Platelet Storage Lesion Research Articles

Platelet Storage-Problems, Improvements, and New Perspectives.

Platelet transfusions are routine procedures in clinical treatment aimed at preventing bleeding in critically ill patients, including those with cancer, undergoing surgery, or experiencing trauma. However, platelets are susceptible blood cells that require specific storage conditions. The availability of platelet concentrates is limited to five days due to various factors, including the risk of bacterial contamination and the occurrence of physical and functional changes known as platelet storage lesions. In this article, the problems related to platelet storage lesions are categorized into four groups depending on research areas: storage conditions, additive solutions, new testing methods for platelets (proteomic and metabolomic analysis), and extensive data modeling of platelet production (mathematical modeling, statistical analysis, and artificial intelligence). This article provides extensive information on the challenges, potential improvements, and novel perspectives regarding platelet storage.

The effect of agitating buffy coats on platelet quality before soft spin

BackgroundPlatelet plays a vital role in both physiological and pathological processes. However, the limited storage time of platelet in vitro poses an immense challenge for its applications because of the increased risk of bacterial contamination and platelet storage lesions. Agitation can inhibit lesions by facilitating continuous oxygenation of platelets and permitting excess carbon dioxide to be removed during storage. However, it is still not known whether agitating BCs gives a positive effect on platelet quality. Objectives: To evaluate the quality difference between platelet concentrates (PCs) from buffy coats (BCs) held rest and agitation. MethodsSamples were withdrawn for cell count, blood gas analysis, free hemoglobin level, hypotonic shock response, maximum aggregation rate, activation marker expression (CD62P and CD42b) and coagulation function. Results: We found the PCs prepared from the agitating BCs had fewer residual WBCs, exhibited a better gas exchange ability, slower metabolism (higher pH, higher content glucose, and lower lactic acid levels), better hypotonic shock response, and lower levels of CD62P. The TEG-PC assays showed no difference in coagulation function. Conclusion: Our findings showed that BC can be agitated overnight before a soft spin.

Shedding light on GPIbα shedding.

Ectodomain shedding has been investigated since the late 1980s. The abundant and platelet specific GPIbα receptor is cleaved by ADAM17 resulting in the release of its ectodomain called glycocalicin. This review will address the role of glycocalicin as an end-stage marker of platelet turnover and storage lesion and will consider a potential function as effector in processes beyond hemostasis. Glycocalicin has been described as a marker for platelet senescence, turnover and storage lesion but is not routinely used in a clinical setting because its diagnostic value is nondiscriminatory. Inhibition of glycocalicin shedding improves posttransfusion recovery but little is known (yet) about potential hemostatic improvements. In physiological settings, GPIbα shedding is restricted to the intracellular GPIbα receptor subpopulation suggesting a role for shedding or glycocalicin beyond hemostasis. So far, all evidence represents glycocalicin as an end-stage biomarker of platelet senescence and a potential trigger for platelet clearance. The extensive list of interaction partners of GPIbα in fields beyond hemostasis opens new possibilities to investigate specific effector functions of glycocalicin.

The Effect of Leukocyte Removal and Matrix Metalloproteinase Inhibition on Platelet Storage Lesions.

The reasons for unfavorable changes in platelet concentrate (PC) quality during storage are not fully understood yet. We aimed to evaluate whether leukocytes and matrix metalloproteinases (MMPs) lead to a decrease in the quality of PCs and examine whether MMP inhibition will slow down the platelets' aging. Nine PCs were divided into three parts: (1) leukocyte-depleted (F) PCs, (2) PCs with no additional procedures (NF), and (3) PCs with the addition of an MMP inhibitor-doxycycline (D). Each PC was stored for 144 h, and a sample for testing was separated from each part on the day of preparation and after 24, 48, 72 and 144 h of storage. Blood morphological analysis, platelet aggregation, and the expression of activation markers were evaluated. MMP-2 and MMP-9 concentration, activity, and gene expression were assessed. Platelet aggregation decreased, and platelet activation marker expression increased during the storage. D concentrates showed the lowest level of platelet activation. In turn, leukocyte-depleted PCs showed the highest level of platelet activation in general. MMP-9 platelet activity was higher in leukocyte-containing concentrates at the end of the storage period. We concluded that the filtration process leads to a higher platelet activation level. The presence of doxycycline in PCs reduces the expression of the activation markers as compared to leukocyte-depleted concentrates.

