S. Holme

The expression of p-selectin during collection, processing, and storage of platelet concentrates: relationship to loss of in vivo viability

BACKGROUND: Recent studies suggested that platelet activation with surface expression of p‐selectin on stored platelets may be related to a loss of viability. At present, there has been no thorough investigation of the extent or significance of p‐selectin expression during the collection, processing, and storage of platelet concentrates (PCs) under various conditions.

In vivo regeneration of red cell 2, 3-diphosphoglycerate following transfusion of DPG-depleted AS-1, AS-3 and CPDA-1 red cells

Regeneration of 2, 3‐diphosphoglycerate (DPG) was determined following transfusion of DPG‐depleted group O red cells into group A recipients. Blood from five donors was stored in the adenine‐containing solutions CPDA‐1, AS‐1 or AS‐3 for 35 d at 4°C. Post‐transfusion red cell DPG and ATP were measured in separated group O red cells over a 7 d period. The studies confirmed rapid in vivo DPG regeneration with ≥ 50% of the maximum level being achieved within 7 h. An average of 95% of the recipients’ pre‐transfusion DPG level was achieved by 72 h and by 7 d mean (±SEM) DPG levels relative to recipients pre‐transfusion DPG averaged 84% (± 13%), 92% (± 17%) and 84% (± 21%) for CPDA‐1, AS‐1 and AS‐3 red cells, respectively. Results were comparable to those previously reported for blood stored in ACD for 15‐20 d (Valeri & Hirsch, 1969; Beutler & Wood, 1969). The immediate regeneration rate, V, closely approximated first order regeneration kinetics with AS‐3 red cells exhibiting double the rate of CPDA‐1 red cells (P<0·001). AS‐1 red cells exhibited an intermediate rate of regeneration which was not significantly different compared to either CPDA‐1 or AS‐3 (P>0·05). V exhibited a significant (P<0·05) positive correlation with ATP levels 5‐7 h post‐infusion. ATP regeneration of the infused cells was rapid with a mean increase of 1·2 μmol/g Hb above post‐storage levels being achieved 1 h following transfusion.

Improved in vivo and in vitro viability of platelet concentrates stored for seven days in a platelet additive solution

An additive solution has been developed for storage of platelet concentrates (PC) which sustains improved in vivo and in vitro viability after 7 d of storage in second generation oxygen permeable containers. This platelet additive solution is a protein‐free physiologic salt solution fortified with citrate, bicarbonate and glucose. The in vivo quality of the PC was evaluated by autologous radiolabelling with Indium‐111‐oxine to measure recovery and survival by multiple hit analysis. The in vitro quality was evaluated by total ATP content, hypotonic shock response and extent of shape change with ADP. Ten paired studies were performed with PC from the same donor being stored for 7 d at 22°C in both CPDA‐1 plasma and the additive solution. Mean recoveries and survivals were found to be substantially higher with PC stored in the additive solution than with PC stored in CPDA‐I plasma (51·0±7·8% and 144·1 ± 15·9 h versus 36·6±10·7% and 110·4±31·6 h). The differences were statistically significant (P< 0·001). The results of the in vitro assays described above parallelled the in vivo results, with statistically significantly superior results (P<0·01) for all parameters of PC stored in the additive solution. This study is the first to show that PC quality may be improved and storage extended using an additive solution.

Platelet storage lesion in second-generation containers: correlation with platelet ATP levels.

Abstract. The nature of platelet lesion occurring with storage of platelet concentrates (PC) in second‐generation containers was investigated using various storage media and storage periods up to 14 days. In CPD‐plasma (control medium), the changes which occurred progressively during storage were loss of discoid shape, microscopic platelet aggregate formation, fragmentation and the appearance of disintegrated, ‘balloon’ forms. By day 14 less than 10% of the platelets were discoid in shape, the platelet count had decreased by 23%, and there was a 5‐fold increase in the amount of lactate dehydrogenase in plasma. Associated with this was a decrease in the platelet oxygen consumption rate, D(O2), loss of cellular ATP and extent of ADP‐induced shape change, and a decrease in the hypotonic shock response. These parameters decreased at a similar rate, with a 50% decrease (t1/2) at days 7–9. They correlated highly with each other during storage and also with a fall in pH. At day 14 of storage, mean pH was 6.1 ± 0.3. To evaluate the effect of pH stabilization during storage, 4 mEq sodium bicarbonate was added to PC in CPD‐plasma. Although pH maintenance was much improved, 7.2–6.6 during 14 days of storage, the same in vitro lesions developed, although more slowly. The t1/2 of the same parameters was prolonged for approximately two days. When PC were stored in a plasma‐free physiologic salt solution whose salt composition was similar to CPD‐plasma, the t1/2 of the parameters increased to 11–15 days of storage, although the platelets eventually developed the same in vitro lesions. In all media there was a high correlation between platelet ATP level and the other in vitro parameters during storage. Furthermore, energy depletion of the platelets produced by using the physiologic salt solution without glucose and by addition of metabolic inhibitors caused a similar deterioration of in vitro functions, but at a significantly faster rate which correlated with the rate of decrease in platelet ATP. This suggests that the platelet lesion observed with prolonged storage in second‐generation containers might be due to a failure in platelet energy metabolism.

Viability and in vitro properties of AS-1 red cells after gamma irradiation

BACKGROUND: Irradiation has been shown to adversely affect both in vivo 24‐hour recovery (recovery [%]) and in vitro properties of stored red cells (RBCs). There is uncertainty as to how these changes are related to the day of irradiation and the length of storage after irradiation.

