Manipulation of Lipid Metabolism During Normothermic Machine Perfusion: Effect of Defatting Therapies on Donor Liver Functional Recovery.

Strategies to increase the use of steatotic donor livers are required to tackle the mortality on the transplant waiting list. We aimed to test the efficacy of pharmacological enhancement of the lipid metabolism of human livers during ex situ normothermic machine perfusion to promote defatting and improve the functional recovery of the organs. Because of steatosis, 10 livers were discarded and were allocated either to a defatting group that had the perfusate supplemented with a combination of drugs to enhance lipid metabolism or to a control group that received perfusion fluid with vehicle only. Steatosis was assessed using tissue homogenate and histological analyses. Markers for lipid oxidation and solubilization, oxidative injury, inflammation, and biliary function were evaluated by enzyme-linked immunosorbent assay, immunohistochemistry, and in-gel protein detection. Treatment reduced tissue triglycerides by 38% and macrovesicular steatosis by 40% over 6 hours. This effect was driven by increased solubility of the triglycerides (P = 0.04), and mitochondrial oxidation as assessed by increased ketogenesis (P = 0.008) and adenosine triphosphate synthesis (P = 0.01) were associated with increased levels of the enzymes acyl-coenzyme A oxidase 1, carnitine palmitoyltransferase 1A, and acetyl-coenzyme A synthetase. Concomitantly, defatted livers exhibited enhanced metabolic functional parameters such as urea production (P = 0.03), lower vascular resistance, lower release of alanine aminotransferase (P = 0.049), and higher bile production (P = 0.008) with a higher bile pH (P = 0.03). The treatment down-regulated the expression of markers for oxidative injury as well as activation of immune cells (CD14; CD11b) and reduced the release of inflammatory cytokines in the perfusate (tumor necrosis factor α; interleukin 1ß). In conclusion, pharmacological enhancement of intracellular lipid metabolism during normothermic machine perfusion decreased the lipid content of human livers within 6 hours. It also improved the intracellular metabolic support to the organs, leading to successful functional recovery and decreased expression of markers of reperfusion injury.

Cold storage of platelets in additive solution: the impact of residual plasma in apheresis platelet concentrates.

Platelet transfusion has become essential therapy in modern medicine. Although the clinical advantage of platelet transfusion has been well established, adverse reactions upon transfusion, especially transmission of bacterial infection, still represent a major challenge. While bacterial contamination is favored by the storage of platelets at room temperature, cold storage may represent a solution for this important clinical issue. In this study, we aimed to clarify whether plasma has protective or detrimental effects on cold-stored platelets. We investigated the impact of different residual plasma contents in apheresis-derived platelet concentrates, stored at 4°C or room temperature, on platelet function and survival. We found that platelets stored at 4°C have higher expression of apoptosis marker compared to platelets stored at room temperature, leading to accelerated clearance from the circulation in a humanized animal model. While cold-induced apoptosis was independent of the residual plasma concentration, cold storage was associated with better adhesive properties and higher response to activators. Interestingly, delta (δ) granule-related functions, such as ADP-mediated aggregation and CD63 release, were better preserved at 4°C, especially in 100% plasma. An extended study to assess cold-stored platelet concentrates produced under standard care Good Manufacturing Practice conditions showed that platelet function, metabolism and integrity were better compared to those stored at room temperature. Taken together, our results show that residual plasma concentration does not have a cardinal impact on the cold storage lesions of apheresis-derived platelet concentrates and indicate that the current generation of additive solutions represent suitable substitutes for plasma to store platelets at 4°C.

Systems analysis of metabolism in platelet concentrates during storage in platelet additive solution.