A tight interplay between platelet activation and mitochondrial DNA release promotes platelet storage lesion in platelet concentrates.

Platelet storage lesion (PSL) adversely affects the quality of platelet concentrates (PCs). Platelets are prone to activation during storage. Moreover, elevated free mitochondrial DNA (mtDNA) levels in PCs are associated with a higher risk of adverse transfusion reactions. Therefore, we aimed to evaluate the correlation between platelet activation markers and mtDNA release during PC storage. Six PCs prepared by the platelet-rich plasma method were assessed for free mtDNA copy number using quantitative real-time PCR and CD62P (P-selectin) expression by flow cytometry on days 0 (PC collection day), 3, 5 and 7 of storage. Lactate dehydrogenase (LDH) activity, pH, platelet count, mean platelet volume (MPV) and platelet distribution width (PDW) were measured as well. The correlation between free mtDNA and other PSL parameters, and the correlation between all parameters, was determined. Significant increases in free mtDNA, MPV and PDW, and a significant decrease in platelet count and pH were observed. CD62P expression and LDH activity elevated significantly, particularly on storage days 5-7 and 0-3, respectively. Moreover, a moderate positive correlation (r = 0.61) was observed between free mtDNA and CD62P expression. The r values between free mtDNA and LDH, pH, platelet count, MPV and PDW were 0.81, -0.72, -0.49, 0.81 and 0.77, respectively. The interplay between platelet activation and mtDNA release in promoting PSL in PCs may serve as a promising target for future research on applying additive solutions and evaluating the quality of PCs to improve transfusion and clinical outcomes.

A comparative study of platelet storage lesion in platelet-rich plasma under cryopreservation.

Platelet-rich plasma (PRP) has significant potential for various applications and holds clinical value in regenerative medicine. Cryopreservation is used to extend the preservation period of PRP, facilitating its clinical application. However, the potential negative effects of long-term cryopreservation on platelet storage lesion are still uncertain. In this study, PRP was stored at - 30°C or - 80°C. Platelet count, apoptosis, reactive oxygen species (ROS) content, and CD62P expression were assessed on the 14th and 28th days. The study also evaluated platelet mitochondria morphology and function, serotonin (5-HT) secretion by platelets, and the inflammatory activating effect of cryopreserved platelets in PRP. The results showed that there were no significant differences in platelet count, the content of 5-HT, and inflammatory effects between fresh PRP and PRP cryopreserved at both - 30°C and - 80°C. However, there was an increase in ROS level, apoptosis, and CD62P level after cryopreservation at both temperatures. Additionally, the levels of ROS, apoptosis, and CD62P in platelets were similar after storage at - 30°C and - 80°C. The main difference observed was that the morphology and function of mitochondria were severely damaged after storage at - 30°C, while they were less affected at - 80°C. Based on these findings, it can be concluded that storing PRP at - 80°C is more suitable for achieving a better therapeutic effect in clinical applications, but cryopreservation could not replace the current standard.

The platelet storage lesion, what are we working for?

Platelet concentrate (PC) transfusions are crucial in prevention and treatment of bleeding in infection, surgery, leukemia, and thrombocytopenia patients. Although the technology for platelet preparation and storage has evolved over the decades, there are still challenges in the demand for platelets in blood banks because the platelet shelf life is limited to 5 days due to bacterial contamination and platelet storage lesions (PSLs) at 20-24°C under constant horizontal agitation. In addition, the relations between some adverse effects of platelet transfusions and PSLs have also been considered. Therefore, understanding the mechanisms of PSLs is conducive to obtaining high quality platelets and facilitating safe and effective platelet transfusions. This review summarizes developments in mechanistic research of PSLs and their relationship with clinical practice, providing insights for future research. Authors conducted a search on PubMed and Web of Science using the professional terms "PSL" and "platelet transfusion." The obtained literature was then roughly categorized based on their research content. Similar studies were grouped into the same sections, and further searches were conducted based on the keywords of each section. Different studies have explored PSLs from various perspectives, including changes in platelet morphology, surface molecules, biological response modifiers (BMRs), metabolism, and proteins and RNA, in an attempt to monitor PSLs and identify intervention targets that could alleviate PSLs. Moreover, novel platelet storage conditions, including platelet additive solutions (PAS) and reconsidered cold storage methods, are explored. There are two approaches to obtaining high-quality platelets. One approach simulates the invivo environment to maintain platelet activity, while the other keeps platelets at a low activity level invitro under low temperatures. Understanding PSLs helps us identify good intervention targets and assess the therapeutic effects of different PSLs stages for different patients.