Effects of 3‐5 log10 pre‐storage leucocyte depletion on red cell storage and metabolism

SUMMARY. A new, in‐line high‐efficiency 3‐5 log10 leucodepletion filter system (Leukotrap° RC system) was used to investigate the effect of pre‐storage white cell removal on the quality of AS‐3 red cell concentrates stored for 42 d at 4°. Median residual white cell content was 4 × 105 when filtration was performed at 22° within 8 h of phlebotomy (n= 20) and 3.2 × 104 when filtration was performed at 4° 12‐24 h after phlebotomy (n= 24). None exceeded 1 × 106 WBC per red cell product. Filtration was rapid (median 28 min), and red cell loss averaged (mean ± 1 SD) 6.4 ± 0.7%. In a paired study design, post‐transfusion recoveries of 42 d stored red cells in the filtered units averaged 84 ± 6% v 82 ± 8% for unfiltered units (P < 0.05) and post‐storage haemolysis, ATP, osmotic fragility, K+ and pH were significantly (P < 0.05) better in the filtered units. Reduced glycolytic activity was also observed in the filtered units, and there was a correlation between osmotic fragility, glucose consumption, and lactate produced in standard units that was not present in leucodepleted units. In conclusion, this study suggests that leucodepletion of AS‐3 red cell concentrates prior to storage results in better maintenance of the integrity of the red cell membrane with reduced glycolytic activity. There was a modest improvement in post‐infusion viability sufficient to offset the filtration‐induced loss and to result in an equivalent red cell product.

Post‐transfusion recovery of function of 5‐day stored platelet concentrates

Summary Platelets show a rapid reduction in their responsiveness to aggregating agents during storage for transfusion, but little is known about reversal of this defect in vivo after transfusion. In this study, fresh and stored platelets from the same donor (n=12) were labelled with 111In or 51Cr, respectively, mixed, and simultaneously infused. Blood samples were taken for up to 5 d post‐infusion, and the functional behaviour of the labelled platelets ex vivo was measured by retention on glass bead columns, and by whole blood aggregability to ADP, epinephrine and ristocetin. Aggregation was determined by filtering aggregated samples through a column of cotton wool to remove the aggregates, and quantitated as per cent decrease in radioactive counts. The study showed that, although infused radiolabelled 5 d stored platelets had a significantly lower aggregability towards ADP and epinephrine immediately (1 h) after infusion (32% and 29%, respectively, of fresh platelet values), a complete restoration to fresh platelet levels was found 24–72 h post‐infusion, with no further change observed over the ensuing 5 d with either fresh or stored labelled platelets. A slightly (6–9%) lower adhesion to both uncoated and collagen‐coated beads was found for the stored platelets throughout the 5 d period of study post‐infusion.

Paired comparison of platelet concentrates prepared from platelet‐rich plasma and buffy coats using a new technique with 111In and 51Cr

Two techniques for the preparation of platelet concentrate (PC), the standard platelet‐rich plasma (PRP) and buffy coat (BC) methods, were compared in nine paired studies with regard to platelet harvest, white cell (WBC) contamination, and PC quality after 5 days of 22°C storage. Platelet harvest using the BC method averaged approximately 56 percent of the whole blood level (6.2 × 1010/concentrate), which was less than the 76 percent achieved with the PRP‐PC method (8.7 × 1010/concentrate). An additional 5 units collected into an experimental siphon bag for BC‐PC processing showed improved platelet harvest (6.7 × 1010/concentrate, or approx. 70% of whole blood). WBCs remaining in the BC‐PC averaged 0.19 × 108 per unit compared to 3.6 × 108 per unit for PRP‐PC. Buffy coat processing produced red cell (RBC) units with 50 percent of the WBC contamination of conventionally prepared units (9.8 ± 6.2 × 108/unit vs. 18.9 ± 7.1 × 108/unit). The siphon bag further reduced WBC levels in the AS‐3 RBC units (6.4 ± 3.7 × 108/unit). In vitro studies performed on Days 1 and 5 after collection showed no significant differences in platelet metabolic and biologic function or cell integrity. β‐thromboglobulin and surface glycoprotein levels, indicators of platelet activation and membrane alteration, respectively, did not differ significantly in the PRP‐PC and BC‐PC; nor was lactate production higher in PRP‐PC, despite the substantially higher WBC counts. Autologous in vivo platelet viability determinations were performed by using concurrent transfusion of 111In‐labeled freshly drawn platelets and 51Cr‐labeled stored platelets. Paired f test analysis of BC‐PC versus PRP‐PC indicated no significant differences in platelet recovery and survival after 5 days of 22°C storage in polyolefin containers. Therefore, these studies confirm the equivalence of PC quality, comparable platelet harvest with the siphon bag, and decreased WBC contamination with the BC method.

Concurrent label method with 111In and 51Cr allows accurate evaluation of platelet viability of stored platelet concentrates.

Summary. The precision and reproducibility of 111In and 51Cr platelet radiolabel agents for in vivo kinetic studies of stored platelet concentrates (PC) were investigated. The objective was to develop a precise method with concurrent labelling of two platelet populations using different isotopes, which would allow identification of small differences in in vivo platelet quality. Identical labelling procedures were used to investigate the effects of PC storage age, different methods of red cell (RBC) and white cell (WBC) contamination correction, and label elution correction on the results of 111In and 51Cr kinetic studies.

Storage of apheresis platelets after gamma radiation

BACKGROUND: There are conflicting data on the effect of irradiation and subsequent storage on the quality of platelet components.