Releasates of riboflavin/UV-treated platelets: Microvesicles suppress cytokine-mediated endothelial cell migration/proliferation.

BACKGROUND: Accelerated development of the platelet (PLT) storage lesion upon pathogen inactivation (PI) is associated with the release of proteins from granules and platelet microvesicles (PMVs). Whether PI treatments alter the interaction between PLT factors and the vessel endothelium is of interest in understanding the risk profile of these technologies. STUDY DESIGN AND METHODS: In a pool-and-split study, one platelet concentrate (PC) was treated with riboflavin/UV (RF/UV) light, while the other one was kept as an untreated control. Releasates and PMV-depleted releasates were prepared by differential centrifugation steps on days 0, 1, 5, and 7 of storage. Cytokine/chemokine release following PI treatment was analyzed by an antibody array, and results were verified by the enzyme-linked immunosorbent assay. PMVs were enumerated by CD41 labeling and flow cytometry. Wound scratch assays were performed using cultured Ea.hy926 cells exposed to the differently prepared releasates. Effects of releasates on the phosphorylation levels of kinases ERK and p38 expressed by endothelial cells were analyzed by immunoblot. RESULTS: Cytokine/chemokine assays identified a 2-fold increase in epidermal growth factor released from PCs treated with RF/UV light compared with control. PMV count increased ~100-fold following PI treatment. Unmodified releasates and PMV-depleted releasates displayed different contributions to the kinetics of endothelial cell wound closure. This observation was associated with an increased ERK versus unaltered p38 activation in the endothelial cells. CONCLUSION: This study identified an inhibitory impact of PMVs on endothelial cell migration/proliferation upon stimulation by released cytokines and PMVs from PLTs treated with RF/UV light for endothelial cell wound closure.

Hemostatic efficacy of pathogen-inactivated vs untreated platelets: a randomized controlled trial.

Pathogen inactivation of platelet concentrates reduces the risk for blood-borne infections. However, its effect on platelet function and hemostatic efficacy of transfusion is unclear. We conducted a randomized noninferiority trial comparing the efficacy of pathogen-inactivated platelets using riboflavin and UV B illumination technology (intervention) compared with standard plasma-stored platelets (control) for the prevention of bleeding in patients with hematologic malignancies and thrombocytopenia. The primary outcome parameter was the proportion of transfusion-treatment periods in which the patient had grade 2 or higher bleeding, as defined by World Health Organization criteria. Between November 2010 and April 2016, 469 unique patients were randomized to 567 transfusion-treatment periods (283 in the control arm, 284 in the intervention arm). There was a 3% absolute difference in grade 2 or higher bleeding in the intention-to-treat analysis: 51% of the transfusion-treatment periods in the control arm and 54% in the intervention arm (95% confidence interval [CI], -6 to 11; P = .012 for noninferiority). However, in the per-protocol analysis, the difference in grade 2 or higher bleeding was 8%: 44% in the control arm and 52% in the intervention arm (95% CI -2 to 18; P = .19 for noninferiority). Transfusion increment parameters were ∼50% lower in the intervention arm. There was no difference in the proportion of patients developing HLA class I alloantibodies. In conclusion, the noninferiority criterion for pathogen-inactivated platelets was met in the intention-to-treat analysis. This finding was not demonstrated in the per-protocol analysis. This trial was registered at The Netherlands National Trial Registry as #NTR2106 and at www.clinicaltrials.gov as #NCT02783313.

Improved in vitro quality of stored red blood cells upon oxygen reduction prior to riboflavin/UV light treatment of whole blood.