Liposomal Resveratrol Alleviates Platelet Storage Lesion via Antioxidation and the Physical Buffering Effect.

Platelet transfusion is essential in the treatment of platelet-related diseases and the prevention of bleeding in patients with surgical procedures. Platelet transfusion efficacy and shelf life are limited mainly by the development of platelet storage lesion (PSL). Mitigating PSL is the key to prolonging the platelet shelf life and reducing wastage. Excess intracellular reactive oxygen species (ROS) are one of the main factors causing PSL. In this study, we explored a nanomedicine strategy to improve the quality and functions of platelets in storage. Resveratrol (Res), a natural plant product, is known for its antioxidative effect. However, medical applications of Res are limited due to its low water solubility and stability. Therefore, we used a resveratrol-loaded liposomal system (Res-Lipo) to better utilize the antioxidant effect of the drug. This study aimed to evaluate the effect of Res-Lipo on platelet oxidative stress and alleviation of PSL during the storage time. Res-Lipo scavenged intracellular ROS and inhibited platelet apoptosis and activation during storage. Res-Lipo not only maintained mitochondrial function but also improved platelet aggregation in response to adenosine 5'-diphosphate. These results revealed that Res-Lipo ameliorated PSL and prolonged the platelet survival time in vivo. The strategy provides a potential method for extending the platelet storage time and might be considered a potential and safe additive to alleviate PSL.

Evaluation of Platelet Storage Lesion (PSL) Markers in SSP+ Platelet Additive Solution Added Canine Platelet Concentrates (CPC)

Background: To evaluate the Platelet Storage Lesion (PSL) markers and extended storage time for SSP+ added Canine Platelet Concentrates (CPC) beyond five days at 22°C.Fresh Canine CPCundergo certain metabolic and morphologic changes during storage. Estimation of PSL markers helps us better understand quality of the canine PC and to estimate the efficacy of SSP+ in increasing the life span of stored CPC. Method:CPC was prepared from 350 ml of whole blood, collected from 6 apparently healthy donor dogs by buffy coat methodin a quadruple bag closed system. CPC was divided into four aliquots of 15 ml each with one control and three test groups with SSP+ additive solution in different concentrations (65%, 75%, 85%). TheIn vitro PSL markers including swirling, pH, glucose, lactate, bicarbonate, pO2, pCO2, platelet concentration, MPV and PDWwere evaluated on day 1, 5, 9 and 13 of storage. Result:The results were analysed by Two-way ANOVA wherein swirling, pH, glucose, lactate, bicarbonate, pO2, pCO2, platelet concentration, MPV showed significant difference (p less than 0.01) between the groups. Interaction between the additive concentrations and the days of storage revealed a pH on day 9 for 65% SSP+ to be similar to day 5 for Plasma stored CPC while swirling, lactate and bicarbonate were also better maintained for 65% SSP+ PAS added CPC till day 9.Addition of 65% SSP+ PAS to CPC evidenced increased shelf life up to 9 days at 22°C under agitation without significant deterioration in product quality.

