Transfusion

BackgroundTo identify blood donors eligible to donate Coronavirus Disease-2019 (COVID-19) Convalescent Plasma (CCP), a large blood center began testing for antibodies to SARS-CoV-2, the etiologic agent of COVID-19. We report the seroprevalence of total immunoglobulin directed against the S1 spike protein of SARS-CoV-2 in US blood donors. MethodsUnique non-CCP donor sera from June 1-July 31, 2020 were tested with the Ortho VITROS Anti-SARS-CoV-2 total immunoglobulin assay (positive: signal-to-cutoff (S/C) [≥]1). Donor age, sex, race/ethnicity, ABO/RhD, education, and experience were compared to June and July 2019. Multivariate regressions were conducted to identify demographics associated with the presence of antibodies and with S/C values. ResultsUnique donors (n=252,882) showed an overall seroprevalence of 1.83% in June (1.37%) and July (2.26%), with the highest prevalence in northern New Jersey (7.3%). In a subset of donors with demographic information (n=189,565), higher odds of antibody reactivity were associated with non-Hispanic Native American/Alaskan (NH-NAA/A) and Black (NH-B), and Hispanic (H) race/ethnicity, age 18-64, middle school or lesser education, blood Group A, and never or non-recent donor status. In positive donors (n=2,831), antibody signal was associated with male sex, race/ethnicity (NH-NAA/A, NH-B and H) and geographic location. ConclusionsSeroprevalence remains low in US blood donors but varies significantly by region. Temporal trends in reactivity may be used to gauge the effectiveness of public health measures. Before generalizing these data from healthy donors to the general population however, rates must be corrected for false positive test results among low prevalence test subjects and adjusted to match the wider demography.

Hema-Quebec, the blood supplier in the Province of Quebec, Canada, collects and tests convalescent plasma used in a clinical trial to determine the clinical efficacy of this product for the treatment of hospitalized COVID-19 patients. So far, we have collected 1159 plasma units from 282 COVID-19 convalescent donors. The presence of antibodies to the receptor binding domain (RBD) of SARS-CoV-2 spike protein in convalescent donors was established at the first donation. Seropositive donors were asked to donate additional plasma units every six days. Until now, 15 donors have donated at least four times and, in some cases, up to nine times. This allowed us to perform a longitudinal analysis of the persistence of SARS-CoV-2 RBD-specific antibodies in these repeat donors, with the first donation occurring 33-77 days after symptoms onset and donations up to 71-114 days after symptoms onset thereafter. In all donors, the level of antibodies remained relatively stable up to about 76 days after symptoms onset but then started to decrease more rapidly to reach, in some convalescent donors, a seronegative status within 100-110 days after symptoms onset. The decline in anti-RBD antibodies was not related to the number of donations but strongly correlated with the numbers of days after symptoms onset (r = 0.821). This suggests that de novo secretion of SARS-CoV-2 RBD antibodies by short-lived plasma cells stopped about 2-3 months after disease onset, an observation that has important implications for convalescent plasma collection and seroprevalence studies undertaken several months after the peak of infection.

Structured AbstractO_ST_ABSBackgroundC_ST_ABSCivilian walking blood banks (WBBs) transfuse fresh whole blood from mobilized donors, screened using rapid diagnostic testing (RDT) for transfusion transmissible infections (TTIs), to preserve life when banked blood is unavailable. However, concerns regarding TTI risk using a RDT process instead of a more traditional, laboratory-based test persist. We aimed to understand the marginal risk of TTI using an RDT-based strategy compared to a laboratory-based test through development of a simulation model and accompanying online tool. Study Design and MethodsWe modeled expected TTIs per 100,000 donations from initial collection to transfusion and seroconversion. Parameters included TTI prevalence, donor risk-stratification, efficacy of stratification tools, TTI testing rates, platform test performance, and probability of seroconversion. ResultsA baseline TTI prevalence of 1% (95% CI: 0.25%, 1.75%) resulted in 56 TTIs (95% CI: 23, 91) when the RDT sensitivity was 90% (95% CI: 88%, 92%), 30 TTIs (95% CI: 12, 52) when the RDT sensitivity was 95% (95% CI: 93%, 97%), and 12 TTIs (95% CI: 4, 23) when the RDT sensitivity was 99% (95% CI: 97, 100%) per 100,000 donations. Compared to lab-based testing, 15,351 donations would need to be made under a high-sensitivity RDT testing strategy in order to incur one additional TTI. DiscussionIn a simulated WBB model, modern RDT platforms demonstrated favorable test characteristics, with low absolute rates of TTI, particularly when low-risk donors are selected. These findings support WBB implementation as an emergency transfusion strategy in settings lacking banked blood.