BACKGROUND: The application of riboflavin/UV-based pathogen inactivation (PI) to whole blood (WB) is currently limited by its negative impact on red blood cell (RBC) quality. The generation of reactive oxidative species in RBC products contributes to increased hemolysis. This study evaluated the impact of deoxygenation of WB prior to riboflavin/UV light treatment versus deoxygenation of RBC concentrates after PI treatment by monitoring RBC in vitro quality parameters. STUDY DESIGN AND METHODS: Six ABO-matched WB units were pooled and split. Within three pairs, one unit was treated with riboflavin/UV light while the other was kept as an untreated control prior to manufacture into red cell concentrates (RCCs). The first pair (Cntr; Cntr-PI) served as the normoxic controls. Deoxygenation was performed at the RCC level for the second pair (RCCdeox; PI-RCCdeox), and at the WB level of the third pair (WBdeox; WBdeox-PI). In vitro qualities of the respective RBC units were assessed throughout storage. RESULTS: The data for the Cntr and Cntr-PI units were comparable to previous reports. The PI-RCCdeox units exhibited worse in vitro quality for most parameters tested compared to Cntr-PI and WBdeox-PI units throughout storage. Hemolysis and microvesicle release was significantly (p < 0.05) higher on Days 21 and 42 in Cntr-PI units compared to WBdeox-PI units. CONCLUSION: WB deoxygenation may help to decrease the accelerated deterioration in RCC in vitro quality caused by treatment with riboflavin/UV light. Treatment of WB under reduced oxygen levels needs to be assessed for PI effectiveness.

Red Blood Cells Derived from Whole Blood Treated with Riboflavin and UV Light Maintain Adequate Cell Quality through 21 Days of Storage.

BACKGROUND: The Mirasol system for whole blood (WB) is a non-toxic, non-mutagenic pathogen reduction technology (PRT) that treats WB units with riboflavin (vitamin B2) and ultraviolet (UV) light to alter nucleic acids, thereby reducing pathogen infectivity and inactivating white blood cells. This study evaluates the quality of red blood cells (RBCs) derived from WB treated with the Mirasol system. STUDY DESIGN AND METHODS: Paired units of WB were collected from 61 healthy donors. One unit per donor was treated with riboflavin and UV light and the other was used as an untreated control. RBCs were processed from the WB units and stored in AS-3 at 1-6°C for 21 days and sampled for in vitro analyses of RBC quality parameters. RESULTS: Several statistically significant differences were observed between test and control units, but values were overall within normal clinical ranges. After leukoreduction, the residual leukocyte count and RBC recovery met FDA requirements. The RBC units derived from treated WB maintained haemolysis below 1% through 21 days of storage. CONCLUSION: RBCs derived from WB treated with the Mirasol system meet accepted FDA guidelines for RBC quality through 21 days of storage at 1-6°C.

Special considerations for the use of pathogen reduced blood components in pediatric patients: An overview.

Pediatric patients requiring transfusion constitute one of the most challenging areas of transfusion practice. Due to the limitations posed by their particular physiological conditions they routinely require specialized component support and more personalised transfusion care than what is routinely utilized in the care of adult patients. Pediatric patients, unlike many adult patients requiring transfusion support, also generally have significant lifespans post-transfusion. This combined with the possibility of long term consequences to adverse events related to transfusion such as infection or change in immunological status drives the need to continue to use improved blood components in the transfusion support of pediatric patients. While considerable progress has been made on methods to improve the safety and efficacy of blood components, the use of these products in pediatric patients continues to be a subject of debate and additional considerations. These additional considerations arise due to the changes in blood product quality and function that is observed following treatment with pathogen reduction methods. Additional considerations regarding toxicological and immunological aspects related to these products are heightened in the case of pediatric patients. This manuscript provides an overview of current practice regarding the use of pathogen reduced products in pediatric patients with discussion of issues for consideration in their implementation in routine, including cost/benefit and risk/benefit aspects associated with their use.

Reflections on the dynamics of bacterial and viral contamination of blood components and the levels of efficacy for pathogen inactivation processes.