Platelets (PLTs) deteriorate over time when stored within blood banks through a biological process known as PLT storage lesion (PSL). Here, we describe the refinement of the biochemical model of PLT metabolism, iAT-PLT-636, and its application to describe and investigate changes in metabolism during PLT storage. Changes in extracellular acetate and citrate were measured in buffy coat and apheresis PLT units over 10 days of storage in the PLT additive solution T-Sol. Metabolic network analysis of these data was performed alongside our prior metabolomics data to describe the metabolism of fresh (days 1-3), intermediate (days 4-6), and expired (days 7-10) PLTs. Changes in metabolism were studied by comparing metabolic model flux predictions of iAT-PLT-636 between stages and between collection methods. Extracellular acetate and glucose contribute most to central carbon metabolism in PLTs. The anticoagulant citrate is metabolized in apheresis-stored PLTs and is converted into aconitate and, to a lesser degree, malate. The consumption of nutrients changes during storage and reflects altered PLT activation profiles following their collection. Irrespective of the collection method, a slowdown in oxidative phosphorylation takes place, consistent with mitochondrial dysfunction during PSL. Finally, the main contributors to intracellular ammonium and NADPH are highlighted. Future optimization of flux through these pathways provides opportunities to address intracellular pH changes and reactive oxygen species, which are both of importance to PSL. The metabolic models provide descriptions of PLT metabolism at steady state and represent a platform for future PLT metabolic research.

Preclinical safety assessment of antipyrine combined with lidocaine hydrochloride as ear drops.

This study was designed to assess the preclinical toxicity of antipyrine combined with lidocaine hydrochloride ear drops (ALED) and support the clinical trials of ALED in clinical settings in China. All the experiments including acute toxicity in rodents, skin sensitization toxicity in guinea pigs, skin irritation toxicity in rabbits and chronic toxicity in rats were performed according to China Food and Drug Administration guidelines. The maximum tolerated dose (MTD) of ALED administration for mice and rats was over (400 g antipyrine plus 100 g lidocaine hydrochloride)/kg and (240 g andtipyrine plus 60 g lidocaine hydrochloride)/kg, respectively. No obvious skin sensitization toxicity and skin irritation toxicity were observed. The main changes concentrated in chronic toxicity study in rats. For the chronic toxicity, rats were administrated once a day for 28 consecutive days, and a 14-day recovery period was followed. The side effects of ALED included decreased dietary intake in male rats, increased proportion of reticulocytes, decreased or even inversed granulocyte:erythrocyte ratio, fluctuated alanine aminotransferase and aspartate aminotransferase, and slightly increased relative weight of liver. Conclusively, blood system (especially erythrocyte system) and digestive system, including liver and gastrointestinal tract, might be the toxic targets of ALED.

Investigating the effect of proteoglycan 4 on hyaluronan solution properties using confocal fluorescence recovery after photobleaching.

BACKGROUND: The objective of this study was to use confocal fluorescence recovery after photobleaching (FRAP) to examine the specific and dose-dependent effect of proteoglycan 4 (PRG4) on hyaluronan (HA) solutions of different molecular weight; and assess the effect of reduction and alkylation (R/A) of PRG4 on its effects on HA solutions. METHODS: Confocal FRAP was used to determine the diffusion coefficient of fluorescein isothiocyanate (FITC)-dextran tracer (Dt) through 1500 kDa and 500 kDa HA solutions (0-3.3 mg/ml) ± PRG4 or a control protein, bovine serum albumin (BSA), at physiological (450 µg/ml) or pathophysiological (45 µg/ml) concentrations. The effect of PRG4 or R/A PRG4 on 1500 kDa HA solutions was also investigated. Empirical constants obtained from fitting data to the universal scaling equation were used to calculate the average distribution of apparent mesh sizes. RESULTS: PRG4 at both 45 and 450 µg/ml slowed the diffusion of the FITC-dextran tracer for all concentrations of HA and caused a decrease in the apparent mesh size within the HA solution. This effect was specific to PRG4, not observed with BSA, but not dependent on its tertiary/quaternary structure as the effect remained after R/A of PRG4. CONCLUSIONS: These results demonstrate that PRG4 can significantly alter the solution properties of HA; PRG4 essentially reduced the permeability of the HA network. This effect may be due to PRG4 entangling HA molecules through binding and/or HA crowding PRG4 molecules into a self-assembled network. Collectively these findings contribute to the understanding of PRG4 and HA interaction(s) in solution and therefore the function of SF in diarthroidal joints.

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

BACKGROUND: 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. STUDY DESIGN AND METHODS: 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. RESULTS: 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. CONCLUSION: 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.

Pharmaceutical Perspective on Opalescence and Liquid-Liquid Phase Separation in Protein Solutions.

