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Background: Autologous hematopoietic stem cell transplantation (HSCT) is a potent treatment option for patients with aggressive relapsing-remitting multiple sclerosis (RRMS)., Objective: To evaluate long-term outcomes of HSCT in MS., Methods: National retrospective single-center observational study of patients with aggressive RRMS that underwent HSCT in Norway from January 2015 to January 2018. Criteria for receiving HSCT included at least two clinical relapses the last year while on disease modifying treatment (DMT)., Results: In total, 29 patients, with a mean follow-up time of 70 months (standard deviation:14.3), were evaluated. Twenty patients (69%) had sustained no evidence of disease activity (NEDA-3) status, 24 (83%) were relapse-free, 23 (79%) free of magnetic resonance imaging (MRI) activity, and 26 (90%) free of progression. Number of patients working full-time increased from 1 (3%), before HSCT, to 10 (33%) after 2 years and 15 (52%) after 5 years., Conclusion: HSCT offers long-term disease-free survival with successively increasing work participation in patients with aggressive MS resistant to DMTs., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared the following potential conflicts of interest with respect to the research, authorship, and/or publication of this article: C.E.K. has received an unrestricted grant from Novartis in 2019. A.K.L. has no disclosures. S.A.S.K. has received unrestricted grants from Biogen and Novartis. T.H. has received speakers honoraria and participated in clinical trials organized by Merck, Serono, Biogen, Roche, Novartis, and Alexion. L.H.T. has no disclosures. Å.R.L. has received minor grant from Sanofi. E.K.K. has no disclosures. L.B. has received unrestricted research grants to his institution and/or scientific advisory board or speakers honoraria from Biogen, Genzyme, Merck, Novartis, Roche, and Teva, and has participated in clinical trials organized by Biogen, Merck, Novartis, and Roche. Ø.T. has received speaker honoraria from and served on scientific advisory boards for Biogen, Sanofi-Aventis, Merck, and Novartis.

Objectives: Secretomes of mesenchymal stromal cells (MSC) represent a novel strategy for growth-factor delivery for tissue regeneration. The objective of this study was to compare the efficacy of adjunctive use of conditioned media of bone-marrow MSC (MSC-CM) with collagen barrier membranes vs. adjunctive use of conditioned media of leukocyte- and platelet-rich fibrin (PRF-CM), a current growth-factor therapy, for guided bone regeneration (GBR)., Methods: MSC-CM and PRF-CM prepared from healthy human donors were subjected to proteomic analysis using mass spectrometry and multiplex immunoassay. Collagen membranes functionalized with MSC-CM or PRF-CM were applied on critical-size rat calvaria defects and new bone formation was assessed via three-dimensional (3D) micro-CT analysis of total defect volume (2 and 4 weeks) and 2D histomorphometric analysis of central defect regions (4 weeks)., Results: While both MSC-CM and PRF-CM revealed several bone-related proteins, differentially expressed proteins, especially extracellular matrix components, were increased in MSC-CM. In rat calvaria defects, micro-CT revealed greater total bone coverage in the MSC-CM group after 2 and 4 weeks. Histologically, both groups showed a combination of regular new bone and 'hybrid' new bone, which was formed within the membrane compartment and characterized by incorporation of mineralized collagen fibers. Histomorphometry in central defect sections revealed greater hybrid bone area in the MSC-CM group, while the total new bone area was similar between groups., Conclusion: Based on the in vitro and in vivo investigations herein, functionalization of membranes with MSC-CM represents a promising strategy to enhance GBR., (© 2023 The Authors. Clinical Oral Implants Research published by John Wiley & Sons Ltd.)