BackgroundThe novel coronavirus, SARS-CoV2 that causes COVID-19 has resulted in the death of more than 2.31 million people within the last year and yet no cure exists. Whereas passive immunization with COVID-19 convalescent plasma (CCP) provides a safe and viable option, selection of optimal units for therapy and lack of clear therapeutic benefit from transfusion remain as barriers to the use of CCP. Study design and methodsTo identify plasma that is expected to benefit recipients, we measured anti-SARS-CoV2 antibody levels using clinically available serological assays and correlated with the neutralizing activity of CCP from donors. Neutralizing titer of plasma samples was measured by assaying infectivity of SARS-CoV-2 spike protein pseudotyped retrovirus particles in the presence of dilutions of plasma samples. We also used this assay to identify evidence of passive transfusion of neutralizing activity in CCP recipients. ResultsViral neutralization and anti-spike protein antibodies in 109 samples from 87 plasma donors were highly varied but modestly correlated with each other. Recipients who died of COVID-19 were found to have been transfused with units with lower anti-spike antibody levels and neutralizing activity. Passive transfer of neutralization activity was documented in 62% of antibody naive plasma recipients. ConclusionsSince viral neutralization is the goal of CCP transfusion, our observations not only support the use of anti-spike SARS-CoV2 serology tests to identify beneficial CCP units, but also support the therapeutic value of convalescent plasma with high titers of anti-spike antibodies.

RationaleMean platelet volume (MPV) and platelet count (PLT) are platelet measures that have been linked to cardiovascular disease (CVD) and mortality risk. Identifying protein biomarkers for these measures may yield insights into CVD mechanisms. ObjectiveWe aimed to identify causal protein biomarkers for MPV and PLT among 71 CVD-related plasma proteins measured in Framingham Heart Study (FHS) participants. Methods and ResultsWe conducted integrative analyses of genetic variants associated with PLT and MPV with protein quantitative trait locus (pQTL) variants associated with plasma proteins followed by Mendelian randomization (MR) to infer causal relations of proteins for PLT/MPV, and tested protein-PLT/MPV association in FHS participants. Utilizing induced pluripotent stem cell (iPSC)-derived megakaryocyte (MK) clones that produce functional platelets, we conducted RNA-sequencing and analyzed transcriptome-wide differences between low- and high-platelet producing clones. We then performed small interfering RNA (siRNA) gene knockdown experiments targeting genes encoding proteins with putatively causal platelet effects in MK clones to examine effects on platelet production. Protein-trait association analyses were conducted for MPV (n = 4,348) and PLT (n = 4,272). Eleven proteins were associated with MPV and 31 with PLT. MR identified four putatively causal proteins for MPV and four for PLT. Glycoprotein V (GP5), granulin (GRN), and melanoma cell adhesion molecule (MCAM) were associated with PLT in both protein-trait and MR analyses. Myeloperoxidase (MPO) showed significant association with MPV in both analyses. MK RNA-sequencing analysis results were directionally concordant with observed and MR-inferred associations for GP5, GRN, and MCAM. In siRNA gene knockdown experiments, silencing GP5, GRN, and MPO decreased platelet counts. ConclusionsBy integrating population genomics data, epidemiological data, and iPSC-derived MK experiments, we identified four proteins that are causally linked to platelet counts. These proteins and genes may be further explored for their utility in increasing platelet production in bioreactors for transfusion medicine purposes as well as their roles in the pathogenesis of CVD via a platelet/blood coagulation-based mechanism.