Platelet Storage: Time to Rethink the Cold

Platelet concentrates (PCs) are widely used in transfusion medicine for therapeutic purposes and their demand is constantly on the rise. Current storage regulations make this product highly susceptible to bacterial contaminations and platelet storage lesions (PSL) causing the need for alternative storage methods to be considered. The implementation of cold storage not only reduces unnecessary wastage of valuable donations and overall costs but also decreases both the risk of bacterial contamination and the occurrence of PSL. The current study aimed at determining how a prolonged cold storage may affect PCs. This was accomplished by investigating two different PC cohorts of 10 units each. One of the cohorts, labelled as ‘Room Temperature’, was stored at 22℃±2℃ for 5 days and then transferred to a temperature of 4℃±2℃. The other cohort, labelled as ‘Cold’, was stored directly at 4℃±2℃. Both cohorts were stored for a total of 21 days and platelet indices, platelet counts, pH, and platelet factor IV (PF4) were measured at different time intervals. Sterility was performed on Day 21. The key findings showed no significant difference in mean platelet count, platelet distribution width (PDW), mean platelet volume (MPV), platelet-large cell ratio (P-LCR), and plateletcrit (PCT) between the two cohorts. On the other hand, a significant difference in mean pH and PF4 resulted between the two cohorts. Moreover, no significant difference in mean platelet count, and PCT was found between Days 1, 5, or 10, and Day 21 in both cohorts. However, an overall significant difference in mean PDW, MPV, P-LCR and pH was discovered between Days 1 and 21, Days 5 and 21, and Days 10 and 21 in both cohorts. Regards PF4, a significant difference was detected between Days 1 and 21, and Days 10 and 21; however, no significant difference was found between Days 5 and 21 in both cohorts. Corynebacterium freneyi and Microbacterium liquefaciens, were cultured from 1 unit of the room temperature cohort after it was flagged positive during the sterility testing. In conclusion, through the implementation of a delayed cold storage system, PCs can be safely administered to the patient.

Resveratrol reduces platelet storage lesion by preventing free mitochondrial DNA (mtDNA) accumulation in platelet concentrates during storage.

Platelet activation and mitochondrial damage are among the crucial events leading to the quality reduction of platelet concentrates (PCs) during preparation and storage, called platelet storage lesion. Platelet activation results in the clearance of transfused platelets. Oxidative stress and platelet activation trigger mitochondrial DNA (mtDNA) release into the extracellular milieu which is associated with adverse transfusion reactions. Therefore, we aimed to investigate the effects of resveratrol, an antioxidant polyphenol, on platelet activation markers and mtDNA release. Ten PCs were divided equally into two bags each, one of them was allocated to the control group (n = 10) and another to the case group (resveratrol-treated, n = 10). Free mtDNA level and CD62P (P-selectin) expression level were measured by absolute quantification Real-Time PCR, and flow cytometry on days 0 (the receiving day), 3, 5, and 7 of storage respectively. Moreover, Lactate dehydrogenase (LDH) enzyme activity, pH, platelet count, mean platelet volume (MPV), and platelet distribution width (PDW) were assessed as well. Treatment of PCs with resveratrol can significantly decrease mtDNA release during storage compared to the control. In addition, platelet activation was significantly mitigated. We also observed significantly lower MPV, PDW, and LDH activity in resveratrol-treated PCs compared to the control group on days 3, 5, and 7. Furthermore, resveratrol maintained the pH of PCs on day 7.Resveratrol diminished free mtDNA and maintained biochemical parameters in PCs, possibly by reducing platelet activation. Therefore, resveratrol might be a possible additive solution for improving the quality of stored PCs.

Whole transcriptome analysis of platelet concentrates during storage.

Platelets are anucleated blood cells and contain various RNA species. We investigated the changes in the whole transcriptome expression profile of platelet concentrates (PC) during storage to explore biological functions and biomarkers in platelet storage damage. Platelets were collected by apheresis from eight healthy blood donors and stored from day 0 to day 4. Platelet phenotyping and function analysis were used to detect platelet activity during storage. RNA-sequencing was used to detect changes in expression of mRNA, lncRNA and circRNA in the PC during storage. Gene ontology and KEGG analyses were applied to predict the functional distribution of differential expression of mRNA. Gene set enrichment analysis was used to analyze the differential levels of gene pathways. Finally, polymerase chain reaction (PCR) analysis was performed to verify the expression of three mRNA (POLE2, DCUN1D4, DAD1). In total, 10,767 mRNA, 2,923 lncRNA and 68,550 circRNA were detected in the PC by RNA-sequencing. The expression levels of 222 mRNA changed significantly from day 0 to day 4 of storage: 58 increased continuously and 145 decreased continuously. Differentially expressed mRNA may be involved in physiological processes such as platelet activation, platelet aggregation, endocytosis, and apoptosis. Expression levels of 1,413 lncRNA were obvious. The levels of 42 species increased and the levels of 28 species decreased. The expression levels of 198 species of circRNA changed significantly, with those of 13 species changing continuously. The differential levels of expression of DAD1, DCUN1D4 and POLE1 mRNA, shown by RNA sequencing, were validated by PCR assay. Changes in mRNA, lncRNA and circRNA during platelet storage may be closely related to platelet apoptosis and physiological functions in the platelet storage lesion. The expression levels of DAD1, DCUN1D4 and POLE1 could be biomarkers to monitor platelet status in PC bags.

