Hypotonic Shock Response Research Articles

Comparing quality parameters of apheresis and pooled platelet concentrates

Introduction. Harvesting of pooled platelet concentrates (PC) significantly expands the volume of transfusion support and could be used to create strategic platelet reserve.The aim of work. To compare the quality of pooled and apheresis platelet concentrates.Materials and methods. We studied platelet fractions (PF), isolated from the whole blood of cadre donors with Reveos system, platelet concentrates, obtained by pooling PF (Pooled PC), and platelet concentrates harvested by automatic apheresis. In all cases 2 days passed from the moment of blood donation to the platelet examination. In PC samples we studied cellular composition, morphofunctional parameters of platelets, pH and electrochemical parameters, hypotonic shock response.Results. The morphofunctional platelet rate in PF was significantly reduced, comparing to the normal levels. At the same time, the morphofunctional parameters of platelets did not significantly differ in PF and pooled PC. The pH values, the total content of platelets and leukocytes in the pooled PC corresponded to the requirements of The rules for procurement, storage, transportation and clinical use of donated blood and its components (2019). The total platelet count, the level of annexin-positive platelets, did not significantly differ in apheresis PC and pooled PC, hypotonic shock response was correct in apheresis PC and pooled PC. On the other hand, the level of platelets with granules in pooled PC was 1,8 times lower than in apheresis PC, other morphofunctional parameters of platelets in pooled PC were also reduced. Pooled PC were characterized by more positive level of open circuit potential and lower antioxidant capacity compared with apheresis PC on the same storage time.Conclusions. Platelet pooling with Reveos system is acceptable for obtaining PC doses that comply with Technical Regulations. Nevertheless, methods of pooled PC harvesting need further optimization.

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.

Modeling Calcium Signaling in S. cerevisiae Highlights the Role and Regulation of the Calmodulin-Calcineurin Pathway in Response to Hypotonic Shock

Calcium homeostasis and signaling processes in Saccharomyces cerevisiae, as well as in any eukaryotic organism, depend on various transporters and channels located on both the plasma and intracellular membranes. The activity of these proteins is regulated by a number of feedback mechanisms that act through the calmodulin-calcineurin pathway. When exposed to hypotonic shock (HTS), yeast cells respond with an increased cytosolic calcium transient, which seems to be conditioned by the opening of stretch-activated channels. To better understand the role of each channel and transporter involved in the generation and recovery of the calcium transient—and of their feedback regulations—we defined and analyzed a mathematical model of the calcium signaling response to HTS in yeast cells. The model was validated by comparing the simulation outcomes with calcium concentration variations before and during the HTS response, which were observed experimentally in both wild-type and mutant strains. Our results show that calcium normally enters the cell through the High Affinity Calcium influx System and mechanosensitive channels. The increase of the plasma membrane tension, caused by HTS, boosts the opening probability of mechanosensitive channels. This event causes a sudden calcium pulse that is rapidly dissipated by the activity of the vacuolar transporter Pmc1. According to model simulations, the role of another vacuolar transporter, Vcx1, is instead marginal, unless calcineurin is inhibited or removed. Our results also suggest that the mechanosensitive channels are subject to a calcium-dependent feedback inhibition, possibly involving calmodulin. Noteworthy, the model predictions are in accordance with literature results concerning some aspects of calcium homeostasis and signaling that were not specifically addressed within the model itself, suggesting that it actually depicts all the main cellular components and interactions that constitute the HTS calcium pathway, and thus can correctly reproduce the shaping of the calcium signature by calmodulin- and calcineurin-dependent complex regulations. The model predictions also allowed to provide an interpretation of different regulatory schemes involved in calcium handling in both wild-type and mutants yeast strains. The model could be easily extended to represent different calcium signals in other eukaryotic cells.