The novel coronavirus SARS-CoV2, which causes COVID-19, has resulted in the death of nearly 4 million people within the last 18 months. While preventive vaccination and monoclonal antibody therapies have been rapidly developed and deployed, early in the pandemic the use of COVID-19 convalescent plasma (CCP) was a common means of passive immunization, with the theoretical risk of antibody-dependent enhancement (ADE) of viral infection remaining undetermined. Though vaccines elicit a strong and protective immune response, and transfusion of CCP with high titers of neutralization activity are correlated with better clinical outcomes, the question of whether antibodies in CCP can enhance infection of SARS-CoV2 has not been directly addressed. In this study, we analyzed for and observed passive transfer of neutralization activity with CCP transfusion. Furthermore, to specifically understand if antibodies against the spike protein (S) enhance infection, we measured the anti-S IgG, IgA, and IgM responses and adapted retroviral-pseudotypes to measure virus neutralization with target cells expressing the ACE2 virus receptor and the Fc alpha receptor (FcR) or Fc gamma receptor IIA (Fc{gamma}RIIA). Whereas neutralizing activity of CCP correlated best with higher titers of anti-S IgG antibodies, the neutralizing titer was not affected when Fc receptors were present on target cells. These observations support the absence of antibody-dependent enhancement of infection (ADE) by IgG and IgA isotypes found in CCP. The results presented, therefore, support the clinical use of currently available antibody-based treatment including the continued study of CCP transfusion strategies.

BackgroundCharacterizing the kinetics of the antibody response to SARS{square}CoV{square}2 is of critical importance to developing strategies that may mitigate the public health burden of COVID-19. We sought to determine how circulating antibody levels change over time following natural infection. Methods/MaterialsWe conducted a prospective, longitudinal analysis of COVID-19 convalescent plasma (CCP) donors at multiple time points over a 9-month period. At each study visit, subjects either donated plasma or only had study samples drawn. In all cases, anti-SARS-CoV-2 donor testing was performed using semi-quantitative chemiluminescent immunoassays (ChLIA) targeting subunit 1 (S1) of the SARS-CoV-2 spike (S) protein, and an in-house fluorescence reduction neutralization assay (FRNA). ResultsFrom April to November 2020 we enrolled 202 donors, mean age 47.3 {+/-}14.7 years, 55% female, 75% Caucasian. Most donors reported a mild clinical course (91%, n=171) without hospitalization. One hundred and five (105) (52%) donors presented for repeat visits with a median 42 (12-163) days between visits. The final visit occurred at a median 160 (53-273) days post-symptom resolution. Total anti-SARS-CoV-2 antibodies (Ab), SARS-CoV-2 specific IgG and neutralizing antibodies were detected in 97.5%, 91.1%, and 74% of donors respectively at initial presentation. Neutralizing Ab titers based on FRNA50 were positively associated with mean IgG levels (p = <0.0001). Mean IgG levels and neutralizing titers were positively associated with COVID-19 severity, increased donor age and BMI (p=0.0006 and p=0.0028, p=0.0083 and p=0.0363, (p=0.0008 and p=0.0018, respectively). Over the course of repeat visits, IgG decreased in 74.1% of donors; FRNA50 decreased in 44.4% and remained unchanged in 33.3% of repeat donors. A weak negative correlation was observed between total Ab levels and number of days post-symptom recovery (r = 0.09). ConclusionAnti-SARS-CoV-2 antibodies were identified in 97% of convalescent donors at initial presentation. In a cohort that largely did not require hospitalization. IgG and neutralizing antibodies were positively correlated with age, BMI and clinical severity, and persisted for up to 9 months post-recovery from natural infection. On repeat presentation, IgG anti-SARS-CoV-2 levels decreased in 56% of repeat donors. Overall, these data suggest that CP donors possess a wide range of IgG and neutralizing antibody levels that are proportionally distributed across demographics, with the exception of age, BMI and clinical severity.

Purposevon Willebrand factors (vWFs), hemostatic factors, are produced as large multimers and are shear stress-dependently cleaved to become the appropriate size. A reduction in vWF large multimers develops in various conditions including the use of extracorporeal life support, which can cause excessive-high shear stress in the blood flow and result in hemostatic disorders. The objective of this prospective study was to investigate the impact of venovenous extracorporeal membrane oxygenation (VV ECMO) use on the status of vWF large multimers and hemostatic disorders during single lung transplantation (SLT). MethodsWe prospectively enrolled 12 patients who underwent SLT at our center. Among them, seven patients were supported by VV ECMO intraoperatively (ECMO group) and the remaining five patients underwent SLT without ECMO support (control group). The vWF large multimer index (%) was defined as the ratio of the large multimer proportion in total vWF (vWF large multimer ratio) derived from a patients plasma to that from the standard human plasma. ResultsThe vWF large multimer index at the end of the surgery was significantly lower in the ECMO group than in the control group (112.6% vs. 75.8%, respectively; p < 0.05). The intraoperative blood loss and the amounts of intraoperative transfusion products in the ECMO group tended to be greater than those in the control group; however, the differences were not significant. ConclusionDuring SLT, the intraoperative use of VV ECMO caused a decrease in the vWF large multimer index. However, the vWF large multimer index was maintained at > 75% in average at the end of SLT, which did not affect the bleeding complications.

