Mitsuaki Akino

Reduction in adverse reactions to platelets by the removal of plasma supernatant and resuspension in a new additive solution (M-sol).

BACKGROUND: Leukodepletion reduces but does not eliminate adverse reactions to platelet concentrate (PC). As an alternative strategy, plasma reduction or washing of platelets should be considered. However, the efficacy of this strategy is still unclear.

Comparison between in vitro qualities of platelets washed with commercially available additive solutions and those washed with M-sol.

Background and Objectives  We previously developed a novel additive solution (M‐sol) with a high ability to preserve the in vitro qualities of platelets (PLTs) in washed PLTs Here, we compared the ability of M‐sol with that of commercially available additive solutions (ASs) to preserve the in vitro qualities (pH, mean PLT volume, %disc, P‐selectin, %hypotonic shock response and aggregation) of PLTs at a low plasma concentration.

Replaced platelet concentrates containing a new additive solution, M-sol: safety and efficacy for pediatric patients

Allergic transfusion reactions (ATRs), particularly those caused by plasma‐rich platelet concentrates (P‐PCs), are an important concern in transfusion medicine. Replacing P‐PCs with PCs containing M‐sol (M‐sol‐R‐PCs) is expected to prevent ATRs. However, this has not yet been verified by sufficient clinical evidence.

Platelet additive solution - electrolytes.

Recent attention to solutions that replace most or all plasma in platelet concentrates, while maintaining satisfactory platelet function, is motivated by the potential of plasma reduction or depletion to mitigate various transfusion-related adverse events. This report considers the electrolytic composition of previously described platelet additive solutions, in order to draw general conclusions about what is required for platelet function and longevity. The optimal concentrations of Na(+) and Cl(-) are 69-115 mM. The presence of both K(+) and Mg(2+) in platelet suspension at nearly physiological concentrations (3-5mM and 1.5-3mM, respectively) is indispensable for good preservation capacity because both electrolytes are required to prevent platelet activation. In contrast to K(+) and Mg(2+), Ca(2+) may not be important because no free Ca(2+) is available in M-sol, which showed excellent platelet preservation capacity at less than 5% plasma concentration. The importance of bicarbonate (approximately 40 mM) can be recognized when the platelets are suspended in additive solution under less than 5% residual plasma concentration.

Aggregates in platelet concentrates

P. F. van der Meer, L. J. Dumont, M. Lozano, N. Bondar, J. Wong, S. Ismay, J. Pink, W. Nussbaumer, J. Coene, H. B. Feys, V. Compernolle, D. V. Devine, D. Howe, C. K. Lin, J. Sun, J. Ringwald, E. F. Strasser, R. Eckstein, A. Seltsam, P. Perseghin, P. Proserpio, S. Wakamoto, M. Akino, S. Takamoto, K. Tadokoro, D. Teo, P. H. Shu, S. S. Chua, T. Jimenez-Marco, J. Cid, E. Castro, I. Mu~ noz, H. Gulliksson, P. Sandgren, S. Thomas, J. Petrik, K. McColl, H. Kamel, J. Dugger, J. D. Sweeney, J. B. Gorlin, L. J. Sutor, D. Heath & M. H. Sayers.

Clump formation in apheresis platelet concentrates

Number of Storage period (hours) Apheresis system apheresis PCs examined 0 24 48 72 CS-3000 Plus Blood center 3,203 30.5 12.3 1.4 0.8 Branches 4,375 48.5 15.2 1.6 0.9 Total 7,578 40.9 14.0 1.6 0.8 MCS-3P Blood center 2,933 6.4 0.3 0.0 0.0 Branches 6,723 27.6 2.5 0.0 0.0 Total 9,656 21.1 1.8 0.0 0.0 Amicus Blood center 2,005 22.2 11.5 1.1 0.8 Spectra Blood center 1,426 9.8 3.1 0.1 0.0 Overall total 20,665 27.7 7.3 0.7 0.4 Clump formation in apheresis platelet concentrates

Comparison between bacterial growth in platelets (PLTs) washed with M‐sol and that in PLT‐rich plasma