Divalent magnesium restores cytoskeletal storage lesions in cold-stored platelet concentrates

Cold storage of platelet concentrates (PC) has become attractive due to the reduced risk of bacterial proliferation, but in vivo circulation time of cold-stored platelets is reduced. Ca2+ release from storage organelles and higher activity of Ca2+ pumps at temperatures < 15 °C triggers cytoskeleton changes. This is suppressed by Mg2+ addition, avoiding a shift in Ca2+ hemostasis and cytoskeletal alterations. We report on the impact of 2–10 mM Mg2+ on cytoskeleton alterations of platelets from PC stored at room temperature (RT) or 4 °C in additive solution (PAS), 30% plasma. Deformation of platelets was assessed by real-time deformability cytometry (RT-DC), a method for biomechanical cell characterization. Deformation was strongly affected by storage at 4 °C and preserved by Mg2+ addition ≥ 4 mM Mg2+ (mean ± SD of median deformation 4 °C vs. 4 °C + 10 mM Mg2+ 0.073 ± 0.021 vs. 0.118 ± 0.023, p < 0.01; n = 6, day 7). These results were confirmed by immunofluorescence microscopy, showing that Mg2+ ≥ 4 mM prevents 4 °C storage induced cytoskeletal structure lesion. Standard in vitro platelet function tests showed minor differences between RT and cold-stored platelets. Hypotonic shock response was not significantly different between RT stored (56.38 ± 29.36%) and cold-stored platelets with (55.22 ± 11.16%) or without magnesium (45.65 ± 11.59%; p = 0.042, all n = 6, day 1). CD62P expression and platelet aggregation response were similar between RT and 4 °C stored platelets, with minor changes in the presence of higher Mg2+ concentrations. In conclusion, increasing Mg2+ up to 10 mM in PAS counteracts 4 °C storage lesions in platelets, maintains platelet cytoskeletal integrity and biomechanical properties comparable to RT stored platelets.

The strategy of modulation blood responses by surface modification with different functional groups on polyester film.

A main problem in the design of blood-contacting biomaterials has been the deficiency of a systematic understanding of blood-biomaterial interactions and the strategy to modulate blood responses. In this work, different functional groups including carboxyl (COOH), hydroxyl (OH) and zwitterionic sulfobetaine group (⊕N((CH3 )2 )(CH2 )3 SO3-○- , SMDB) were grafted on the poly (butylene terephthalate) (PBT) film to study how the functional groups modulate blood responses and in terms of interaction with the coagulation system, the complement system, and platelets. The results showed protein absorption and platelet adhesion was stronger on the PBT bearing COOH group than PBT films bearing OH and zwitterionic sulfobetaine groups (total protein (μg/cm2 ): 32.92 ± 5.89 vs. 22.02 ± 1.44 vs. 19.09 ± 1.59; platelet adhesion (/mm2 ): 1,626.7 ± 120.1 vs. 1,395.6 ± 363.3 vs. 1,102.2 ± 373.7), which had a rougher and negatively charged surface, and the coagulation system was inhibited by binding fibrinogen (Fg) and coagulation factors. Meanwhile, PBT-PSMDB showed anticoagulant property and induced platelet activation. As a result, complement formation on these two films were less than PBT bearing OH groups by inhibiting the coagulation system (C3a (ng/ml): 3,745.4 ± 143.9 vs. 3,290.9 ± 249.7 vs. 4,887.9 ± 88.9; C5a (ng/ml): 22.1 ± 2.6 vs. 22.3 ± 1.8 vs. 27.9 ± 2.0). On the other hand, PBT bearing OH groups did not facilitate remarkable platelet adhesion and activation, and had no influence on platelet aggregation, hypotonic shock response, and coagulation system. The above results showed that the blood responses were highly interlinked, and could be modulated by grafting with different functional groups on the biomaterial surfaces. These findings may help identify a strategy to design materials with better hemocompatibility for blood contact, filtration, and purification applications.