BackgroundTranslational research driven by large-scale biological testing requires significant volumes of blood for research testing. However, blood is also required for clinical management of research subjects, which must take priority. Paradoxically, much of the blood drawn for clinical management goes unused. Here, we present our approach for retrieving unused blood samples collected for clinical management and recycling them for research purposes. MethodsClinical Blood samples were collected for 60 days during a 61-day xenotransplantation experiment in a brain-dead decedent. Twice weekly, research staff went to the chemistry and hematology laboratories and collected stored blood, serum, and plasma samples that were >12 hours old. Sample collection and storage before retrieval was per standard clinical protocols. Samples were de-identified and relabeled and brought to a central biorepository for processing and storage. The quantity of plasma, serum, red blood cells (RBCs), and peripheral blood mononuclear cells (PBMCs) collected from clinical labs and bespoke research blood draws were compared. ResultsUnused blood from clinical samples yielded a minimum of 6.0 ml per day of plasma, representing 62% of all plasma obtained. Serum was only recoverable on 13 days (22%), with a mean 2.3 ml collected on those days, representing 8% of all serum obtained. PBMCs were only recoverable on six days (10%). ConclusionsOverdrawn clinical laboratory samples represent an untapped resource of blood samples for research and can help augment samples collected explicitly for research purposes. With careful planning, this represents an opportunity to minimize iatrogenic blood loss in clinical-translational research.

Although red blood cell (RBC) transfusions save lives, some patients develop clinically-significant alloantibodies against donor blood group antigens, which then have adverse effects in multiple clinical settings. Few effective measures exist to prevent RBC alloimmunization and/or eliminate alloantibodies in sensitized patients. Donor-related factors may influence alloimmunization; thus, there is an unmet clinical need to identify which RBC units are immunogenic. Repeat volunteer blood donors and donors on iron supplements have elevated reticulocyte counts compared to healthy non-donors. Early reticulocytes retain mitochondria and other components, which may act as danger signals in immune responses. Herein, we tested whether reticulocytes in donor RBC units could enhance RBC alloimmunization. Using a murine model, we demonstrate that transfusing donor RBC units with increased reticulocyte frequencies dose-dependently increase RBC alloimmunization rates and alloantibody levels. Transfusing reticulocyte-rich RBC units was associated with increased RBC clearance from the circulation and a robust proinflammatory cytokine response. As compared to previously reported post-transfusion RBC consumption patterns, erythrophagocytosis from reticulocyte-rich units was increasingly performed by splenic B cells. These data suggest that reticulocytes in a donated RBC unit impact the quality of blood transfused, are targeted to a distinct compartment, and may be an underappreciated risk factor for RBC alloimmunization.

BackgroundIn the absence of structured donor registries, social media platforms have become a dominant mechanism for blood donor recruitment in many low-resource settings. However, the implications of this shift for transfusion timeliness and system reliability remain unclear. ObjectiveTo evaluate the impact of social media-sourced donors on transfusion delay, donor reliability, and hemovigilance-related outcomes compared with conventional donor pathways. MethodsThis prospective analytical study included 400 transfusion episodes across tertiary hospitals in Bangladesh. Donor sources were categorized as social media (SM) or conventional (CON). The primary outcome was delay-to-transfusion. Secondary outcomes included donor-related irregularities, documentation completeness, near-miss events, and acute transfusion reactions. Multivariable logistic regression identified predictors of delay [&ge;]4 hours. ResultsSocial media-sourced donors were associated with significantly longer transfusion delays (5.98 vs 2.97 hours; p<0.001). Delay [&ge;]4 hours occurred in 83.6% of SM cases versus 17.6% of CON cases (OR 23.78). Donor-related irregularities were observed in 85% of SM episodes and absent in CON donors. Safety outcomes did not differ significantly between groups. Social media donor sourcing remained the strongest independent predictor of delay (adjusted OR 18.09). ConclusionUnregulated social media-based donor recruitment introduces substantial delays and undermines system reliability without improving access. Integration of digital tools into regulated donor systems is essential to strengthen transfusion timeliness and hemovigilance in resource-limited settings.