Secretomes of mesenchymal stromal cells (MSCs) are emerging as a novel growth factor (GF)-based strategy for periodontal and bone regeneration. The objective of this study was to compare the secretome of human bone marrow MSC (BMSC) to that of leukocyte- and platelet-rich fibrin (L-PRF), an established GF-based therapy, in the context of wound healing and regeneration. Conditioned media from human BMSCs (BMSC-CM) and L-PRF (LPRF-CM) were subjected to quantitative proteomic analysis using liquid chromatography with tandem mass spectrometry. Global profiles, gene ontology (GO) categories, differentially expressed proteins (DEPs), and gene set enrichment (GSEA) were identified using bioinformatic methods. Concentrations of selected proteins were determined using a multiplex immunoassay. Among the proteins identified in BMSC-CM (2157 proteins) and LPRF-CM (1420 proteins), 1283 proteins were common. GO analysis revealed similarities between the groups in terms of biological processes (cellular organization, protein metabolism) and molecular functions (cellular/protein-binding). Notably, more DEPs were identified in BMSC-CM ( n = 550) compared to LPRF-CM ( n = 118); these included several key GF, cytokines, and extracellular matrix (ECM) proteins involved in wound healing. GSEA revealed enrichment of ECM (especially bone ECM)-related processes in BMSC-CM and immune-related processes in LPRF-CM. Similar trends for intergroup differences in protein detection were observed in the multiplex analysis. Thus, the secretome of BMSC is enriched for proteins/processes relevant for periodontal and bone regeneration. The in vivo efficacy of this therapy should be evaluated in future studies.

Background: In Norway, treatment with COVID-19 convalescent plasma has been given through the NORPLASMA project. The treatment was initially offered to critically ill patients after an individual assessment, but from December 2020, the indication was limited to critically ill, immunocompromised patients. In this article we describe clinical characteristics, comorbidity and mortality in patients who received convalescent plasma in these two periods., Material and Method: From 22 April 2020 to 30 March 2022, a total of 79 patients were included in the observational studies NORPLASMA MONITOR and the Norwegian SARS-CoV-2 study. The patients had received a total of 193 units of convalescent plasma at 15 Norwegian hospitals/nursing homes; 62 in South-Eastern Norway Regional Health Authority, 8 in Western Norway Regional Health Authority and 9 in Central Norway Regional Health Authority. Information on immune status, comorbidity and course of infection was retrieved from the patient records after informed written consent was obtained., Results: Of 79 patients with a median age of 65 years (interquartile range 51-⁠73) who were treated with convalescent plasma, 31 (39 %) died during hospitalisation. A total of 59 patients were immunocompromised, and of these, 20 died in hospital compared to 11 of 20 who were assumed to be immunocompetent. Median number of comorbidities was 2 (interquartile range 1-4). The patients received a median of two plasma units (min.-max. 1-21). Two of the patients developed mild allergic skin reactions., Interpretation: Convalescent plasma was well tolerated by patients with COVID-19. Immunocompromised patients may have benefitted from the treatment, with lower mortality than for those assumed to be immunocompetent.

Background: At the start of the pandemic, the Norwegian Directorate of Health and Norwegian blood banks initiated the production of COVID-19 convalescent plasma within the framework of clinical studies. In this article we describe the blood donors who participated., Material and Method: Blood donors who had recovered from COVID-19 were recruited to donate single donor plasma for the purpose of patient treatment. Data on the course of infection, leukocyte antibodies and antibody level against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) per plasma unit were registered after informed consent was obtained. We calculated a disease score defined as the total number of self-reported symptoms/findings and hospitalisation where relevant (score 0-⁠11)., Results: A total of 1644 plasma units were collected from 266 plasma donors at 12 blood banks. Median disease score was 5 (interquartile range 3-⁠6), and 15 donors had recovered from pneumonia and/or been hospitalised. A total of 599/1644 plasma units from 106/266 donors met our requirement for SARS-CoV-2 antibody content (> 60 % inhibition of virus binding to angiotensin-converting enzyme 2 (ACE2)) or positive virus neutralisation test. The antibody level in donors waned over time following infection, and showed no clear correlation with disease score., Interpretation: The number of symptoms and findings in blood donors could not predict antibody response at individual level, and antibody testing was crucial for the production of effective convalescent plasma.