IMPACT OF THE PLATELET WASHING PROCESS ON IN VITRO PLATELET PROPERTIES, AND THE LEVELS OF SOLUBLE CD40 LIGAND AND PLATELET-DERIVED MICROPARTICLES IN THE STORAGE MEDIA

Background A new platelet (PLT) additive solution, bicarbonated Ringer's solution supplemented with acid-citrate-dextrose Formula A, termed BRS-A, as well as a new automated closed system cell processor for washing PLTs have recently been developed. This study evaluated the in vitro properties of PLTs with the automated system versus the manual method, using the BRS-A additive solution for washing and storage. Methods ABO-identical apheresis PLTs in 100% plasma were pooled and split equally for control (in 100% plasma or a manual method) and test (ACP215 automated system) units. In vitro characteristics of PLTs washed with the automated system were compared to those of PLTs in 100% plasma (Study 1) or washed with a manual method (Study 2) during the 7-day storage. Results In Study 1, hypotonic shock response, aggregation response, mitochondrial membrane potential, adenosine triphosphate, and CD42b mean fluorescence intensity were comparable in the control and test groups during the 7-day storage. CD62P expression was lower in the test group than controls on Days 3 and 7. The level of platelet-derived microparticles (PDMPs) in the test group on Days 1 and 2 were higher than those in controls. In contrast, the levels of soluble CD40 ligand (sCD40L) and regulated upon activation of normal T-cell expressed and secreted (RANTES) in the test units were lower than controls. In Study 2, no significant differences were found in all in vitro properties except for PLT count and the levels of PDMPs in the test units were higher than controls during storage. Conclusion Apheresis PLTs washed with the automated system using BRS-A additive solution maintained in vitro properties during storage. Washing methods influenced PDMP levels but not sCD40L and RANTES.

Impact of the platelet washing process on in vitro platelet properties, and the levels of soluble CD40 ligand and platelet-derived microparticles in the storage media.

A new platelet (PLT) additive solution, bicarbonated Ringer's solution supplemented with acid-citrate-dextrose Formula A, termed BRS-A, as well as a new automated closed system cell processor for washing PLTs have recently been developed. This study evaluated the in vitro properties of PLTs with the automated system versus the manual method, using the BRS-A additive solution for washing and storage. ABO-identical apheresis PLTs in 100% plasma were pooled and split equally for control (in 100% plasma or a manual method) and test (ACP215 automated system) units. In vitro characteristics of PLTs washed with the automated system were compared to those of PLTs in 100% plasma (Study 1) or washed with a manual method (Study 2) during the 7-day storage. In Study 1, hypotonic shock response, aggregation response, mitochondrial membrane potential, adenosine triphosphate, and CD42b mean fluorescence intensity were comparable in the control and test groups during the 7-day storage. CD62P expression was lower in the test group than controls on Days 3 and 7. The level of platelet-derived microparticles (PDMPs) in the test group on Days 1 and 2 were higher than those in controls. In contrast, the levels of soluble CD40 ligand (sCD40L) and regulated upon activation of normal T-cell expressed and secreted (RANTES) in the test units were lower than controls. In Study 2, no significant differences were found in all in vitro properties except for PLT count and the levels of PDMPs in the test units were higher than controls during storage. Apheresis PLTs washed with the automated system using BRS-A additive solution maintained in vitro properties during storage. Washing methods influenced PDMP levels but not sCD40L and RANTES.

Platelet storage in more than 90% additive solution containing glucose and bicarbonate has the potential to increase shelf life.

Platelet concentrates (PCs) suspended in more than 90% additive solution (AS) are recommended for patients with reactions to platelets stored in plasma. Next-generation AS containing glucose and bicarbonate might enable storage of these PCs for longer than those in current-generation AS, which was therefore evaluated in this study. Five buffy coat or apheresis-derived PCs were pooled and split into identical units. All units except the control were centrifuged, the plasma removed and replaced with AS (SSP+, PAS-5, or PAS-G), or resuspended in the same plasma and sampled 96 hours after resuspension for analysis. Plasma carryover was less than 10%, total protein less than 1 g/unit, and immunoglobulin A levels lower than 0.1 mg/mL for all PCs in AS. The pH of all the platelets during storage was higher than 6.4. PAS containing glucose maintained superior in vitro platelet function during storage compared with those resuspended in SSP+. Compared with storage in SSP+, storage in PAS-5 or PAS-G resulted in better preservation of platelet adenosine triphosphate and hypotonic shock response, lower annexin V binding, and improved mitochondrial membrane potential. Platelets resuspended in PAS-5 and PAS-G maintained in vitro function and metabolism during storage compared with SSP+ platelets. Elevated platelet metabolic activity was noticed in PAS-G, and higher platelet activation was detected with PAS-5. Platelets resuspended in PAS containing glucose has the potential to increase the shelf life of PC in more than 90% AS.