BackgroundEnvenoming from numerous sources, such as snakes, scorpions, and spiders, is a major health issue across the world, resulting in millions of cases and tens of thousands of deaths annually. Venom induced symptoms ranges from systemic reactions like nausea and vomiting to localised pain and swelling. One major risk is the development of venom induced consumption coagulopathy (VICC), which might result in significant consequences. Plasmapheresis is being investigated as a possible therapy for severe envenoming. ObjectivesWe aim to assess the effectiveness and potential advantages of plasmapheresis in snakebite cases, focusing on clinical results. We seek to find if plasmapheresis improves neurological, renal, and hematological dysfunction and impacts secondary outcomes, including patient discharge rates, morbidity, mortality, duration of hospital stay, and the number of plasmapheresis sessions required. MethodsFollowing PRISMA guidelines, we conducted a systematic search of articles published between 1980 and July 2023 across multiple databases. MeSH terms related to snakebite and plasmapheresis were applied without publication or language type restrictions. Inclusion criteria considered case reports, cross-sectional studies, or case series featuring plasmapheresis in snakebite management. Inclusions were participants aged 18 years or older with confirmed or suspected snakebites, meeting plasmapheresis indications. Exclusions included participants under 18 years, studies reporting only in vitro data, review articles, and redundant reporting. The emphasis was on Emergency Departments or Intensive Care Units. ResultsIn a review of 147 cases (1980 to July 2023), the most common snake was the hump-nosed viper (Hypnale hypnale). Renal, neurological, and hematological dysfunctions improved after plasmapheresis. The mean plasmapheresis sessions were 2.1, and the average hospital stay was 13.13 days. ConclusionOnce the data has been analyzed, the result emphasizes the clinical importance of plasmapheresis in snakebite envenoming. It helps decision-making when standard therapies are insufficient or ineffective, potentially saving lives. Author SummarySnakebites pose a significant global health threat, causing numerous deaths and serious injuries annually. While antivenom is the primary treatment, its not always effective or available. This study explores an alternative treatment called plasmapheresis, a method that filters harmful substances from the blood. We reviewed 147 cases of snake envenoming treated with plasmapheresis between 1980 and 2023. Our findings show that plasmapheresis can improve various complications caused by snake venom, including kidney problems, nerve damage, and blood disorders. On average, patients received about two plasmapheresis treatments and stayed in the hospital for around 13 days. The study suggests that plasmapheresis could be a valuable option when standard treatments arent working well enough. It might help save lives in severe cases of snake envenoming. While more research is needed, this review provides important insights for doctors treating snakebite victims, especially in areas where snakebites are common and resources are limited.

BackgroundTransfusion-transmitted malaria risk in the USA is estimated at less than one per ten million blood donations or about 1 case every 2 years. A total of 18 autochthonous malaria infections have been reported in the USA since 2003: eight cases in 2003 and ten cases in 2023 with fifteen reported from Florida. From May to July 2023 in Sarasota County, Florida, symptomatic P. vivax malaria was detected in seven individuals with no recent malaria travel history. Study Design and MethodsIn the absence of an FDA-approved blood donor screening test for malaria at the time, the local blood center responded to this 2023 cluster by implementing a non-testing risk mitigation strategy of escalating blood donor screening measures and pathogen reduction. Potential blood supply safety enhancement by use of additional nucleic acid testing (NAT) on samples from donors in impacted areas was studied retrospectively using the Procleix Plasmodium Assay, a transcription-mediated amplification detecting five human Plasmodium species 18S ribosomal RNA with a limit of detection ranging from 2-7 infected erythrocytes per mL. ResultsAmong the 435 valid study samples from accepted healthy blood donors, screened with heightened non-testing measures in the 4-6 mile radius of malaria-impacted area, all samples were nucleic acid test non-reactive. DiscussionThe blood center strategy in response to the local 2023 malaria outbreak maintained the blood supply safety with no transfusion-transmitted malaria and no retrospective evidence of Plasmodium in the 435 valid samples collected from blood donors residing in impacted areas.