Opalescence in protein solutions reduces aesthetic appeal of a formulation and can be an indicator of the presence of aggregates or precursor to phase separation in solution signifying reduced product stability. Liquid-liquid phase separation of a protein solution into a protein-rich and a protein-poor phase has been well-documented for globular proteins and recently observed for monoclonal antibody solutions, resulting in physical instability of the formulation. The present review discusses opalescence and liquid-liquid phase separation (LLPS) for therapeutic protein formulations. A brief discussion on theoretical concepts based on thermodynamics, kinetics, and light scattering is presented. This review also discusses theoretical concepts behind intense light scattering in the vicinity of the critical point termed as "critical opalescence". Both opalescence and LLPS are affected by the formulation factors including pH, ionic strength, protein concentration, temperature, and excipients. Literature reports for the effect of these formulation factors on attractive protein-protein interactions in solution as assessed by the second virial coefficient (B2) and the cloud-point temperature (Tcloud) measurements are also presented. The review also highlights pharmaceutical implications of LLPS in protein solutions.

Development of Long-Circulating pH-Sensitive Liposomes to Circumvent Gemcitabine Resistance in Pancreatic Cancer Cells.

PURPOSES: To develop pH-sensitive liposomes (PSL) containing a high content of gemcitabine; and to investigate whether drug loading (DL) would alter the in vitro and pharmacokinetic properties. METHODS: PSL with a high DL were obtained using a modified small-volume incubation method. The DL effects on drug release rate and in vitro cytotoxicity of PSL were evaluated using MIA PaCa-2 pancreatic cancer cells and their pharmacokinetics investigated in rats. RESULTS: The highest DL of 4.5 ± 0.1% was achieved for gemcitabine in PSL with 145 ± 5 nm diameter. DL did not alter the in vitro release rate from PSL. The IC50 (48 h) of PSL (DL 0.5 and 4.5%) and non pH-sensitive liposomes (NPSL, DL 4.2%) were 1.1 ± 0.1, 0.7 ± 0.1 and 37.0 ± 7.5 µM, respectively. The PSL resulted in a 4.2-fold increase in its elimination half-life (6.2 h) compared to gemcitabine solution (1.4 h) in rats. No significant difference in pharmacokinetic parameters was observed between the two PSL (DL 0.5 and 4.5%). CONCLUSION: The PSL offered advantages over NPSL in restoring the sensitivity of pancreatic cancer cells to gemcitabine without requiring a high DL. DL in the PSL did not alter release rate, cytotoxicity or their long-circulating properties. Graphical Abstract ᅟ.

The bioequivalence of frozen plasma prepared from whole blood held overnight at room temperature compared to fresh-frozen plasma prepared within eight hours of collection.

BACKGROUND: Overnight, room temperature hold of whole blood (WB) before leukoreduction and component processing offers significant logistic and cost advantages over WB processed within 8 hours. Plasma prepared from WB held at room temperature overnight (PF24RT24WB) may result in a degradation of plasma coagulation protein activities compared to plasma frozen within 8 hours of collection. In this study, we intended to evaluate the bioequivalence (BE) of PF24RT24WB prepared using a new WB collection, leukoreduction, and storage system compared to fresh-frozen plasma (FFP) after 12 months of frozen storage. STUDY DESIGN AND METHODS: We conducted a three-center, three-arm evaluation of the LEUKOSEP HWB-600-XL test system (Hemerus Medical LLC) compared to the RZ2000 control (Fenwal, Inc.). FFP was prepared from WB held at room temperature more than 6 hours and placed at less than -18 °C by 8 hours for control (n = 60) and test (n = 60) arms. PF24RT24WB (n = 60) was prepared with the test system from WB held at room temperature and then filtered and processed 20 to 24 hours postcollection. Frozen plasma was tested at 3, 6, and 12 months using a comprehensive panel of protein and coagulation factor assays. RESULTS: The test FFP was BE for all coagulation factors and tested proteins at 12 months. As expected, PF24RT24WB had a reduced Factor (F)VIII activity compared to control FFP (87.1%; 90% confidence interval, 79.4%-93.3%) with the lower confidence limit less than 80%. All other factors were within the BE region. CONCLUSION: Leukoreduced FFP and PF24RT24WB prepared using the LEUKOSEP HWB-600-XL system has been shown to be BE to control leukoreduced FFP with an expected decrease in FVIII activity after overnight hold.