Blood transfusion safety has been increasingly improving during the past two decades. However, threats from both known and emerging pathogens require continual improvement and re-assessment of blood safety measures. In this respect, we are currently witnessing the broader implementation of Pathogen reduction technology (PRT) for blood complements. These methods, combined with existing safety measures, have helped to reduce the pathogen risks of transfusion-transmitted infections. Currently multiple reviews have compared levels of inactivation between different commercialized PRTs. However, to analyze levels of pathogen inactivation, it is necessary to understand the dynamics of infectivity as well as the modes of disease transmission by blood transfusion for various pathogens. It is well known that contributing variables include donor characteristics through the processing of blood components to ultimately the recipient characteristics, which create enormous variability in overall outcomes relative to disease transmission. The aim of this paper is to discuss bacterial and viral contamination of blood components in order to determine adequate levels of efficacy and subsequent disease transmission safety of current pathogen inactivation protocols that are designed to reduce the risk of transfusion-transmitted infections. In such a conceptual analysis, however, it is important to understand several contributing factors including the measurement of pathogen load in blood products and the dynamics, infectivity and disease transmission of various pathogens via transfusion of blood components and products. In many cases, the log reduction values observed do not truly reflect the extent of reduction in the levels of infectivity that are observed clinically. Results from clinical trials and hemovigilance programs upon routine implementation of PRT methods provide a more direct insight into effectiveness with regard to clinical relevance of in vitro spiking studies. These issues are briefly addressed in this manuscript.

Effect of Plasmodium inactivation in whole blood on the incidence of blood transfusion-transmitted malaria in endemic regions: the African Investigation of the Mirasol System (AIMS) randomised controlled trial.

BACKGROUND: Transfusion-transmitted malaria is a frequent but neglected adverse event in Ghana. We did a randomised controlled clinical trial to assess the efficacy and safety of a whole blood pathogen reduction technology at preventing transfusion transmission of Plasmodium spp parasites. METHODS: For this randomised, double-blind, parallel-group clinical trial, eligible adult patients (aged ≥ 18 years) with blood group O+, who required up to two whole blood unit transfusions within 3 days of randomisation and were anticipated to remain in hospital for at least 3 consecutive days after initial transfusion, were enrolled from Komfo Anokye Teaching Hospital in Kumasi, Ghana. The main exclusion criteria were symptoms of clinical malaria, antimalaria treatment within 7 days before randomisation, fever, and haemorrhage expected to require transfusion with up to two units of whole blood during the 3 days following study entry. Eligible patients were randomly assigned 1:1 by computer-generated permuted block randomisation (block size four) list to receive transfusion with either pathogen-reduced whole blood (treated) or whole blood prepared and transfused by standard local practice (untreated). Patients, health-care providers, and data collectors were masked to treatment allocation. Patients in both groups received up to two whole blood unit transfusions that were retrospectively tested for parasitaemia. Pre-transfusion and post-transfusion blood samples (taken on days 0, 1, 3, 7, and 28) were tested for presence and amount of parasite genome, and assessed for haematological and biochemical parameters. The primary endpoint was the incidence of transfusion-transmitted malaria in non-parasitaemic recipients exposed to parasitaemic whole blood, defined as two consecutive parasitaemic post-transfusion samples with parasite allelic matching, assessed at 1-7 days after transfusion. Secondary endpoints included haematological parameters and a safety analysis of adverse events in patients. This study is registered with ClinicalTrials.gov, number NCT02118428, and with the Pan African Clinical Trials Registry, number PACTR201406000777310. FINDINGS: Between March 12, 2014, and Nov 7, 2014, 227 patients were enrolled into the study, one of whom was subsequently excluded because she did not meet the inclusion criteria. Of the 226 randomised patients, 113 were allocated to receive treated whole blood and 113 to receive standard untreated whole blood. 223 patients (111 treated and 112 untreated) received study-related transfusions, whereas three patients (two treated and one untreated) did not. 214 patients (107 treated and 107 untreated) completed the protocol as planned and comprised the per-protocol population. Overall, 65 non-parasitaemic patients (28 treated and 37 untreated) were exposed to parasitaemic blood. The incidence of transfusion-transmitted malaria was significantly lower for the pathogen-reduced (treated) patients (1 [4%] of 28 patients) than the untreated group (8 [22%] of 37 patients) in this population (p=0.039). Overall, 92 (41%) of 223 patients reported 145 treatment-related emergent adverse events during the conduct of the study, with a similar incidence of adverse events between groups receiving untreated or treated whole blood. No transfusion-related deaths occurred in the trial. INTERPRETATION: Treatment of whole blood with the Mirasol pathogen reduction system for whole blood reduced the incidence of transfusion-transmitted malaria. The primary endpoint of the study was achieved in the population of non-parasitaemic patients receiving parasitaemic whole blood. The safety profile and clinical outcomes were similar across the two treatment groups. FUNDING: Terumo BCT Inc.