IntroductionAntivenom neutralizes the effects of snake venom toxins. Its timely administration can by critical in reducing the risk of complications, including serious clinical hemorrhage. The objective of this study was to evaluate the relationship between the late administration of antivenom and the persistence of venom-induced consumption coagulopathy (VICC) in patients with Bothrops snakebite envenomation. MethodsWe conducted a retrospective observational study in three hospitals referent in the management of snakebite envenomation in a region of Colombia. Patients were identified through ICD10 codes. The information was extracted from patients clinical records. Late and early antivenom administration patients were compared; "late" was defined as a period between snakebite and antivenom administration >4h. Definition of persistence of VICC was as an International Normalized Ratio >1.5 between 25-72h after snakebite. The clinical bleeding (as reported by clinicians) was also explored. Groups were compared through X2 of association; binary and multiple logistic regressions were ran to estimate the magnitude of the crude and adjusted relative risk (RR) for the analyzed outcomes. Results304 patients with snakebite from Bothrops spp.were analyzed; 138 (45.3%) with late antivenom administration, and 166 (54.7%) early. The persistence of VICC was more frequent in the late antivenom group (2.06 [95% IC 1.11-3.83]), as well as for clinical bleeding and major bleeding (1.97[1.30-2.98]; 3.00 [1.19-7.54], respectively). DiscussionLate antivenom administration was associated with increased risk in the persistence of VICC and major bleeding in patients with envenomation by a Bothrops complex snakebite. The results reported in the literature are diverse; inter and intra species variability and definitions can influence the finding. ConclusionsThe Timely administration of antivenom was associated with an increased risk in persistence of VICC. SynopsisSnakebites can cause different levels of damage to humans, depending on the type of venom. The venom produced by pit vipers, specifically those of the genus Bothrops, can change blood clotting and cause bleeding that can be life-threatening. Most snakebites from these vipers occur in tropical areas where the most affected are farmers and rural people. There is often a lack of access to antivenom treatment, depending on access to and provision of health services. It seems that giving the antivenom at the right time can reduce the risk of complications. However, it is still unclear whether waiting to give the antivenom increases the risk of continued envenoming and bleeding. This study shows that waiting to treat a patient can lead to a higher chance of bleeding. This information can help doctors, decision makers, and lawmakers improve treatment options and access in remote areas.

BACKGROUNDAlloimmunization to transfused red blood cells (RBCs) remains a significant clinical problem. However, the cells that initiate immune responses to transfused RBCs remain incompletely characterized. Recently published work has identified splenic marginal zone B (MZB) cells as being critically required for the production of anti-RBC alloantibodies in response to RBCs. In infectious models, MZB cell activation has been shown to depend on a unique population of marginal zone macrophages (MZMs). We hypothesized that MZMs would capture stored RBCs and present them to MZBs, and ultimately MZMs would be required for generation of anti-RBC alloantibodies in response to stored RBC transfusion. STUDY DESIGN AND METHODSStored GFP+ murine RBCs were utilized to determine the splenic localization and erythrophagocytosis by splenic macrophage populations. To determine the functional impact of MZMs, we compared LXR-KO mice, which have been reported to lack MZMs, with wild type mice. Both innate and adaptive immune responses to stored HOD allogenic RBC transfusion were measured in LXR-KO and wild type mice. RESULTSRBC storage leads to a significant increase in the phagocytosis of transfused RBCs by splenic MZMs. LXR-KO mice demonstrated a lack of MZMs and had significantly decreased rapid phase production of cytokines MCP-1 and KC, but similar levels of IL-6. Surprisingly, anti-RBC alloantibody levels were unaffected by the absence of splenic MZMs. CONCLUSIONSSplenic MZMs are involved in the innate response to transfused stored HOD RBCs, contributing to both MCP-1 and KC cytokine production. However, MZMs are dispensable for anti-RBC alloantibody production.