Platelets (PLTs) washed with additive solution (AS) are useful for the reduction of adverse reactions such as anaphylaxis or febrile nonhemolytic reactions induced by PLT concentrate (PC) transfusion. We previously developed a novel AS (M-sol), which is able to maintain the in vitro qualities of PLTs at a low plasma concentration for a longer period than other commercially available AS. Improvement of AS may lead to extending the shelf life of washed PLTs. Since M-sol can be aseptically prepared by simply mixing commercially available solutions approved for clinical use, this solution is already being used to prepare washed PLTs at some blood centers and hospitals in Japan. Bacterial contamination is currently the most frequent infectious risk related to transfusion. Due to storage at room temperature, PCs are more prone to grow bacteria than red blood cells. Since plasma contains certain factors such as complement components that inhibit bacterial growth, decreasing the plasma concentration by washing of PC may promote bacterial growth in washed PLTs. Here we compared bacterial growth in PLTs washed with M-sol with that in PLT-rich plasma (PRP) during a 7-day storage period. Apheresis PC, which was processed 4 or 5 days after collection, was divided into two equal aliquots (control group and test group). After centrifugation (2560 ¥ g, 10 min) of both aliquots and removal of as much supernatant as possible, the pellet of the control group was resuspended in 170 mL of ABO-identical fresh-frozen plasma that had been prepared from CPD whole blood within 6 hours after donation and stored at -20°C until use. In the test group, the pellet was resuspended in 170 mL of M-sol. The control group (176 8 mL, 109 ¥ 10 18 ¥ 10/mL, 5 units under Japanese standard, plasma concentration 100%) and test group (175 7 mL, 104 ¥ 10 18 ¥ 10/mL, 5 units, plasma concentration 4.54%) were stored at 20 to 24°C on a flatbed shaker (50-60 cycles/min) in a polyolefin bag (KBP600FPN, Kawasumi Co., Ltd, Tokyo, Japan). The bacterial strains used in this study were: Bacillus cereus (ATCC#10876), Propionibacterium acnes (ATCC#6919), Staphylococcus epidermidis (ATCC#49134), Staphylococcus aureus (NBRC#13276), Escherichia coli (ATCC#11775), and Serratia marcescens (ATCC#14756). Fresh bacterial culture was prepared by incubating a single colony in liquid medium (BacT/ALERT BPA or BPN, bioMérieux, Inc., Durham, NC). Bacterial culture diluted with saline was inoculated into the control group and test group so that the final concentration of bacteria in each group was less than 10 colony-forming units (CFUs)/mL (Day 0). The PLT suspensions including bacteria were stored with agitation at 20 to 24°C until Day 7. To count bacterial colonies, aliquots sampled from the test group and the control group were plated as undiluted samples (200 mL/ plate ¥ five plates) and as serial 1-in-10 dilutions (100 mL/ plate) on agar plates. The aerobic bacteria were incubated for 24 hours or more at 37°C under aerobic conditions on agar plates (Trypto-Soya agar plate, Nissui Pharmaceutical Co. Ltd, Tokyo, Japan). P. acnes was incubated on blood agar plates (Brucella HK agar plate, Kyokuto Pharmaceutical Industrial Co. Ltd, Tokyo, Japan) for 7 days at 30°C under anaerobic conditions. Two-tailed paired t test was used for analysis of differences between the control group and test group. Significance was accepted at a p value less than 0.05. Bacterial concentrations of S. epidermidis in the test group were significantly lower than those in the control group on Days 3 and 4 (Table 1). Similarly, bacterial concentrations of S. aureus in the test group were significantly lower than those in the control group on Days 1 and 2 (Table 1). Previously, the decreased iron concentration in PC due to iron chelation was reported to suppress the growth of S. epidermidis and S. aureus, indicating that iron is a critical factor for the proliferation of these two bacteria. Therefore, growth suppression of those bacteria by washing is thought to be due to a reduction of the essential nutritional content. The exhaustion of essential nutrients by plasma removal may have larger effects on the growth of S. epidermidis and S. aureus than the reduction of suppressive effects of plasma components on bacterial growth. Furthermore, it was reported that pH increase in PLT suspension inhibits the proliferation of S. epidermidis. The pH of PLTs washed with M-sol (7.47-7.64) is higher than that of PRP (6.91-7.09) 24 hours after washing. Therefore, the higher pH may be another reason for the growth suppression of S. epidermidis in the test group. Bacterial concentrations of B. cereus in both groups were comparable on Days 2, 3, and 7, while those in the test group were significantly lower than those in the control group on Day 1, which may be due to a delay in entering the vegetative state in M-sol compared to that in plasma. The growth of P. acnes (anaerobic bacteria) was negligible in both groups during 7 days of storage under aerobic conditions (Table 1). In case of E. coli and S. marcescens, bacterial concentrations in the test group were increased in a time-dependent manner, while that in the control group was negligible or under the detectable limit during 7 days of storage (Table 1). Since Gramnegative bacteria are susceptible to growth inhibition

Storage of volume-reduced washed platelets in M-sol additive solution for 7 days

Volume‐reduced washed platelets (VR‐wPLTs), which are prepared by concentrating platelets (PLTs) into a smaller volume of additive solution (AS), may prevent not only circulatory overload, but also adverse reactions caused by plasma components. Although VR‐wPLTs may be quickly degraded due to high PLT concentrations, few studies have examined the effects of storage on VR‐wPLTs. We examined here the in vitro properties of VR‐wPLTs prepared with M‐sol AS during their storage for 7 days.