Quality control of riboflavin-treated platelet concentrates using Mirasol® PRT system: Polish experience.

The quality of platelet concentrates (PCs) is affected by preparation, storage, the type of container, and pathogen reduction technology (PRT). The Mirasol® Pathogen Reduction Technology (PRT) system (Terumo BCT Inc., Lakewood, USA), which uses riboflavin and ultraviolet (UV) light, has recently been proven effective against bacteria, viruses, parasites, and leukocytes. The aim of the study was to evaluate the effect of the Mirasol® PRT system, based on riboflavin and UV light exposure, on the most common in vitro platelet quality parameters of PCs prepared from whole blood-derived buffy coats. The study included 15 trials (n = 15). For each trial, 2 PCs were used: 1 for treatment with the Mirasol® PRT system (M) and 1 for a control (C). In the M group, PCs were illuminated. In the C group, saline solution was added. PCs from groups M and C were stored at 20-24°C, with agitation. Samples were collected on days 1, 3 and 5 to determine platelet concentration, total platelet count/unit, mean platelet volume (MPV), power of hydrogen (pH), glucose and beta-thromboglobulin concentration (BTG), hypotonic shock response (HSR), aggregation, CD42b and CD62P expression, pCO2, and pO2. No significant differences in HSR or CD42b expression were observed between groups M and C. All pH values were stable during the whole storage period (7.1-7.5). On storage day 1, CD62P expression in group C was significantly higher than in group M. In the Mirasol® group, significantly higher glucose consumption was noted on storage days 3 and 5. On day 5, a 2-3-fold increase in BTG was observed in both groups as compared to day 1; on day 5, BTG concentration was 32% higher in group M than in group C. On all storage days, pCO2 was comparable in groups M and C; lower pO2 values were reported for group M. In vitro results demonstrated that pH, HSR, aggregation, CD42b antigen expression, and MPV and platelet count parameters were comparable in groups M and C.

In vitro quality of apheresis platelets divided into paediatric-sized units and stored in PVC bags plasticised with TOTM, BTHC or DINCH.

Two of the predictive factors of the quality of small volumes of platelets suitable for paediatric use are bag size and material. This study evaluated the storage properties of paediatric platelet aliquots in TOTM-, BTHC- or DINCH-PVC bags. (i) Three apheresis platelet concentrates (PC) were pooled and split into three units. One was retained as an adult unit (control; polyolefin bag). The second and third units were split into four MacoPharma TOTM-PVC and BTHC-PVC paediatric bags, respectively. (ii) Two apheresis PC were pooled and split into two units. One PC was retained as an adult unit, and the other was split into four Fresenius DINCH-PVC paediatric bags. Testing was performed on storage for pH, blood gases, hypotonic shock response, soluble CD62P, LDH, glucose and lactate, ATP, CD62P, CD63, platelet-derived microparticles and annexin V. The volumes, platelet yields and pH of all paediatric units met local specifications. The TOTM-PVC bag showed no worse quality than the adult bag up to day 7 for all parameters studied, and it maintained pH higher than BTHC-PVC and DINCH-PVC over storage. The BTHC-PVC bag was shown to be the most gas permeable; however, it had the highest glucose consumption rates and the highest platelet activation. All bags showed an acceptable in vitro quality. Overall, the TOTM-PVC paediatric bag showed better platelet quality compared to the other storage bags, whereas storage in the BTHC-PVC bag resulted in poorer platelet quality.