Overnight, room temperature hold of whole blood followed by 42-day storage of red blood cells in additive solution-7.

BACKGROUND: Overnight, room temperature hold (ONH) of whole blood before component processing offers several benefits. This study evaluated the storage and in vivo recovery characteristics of ONH red blood cells (RBCs) stored in additive solution-7 (AS-7). STUDY DESIGN AND METHODS: We conducted a three-center, three-arm evaluation of a new blood collection system with AS-7 compared to leukoreduced RBCs processed within 8 hours and stored in AS-1 (control). Whole blood (500 ± 50 mL) from healthy research subjects (n = 240) was held at room temperature 0 to 2 hours, 6 to 8 hours, or ONH (18-24 hr) before component processing and storage at 1 to 6 °C. RBCs were evaluated on Days 42 and 56 with a panel of in vitro assays. Subsets of the AS-7-stored RBCs were evaluated for (51) Cr 24-hour in vivo recovery and long-term survival. RESULTS: Adenosine triphosphate (ATP) levels in ONH RBCs were not different than AS-7 RBCs prepared within 8 hours. ATP was higher in the ONH group on Day 42 than control, and ATP was maintained in all AS-7 groups through Day 56. ONH units had 0.36 ± 0.14% on Day 42 hemolysis (60/60 < 0.8%), and 0.54 ± 0.22% on Day 56 (10/60 > 0.8%, 2/60 > 1%). In vivo recoveries of stored RBCs were not different between the AS-7 arms at 42 days (p = 0.16; 27/27 ONH units > 75%), but the Day 56 ONH was significantly less than ONH on Day 42 (p = 0.008; 7/28 < 75%). CONCLUSIONS: Overnight hold of whole blood at room temperature before component processing meets current regulatory requirements when RBCs are stored up to 42 days in AS-7.

Stored red blood cell susceptibility to in vitro transfusion-associated stress conditions is higher after longer storage and increased by storage in saline-adenine-glucose-mannitol compared to AS-1.

BACKGROUND: Biochemical changes induced in red blood cells (RBCs) during storage may impair their function upon transfusion. Transfusion-associated stresses may further amplify storage lesion effects including increased phosphatidylserine (PS) exposure at the RBC membrane, microparticle (MP) release, and adhesion to endothelial cells (ECs). RBC stress susceptibility in vitro was investigated in relation to storage time and additive solution. STUDY DESIGN AND METHODS: Leukoreduced whole blood donations (n = 18) were paired, mixed, and resplit before separating the RBCs for storage in saline-adenine-glucose-mannitol (SAGM) or AS-1. Samples were taken after 3, 21, or 35 days. For oxidative stress treatment, RBCs were exposed to 0.5 mmol/L tert-butylhydroperoxide. Transfusion-associated stress was simulated by overnight culture at 37 °C with plasma containing inflammatory mediators. PS exposure and MPs were measured by flow cytometry and adhesion to ECs was tested under flow conditions. PS specificity of adhesion was tested by blocking with PS-containing lipid vesicles. RESULTS: Oxidative stress induced significantly higher PS exposure and adhesion to ECs in RBCs stored for 35 days compared to 3 days (p < 0.04). PS-containing vesicles blocked RBC-EC adhesion. After overnight culture with or without plasma, PS exposure and EC adhesion were significantly increased (p < 0.05). MP numbers increased with longer RBC storage and after RBC culture with plasma. Culture conditions influenced MP numbers from Day 35 RBCs. RBCs stored in SAGM had significantly higher PS exposure after stress treatment than AS-1 RBCs (p < 0.02). CONCLUSION: Storage for 35 days significantly increased RBC susceptibility to oxidative and in vitro transfusion-associated stresses and was higher for RBCs stored in SAGM compared to AS-1.

Additive solution-7 reduces the red blood cell cold storage lesion.