Reduced MHC alloimmunization and partial tolerance protection with pathogen reduction of whole blood.

BACKGROUND: Allogeneic blood transfusion can result in an immune response against major histocompatibility complex (MHC) antigens, potentially complicating future transfusions or transplants. We previously demonstrated that pathogen reduction of platelet-rich plasma (PRP) with riboflavin and ultraviolet light (UV+R) can prevent alloimmunization in mice. A similar pathogen-reduction treatment is currently under development for the treatment of whole blood using riboflavin and a higher dose of UV light. We sought to determine the effectiveness of this treatment in the prevention of alloimmunization. STUDY DESIGN AND METHODS: BALB/cJ mice were transfused with untreated or UV+R-treated, allogeneic C57Bl/6J whole blood with or without leukoreduction. Mice were evaluated for donor-specific antibodies, ex vivo splenocyte cytokine responses, and changes in the frequency of regulatory T (Treg ) cells. RESULTS: UV+R treatment blocked cytokine priming and reduced anti-MHC alloantibody responses to transfused whole blood. Leukoreduction reduced alloantibody levels in both the untreated and UV+R-treated groups. Mice transfused with UV+R-treated whole blood had reduced alloantibody and cytokine responses when subsequently transfused with untreated blood from the same donor type. This reduction in responses was not associated with increased Treg cells. CONCLUSIONS: Pathogen reduction of whole blood with UV+R significantly reduces, but does not eliminate, the alloimmune response. Exposure to UV+R-treated whole blood transfusion does appear to induce tolerance to alloantigens, resulting in reduced anti-MHC alloantibody and cytokine responses to subsequent exposures to the same alloantigens. This tolerance does not appear to be driven by an increase in Treg cells.

Red blood cells derived from whole blood treated with riboflavin and ultraviolet light maintain adequate survival in vivo after 21 days of storage.

BACKGROUND: Pathogen reduction (PR) of whole blood (WB) may increase blood safety when applied before component separation. This study evaluates the in vivo performance of red blood cells (RBCs) derived from WB treated with the riboflavin and ultraviolet (UV) light PR (Mirasol) system. STUDY DESIGN AND METHODS: This was a prospective, two-center, single-blind, randomized, two-period, crossover clinical trial designed to evaluate autologous 51 Cr/99m Tc-radiolabeled recovery and survival of RBCs derived from Mirasol-treated WB compared to untreated WB. RBCs were stored in AS-3 for 21 days at 1 to 6°C. In vitro RBC variables were characterized. Frequency and severity of treatment-emergent adverse event (TEAE) and neoantigenicity were determined. RESULTS: Twenty-four healthy adult volunteers (n = 12 per site) were evaluated. The Mirasol 24-hr RBC recoveries were 82.5 ± 3.9% with one-sided 95% lower confidence limit of 80.9%, meeting US Food and Drug Administration acceptance criteria, albeit at lower level than controls (91.7 ± 6.8%, p < 0.001). Mean RBC survival and T50 were reduced in the Mirasol group (61 and 23 days, respectively) versus controls (82 and 36 days, respectively; p < 0.001) with a mean area under the curve survival of treated RBCs of 83% of untreated controls. End-of-storage hemolysis in the Mirasol group was 0.22 ± 0.1% (control, 0.15 ± 0.1%; p < 0.001). No neoantigenicity or differences in TEAEs were found. CONCLUSION: RBCs derived from Mirasol WB and stored for up to 21 days in AS-3 maintained acceptable cell quality and recovery, albeit modestly reduced compared with untreated RBCs. Mirasol WB may represent a valid single WB PR platform that allows manufacture of RBC for storage for up to 21 days.