Platelet sequestration with a new-generation autotransfusion device.

Autologous platelet-rich plasma (PRP) can be used either to prevent platelets (PLTs) from lesions during extracorporeal circulation or for wound therapy, when processed into PLT gel. The aim of this study was to evaluate the PLT sequestration abilities of a new-generation autotransfusion device. In this experimental study the discontinuous, new-generation autotransfusion device XTRA was evaluated using fresh donor blood. The blood was processed in four different size bowls (X55, X125, X175, X225 [bowls' size in mL]) using the device's built-in "PLT sequestration program." PLT functionality was tested using aggregometry; for PLT activation, β-thromboglobulin (β-TG) and soluble P-selectin levels were determined. Cell damage was assessed by a morphology score and hypotonic shock response. Additionally, PLTs were tested after 6 hours to identify storage lesions. Platelet recovery in the PRP ranged from 39% to 64% and averaged 6.2-fold PLT enrichment as defined by the increase in PLT concentration. The preparation caused minimal cell damage and a decrease in cell function by only 10%, but a slight activation was observed amounting to 9% of the maximal β-TG release. The efficiency of the preparation, represented by the PLT recovery rate, increased in a linear fashion with the increasing bowl sizes being tested. After 6 hours of storage the prepared PLTs showed an additional 9% loss in function, but only 4% decrease in viability. The autotransfusion device XTRA was capable of high-quality perioperative PRP preparation, and the bowl size was found to have an influence on the efficiency of the preparation.

Comparison of Platelet Concentrates Prepared from Platelet Rich Plasma-Platelet Concentrate and Buffy Coat Poor-Platelet Concentrate on Storage Days 1 and 5

Objective: The purpose of this study is to assess the quality of platelet concentrates on storage days 1 and 5 prepared by platelet rich plasma-platelet concentrate (PRP-PC) and buffy coat poor-platelet concentrate (BC-PC) methods comparing to the American Association of Blood Banks (AABB) recommendations.Material and Method: Totally of 120 platelet concentrates (PC) units on storage days 1 and 5 (60 of PRP-PC triple blood bag and 60 of BC-PC quadruple AS-5 blood bag) were separated from whole blood donations at Songklanagarind Hospital. The prepared PC were assessed with 5 parameters such as volume, platelet count, white blood cell count per unit, pH, swirling phenomenon score and hypotonic shock response. The independent t-tests, paired Student’s t-tests and SPSS program were utilized in statistical analysis step.Results: The mean±standard deviation (S.D.) of each parameter were as follow : (1) Volume of PRP-PC and BC-PC met the standard (40-70 ml). (2) All of the platelet concentrates met the standard (≥ 5.5x1010/unit). The mean±S.D.: PRP-PC and BC-PC (day 1) were 6.820±1.480 x1010 and 7.010±1.300 x1010/unit (p-value=0.260), while PRP-PC and BC-PC (day 5) were 6.620±1.160x1010 and 6.720± 1.150x1010/unit (p-value=0.040). (3) The white blood cell in platelet concentrates met the standard (&lt;0.2x1010/unit). The mean±S.D.: PRP-PC and BC-PC (day 1) were 0.030±0.017 x1010and 0.026±0.019x1010/unit (p-value=0.040), while PRP-PC and BC-PC (day 5) were0.033±0.013x1010 and 0.027±0.019x1010/unit (p-value= 0.580). (4) The pH of all units (PRP-PC and BC-PC) met the standard (≥6.2). The mean±S.D.: PRP-PC and BC-PC (day 1) were 7.430±0.330 and 7.750±0.160 (p-value=0.006), while PRP-PC and BC-PC (day 5) were 7.590±0.350 and 7.620±0.280 (p-value=0.710). The swirling phenomenon score and hypotonic shock response were the same as standard AABB and were not statistically difference.Conclusion: The quality of PRP-PC and BC-PC after storing on days 1 and 5 as follow (1) Volume of PRP-PC and BC-PC met the standard. (2) The platelet count per unit of PRP-PC and BC-PC (day 1), PRP-PC and BC-PC (day 5) were not statistically difference. (3) The white blood cell count per unit of PRP-PC and BC-PC (day 1) were statistically difference, while PRP-PC and BC-PC (day 5) were not statistically difference. (4) The pH of PRP-PC and BC-PC (day 1) were statistically difference, while PRP-PC and BC-PC (day 5) were not statistically difference. The swirling phenomenon score and hypotonic shock response of PRP-PC and BC-PC were not statistically difference. Platelet concentrates of both method storing on days 1 and 5 fulfilled the quality guideline of AABB.