BACKGROUND: Transfusion of long-stored red blood cells (RBCs) is associated with decreased in vivo RBC recovery, delivery of RBC breakdown products, and increased morbidity and mortality. Reducing the burden of this RBC "storage lesion" is a major challenge in transfusion medicine. Additive solution-7 (AS-7) is a new RBC storage solution designed to improve RBC metabolism by providing phosphate and increasing buffering capacity. STUDY DESIGN AND METHODS: Storage quality in AS-7 was measured in a prospective, randomized, three-center trial using units of whole blood from healthy human subjects whose RBCs were stored for up to 56 days in AS-7 (n = 120) or for 42 days in the control solution AS-1 (n = 60). RESULTS: Hemolysis and shedding of protein-containing microvesicles were significantly reduced in RBCs stored in AS-7 for 42 and 56 days compared with RBCs stored in AS-1. Autologous in vivo recoveries of RBCs stored in AS-7 was 88 ± 5% at 42 days (n = 27) and 82 ± 3% at 56 days (n = 27), exceeding recoveries of RBCs stored in currently used solutions. CONCLUSION: Increasing the phosphate, pH range, and buffer capacity of a RBC storage system allowed RBCs to be stored better and longer than currently approved storage systems. AS-7 ameliorates the long-term storage lesion resulting in significantly increased viability in vitro and in vivo.

In vitro evaluation of the quality of blood products collected and stored in systems completely free of di(2-ethylhexyl)phthalate-plasticized materials.

BACKGROUND: The plasticizer di(2-ethylhexyl)phthalate (DEHP) is a common component in blood bags. DEHP is noncovalently bound to polyvinylchloride (PVC) polymer and can leach into the blood product. There are public concerns that exposure to DEHP might induce developmental and reproductive toxicity in humans. The aim of this study was to evaluate an alternative plasticizer, di(isononyl) cyclohexane-1,2-dicarboxylate (Hexamoll DINCH, BASF SE), for its use in blood bags. STUDY DESIGN AND METHODS: Whole blood (WB) was collected into DEHP-containing and DEHP-free collection systems. After overnight hold, WB was centrifuged and separated in plasma, buffy coat, and red blood cells (RBCs). Buffy coats and plasma were used to make platelet (PLT) concentrates in DEHP-free systems. After addition of additive solution (AS), SAG-M, PAGGS-M, AS-3, or PAGGG-M, RBCs were leukoreduced and analyzed for in vitro characteristics and plasticizer levels during storage. RESULTS: The use of DINCH-based systems had no effect on WB composition, blood processing, and plasma quality. PLT in vitro quality variables were maintained during storage in DEHP-free systems. During storage in SAG-M, hemolysis was significantly higher in DINCH-PVC while potassium leakage and adenosine triphosphate content were comparable. During storage in alternative ASs, hemolysis was reduced compared to storage in SAG-M. CONCLUSIONS: The complete absence of DEHP in the collection system had no effect on WB composition, processing, or plasma and PLT quality. During storage in SAG-M, the absence of DEHP resulted in increased hemolysis. With alternative ASs like PAGGS-M, AS-3, or PAGGG-M, the absence of DEHP had no effect on hemolysis. Leakage of DINCH into the blood product was less pronounced than that of DEHP.

Thiol-based regulation of glyceraldehyde-3-phosphate dehydrogenase in blood bank-stored red blood cells: a strategy to counteract oxidative stress.

BACKGROUND: Red blood cell (RBC) glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a glycolytic enzyme normally inhibited upon binding to the membrane-spanning protein Band 3, but active when free in the cytosol. Accumulating evidence in other cells indicates that oxidative thiol modifications in cytosolic GAPDH drive this molecule into functional avenues that deviate from glycolysis. This study aimed to investigate the role of GAPDH in oxidative stress-dependent metabolic modulations occurring in SAGM-stored RBCs, to increase the knowledge of the molecular mechanisms affecting RBC survival and viability under blood banking conditions. STUDY DESIGN AND METHODS: Membranes and cytosol from CPD SAGM-stored RBCs were subjected to Western blotting with anti-GAPDH at 0, 7, 14, 21, 28, 35, and 42 days of preservation. Immunoreactive bands were excised, digested with trypsin, and analyzed by mass spectrometry for the presence of oxidative posttranslational modifications. GAPDH enzymatic activity was also measured in the cytosolic fraction during storage. RESULTS: At 21 days of storage, we demonstrated that cytosolic GAPDH undergoes temporary inactivation due to the formation of an intramolecular disulfide bond between the active-site Cys-152 and nearby Cys-156, a mechanism to rerouting glucose flux toward the pentose phosphate pathway. In addition, an increase in the membrane-bound GAPDH was detected in long-stored RBCs. CONCLUSION: Reversible inhibition or activation of cytosolic GAPDH may represent a protective strategy against oxidative stress to favor NADPH production in stored RBCs.