Riboflavin-ultraviolet light pathogen reduction treatment does not impact the immunogenicity of murine red blood cells.

BACKGROUND: Ultraviolet (UV) illumination/pathogen reduction effectively inactivates white blood cells (WBCs) in whole blood. Given that cotransfused WBCs may impact recipient immune responses, we hypothesized that pathogen reduction of whole blood may alter responses to RBC antigens. STUDY DESIGN AND METHODS: Transgenic mice expressing a model (HOD) antigen, authentic human (hGPA or KEL) antigens, or natural fluorescence (uGFP) on their RBCs were utilized as blood donors. Recipients were transfused with fresh whole blood to which riboflavin had been added or fresh whole blood treated by UV illumination/pathogen reduction treatment after the addition of riboflavin. Posttransfusion RBC recovery, survival, and alloimmunization were measured by flow cytometry. RESULTS: UV illumination/pathogen reduction treatment did not alter RBC antigen expression, and recipients of treated syngeneic RBCs had persistently negative direct antiglobulin tests. Greater than 75% of treated and untreated syngeneic RBCs were recovered 24 hours posttransfusion in all experiments, although alterations in the long-term posttransfusion survival of treated RBCs were observed. Treated and untreated KEL RBCs induced similar recipient alloimmune responses, with all recipients making anti-KEL glycoprotein immunoglobulins (p > 0.05). Alloimmune responses to treated HOD or hGPA RBCs were no different from untreated RBCs (p > 0.05). CONCLUSION: Pathogen inactivation treatment of fresh whole murine blood with riboflavin and UV illumination does not impact the rate or magnitude of RBC alloimmunization to three distinct RBC antigens. Further, UV illumination/pathogen reduction appears safe from an immunohematologic standpoint, with no immunogenic neoantigens detected on treated murine RBCs. Future studies with fresh and stored human RBCs are warranted to confirm these findings.

Reduced alloimmunization in mice following repeated transfusion with pathogen-reduced platelets.

BACKGROUND: Allogeneic transfusion can result in alloimmunization, leading to platelet (PLT) refractoriness and rejection of solid organ transplants. Previously we demonstrated that pathogen reduction using UV light and riboflavin (UV + R) eliminates the immunogenicity of white blood cells (WBCs) in vitro, blocks alloimmunization from transfusion in mice, and results in reduced ex vivo cytokine responses to subsequent untreated transfusions. We sought to determine if repeated transfusion with pathogen-reduced PLT-rich plasma (PRP) would eventually cause breakthrough alloimmunization or enhanced tolerance. STUDY DESIGN AND METHODS: BALB/cJ mice were transfused weekly for 2, 4, or 8 weeks with C57Bl/6J PRP that was either untreated or pathogen reduced with UV + R and leukoreduced or not. Alloimmunization was determined by measuring donor antibody levels, ex vivo cytokine responses, and 24-hour donor PLT recovery. The role of donor antibodies in PLT refractoriness was also assessed by transfer of diluted immune sera into naïve recipients. RESULTS: Donor antibody levels increased with the number of transfusions, but levels were significantly reduced using either UV + R or leukoreduction, and combining UV + R and leukoreduction gave the best protection. Priming of ex vivo cytokine responses required WBCs and remained suppressed with repeated UV + R-treated transfusion. PLT recovery was reduced with UV + R in naïve mice, and multiply transfused mice had poor PLT recovery even when antibody levels were relatively low. Approximately 1/100 dose of serum from a multiply transfused mouse was sufficient for complete rejection of donor PLTs. CONCLUSIONS: Pathogen reduction significantly reduces alloimmunization in repeatedly transfused mice and combined with leukoreduction provides a high level of protection from alloimmunization.