Effects of helicopter transport on red blood cell components

BACKGROUND There are no reported studies on whether a helicopter flight affects the quality and shelf-life of red blood cells stored in mannitol-adenine-phosphate. MATERIALS AND METHODS Seven days after donation, five aliquots of red blood cells from five donors were packed into an SS-BOX-110 container which can maintain the temperature inside the container between 2 °C and 6 °C with two frozen coolants. The temperature of an included dummy blood bag was monitored. After the box had been transported in a helicopter for 4 hours, the red blood cells were stored again and their quality evaluated at day 7 (just after the flight), 14, 21 and 42 after donation. Red blood cell quality was evaluated by measuring adenosine triphosphate, 2,3-diphosphoglycerate, and supernatant potassium, as well as haematocrit, intracellular pH, glucose, supernatant haemoglobin, and haemolysis rate at the various time points. RESULTS During the experiment the recorded temperature remained between 2 and 6 °C. All data from the red blood cells that had undergone helicopter transportation were the same as those from a control group of red blood cell samples 7 (just after the flight), 14, 21, and 42 days after the donation. Only supernatant Hb and haemolysis rate 42 days after the donation were slightly increased in the helicopter-transported group of red blood cell samples. All other parameters at 42 days after donation were the same in the two groups of red blood cells. DISCUSSION These results suggest that red blood cells stored in mannitol-adenine-phosphate are not significantly affected by helicopter transportation. The differences in haemolysis by the end of storage were small and probably not of clinical significance.

Influence of a 24-hour interruption of agitation on in vitro properties of platelets washed with M-sol during 7-day storage

We would like to make a contribution toward the recent publication in TRANSFUSION by Mestra and coworkers. Firstly, the authors said that this is the first report of transfusion-transmitted leishmaniasis in an immunocompromised patient in Colombia. However, identification of the Leishmania infection was demonstrated neither in transfused blood components nor in blood donors. Antibodies against Leishmania spp. were detected in the patient’s serum samples as part of a study to rule out renal transplant-transmitted visceral leishmaniasis. However, it is not clear if the serologic Leishmaniapositive result of the patient’s serum corresponded to a sample taken before or after receiving transfusions. To our understanding, if no data related to the patient’s infection status before the transfusions have been clearly provided and, additionally, as Leishmania infection in transfused blood components and/or blood donors has not been demonstrated by the authors, it is very difficult to agree that this certainly represents a case of transfusiontransmitted visceral leishmaniasis. From the transfusion point of view it would have been interesting to know whether or not the transfused blood components were leukoreduced and, if so, when the leukoreduction was performed, that is, before storage or at the bedside, since various studies have been published on the efficacy of whole blood, fresh plasma, and red blood cell filters used for leukoreduction in reducing the number of Leishmania spp. parasites in blood components and thereby minimizing the potential risk of Leishmania transmission through blood transfusions. This information could have thrown light on the probability of the parasite being transmitted to the patient through the transfusion of a contaminated blood component taken from an asymptomatic donor. This case was also published in 2006 as allograft kidney dysfunction associated with infection with amastigotes of Trypanosoma cruzi. However, neither polymerase chain reaction (PCR) nor any other antigen detection technique was performed to demonstrate that the amastigotes in the renal parenchymal corresponded to T. cruzi. To distinguish Chagas disease from Leishmania infection, it would have been helpful to perform Leishmaniaand T. cruzi– specific PCRs on the renal biopsy since it is difficult to differentiate the amastigotes from these two genera by direct microscopic examination. This observation has implications related to the article being commented on because it would explain the patient’s clinical symptoms showing after the renal transplant and immediately before initiating the transfusions. Nifurtimox was given to treat Chagas disease. This may have also contributed to delaying the diagnosis of visceral leishmaniasis, since this medication is partially effective against Leishmania spp. as in vivo studies have shown. All these aspects could also be compatible with the reactivation of a latent Leishmania infection by immunosuppression. We believe that this is a very interesting report concerning a case of visceral leishmaniasis caused by L. (L.) mexicana in an immunocompromised patient in Colombia. This is already in itself a very remarkable contribution made by the authors because, as mentioned in the article, L. (L.) mexicana is a species that has been associated mainly with cutaneous leishmaniasis. There are, however, various aspects that might support the fact that this case was due to the reactivation of a latent Leishmania infection, that is, the patient’s clinical symptoms, the lack of data related to the patient’s infection status before the transfusion, the blood components transfused, and their donors. Similarly, other aspects, which are primarily based on ruling out the transmission of visceral leishmaniasis through solid-organ transplantation, and the fact that the patient’s medical interview did not reveal any prior exposure to Leishmania, all support the hypothesis that the visceral leishmaniasis was transmitted through blood transfusion. Since there are arguments both for and against the two options, we would suggest that this report be considered as a possible case of transfusion-transmitted visceral leishmaniasis.