Five-year longitudinal assessment (2008 to 2012) of E-101 solution activity against clinical target and antimicrobial-resistant pathogens.

This study summarizes the topical E-101 solution susceptibility testing results for 760 Gram-positive and Gram-negative target pathogens collected from 75 U.S. sites between 2008 and 2012 and 103 ESKAPE pathogens. E-101 solution maintained potent activity against all bacterial species studied for each year tested, with MICs ranging from <0.008 to 0.25 µg porcine myeloperoxidase (pMPO)/ml. These results confirm that E-101 solution retains its potent broad-spectrum activity against U.S. clinical isolates and organisms with challenging resistance phenotypes.

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

BACKGROUND: 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. STUDY DESIGN AND METHODS: Platelet concentrates (PCs) were divided into two equal aliquots (control group and test group). After the centrifugation of both aliquots and removal of as much supernatant as possible, the pellet of the control group was resuspended in 160 mL of M-sol while that of the test group was resuspended in 80 or 40 mL of M-sol. The wPLTs of both groups were stored in polyolefin bags with agitation at 20 to 24°C for 7 days. RESULTS: The pH values of both groups were maintained at higher than 7.0 during the 7-day storage. Differences in %disk, CD62P, annexin V, percent hypotonic shock response, and aggregation values between the test group and control group were small for at least 2 days after washing. CONCLUSIONS: The in vitro properties of VR-wPLTs were not markedly degraded for at least 2 days. Therefore, the storage properties of PLTs may be maintained in VR-wPLTs prepared at blood centers until they are administered to patients in hospitals.

The granulocytes in neutropenia 1 (GIN 1) study: a safety study of granulocytes collected from whole blood and stored in additive solution and plasma.

OBJECTIVE/AIM: To evaluate the safety of transfusing pooled, whole blood-derived granulocytes in additive solution and plasma (GASP) in 30 recipients. BACKGROUND: Demand for granulocytes in England has increased five-fold. With the advantages of reduced red cell, plasma and overall volume, GASP maintains function in vitro. METHODS AND MATERIALS: Observations were recorded prior to and post transfusion. Increments were recorded at 1 h and the following morning. Leucocyte antibody screening was undertaken prior to and at 1-6 months following transfusion. RESULTS: Thirty patients aged between 8 months and 68 years received 221 GASP in 148 transfusion episodes. GASP contained an average of 1.0 × 10(10) granulocytes in 207 mL. Adults usually received two packs and children 10-20 mL kg(-1). Children and adults received a median [interquartile range (IQR)] dose of 12.5 (9.1-25.3) and 19.7 (12.0-25.8) × 10(9) granulocytes per transfusion, respectively. There was one episode of transfusion-associated circulatory overload (TACO) in a patient with chronic cardiac failure following 600 mL of unpooled granulocytes, other fluids and one GASP. New leucocyte alloimmunisation occurred in 3/30 recipients 10%. No other significant reactions were reported. Median peripheral blood neutrophil increments at 1 h post transfusion were 0.06 (IQR, 0.01-0.17) in children and (0.03) (IQR, 0-0.16) in adults. CONCLUSION: GASP has a similar safety profile to other sources of granulocytes for patients with refractory infection or in need of secondary prophylactic transfusion. Further studies are required to clarify the role of GASP in the treatment of neutropenic patients.

[Intoxication syndrome concomitant with recurrent streptococcal tonsillitis and its correction].