Metastatic castration-resistant prostate cancer (mCRPC) is an immunologically cold disease with dismal outcomes. Cryoablation destroys cancer tissue, releases tumor-associated antigens and creates a pro-inflammatory microenvironment, while dendritic cells (DCs) activate immune responses through processing of antigens. Immunotherapy combinations could enhance the anti-tumor efficacy. This open-label, single-arm, single-center phase I trial determined the safety and tolerability of combining cryoablation and autologous immature DC, without and with checkpoint inhibitors. Immune responses and clinical outcomes were evaluated. Patients with mCRPC, confirmed metastases and intact prostate gland were included. The first participants underwent prostate cryoablation with intratumoral injection of autologous DCs in a 3 + 3 design. In the second part, patients received cryoablation, the highest acceptable DC dose, and checkpoint inhibition with either ipilimumab or pembrolizumab. Sequentially collected information on adverse events, quality of life, blood values and images were analyzed by standard descriptive statistics. Neither dose-limiting toxicities nor adverse events > grade 3 were observed in the 18 participants. Results indicate antitumor activity through altered T cell receptor repertoires, and 33% durable (> 46 weeks) clinical benefit with median 40.7 months overall survival. Post-treatment pain and fatigue were associated with circulating tumor cell (CTC) presence at inclusion, while CTC responses correlated with clinical outcomes. This trial demonstrates that cryoimmunotherapy in mCRPC is safe and well tolerated, also for the highest DC dose (2.0 × 10 8 ) combined with checkpoint inhibitors. Further studies focusing on the biologic indications of antitumor activity and immune system activation could be considered through a phase II trial focusing on treatment responses and immunologic biomarkers., (© 2023. The Author(s).)

Background and Objectives: Based on previous success using apheresis platelets, we wanted to investigate the in vitro quality and platelet function in continuously cold-stored and delayed cold-stored platelet concentrates (PCs) from interim platelet units (IPUs) produced by the Reveos system., Materials and Methods: We used a pool-and-split design to prepare 18 identical pairs of PCs. One unit was stored unagitated and refrigerated after production on day 1 (cold-stored). The other unit was stored agitated at room temperature until day 5 and then refrigerated (delayed cold-stored). Samples were taken after pool-and-split on day 1 and on days 5, 7, 14 and 21. Swirling was observed and haematology parameters, metabolism, blood gas, platelet activation and platelet aggregation were analysed for each sample point., Results: All PCs complied with European recommendations (EDQM 20th edition). Both groups had mean platelet content >200 × 10 9 /unit on day 21. The pH remained above 6.4 for all sample points. Glucose concentration was detectable in every cold-stored unit on day 21 and in every delayed cold-stored unit on day 14. The cold-stored group showed a higher activation level before stimulation as measured by flow cytometry. The activation levels were similar in the two groups after stimulation. Both groups had the ability to form aggregates after cold storage and until day 21., Conclusion: Our findings suggest that PCs from IPUs are suitable for cold storage from day 1 until day 21 and delayed cold storage from day 5 until day 14., (© 2023 International Society of Blood Transfusion.)

Introduction: In this report, we describe a training program in emergency whole blood collection and transfusion for medical students at the University of Bergen. The overall aim of the program is to improve the availability of early balanced blood transfusion for the treatment of patients with life-threatening bleeding in rural health care services., Study Design and Methods: The voluntary training program provides the knowledge needed to practice emergency whole blood transfusions and understand the system for emergency whole blood collection in the framework of a civilian walking blood bank (WBB). It includes theoretical and practical sessions. In-person teaching and web-based learning resources are provided. An anonymous survey of the students attending the training course in the autumn of 2022 and spring 2023 was performed., Results: 128 of 178 students participated in the practical training. 88 of 128 (69%) responded to the survey. 82 (93%) performed blood typing, 71 (81%) performed donor interviews, 61 (69%) partially performed whole blood collection (up to blood in bag) and 27 (30%) participated in complete whole blood collection and performed autologous reinfusion. No complications occurred during training. The students reported that the training course increased their understanding of how to ensure access to emergency blood transfusion by the use of a WBB., Discussion: Structured theoretical and practical training in emergency whole blood collection and emergency transfusion is feasible and of interest to medical students. A multidisciplinary approach to student training in emergency whole blood collection and transfusion should be considered., (© 2023 AABB.)