Reduction of bacteria and human immunodeficiency virus Type 1 infectivity of platelet suspension in plasma using xenon flash-pulse light in a bench-scale trial.

Current pathogen reduction systems for platelet concentrates (PCs) require addition of chemical compounds and/or reduction of plasma content in PCs. We have investigated a new method using xenon (Xe) flash-pulse light without additional compounds or plasma replacement. An aliquot of apheresis platelets (PLTs) in plasma inoculated with bacteria or human immunodeficiency virus Type 1 (HIV-1) was irradiated with Xe flash-pulse light (Xe flash phototreatment). Bacterial growth was monitored up to 6 days of storage, whereas HIV-1 infectivity was assayed just after treatment. Pairs of Xe flash-phototreated and untreated PCs were examined for PLT lesion during the storage period. Under the current conditions, a low titer (1.8 colony-forming units [CFUs]/mL) of Staphylococcus aureus did not proliferate during the 6-day storage period, but grew in some cases at high-titer (24.0 CFUs/mL) inoculation. HIV-1 infectivity was reduced by 1.8 log. PLT recovery of the treated PCs was lower than untreated ones. An increase of mean PLT volume and glucose consumption, together with a decrease of hypotonic shock response and pH, were enhanced by the treatment. CD62P- and PAC-1-positive PLTs increased after the treatment, indicating the induction of PLT activation. Among biologic response modifiers, soluble CD40 ligand was significantly increased in the treated PCs on Day 6. Xe flash phototreatment could prevent bacterial proliferation and reduce HIV-1 infectivity in 100% plasma PCs without any additional compounds, but enhanced PLT storage lesions. Further improvement is required to increase the potency of pathogen inactivation with reducing PLT damage.

異なる2種類の重炭酸リンゲル液で調製した血小板保存液BRS-Aによる血小板の洗浄と保存

日本赤十字社血液センターは血小板製剤の有害反応を減らすために血小板の洗浄をしばしば求められるが，国内には臨床使用が承認された血小板保存液は無い．最近，我々は新規血小板保存液であるBRS-A（BRS supplemented with ACD-A）を開発した．BRS-Aは臨床で入手可能なビカネイトⓇ輸液（重炭酸リンゲル液，BRS）とACD-A液を用いて調製することができ，5%未満の血漿濃度で7日間血小板機能を維持することができる．本研究の目的は，ビカネイトⓇ輸液とは電解質濃度が異なる別のBRSであるビカーボンⓇ輸液を用いて調製したBRS-Aの性能を評価することである．

In vitro Quality of Platelets with Low Plasma Carryover Treated with Ultraviolet C Light for Pathogen Inactivation

Background: The THERAFLEX UV-Platelets system uses shortwave ultraviolet C light (UVC, 254 nm) to inactivate pathogens in platelet components. Plasma carryover influences pathogen inactivation and platelet quality following treatment. The plasma carryover in the standard platelets produced by our institution are below the intended specification (<30%). Methods: A pool and split study was carried out comparing untreated and UVC-treated platelets with <30% plasma carryover (n = 10 pairs). This data was compared to components that met specifications (>30% plasma). The platelets were tested over storage for in vitro quality. Results: Platelet metabolism was accelerated following UVC treatment, as demonstrated by increased glucose consumption and lactate production. UVC treatment caused increased externalization of phosphatidylserine on platelets and microparticles, activation of the GPIIb/IIIa receptor (PAC-1 binding), and reduced hypotonic shock response. Platelet function, as measured with thrombelastogram, was not affected by UVC treatment. Components with <30% plasma were similar to those meeting specification with the exception of enhanced glycolytic metabolism. Conclusion: This in vitro analysis demonstrates that treatment of platelets with <30% plasma carryover with the THERAFLEX UV-Platelets system affects some aspects of platelet metabolism and activation, although in vitro platelet function was not negatively impacted. This study also provides evidence that the treatment specifications of plasma carryover could be extended to below 30%.