The objective of the present work was to study selected features of intoxication in the patients presenting with recurrent tonsillitis (RT) and managed by means of the combination of basal therapy with reamberine, a preparation showing the detoxication activity. A total of 70 patients with moderately severe recurrent streptococcal tonsillitis induced by infection with group A beta-hemolytic streptococci were available for the examination. Characteristics of endogenous intoxication were explored. The study has demonstrated that recurrent tonsillitis is associated with the development of the pronounced symptoms of endogenous intoxication (EI) as suggested by the enhanced leukocytic index of intoxication (LII) and the elevated levels of medium molecular weight molecules (MWM) with the simultaneous impairment of detoxication properties of albumin. Collectively, these changes are indicative of incompleteness of the pathological process that may be regarded as an unfavourable prognostic factor and suggests the risk of development of tonsillogenic pathology, its relapses, and chronization of the process. Taken together, these findings indicate the necessity of developing novel approaches to the treatment of recurrent tonsillitis with special reference to the use of pharmaceutical products possessed of detoxication activity. Reamberine introduced in the combined treatment of RT was shown to decrease the severity of intoxication as confirmed by normalization of LII and MWM levels as well as the improvement of detoxication properties of albumin. It is concluded that therapeutic efficacy of reamberine opens up wide prospects for its clinical applications.

Hemostatic function of buffy coat platelets in additive solution treated with pathogen reduction technology.

BACKGROUND: Pathogen reduction technologies (PRTs) may influence the hemostatic potential of stored platelet (PLT) concentrates. To investigate this, buffy coat PLTs (BCPs) stored in PLT additive solution (SSP+) with or without Mirasol PRT treatment (CaridianBCT Biotechnologies) were compared by functional hemostatic assays. STUDY DESIGN AND METHODS: We performed in vitro comparison of PRT (PRT-BCP) and control pooled-and-split BCPs (CON-BCP) after 2, 3, 6, 7, and 8 days' storage. Hemostatic function was evaluated with thrombelastography (TEG) and impedance aggregometry (Multiplate), the latter also in a sample matrix (Day 2) with or without addition of red blood cells (RBCs), control plasma, and/or PRT-treated plasma. RESULTS: PRT treatment of 8-day-stored BCPs influenced clot formation (TEG) minimally, with reductions in maximum clot strength (maximum amplitude, p = 0.014) but unchanged initial fibrin formation (R), clot growth rate (α), and fibrinolysis resistance. In the absence of RBCs and plasma, PRT impaired aggregation (Multiplate) in stored BCPs, with reduced aggregation against thrombin receptor activating peptide-6 (p < 0.001), collagen (p = 0.014), adenosine 5'-diphosphate (p = 0.007), and arachidonic acid (p = 0.070). Addition of RBCs and PRT-treated or untreated plasma to PRT-BCP and CON-BCP, respectively, enhanced aggregation in both groups. CONCLUSIONS: Mirasol PRT treatment of BCPs had a minimal influence on clot formation, whereas aggregation in the absence of RBCs and plasma was significantly reduced. Addition of RBCs and plasma increased agonist-induced responses resulting in comparable aggregation between PRT-BCP and CON-BCP. The clinical relevance for PLT function in vivo of these findings will be investigated in a clinical trial.

The effect of pathogen reduction technology (Mirasol) on platelet quality when treated in additive solution with low plasma carryover.

BACKGROUND AND OBJECTIVES: Pathogen reduction technologies (PRT) for platelets are now compatible with both plasma and platelet additive solutions (PAS). The aim of this study was to examine the effect of PRT on the platelet storage lesion, in the presence of PAS with low plasma carryover. MATERIALS AND METHODS: PRT-treated (Mirasol) and untreated buffy coat-derived platelet concentrates prepared in 28% plasma/PAS-IIIM were evaluated using in vitro cell quality parameters on days 1, 2, 5, and 7 post-collection. RESULTS: At day 5, there were no significant differences between control and PRT treated platelets for swirl, viability, pO(2) , pCO(2) , mean platelet volume and adenosine diphosphate-induced aggregation. PRT treatment did not affect the functional integrity of the mitochondria. However, PRT resulted in a decrease in pH and enhancement of platelet glycolysis and activation, evidenced by increased glucose consumption and lactate production rates, increased expression of CD62P, CD63, annexin V staining and increased secretion of cytokines (P < 0.05). Hypotonic shock response and aggregation in response to collagen were also significantly reduced in PRT treated platelets (P < 0.05). CONCLUSION: Despite the observed differences in platelet metabolism and activation observed following PRT treatment in PAS and low plasma carryover, the results suggest that treatment and storage of platelets in PAS is no more detrimental to platelets than treatment and storage in plasma.