Functionalizing biomaterials with conditioned media (CM) from mesenchymal stromal cells (MSC) is a promising strategy for enhancing the outcomes of guided bone regeneration (GBR). This study aimed to evaluate the bone regenerative potential of collagen membranes (MEM) functionalized with CM from human bone marrow MSC (MEM-CM) in critical size rat calvarial defects. MEM-CM prepared via soaking (CM-SOAK) or soaking followed by lyophilization (CM-LYO) were applied to critical size rat calvarial defects. Control treatments included native MEM, MEM with rat MSC (CEL) and no treatment. New bone formation was analyzed via micro-CT (2 and 4 weeks) and histology (4 weeks). Greater radiographic new bone formation occurred at 2 weeks in the CM-LYO group vs. all other groups. After 4 weeks, only the CM-LYO group was superior to the untreated control group, whereas the CM-SOAK, CEL and native MEM groups were similar. Histologically, the regenerated tissues showed a combination of regular new bone and hybrid new bone, which formed within the membrane compartment and was characterized by the incorporation of mineralized MEM fibers. Areas of new bone formation and MEM mineralization were greatest in the CM-LYO group. Proteomic analysis of lyophilized CM revealed the enrichment of several proteins and biological processes related to bone formation. In summary, lyophilized MEM-CM enhanced new bone formation in rat calvarial defects, thus representing a novel 'off-the-shelf' strategy for GBR.

Background: Early balanced transfusion is associated with improved outcome in haemorrhagic shock patients. This study describes the implementation and evaluates the safety of a whole blood transfusion program in a civilian helicopter emergency medical service (HEMS)., Methods: This prospective observational study was performed over a 5-year period at HEMS-Bergen, Norway. Patients in haemorrhagic shock receiving out of hospital transfusion of low-titre Group O whole blood (LTOWB) or other blood components were included. Two LTOWB units were produced weekly and rotated to the HEMS for forward storage. The primary endpoints were the number of patients transfused, mechanisms of injury/illness, adverse events and survival rates. Informed consent covered patient pathway from time of emergency interventions to last endpoint and subsequent data handling/storage., Results: The HEMS responded to 5124 patients. Seventy-two (1.4%) patients received transfusions. Twenty patients (28%) were excluded due to lack of consent (16) or not meeting the inclusion criteria (4). Of the 52 (100%) patients, 48 (92%) received LTOWB, nine (17%) received packed red blood cells (PRBC), and nine (17%) received freeze-dried plasma. Of the forty-six (88%) patients admitted alive to hospital, 35 (76%) received additional blood transfusions during the first 24 h. Categories were blunt trauma 30 (58%), penetrating trauma 7 (13%), and nontrauma 15 (29%). The majority (79%) were male, with a median age of 49 (IQR 27-70) years. No transfusion reactions, serious complications or logistical challenges were reported. Overall, 36 (69%) patients survived 24 h, and 28 (54%) survived 30 days., Conclusions: Implementing a whole blood transfusion program in civilian HEMS is feasible and safe and the logistics around out of hospital whole blood transfusions are manageable. Trial registration The study is registered in the ClinicalTrials.gov registry (NCT02784951)., (© 2022. The Author(s).)

Culturing mesenchymal stromal cells (MSC) in human platelet lysate (HPL) supplemented media can enhance their osteogenic differentiation potential. The objective of this study was to test the hypothesis that conditioned media (CM) derived from HPL-cultured MSC also have pro-osteogenic effects. Pooled CM was prepared from HPL-cultured human bone marrow MSC (BMSC) of multiple donors and applied on BMSC of different donors (than those used for CM preparation), with or without additional supplementation [HPL, fetal bovine serum (FBS)] and osteogenic stimulation. At various time-points, cell proliferation, alkaline phosphatase (ALP) activity, osteogenic gene expression and in vitro mineralization were assessed. BMSC in standard unstimulated growth media served as controls. After 3-7 days, CM alone did not promote BMSC proliferation or ALP activity; supplementation of CM with HPL slightly improved these effects. After 2 and 7 days, CM alone, but not CM supplemented with HPL, promoted osteogenic gene expression. After 14 days, only CM supplemented with FBS and osteogenic stimulants supported in vitro BMSC mineralization; CM alone and CM supplemented with HPL did not support mineralization, regardless of osteogenic stimulation. In summary, CM from HPL-cultured BMSC promoted osteogenic gene expression but not in vitro mineralization in allogeneic BMSC even when supplemented with HPL and/or osteogenic stimulants. Future studies should investigate the role and relevance of supplementation and osteogenic induction in in vitro assays using CM from MSC., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Shanbhag, Al-Sharabi, Mohamed-Ahmed, Gruber, Kristoffersen and Mustafa.)