Effect of 25 Gy (60)Co Irradiation on the Physico-chemical Property and Functions of the Platelets During Storage

To evaluate the effects of the 25 Gy ⁶⁰Co irradiation on the physiological and biochemical properties and the functions of the platelets during storage. A total of 15 bags of platelets were apheresis-collected from 15 healthy donors, and each bag of platelets were divided into 2 parts, then the platelets were divided into the control group (without 25 Gy ⁶⁰Co irradiation) and the irradiated group (with 25 Gy ⁶⁰Co irradiation) groups. The two groups of platelets were kept under the condition of (22 ± 2) °C and shaken. The Platelet count and pH value were detected on the d 1, d 2, d 3, d 4 and d 5. The variables such as R, K values, α angle and maximal amplitude (MA) were measured by thrombelastography on the same days. Hypotonic shock response (HSR), morphological score were devised. There were no statistically significant difference in Plt counts, mean platelet volume (MPV), platelet distribute width (PDW) and pH between the two groups (P > 0.05), and Plt count decreased on the end of storage. There were no marked changes in HSR level and morphological score between the two groups during storage, and there were no significant difference between the two groups (P > 0.05). In the TEG analysis there were no significant difference of the R, K, α angle and MA values between the two groups (P > 0.05). R value showed upward trend increased along with prolongation of preserved time (P < 0.01), no significant changes in α angle (P > 0.05), K value was slightly higher and MA value was lower in the last day of storage than the days 1-4 (P < 0.01), respectively. 25 Gy ⁶⁰Co gamma-ray irradiation can not damage the physiological, biochemical properties and the functions of the platelets during storage. In order to ensure the best curative effect, it is suggested that no matter the platelets were irradiated or not, the platelets should be used as soon as possible.

Correlation between the In Vitro Functionality of Stored Platelets and the Cytosolic Esterase-Induced Fluorescence Intensity with CMFDA.

It has been hypothesized that the cytosolic esterase-induced fluorescence intensity (CEIFI) from carboxy dimethyl fluorescein diacetate (CMFDA) in platelets may related to platelet functions. In the present study, we measured the change of CEIFI in platelets during storage, and examined the correlations of CEIFI with the in vitro functionality of stored platelets, including the ADP-induced aggregation activity, hypotonic shock response, expression of CD62P as well as platelet apoptosis. The CEIFI of fresh platelets, when tested at 10 μM CMFDA, the mean fluorescence intensity index (MFI) was 305.9 ± 49.9 (N = 80). After 1-day storage, it was 203.8 ± 34.4, the CEIFI of the stored platelets started to decline significantly, and reduced to 112.7 ±27.7 after 7-day storage. The change in CEIFI is highly correlated to all four functional parameters measured, with the correlation coefficients being 0.9813, 0.9848, -0.9945 and -0.9847 for the ADP-induced aggregation activity, hypotonic shock response (HSR), expression of CD62P and platelet apoptosis respectively. The above results show that the CEIFI measurement of platelets represents well the viability and functional state of in vitro stored platelets. This may be used as a convenient new method for quality evaluation for stored platelets if this result can be further validated by the following clinical trials.