Characterization of posttransfusion Plasmodium falciparum infection in semi-immune nonparasitemic patients.

BACKGROUND: Transfusion-transmitted malaria (TTM) has not been studied with molecular means in hyperendemic areas where it is assumed to occur frequently. The African Investigation of the Mirasol System (AIMS) trial provided the opportunity to study TTM from standard whole blood (WB) units. STUDY DESIGN AND METHODS: The Plasmodium genome in transfused WB units and patient samples both before transfusion and 1, 3, 7, and 28 days after transfusion was screened and quantified using real-time polymerase chain reaction. Parasitemic samples were confirmed, three alleles were sequenced, and the percentage homology was determined between paired WB units and patient samples. Anti-Plasmodium titers were quantified by serial dilution. Clinical symptoms and microscopic detection of malaria were monitored. RESULTS: Microscopy was negative below 3 × 10(6) genome copies/mL. Thirty-seven patients who were nonparasitemic before transfusion were exposed to parasitemic WB. The amount of Plasmodium genome load transfused ranged between 0.1 × 10(6) and 2.0 × 10(9) copies. The parasite load received through transfusion by 13 patients with TTM was higher than that received by patients without TTM (p = 0.01). Patients with a single parasitemic posttransfusion sample were not considered to have TTM. Four elements critical to predict outcome emerged: parasite load, patient anti-Plasmodium titer pretransfusion, percentage clearance of parasites at Day 1, and level of anti-Plasmodium humoral immune response. Four patients with TTM became parasite-free at Day 28 (effective control), four patients with TTM maintained relatively stable levels of parasitemia (uncertain control), and five patients reached high levels of parasitemia at Day 28 posttransfusion, indicating ineffective control of malarial infection by semi-immune individuals. CONCLUSIONS: TTM in endemic areas is relatively frequent (13 of 112 donations; 11.6%) and, although largely controlled by semi-immunity in recipient patients, may require antimalarial treatment.

Treatment of blood with a pathogen reduction technology using ultraviolet light and riboflavin inactivates Ebola virus in vitro.

BACKGROUND: Transfusion of plasma from recovered patients after Ebolavirus (EBOV) infection, typically called "convalescent plasma," is an effective treatment for active disease available in endemic areas, but carries the risk of introducing other pathogens, including other strains of EBOV. A pathogen reduction technology using ultraviolet light and riboflavin (UV+RB) is effective against multiple enveloped, negative-sense, single-stranded RNA viruses that are similar in structure to EBOV. We hypothesized that UV+RB is effective against EBOV in blood products without activating complement or reducing protective immunoglobulin titers that are important for the treatment of Ebola virus disease (EVD). STUDY DESIGN AND METHODS: Four in vitro experiments were conducted to evaluate effects of UV+RB on green fluorescent protein EBOV (EBOV-GFP), wild-type EBOV in serum, and whole blood, respectively, and on immunoglobulins and complement in plasma. Initial titers for Experiments 1 to 3 were 4.21 log GFP units/mL, 4.96 log infectious units/mL, and 4.23 log plaque-forming units/mL. Conditions tested in the first three experiments included the following: 1-EBOV-GFP plus UV+RB; 2-EBOV-GFP plus RB only; 3-EBOV-GFP plus UV only; 4-EBOV-GFP without RB or UV; 5-virus-free control plus UV only; and 6-virus-free control without RB or UV. RESULTS: UV+RB reduced EBOV titers to nondetectable levels in both nonhuman primate serum (≥2.8- to 3.2-log reduction) and human whole blood (≥3.0-log reduction) without decreasing protective antibody titers in human plasma. CONCLUSION: Our in vitro results demonstrate that the UV+RB treatment efficiently reduces EBOV titers to below limits of detection in both serum and whole blood. In vivo testing to determine whether UV+RB can improve convalescent blood product safety is indicated.