Different Expression Profiles of Exosomal circRNAs from Apheresis Platelets during Storage

Introduction: Bioactive substances of stored platelets change during the stored periods. Exosomes are reported to be increased during platelet storage. Circular RNAs (circRNAs) are one of the main components in exosomes. It is the purpose of this study to investigate the different expression of exosomal circRNAs during storage. Methods: Apheresis platelets were collected from 7 healthy volunteers and stored in platelet storage bags for 1 day or 5 days. We isolated exosomes by ultracentrifugation and characterized exosomes by transmission electron microscopy, nano-flow cytometry, and Western blot. We conducted microarray analysis to detect changes in the exosomal circRNAs from apheresis platelets during storage, and qRT-PCR to validate their expressions. To analyze the competing endogenous RNA (ceRNA) of circRNAs, microRNAs (miRNAs) targets were predicted based on interactions of circRNAs/miRNAs and miRNAs/mRNAs, using TargetScan and miRanda. A ceRNA network was constructed by Cytoscape. The targeted mRNAs were performed for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genome (KEGG) pathway analysis by the DAVID. Results: Microarray analysis revealed that 61 differentially expressed circRNAs between day 1 and day 5. Thirty-one circRNAs of these are upregulated, while 30 circRNAs are downregulated. A ceRNA visualized network includes 9 circRNAs, 48 miRNAs, and 117 mRNAs. There were 117 mRNAs enriched in 203 GO terms and 9 KEGG pathways based on the GO and KEGG pathway enrichment analyses. Conclusion: We identified 61 dysregulated exosomal circRNAs from apheresis platelets during storage. The study provided insights into the underlying mechanisms of platelet storage lesion.

The evaluation of reactive oxygen species generation and free mitochondrial DNA in platelet concentrates during 5 days of storage.

Oxidative stress and mitochondrial damage are causes of platelet storage lesions (PSLs). Mitochondrial damage causes mitochondrial DNA (mtDNA) to be released into the extracellular space. MtDNA in platelet concentrates is considered damage-associated molecular patterns (DAMPs) and is one of the major causes of PSLs. The mechanism of mtDNA release in platelet concentrates has not been thoroughly investigated. This study aimed to determine the effect of reactive oxygen species (ROS) on mtDNA release in platelet concentrates during storage. Ten platelet concentrates from healthy donors were obtained in this investigation. Platelet concentrates were prepared by platelet-rich plasma (PRP) and stored at 22 ± 2 C° with gentle agitation. Platelet concentrates were subjected to flow cytometry and real-time PCR to evaluate total ROS and free mtDNA on days 0, 3, and 5 of platelet concentrate storage. Total ROS detected significantly increased from day 0 to day 5 of platelet concentrate storage (P = 0.0079). The mean of copy numbers of free mtDNA on day 0 increased from 3.43 × 106 ± 1.57 × 106 to 2.85 × 107 ± 1.51 × 107 (molecules/μl) on the fifth day of platelet concentrate storage, and it was statistically significant (P = 0.0039). In addition, LDH enzyme activity significantly increased during platelet concentrate storage (P < 0.0001). Also, releasing mtDNA in platelet concentrates was directly correlated with total ROS generation (P = 0.021, r = 0.61) and LDH activity (P = 0.04, r = 0.44). The evidence from this study confirmed the increasing level mtDNA copy numbers in platelet concentrates during storage, and the amount of free mtDNA is directly correlated with ROS generation and platelet lysis during 5 days of platelet concentrate storage. Finally, these changes may be related to DAMPs in the platelet concentrates.

L-carnitine modulates free mitochondrial DNA DAMPs and platelet storage lesions during storage of platelet concentrates.