Analysis of the Physiological Activities and Functions in Vitro of Apheresis Platelets during Storage

To investigate the changes of physiological activities and functions in vitro of apheresis platelets during storage. 17 units of apheresis platelets were randomly chosen and stored at 20 °C to 24 °C with agitation. Platelet counting (Plt), mean platelet volume (MPV), blood gases, pH value, glucose (Glu) concentration, lactate (LA) concentration, LDH concentration, thromboelastogram (TEG), hypotonic shock response (HSR), CD62p expression rate and anew expression rate were measured on days 0, 1, 3, 5 after platelet storage. Changes of physiological activities and functions in vitro were systematically evaluated by above-mentioned indexes. During storage, Plt, MPV and HSR were not significantly changed; but pH value, blood gases, Glu, LA, LDH, HSR, expression rate of CD62p and anew expression rate were significant differenty. Among thromboelastogram indexes, R value increased obviously with prolongation of storage time; K value and αAngle were not significantly changed; MA was not significantly changed on day 1 and 3, but was slightly increase on day 5. The physiological activities and functions in vitro of apheresis platelets are kept well during storage. For clinical transfusion of apheresis platelet during storage, clinical effect of transfusione is not influenced.

Storage of washed platelets in BRS-A platelet additive solutions based on two types of clinically available bicarbonated Ringer's solutions with different electrolyte concentrations

BackgroundIn Japan, no platelet (PLT) additive solutions (PASs) are officially approved for clinical use although blood centers often receive requests for washed PLTs to reduce adverse reactions. Recently, we developed a novel PAS called BRS-A based on clinically available bicarbonated Ringer's solution (BRS), Bicanate and acid–citrate–dextrose formula A (ACD-A), which has been shown to maintain the in vitro properties of PLTs in the condition of <5% residual plasma during 7-day storage. The aim of this study was to evaluate whether another clinically available BRS, Bicarbon with different electrolyte concentrations can be used as a PAS. Study design and methodsTwo types of BRS-As were prepared by adding 25 mL of ACD-A to 500 mL of Bicanate or Bicarbon BRSs. Bicanate-based BRS-A and Bicarbon-based BRS-A contain 0.9 or 0.5 mmol/L of magnesium chloride, 95.2 or 100.1 mmol/L of sodium chloride, 4.2 or 5.1 mmol/L of trisodium citrate, and 26.6 or 23.8 mmol/L of sodium bicarbonate, respectively; the other components were identical. Apheresis PLTs stored in these solutions with less than 5% plasma for 7-day storage were compared with regard to their in vitro properties. ResultsThe pH levels of all units were above 7 throughout storage. The mean PLT volume, hypotonic shock response, glucose consumption, lactate production, swirling, and CD62P and CD42b expression were similar during 7-day storage. The bicarbonate levels in Bicarbon-based BRS-A were lower than those in Bicanate-based BRS-A. ConclusionDifferences in concentrations of electrolytes such as magnesium, sodium, citrate, and bicarbonate salts in BRS-A do not affect the in vitro properties of PLTs during 7-day storage. These results indicate that the use of another type of BRS-A based on Bicarbon as a PAS is feasible. Thus, BRS-A can be used in hospitals that do not stock Bicanate but have Bicarbon.

A hollow-fibre column system to effectively prepare washed platelets.

We developed a hollow-fibre column system specifically adapted to prepare washed platelet concentrates (WPCs). This study was performed to evaluate the efficacy of the hollow-fibre column system for preparing WPCs. First, the percentages of platelet (PLT) recovery and remaining plasma proteins were calculated by determining the PLT count, volume and plasma protein levels in both the prewash and postwash. Secondly, washed PLTs and unwashed control PLTs were stored for 5 days, and the changes during this 5-day storage of in vitro PLT characteristics were determined. The hollow-fibre column system effectively removed >98% of plasma in platelet concentrates (PCs), and the PLT recovery was 97% on an average. The CD62P-expression level on washed PLTs immediately after washing was approximately twofold higher than that on prewashed PLTs as well as on PLTs washed via manual methods or cell washing devices. Until day 5 during storage, PLT aggregability, hypotonic shock response and swirling scores of washed PLTs were not significantly different from those of the control PCs. Our novel hollow-fibre column system proved valuable in preparing washed PLTs with <2% of residual plasma proteins and high recovery of PLTs.