Reduction of Leishmania donovani infectivity in whole blood using riboflavin and ultraviolet light.

BACKGROUND: Leishmaniasis is a vector-borne disease caused by the protozoan parasite Leishmania sp. that is transmitted by sandflies. Travelers to endemic areas, and US military personnel stationed in the Middle East, are at risk for contracting the disease. STUDY DESIGN AND METHODS: Whole blood (WB) units were spiked with human monocytes infected with L. donovani amastigotes to a final concentration of approximately 10(5) infected cells/mL. After riboflavin (RB) addition, units were exposed to 80 J/mLRBCs ultraviolet (UV) light. One pretreatment (collected after RB addition) and one posttreatment sample were collected, serially diluted in culture medium, and incubated at 22°C for up to 5 weeks. Parasite viability was determined by microscopic observation for replicating promastigote forms. RESULTS: Mirasol treatment of 3 units of L. donovani-infected WB with RB and UV light resulted in a parasite reduction of 2.3 ± 0.12 log. CONCLUSIONS: Partial reduction of L. donovani can be achieved in WB using RB and UV light. This technology may be useful when potential donors are exposed to Leishmania sp. during residence, travel, or military deployment to an endemic area.

Whole blood treated with riboflavin and ultraviolet light: quality assessment of all blood components produced by the buffy coat method.

BACKGROUND: Pathogen inactivation (PI) technologies are currently licensed for use with platelet (PLT) and plasma components. Treatment of whole blood (WB) would be of benefit to the blood banking community by saving time and costs compared to individual component treatment. However, no paired, pool-and-split study directly assessing the impact of WB PI on the subsequently produced components has yet been reported. STUDY DESIGN AND METHODS: In a "pool-and-split" study, WB either was treated with riboflavin and ultraviolet (UV) light or was kept untreated as control. The buffy coat (BC) method produced plasma, PLT, and red blood cell (RBC) components. PLT units arising from the untreated WB study arm were treated with riboflavin and UV light on day of production and compared to PLT concentrates (PCs) produced from the treated WB units. A panel of common in vitro variables for the three types of components was used to monitor quality throughout their respective storage periods. RESULTS: PCs derived from the WB PI treatment were of significantly better quality than treated PLT components for most variables. RBCs produced from the WB treatment deteriorated earlier during storage than untreated units. Plasma components showed a 3% to 44% loss in activity for several clotting factors. CONCLUSION: Treatment of WB with riboflavin and UV before production of components by the BC method shows a negative impact on all three blood components. PLT units produced from PI-treated WB exhibited less damage compared to PLT component treatment.

The effect of riboflavin and ultraviolet light on the infectivity of arboviruses.

BACKGROUND: Arboviruses are an emerging threat to transfusion safety and rates of infection are likely to increase with the increased rainfall associated with climate change. Arboviral infections are common in Australia, where Ross River virus (RRV), Barmah Forest virus (BFV), and Murray Valley encephalitis virus (MVEV), among others, have the potential to cause disease in humans. The use of pathogen reduction technology (PRT) may be an alternative approach for blood services to manage the risk of arboviral transfusion transmission. In this study, the effectiveness of the Mirasol PRT (Terumo BCT) system at inactivating RRV, BFV, and MVEV in buffy coat (BC)-derived platelets (PLTs) was investigated. STUDY DESIGN AND METHODS: BC-derived PLT concentrates in additive solution (SSP+) were spiked with RRV, BFV, or MVEV and then treated with the Mirasol PRT system. The level of infectious virus was determined before and after treatment, and the reduction in viral infectivity was calculated. RESULTS: Treatment with PRT (Mirasol) reduced the amount of infectious virus of all three arboviruses. The greatest level of inactivation was observed for RRV (2.33 log; 99.25%), followed by BFV (1.97 log; 98.68%) and then MVEV (1.83 log; 98.42%). CONCLUSION: Our study demonstrates that treatment of PLT concentrates with PRT (Mirasol) reduces the infectious levels of RRV, BFV, and MVEV. The relevance of the level of reduction required to prevent disease transmission by transfusion has not been fully defined and requires further investigation. In the face of a changing climate, with its associated threat to blood safety, PRT represents a proactive approach for maintaining blood safety.

