation and storage systems, unlike those for RBC, lack an objective, absolute performance criterion to determine acceptability. Recently, a criterion based on paired comparison with the radiolabeled recovery and survival of "fresh" platelets has been proposed, namely, recovery = two-thirds and survival = half of "fresh" platelets. STUDY DESIGN AND METHODS: Eleven normal subjects donated a unit of leukoreduced apheresis platelets using a standard, approved system. They received an aliquot radiolabeled with 111In or 51Cr (random selection) 4 to 20 hours after donation and, using the other radioisotope, on Day 5 of storage. The recovery was calculated based on the injectate radioactivity. The survival was determined using the multiple-hit model. The area under the platelet survival curve was calculated using the COST program. RESULTS: Reinfusion of platelets less than 20 hours after collection resulted in a recovery of 74.7 +/- 12.3 percent and a survival time of 7.5 +/- 1.1 days. Reinfusion on Day 5 resulted in a recovery of 58.2 +/- 12.0 percent and a survival time of 6.9 +/- 1.4 days, values that were 77.9 +/- 9.5 percent and 91.8 +/- 16.1 percent of the observation using "fresh" platelets, respectively. The area under the curve using Day 5 platelets was 67.8 +/- 11.5 percent of that using "fresh" platelets. CONCLUSION: The proposed criterion for objective evaluation of platelet preparation and storage systems appears applicable to a commonly accepted approach, leukoreduced apheresis platelets stored in plasma for 5 days, and merits evaluation using other collection, treatment, and storage systems.

A method for increased dose conformity and segment reduction for SMLC delivered IMRT treatment of the prostate.

PURPOSE: The focus of this work is to develop a practical planning method that results in increased dose conformity and reduced treatment time for segmental multileaf collimation (sMLC) based intensity-modulated radiation therapy (IMRT) delivery. METHODS AND MATERIALS: Additional regions for dose constraint are introduced within the normal tissue during the planning process by designing a series of concentric ellipsoids around the target. A dose gradient is then defined by assigning dose constraints to each concentric region. The technique was tested at two centers and data for 26 and 10 patients, respectively, are presented allowing for differences in treatment technique, beam energy, ellipsoid definition, and prescription dose. At both centers, a series of patients previously treated for prostate cancer with IMRT were selected, and comparisons were made between the original and new plans. RESULTS: While meeting target dose specifications and normal tissue constraints, the average number of beam directions decreased by 1.6 with a standard error (SE) of 0.1. The average time for delivery at center 1 decreased by 29.0% with an SE of 2.0%, decreasing from 17.5 min to 12.3 min. The average time for delivery at center 2 decreased by 29.9% with an SE of 3.8%, decreasing from 11 min to 7.7 min. The amount of nontarget tissue receiving D(100) decreased by 15.7% with an SE of 2.4%. Nontarget tissue receiving D(95), D(90), and D(50) decreased by 16.3, 15.1, and 19.5%, respectively, with SE values of approximately 2% at center 1. Corresponding values for D(100), D(95), D(90), and D(50) decreased by 13.5, 16.7, 17.1, and 5.1%, respectively, with SE values of less than 3% at center 2. CONCLUSION: By designating subsets of tissue as concentric regions around the target(s) and carefully defining each region's dose constraints, we have gained an increased measure of control over the region outside the target boundaries. This increased control manifests as two distinct endpoints that are beneficial to the IMRT process: increased dose conformity and decreased treatment time.

The role of DNA deformation energy at individual base steps for the identification of DNA-protein binding sites.

We examine the use of deformation propensity at individual base steps for the identification of DNA-protein binding sites. We have previously demonstrated that estimates of the total energy to bend DNA to its bound conformation can partially explain indirect DNA-protein interactions. We now show that the deformation propensities at each base step are not equally informative for classifying a sequence as a binding site, and that applying non-uniform weights to the contribution of each base step to aggregate deformation propensity can greatly improve classification accuracy. We show that a perceptron can be trained to use the deformation propensity at each step in a sequence to generate such weights.