Platelet storage lesions may occur in Platelet concentrates (PCs) storage time, reducing PCs' quality. Mitochondrial damage causes mitochondrial DNA (mtDNA) to be released into the extracellular space. In this study, we evaluated the effect of L-carnitine (LC) as an antioxidant on free mtDNA DAMPs release in PCs during storage. Ten PCs prepared by the PRP method were studied. The copy numbers of free mtDNA, total reactive oxygen species (ROS), lactate dehydrogenase (LDH) enzyme activity, pH, and platelet counts were measured on days 0, 3, 5, and 7 of PCs storage in LC-treated and untreated platelets. LDH activity was significantly lower than the control group during 7 days of PCs storage (p = 0.041). Also, ROS production decreased in LC-treated PCs compared to the control group during storage (p = 0.026), and the difference mean of ROS between the two groups was significant on day 3, 5, and 7 (Pday3 = 0.02, Pday5 = 0.0001, Pday7 = 0.031). Moreover, LC decreased the copy numbers of free mtDNA during 7 days of storage (p = 0.021), and the difference mean of the copy numbers of free mtDNA in LC-treated PCs compared to the control group was significant on day 5 and 7 (Pday5 = 0.041، Pday7 = 0.022). It seems that LC can maintain the metabolism and antioxidant capacity of PCs and thus can reduce mitochondrial damage and mtDNA release; consequently, it can decrease DAMPs in PCs. Therefore, it may be possible to use this substance as a platelet additive solution in the future.

Contamination of platelet concentrates with Staphylococcus aureus induces significant modulations in platelet functionality

Platelet concentrates (PCs) contaminated with Staphylococcus aureus can escape detection during PC screening, causing septic transfusion reactions. This study aimed to determine the impact of S. aureus contamination on platelet metabolism and functionality during PC storage. Targeted metabolomics (N=3) was performed on non-spiked PCs and PCs inoculated with 10-20 colony-forming units (CFU)/bag of S. aureus. Metabolites were quantified at 0, 48 and 144 h using high-performance mass spectrometry (MS). Additionally, PCs spiked with approximately 20 CFU/bag of S. aureus were sampled every 24 h for up to 144 h to evaluate platelet functionality using flow cytometry (N=2). Eight metabolites had significantly different levels in spiked PCs (log2 fold-change ≤ or ≥±1) versus non-spiked units at 48 and 144 h. Xanthine, uridine, serine, glutamine and threonine were increased, whereas orotic acid, dihydroorotic acid and aspartic acid were decreased. Flow cytometry showed a significant decrease in expression of GPIIb while P-selectin expression was significantly increased in spiked PCs after 72 h of storage when S. aureus concentration was ≥10E+08 CFU/ml. Additionally, phosphatidylserine exposure was significantly increased after 48 h of PC storage, when S. aureus had reached a concentration of 2E+06. Contamination with S. aureus exacerbates platelet storage lesions in contaminated PCs but only when the bacterium has reached clinically significant levels.

CPT2 K79 acetylation regulates platelet life span

AbstractThe short life span of platelets is a major challenge to platelet transfusion services because of the lack of effective intervention. Here, we found that the accumulation of long-chain acylcarnitines (LCACs) is responsible for mitochondrial damage and platelet storage lesion. Further studies showed that the blockade of fatty acid oxidation and the activation of AMP-activated protein kinase (AMPK)/acetyl-CoA carboxylase/carnitine palmitoyltransferase 1 (CPT1) pathways that promote fatty acid metabolism are important reasons for the accumulation of LCACs. The excessive accumulation of LCACs can cause mitochondrial damage and a short life span of stored platelets. The mechanism study elucidated that NAD+ exhaustion and the subsequent decrease in sirtuin 3 (Sirt3) activity caused an increase in the level of CPT2 K79 acetylation, which is the primary cause of the blockade of fatty acid oxidation and the accumulation of LCACs. Blocking LCAC generation with the inhibitors of AMPK or CPT1, the agonists of Sirt3, and antioxidants tremendously retarded platelet storage lesion in vitro and prolonged the survival of stored platelets in vivo posttransfusion with single or combined use. In summary, we discovered that CPT2 acetylation attenuates fatty acid oxidation and exacerbates platelet storage lesion and may serve as a new target for improving platelet storage quality.