Inactivation of viruses in platelet and plasma products using a riboflavin-and-UV-based photochemical treatment.

BACKGROUND: Multilayered blood safety programs reduce the risk of transfusion-transmitted diseases; however, there remains a risk of window period transmission of screened viruses and transmission of unscreened and emerging viruses from asymptomatic donors. To reduce this risk, a riboflavin-and-UV-light-based pathogen reduction process was evaluated against eight viral agents. STUDY DESIGN AND METHODS: Riboflavin and UV light was evaluated against the following eight viral agents: encephalomyocarditis virus (EMC), hepatitis A virus (HAV), hepatitis C virus (HCV), influenza A (FLUAV), La Crosse virus (LACV), pseudorabies virus (PRV), sindbis virus (SINV), and vesicular stomatitis virus (VSV). Before treatment, a sample was removed to determine the product's initial viral load. After treatment the product's viral load was reevaluated and the log reduction was calculated. RESULTS: Virus reduction after treatment with riboflavin and UV light is equivalent in platelet (PLT) and plasma units, as demonstrated by a 3.2-log reduction of EMC in plasma, PLTs, and PLT additive solution containing 35% plasma. Additionally, the following viral reductions values were observed: HAV 1.8 log, HCV at least 4.1 log, FLUAV at least 5.0 log, LACV at least 3.5 log, PRV 2.5 log, SINV 3.2 log, and VSV at least 6.3 log. CONCLUSIONS: The results observed in this study suggest that treating PLT and plasma products with a riboflavin-and-UV-light-based pathogen reduction process could potentially eliminate window period transmission of screened viruses and greatly reduce the risk of transfusion transmission of unscreened viruses.

Riboflavin and ultraviolet light treatment of platelets triggers p38MAPK signaling: inhibition significantly improves in vitro platelet quality after pathogen reduction treatment.

BACKGROUND: Pathogen reduction technologies (PRTs) significantly reduce the risk of transmission of infectious agents in platelet (PLT) concentrates; however, in vitro studies reveal a negative impact on PLT quality after PRT treatment including effects on PLT aggregation, integrin αIIbß3 conformation, and actin dynamics. Clinically, the interval between transfusions is shortened. STUDY DESIGN AND METHODS: Seeking to understand the biochemical mechanisms underlying these observed effects, we analyzed signal transduction in PLT concentrates after riboflavin and ultraviolet light (UV; Mirasol) treatment and subsequent storage focusing on the phosphorylation levels of selected protein kinases. RESULTS: Among identified candidates, p38MAPK increased fourfold in phosphorylation after PRT. Incubation of PLT concentrates with a p38MAPK-specific inhibitor before PRT significantly improved numerous PLT quality measures. Phosphorylation levels of the p38MAPK substrates AKT, VASP, and HSP27 also decreased with inhibitor treatment. Phospho-HSP27 decrease in the presence of the inhibitor correlated with a reduction in PLT activation determined by surface expression of P-selectin. CONCLUSION: These findings support a model of one dominant underlying molecular signaling mechanism that is impacted by the riboflavin and UV (Mirasol) PRT process resulting in alterations in PLT quality. The identification of such a target should assist in the development of strategies to ameliorate this negative aspect of an otherwise beneficial and important safety development for transfusion medicine.