Background: Blood products are frequently exposed to room temperature or higher for longer periods than permitted by policy. We aimed to investigate if this resulted in a measurable effect on common quality parameters and viscoelastic hemostatic function of cold stored CPDA-1 whole blood., Study Design and Methods: 450 ml of whole blood from 16 O Rh(D) positive donors was collected in 63 ml of CPDA-1 and stored cold. Eights bags were exposed to five weekly 4-h long transient temperature changes to 28°C. Eight bags were stored continuously at 4°C as a control. Samples were collected at baseline on day 1, after the first cycle on day 1 and weekly before each subsequent cycle (day 7, 14, 21, 28 and 35). Hemolysis, hematological parameters, pH, glucose, lactate, potassium, thromboelastography, INR, APTT, fibrinogen, and factor VIII were measured., Results: CPDA-1 whole blood repeatedly exposed to 28°C did not show reduced quality compared to the control group on day 35. Two units in the test group had hemolysis of 1.1% and 1.2%, and two in the control group hemolysis of 0.8%. Remaining thromboelastography clot strength (MA) on day 35 was 51.7 mm (44.8, 58.6) in the test group and 46.1 (41.6, 50.6) in the control group (p = .023). Platelet count was better preserved in the test group (166.7 [137.8, 195.6] vs. 117.8 [90.3, 145.2], p = .018). One sample in the test group was positive for Cutibacterium acnes on day 35 + 6., Conclusion: Hemolysis findings warrant further investigation. Other indicators of quality were not negatively affected., (© 2022 The Authors. Transfusion published by Wiley Periodicals LLC on behalf of AABB.)

Background: To increase preparedness and mitigate the risk of platelet shortage without increasing the number of collections, we introduced a dual platelet inventory with cold-stored platelets (CSP) with 14-days shelf life for actively bleeding patients during the COVID-19 pandemic., Study Design and Methods: We collected apheresis platelet concentrates with blood type O or A. All patients receiving CSP units were included in a quality registry. Efficacy was evaluated by total blood usage and laboratory analysis of platelet count, hemoglobin, and TEG 6s global hemostasis assay. Feasibility was evaluated by monitoring inventory and a survey among laboratory staff., Results: From 17 March, 2020, to 31 December, 2021, we produced 276 CSP units and transfused 186 units to 92 patients. Main indication for transfusion was surgical bleeding (88%). No transfusion reactions were reported. 24-h post-transfusion patient survival was 96%. Total outdate in the study period was 33%. The majority (75%) of survey respondents answered that they had received sufficient information and training before CSP was implemented. Lack of information about bleeding status while issuing platelets, high workload, and separate storage location was described as main reasons for outdates., Discussion: CSP with 14-days shelf life is a feasible alternative for the treatment of patients with bleeding. Implementation of a dual platelet inventory requires thorough planning, including information and training of clinical and laboratory staff, continuous follow-up of practice and patients, and an easy-to-follow algorithm for use of CSP units. A dual platelet inventory may mitigate the risk of platelet shortage during a pandemic situation., (© 2022 The Authors. Transfusion published by Wiley Periodicals LLC on behalf of AABB.)

Background: Civilian and military guidelines recommend early balanced transfusion to patients with life-threatening bleeding. Low titer group O whole blood was introduced as the primary blood product for resuscitation of massive hemorrhage at Haukeland University Hospital, Bergen, Norway, in December 2017. In this report, we describe the whole blood program and present results from the first years of routine use., Study Design and Methods: Patients who received whole blood from December 2017 to April 2020 were included in our quality registry for massive transfusions. Post-transfusion blood samples were collected to analyze isohemagglutinin (anti-A/-B) and hemolysis markers. Administration of other blood products, transfusion reactions, and patient survival (days 1 and 30) were recorded. User experiences were surveyed for both clinical and laboratory staff., Results: Two hundred and five patients (64% male and 36% female) received 836 units in 226 transfusion episodes. Patients received a mean of 3.7 units (range 1-35) in each transfusion episode. The main indications for transfusion were trauma (26%), gastrointestinal (22%), cardiothoracic/vascular (18%), surgical (18%), obstetric (11%), and medical (5%) bleeding. There was no difference in survival between patients with blood type O when compared with non-group O. Haptoglobin level was lower in the transfusion episodes for non-O group patients, however no clinical hemolysis was reported. No patients had conclusive transfusion-associated adverse events. Both clinical and laboratory staff preferred whole blood to component therapy for massive transfusion., Discussion: The experience from Haukeland University Hospital indicates that whole blood is feasible, safe, and effective for in-hospital treatment of bleeding., (© 2021 The Authors. Transfusion published by Wiley Periodicals LLC. on behalf of AABB.)