Background and AimsAcetaminophen (APAP) overdose is the leading individual cause of acute liver failure (ALF) in the United States, with many patients rapidly progressing to hyperacute liver failure. While hepatocytes are the main target of APAP toxicity, endothelial cells (ECs) are also affected. However, the efficacy of an endothelial-specific biomarker to predict patient outcomes remains unknown. This study aimed to evaluate angiopoietin-2 (ANGPT2) as a prognostic biomarker for poor outcomes in APAP-induced ALF. Approach and ResultsUsing human and mouse single-cell RNA sequencing (scRNAseq) data, we found that ANGPT2 expression was significantly elevated in ECs following APAP exposure. We measured circulating ANGPT2 levels from two independent APAP-ALF cohorts: a Phoenix cohort (n=43) and a cohort from the ALF Study Group (n=80). In the Phoenix cohort, ANGPT2 levels were significantly higher in non-survivors with an AUROC of 0.938. In the ALFSG cohort, we stratified patients based on time of symptom onset finding that ANGPT2 had improved prognostic value in early-presenting patients, with day 1 and day 3 AUC values of 0.825 and 0.918, respectively. Lastly, we combined the patient cohorts (n=110) finding that ANGPT2 alone or in combination with MELD outperformed MELD alone based on AUC (ANGPT2: 0.87, MELD 0.83, ANGPT2+MELD 0.90). ConclusionsANGPT2 is a promising prognostic biomarker for APAP-induced ALF, reflecting endothelial stress and offering superior predictive value compared to MELD alone, especially in early-presenting patients. Its capacity for predicting poor outcomes underscores its value in improving patient prognosis and therapeutic intervention strategies in APAP overdose cases. Lay SummaryAccidental or intentional overdosing on acetaminophen can cause liver injury and in severe cases acute liver failure. Under these circumstances, receiving a liver transplant may be the only remaining therapeutic option. However, a liver transplant is a major surgery and commits the patient to a lifetime of anti-rejection medication. Because there is only a limited time window to decide who will recover and who needs a transplant to survive, prognostic biomarkers are essential to identify transplant candidates as early as possible after the overdose. In this study we discovered that plasma levels of the endothelial growth factor angiopoietin-2 can accurately predict at the peak of injury who will need a liver transplant to survive. In addition, this biomarker can be rapidly measured, which allows the data to be available for clinical decision making. HighlightsO_LIAcetaminophen-induced liver injury can cause hyper-acute liver failure within 3 to 7 days with a high probability of negative outcome. C_LIO_LIUnder these conditions, a liver transplant may be the only therapeutic option. C_LIO_LIIn two independent cohorts, angiopoietin 2 was identified as an early prognostic biomarker for poor outcome. C_LIO_LIAngiopoietin can more accurately inform clinical management during the initial stages of hospital presentation than the MELD score. C_LI

The clinical and scientific communities rely on serology testing to analyze the degree of antibody-mediated immunity afforded to recovered patients from SARS-CoV-2 infection. Neutralizing antibodies present in COVID-19 convalescent plasma (CCP) remains a practical therapy to treat COVID-19 patients requiring hospitalization. However, it remains unclear how long antibody levels persist in CCP donors after recovery. An accurate estimation of antibody kinetics in CCP donors provide an important observation to further define the extent of long-term immunity in recovered patient and simultaneously inform CCP collection processes in efforts to improve CCP dosing and therapeutic outcome. In this study, we analyzed 63 donors and measured antibody levels using two high throughput screening assays (HTSA) designed to detect antibodies targeting the spike protein (S1) and nucleocapsid protein (NP) of SARS-CoV-2 and monitored antibody levels between 2-8 consecutive donations. We show that anti-S1 antibody levels, as measured using the Ortho Total Ig HTSA, increased over time in repeat CCP donors while anti-NP antibody levels, as measured using the Abbott IgG HTSA, were unchanged or decreased over time. When we normalized these data, we found that both the absolute levels of anti-S1 antibodies and the ratio between S1 and NP antibodies tends to increase over time. These data have important implications for the convalescent donation process, patient protection from future infection and characterization of the SARS-CoV-2 immune response.

BackgroundPlatelet microparticles (PMP) are elevated in and associated with disease activity in a number of diseases including cancer, neurological conditions, autoimmune diseases, and infectious diseases. This finding raises the possibility of a removal of these microparticles as a therapeutic strategy. The Hemopurifier is an experimental device consisting of a plasma separator and an affinity resin containing Galanthus nivalis agglutinin (GNA) affinity resin that has previously been shown to remove extracellular vesicles in vitro and in vivo. In this proof on concept study, we sought to determine ex vivo removal of platelet-derived microparticles from healthy human plasma by the Hemopurifier. MethodsTwo hundred milliliters of thawed healthy human plasma were circulated over the Hemopurifier device at a rate of 100 mL/minute. Plasma samples were taken at time points equivalent to a 4-, 6- and 8-hour clinical Hemopurifier session in a healthy adult. Microparticles were isolated from these timepoints and analyzed for treatment concentration changes. Platelet microparticle counts were determined by binding to antiCD41. ResultsThe Hemopurifier removed 98.5% of platelet microparticles during the equivalent of a 4-hour clinical session. ConclusionsWe demonstrated that an extracorporeal device with a GNA affinity resin removes platelet microparticles from normal healthy plasma. Next steps would be demonstration of the removal of PMPs from plasma by the Hemopurifier in different disease states and characterization of the cargo within removed PMPs.