Role of MicroRNA-326 and its Target Genes Bcl-xL and Bak as Potential Markers in Platelet Storage Lesion in Blood Banks

Platelet transfusion is crucial in the management of various conditions such as quantitative and qualitative platelet disorders. A serious problem that impacts public health is the shortage of Platelet concentrates (PCs) that frequently affect few blood donors’ countries, such as Egypt. This has necessitated the need to establish novel standards for determining the quality of PC during storage. It was found that microRNAs (miRNA) differential expression profile is a helpful tool for recognition of physiological platelet changes during storage. The aim of the current study was to highlight the role of platelet miRNA-326 and its putative target apoptotic genes, Bcl-xL and Bak, and their role in platelet storage lesion (PSL). Differential expression of miRNA-326 and its target genes in the apoptotic pathway, Bcl-xL and Bak was done using quantitative real time PCR (QR-PCR) on different storage points at day 0, day 3 and day 5 in blood bank. The results of the current study revealed over expression of miRNA-326 throughout days of storage resulted in down regulation of Bcl-xL gene and subsequently up regulation of Bak gene. MiRNA-326 contributes to platelet apoptosis and PSL through inhibition of anti-apoptotic Bcl-xL expression and enhancing pro-apoptotic Bak expression. Differential miRNA-326 and its target gene, Bcl-xL and Bak, expression levels at different points of platelets storage are promising tools as biomarkers for platelets undergoing PSL in blood banks.

The effects of additive solutions on the development of storage lesions in canine platelet concentrates stored at 4°C.

To assess platelet storage lesion development as evaluated by measurement of metabolic markers, platelet activation markers, and aggregometry, and determine the occurrence of bacterial growth in platelets stored in platelet additive solution (PAS) at 4°C for 7 days. Prospective, ex vivo experimental controlled study. Research laboratory of a university veterinary teaching hospital. Ten units of canine platelet concentrate collected from blood bank donations. Concentrates were aliquoted into 4 separate bags containing 100% plasma (control) or 30% plasma and 70% of a PAS (Plasma-Lyte A, Isoplate, or InterSol). Samples were stored at 4°C without agitation. At days 0, 3, 5, and 7, samples were analyzed for platelet count, mean platelet volume, glucose, lactate, lactate dehydrogenase, Po2 , Pco2 , degree of swirling, aggregate formation, aggregation via light aggregometry, surface P-selectin via flow cytometry, and bacterial contamination via culture. Development of storage lesions was minimal, demonstrated by maintenance of a mean pH>7.2 and mean lactate values <6mmol/L at day 7 in all solutions. Glucose utilization did not vary significantly between any of the solutions.No significant difference was found between plasma and PAS for Po2 and Pco2 . P-selectin expression measured via flow cytometry showed a low platelet activation percent in all the solutions. InterSol had the lowest mean maximum percent aggregation (P<0.001) and Isoplate the highest (P<0.05). The mean maximum percent aggregation increased between day 0 and day 7 in all solutions. No bacterial growth was found in any of the solutions. Overall, PASs were comparable to plasma for the cold storage of platelets. Cold-stored platelets showed minimal storage lesion development with no bacterial growth. Plasma-, Plasma-Lyte A-, and Isoplate-stored platelets maintained function for up to 7 days at 4°C.

Abstract 158: Bioactive Lipids As Biomarkers Of Severe Adverse Reaction Associated With Apheresis Platelet Concentrate Transfusion

Blood transfusion is a life-saving procedure in which whole blood, or blood components are provided to a patient. Platelet concentrates (PC) may sometimes induce adverse reactions, that are occasionally severe. PC comprise active biomolecules such as cytokines and lipid mediators. Platelet storage lesions are structural and biochemical changes in PC and vary in collection and processing conditions. We investigated lipid mediators as bioactive molecules along storage conditions and in adverse reactions after transfusion. The PC left over were analysed after transfusion, classified with/out severe reaction. Our data presented a decrease of lysophosphatidylcholine species to produce lysophosphatidic acid species. This balance could lead and be correlated with complication symptoms, as other lipids. Lysophosphatidic acid increased with primarily platelet-inhibitor-lipids when single donor apheresis platelet upon storage. Anti-inflammatory and platelet-induced-inhibition lipids were weakly expressed in case of severe adverse reaction. Thus, a decrease in lysophosphatidylcholine and an increase in lysophosphatidic acid could be reliable predictors of serious adverse transfusion reactions transfusion. These data highlighted the importance of PC manufacturing and storage processing, not only to maximize clinical efficacy but also to minimize adverse reactions.