Irradiated allogeneic cells enhance umbilical cord blood stem cell engraftment in immunodeficient mice.

Umbilical cord blood (UCB) is a readily available source of hematopoietic stem cells for transplantation. UCB hematopoietic SCT for average- and large-sized patients is often limited by the number of cells available in a single unit. To address this limitation, we performed experiments to determine if adjunctive therapy with third-party human allogeneic cells enhances the engraftment of human UCB in immunodeficient mice. UCB cells with or without sequential infusion of irradiated third-party allogeneic cells were used in transplantation studies of NOD/SCID and NOD/SCID-IL2Rγ null mice. We studied the impact of irradiated allogeneic cells on colony formation in vitro using long-term culture assays also. Our studies demonstrate that short- and long-term UCB engraftment of immunodeficient mice is enhanced by irradiated allogeneic cells. Secondary transplants demonstrate the durability of engraftment. These preclinical studies support the further development of irradiated allogeneic cells as an adjunct to single UCB transplantation when limiting numbers of cells are available.

Apyrase, ascorbic acid and aprotinin ameliorate the storage lesion in pelleted platelet preparations.

To evaluate the effect of apyrase, ascorbic acid and aprotinin (AAA) in preventing platelet activation during storage, 12 sets of platelet concentrates (PCs), were treated with AAA and evaluated at days 1, 3, and 5 utilizing platelet functional and morphological assays. Platelets treated with AAA demonstrated significantly enhanced response to ADP-induced platelet aggregation, higher morphology scores, and evaluated ATP levels compared to control samples after 5 days of storage. Similarly, platelet specimens treated with AAA had significantly reduced PF4 secretion and P-selectin expression compared to controls. Finally, Western blots of aggregated platelets at day 5 demonstrated that AAA-treated PCs continue to express the platelet membrane GPIb whereas specimens from control PCs do not. These results show that PCs treated with AAA have reduced platelet activation and enhanced functional platelet activity.

Buffy coat platelets stored in apyrase, aprotinin, and ascorbic acid in a suspended bag: combined strategies for reducing platelet activation during storage.

BACKGROUND: Platelet activation is an important factor impeding the clinical effectiveness of platelet transfusions. In this study, platelet concentrates (PCs) were prepared by a novel suspended-bag buffy coat technique that was followed by the addition of a mixture of platelet activation inhibitors to the storage bag. STUDY DESIGN AND METHODS: In vitro platelet function was evaluated in PCs prepared by the suspended-bag buffy coat technique and stored at 22 degrees C for 5 days in the presence of (n = 12) or absence (n = 12) of apyrase, ascorbic acid, and aprotinin (AAA). RESULTS: Platelets from AAA-incubated PCs demonstrated mean ATP levels 17 percent (p < 0.004), 13 percent (p < 0.02), and 22 percent (p < 0.003) higher than those measured in parallel control PCs on Days 1, 3, and 5, respectively. Similarly, on Days 3 and 5 of storage, respectively, 45-percent (p < 0.001) and 50-percent (p < 0.001) greater ADP-induced maximum aggregation was observed in AAA-incubated PCs than was seen in control preparations. AAA-incubated PCs demonstrated alpha-granule membrane protein-140 expression 92 percent (p < 0.01), 133 percent (p < 0.003), and 104 percent (p < 0.001) below that in control PCs on Days 1, 3, and 5, respectively. At similar intervals, a significant increase in recovery from hypotonic shock also was observed in AAA-incubated PCs. Further, Day 5 AAA-PCs demonstrated significantly higher morphology scores and O2 consumption than did control preparations. CONCLUSION: Buffy coat platelets prepared in suspended bags and stored in the presence of AAA demonstrate significantly reduced activation and enhanced functional and metabolic activity.

A novel technique for preparing improved buffy coat platelet concentrates.

We evaluated in vitro platelet function of platelet concentrates stored at 22 C for 5 days prepared either by the conventional pelleting procedure or platelet concentrates prepared from buffy coats by utilizing a novel bucket designed to support a suspended bag. For platelet concentrates from buffy coat, whole blood was centrifuged at 3,000 x g for 13 min, with all but 30cc of the cell poor plasma transferred to a satellite bag, followed by a second centrifugation at 170 x g for 5 min utilizing our novel centrifugation device. For pelleted platelets, whole blood was centrifuged at 2,000 x g for 3 min, platelet rich plasma removed, centrifuged, and the pellet resuspended in plasma. Leukocyte contamination in buffy coat platelet concentrates was reduced by 95% (p < 0.001) in comparison to pelleted platelets. Further, platelets from buffy coat platelet concentrates demonstrated significantly enhanced ADP-induced aggregation, increased recovery from hypotonic shock, higher morphology scores, and reduced GMP-140 expression in comparison to pelleted preparations. No differences in O2 consumption, CO2 production, pH and total ATP were observed between the two types of preparations at day 5 of storage. Our results indicate that platelet concentrates from buffy coat, prepared by a suspended storage bag centrifugation technique, are superior with respect to in vitro platelet function when compared to pelleted platelets.

[Clinical usefulness of blood platelets cryopreserved in liquid nitrogen]. / Ocena klinicznej przydatnosci krwinek plytkowych kriokonserwowanych W temperaturze par azotu.

The hemostatic effectiveness of platelet concentrates cryopreserved in a vapor phase of liquid nitrogen were evaluated after transfusion of 14 concentrates to 6 patients. Transfusion of AB0 and Rh-compatible platelet concentrates elevated the number of circulated platelets in donors by about 24 G/L. The mean corrected count increment (C.I.) after 001 and 24 hours post transfusion was 14.694 and 7.735/microliters/m2/10(11) respectively. Decrease of haemorrhagic diathesis, and correction of bleeding time in all cases have been observed.

The influence of CS-3000 plateletapheresis on blood donors and on platelet concentrates.

Platelet concentrates were prepared from blood of donors by means of Fenwal CS-3000, or by double centrifugation method. Platelets from CS-3000 were cryopreserved with 5% DMSO for 2-4 weeks. After thawing and washing platelets were stored in room temperature for 6 hours. Immediately after plateletapheresis the platelet count decreased by 24% and leukocyte count, Ht and Hb diminished on average 30%, 22% and 18% respectively. The efficiency of separation procedure was 45% and white cells contamination 5.8 +/-8.1 x 10(11). Significant morphological changes and the decrease in total ATP comparing to double centrifugation method were observed. Storage of platelets for 6 hours in room temperature didn't cause their quality changes. Single donor plateletapheresis method can prevent alloimmunization and diminish the risk of transmission of the viruses.

Donors leukapheresis on Fenwal CS-3000.

Leukocytes were collected from 24 donors using Fenwal CS-3000 cell separator. Two different sedimentation agents in citric acid were used. After collection donors hematologic data were evaluated. No significant differences in product yield and in donors blood test were observed when in product yield and in donors blood test were observed when Dextran 110 or HES were used. The average donor leukocyte count declined by 30% after the completion of the procedure. Mean donor platelet count decrease an average by 23% (from 203 to 156 x 10(3)/microliters). Other hematologic data were consistent with hemodilution due to anticoagulant and saline solutions.