Background and ObjectivesLiquid plasma (LQP) stands out as an alternative to thawed plasma (TP) for emergent transfusions due to longer shelf life. We aim to measure fibrinogen, Protein C, Protein S, FV, FVII, and FVIII activity in LQP, quantify how these factors levels change during storage, and characterize how they compare in LQP to TP. Materials and MethodsCoagulation factor activities were measured on Days 15, 26, and 27 for LQP (n=26) and Day 5 for TP (n=31). Bayesian statistics was used to compare coagulation factor activity and quantify changes in activity during storage. ResultsFibrinogen and Protein C activity in Day 26 LQP (LQP26) was comparable to Day 5 TP (TP5) with posterior mean activity of 257 mg/dL vs 246 mg/dL and 100.4% vs 108.7%, respectively. FV, FVII, and FVIII had lower activity in LQP26 vs TP5 with posterior mean activities of 42.6% vs 72.0%, 55.0% vs 59.7%, and 48.8% vs 59.2%, respectively. Protein S in LQP26 was low with posterior mean activity of 28.0%, which was less than half that of TP5 at 66.4%. From Day 15 to Day 26, FVII in LQP decreased at a rate of -3.49% per day whereas fibrinogen, Protein C, Protein S, FV, and FVIII activity in LQP remained relatively stable. ConclusionCompared to TP5, LQP26 has comparable activities of fibrinogen, Protein C, FVII, lower activities of FV and Protein S, and slightly lower activity of Factor VIII. LQP is a viable alternative for use in emergency transfusions and massive transfusion protocols. Highlights- Liquid plasma has comparable activities to thawed plasma for fibrinogen, Protein C, and Factor VII, which is advantageous for emergency use due to its extended shelf life. - Liquid plasma has adequate fibrinogen and Protein C levels on the last day of expiration. - Liquid plasma has [~]50% FV, FVII and FVIII activity levels on the last day of expiration, making it a sufficient replacement option to TP for active bleeding.

ObjectiveThis study aimed to evaluate the impact of combined use of chemiluminescent immunoassay (CLIA) and nucleic acid amplification testing (NAAT) to improve transfusion transmitted Hepatitis B Virus (HBV), Hepatitis C virus (HCV) and Human Immunodeficiency Virus-1 (HIV-1) among blood donors in Pakistan. DesignRetrospective, single center observational. SettingRegional blood center at tertiary care hospital in urban Pakistan. ParticipantsAll adults of 18 years or above who were eligible to donate blood after meeting the pre-donation screening criteria during the study period (n = 26,778). Outcome MeasuresPrimary outcome measure was to estimate the incremental yield of HBV, HCV, and HIV-1 infections using combined CLIA+NAAT compared to CLIA-alone per 100,000 donations. Secondary outcome was to calculate inter-test agreement and discordance between CLIA and NAAT methods. ResultsAmong 26,778 donors, the combined CLIA+NAAT testing detected a total of 2,423.6 viral infections per 100,000 donations, NAAT alone contributed 739.7/100,000 of these. For HBV, NAAT uniquely detected 489/100,000 additional cases missed by CLIA; the combined detection rate was 1,561/100,000. For HCV, NAAT-only yield was lower (247/100,000), with total detection of 825.3/100,000. HIV-1 was rare in the donor pool; incremental NAAT yield was 3.7/100,000, with a combined detection rate of 37.3/100,000. Agreement between tests was substantial for HBV ({kappa} = 0.63) and moderate to fair for HCV ({kappa} = 0.47) and HIV-1 ({kappa} = 0.40). The discordant cases detected by NAAT alone for HBV, HCV and HIV-1 were 183, 431 and 37 respectively. McNemars test showed statistically significant differences (p < 0.001) across all markers, with large effect sizes for HIV-1 (0.92, 95% CI: 0.80-1.00) and HCV (0.69, 95% CI: 0.65-0.73). ConclusionIntegrating CLIA with NAAT enhanced the detection of HBV, including occult and window period infections, and refined estimates of active HCV and HIV-1 infections which significantly improved blood transfusion safety. STRENGTHS AND LIMITATIONS OF THIS STUDYO_LIThe selection bias was reduced by including all consecutive eligible blood donors during study period. C_LIO_LIThe information bias was minimized through uniform, fully automated, and validated protocols for combined CLIA and NAAT testing of blood donors. C_LIO_LIThe single center retrospective study design restricts generalizability and causal inference. C_LIO_LIThe false positives/negatives and genetic variability were not confirmed. C_LI