Background: Hematopoietic stem cell treatment (HSCT) is a promising treatment option for multiple sclerosis (MS), but detailed safety and efficacy measures are still scarce., Objective: To evaluate the efficacy and safety of HSCT in MS., Methods: Retrospective single-center observational study of all MS patients that underwent HSCT in Norway during January 2015 to January 2018. The primary outcome was no evidence of disease activity (NEDA-3) status., Results: A total of 30 patients with a median follow-up time of 26 months (range: 11-48) were evaluated. In total, 25 (83%) achieved NEDA-3 status, and none received disease-modifying treatment after HSCT. For 13 (43%) of the patients, there were sustained improvement in Expanded Disability Status Scale (EDSS) score, and 10 (33%) were working full time after the treatment, compared to only 1 (3%) before treatment. There were no serious treatment-related complications and was no mortality. Five patients (17%) were diagnosed with an autoimmune thyroid disease after the procedure, and 10 (43%) of the women had amenorrhea lasting >12 months and symptoms of ovarian failure., Conclusion: HSCT in MS is an effective and relatively safe treatment option, with few serious complications and no mortality in Norway, so far. However, long-term adverse event with amenorrhea is a common problem.

Civilian and military guidelines recommend balanced transfusion to patients with life-threatening bleeding. Early start of transfusion has shown improved survival. Thus, a balanced blood inventory must be available in all levels of health care to ensure early stabilization and damage control resuscitation of patients with bleeding. Whole blood has been reintroduced as a blood product for massive bleeding situations because it affords plasma, red blood cells, and platelets in a balanced ratio in a logistically advantageous way. In this article, we describe how to establish a whole blood-based blood preparedness program in a small rural hospital with limited resources. We present an implementation tool kit, which includes discussions on whole blood program strategies and the process of developing detailed procedures on donor selection, collection, storage, and transfusion management of whole blood. The importance of training and audit of the routines is highlighted, and establishment of an emergency walking blood bank is discussed. We conclude that implementation of a whole blood program is achievable in small rural hospitals and recommend that rural health care facilities at all treatment levels enable early balanced transfusion for patients with life-threatening bleeding by establishing protocols for whole blood-based preparedness., (© 2020 The Authors. Transfusion published by Wiley Periodicals LLC. on behalf of AABB.)

Background: This pilot trial focused on feasibility and safety to provide preliminary data to evaluate the hemostatic potential of cold-stored platelets (2° to 6°C) compared with standard room temperature-stored platelets (20° to 24°C) in adult patients undergoing complex cardiothoracic surgery. This study aimed to assess feasibility and to provide information for future pivotal trials., Methods: A single center two-stage exploratory pilot study was performed on adult patients undergoing elective or semiurgent complex cardiothoracic surgery. In stage I, a two-armed randomized trial, platelets stored up to 7 days in the cold were compared with those stored at room temperature. In the subsequent single-arm stage II, cold storage time was extended to 8 to 14 days. The primary outcome was clinical effect measured by chest drain output. Secondary outcomes were platelet function measured by multiple electrode impedance aggregometry, total blood usage, immediate and long-term (28 days) adverse events, length of stay in intensive care, and mortality., Results: In stage I, the median chest drain output was 720 ml (quartiles 485 to 1,170, n = 25) in patients transfused with room temperature-stored platelets and 645 ml (quartiles 460 to 800, n = 25) in patients transfused with cold-stored platelets. No significant difference was observed. The difference in medians between the room temperature- and cold-stored up to 7 days arm was 75 ml (95% CI, -220, 425). In stage II, the median chest drain output was 690 ml (500 to 1,880, n = 15). The difference in medians between the room temperature arm and the nonconcurrent cold-stored 8 to 14 days arm was 30 ml (95% CI, -1,040, 355). Platelet aggregation in vitro increased after transfusion in both the room temperature- and cold-stored platelet study arms. Total blood usage, number of adverse events, length of stay in intensive care, and mortality were comparable among patients receiving cold-stored and room temperature-stored platelets., Conclusions: This pilot trial supports the feasibility of platelets stored cold for up to 14 days and provides critical guidance for future pivotal trials in high-risk cardiothoracic bleeding patients., (Copyright © 2020, the American Society of Anesthesiologists, Inc. All Rights Reserved.)

Background: Increasing numbers of emergency medical service agencies and hospitals are developing the capability to administer blood products to patients with hemorrhagic shock. Cold-stored whole blood (WB) is the only single product available to prehospital providers who aim to deliver a balanced resuscitation strategy. However, there are no data on the safety and in vitro characteristics of prehospital stored WB. This study aimed to describe the effects on in vitro quality of storing WB at remote helicopter bases in thermal insulating containers., Study Design and Methods: We conducted a two-armed single-center study. Twenty units (test) were stored in airtight thermal insulating containers, and 20 units (controls) were stored according to routine procedures in the Haukeland University Hospital Blood Bank. Storage conditions were continuously monitored during emergency medical services missions and throughout remote and blood bank storage. Hematologic and metabolic variables, viscoelastic properties, and platelet (PLT) aggregation were measured on Days 1, 8, 14, and 21., Results: Storage conditions complied with the EU guidelines throughout remote and in-hospital storage for 21 days. There were no significant differences in PLT aggregation, viscoelastic properties, and hematology variables between the two groups. Minor significantly lower pH, glucose, and base excess and higher lactate were observed after storage in airtight containers., Conclusion: Forward cold storage of WB is safe and complies with EU standards. No difference is observed in hemostatic properties. Minor differences in metabolic variables may be related to the anaerobic conditions within the thermal box., (© 2020 The Authors. Transfusion published by Wiley Periodicals, Inc. on behalf of AABB.)

Background: COVID-19 can lead to life-threatening disease. While awaiting vaccines or documented specific therapeutic agents, several alternative treatment options are under investigation. This is a case report of the first COVID-19 patient treated with convalescent plasma in Norway., Case Presentation: A patient with severe COVID-19 on prolonged mechanical ventilation, who was PCR SARS-Cov-2 positive on day 22, was transfused with convalescent plasma on day 31 and tested negative for SARS-CoV-2 the following day. The patient gradually improved and was weaned from the ventilator and discharged alive from the ICU on day 63., Interpretation: This case report concerns one patient with clinical improvement after convalescent plasma transfusion. A SARS-CoV-2 test was not performed immediately before transfusion and the complexity of intensive care treatment makes it difficult to draw any conclusions on the potential effectiveness of this treatment. However, this case report is encouraging with regard to planned trials with convalescent plasma.

Objectives: The aim of this study was to measure blood concentrations of environmental pollutants in Norwegian donors and evaluate the risk of pollutant exposure through blood transfusions., Background: Transfused blood may be a potential source of exposure to heavy metals and organic pollutants and presents a risk to vulnerable patient groups such as premature infants., Methods/materials: Donors were randomly recruited from three Norwegian blood banks: in Bergen, Tromsø and Kirkenes. Selected heavy metals were measured in whole blood using inductively coupled plasma mass spectrometry (ICP-MS), and perfluoroalkyl substances (PFAS) were measured in serum by ultrahigh-pressure liquid chromatography coupled with a triple-quadrupole mass spectrometer (UHPLC-MS/MS)., Results: Almost 18% of blood donors had lead concentrations over the limit suggested for transfusions in premature infants (0.09 μmol/L). About 11% of all donors had mercury concentrations over the suggested limit of 23.7 nmol/L. Cadmium was higher than the limit, 16 nmol/L, in 4% of donors. Perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS) concentrations were over the suggested limit of 0.91 ng/mL in 68% and 100% of the donors, respectively. PFAS concentrations and heavy metal concentrations increased with donor's age., Conclusion: A considerable percentage of donors had lead, PFOS and PFOA concentrations over the suggested limits. In addition, at each study site, there were donors with high mercury and cadmium concentrations. Selecting young donors for transfusions or measurements of pollutants in donor blood may be a feasible approach to avoid exposure through blood transfusions to vulnerable groups of patients such as premature infants., (© 2020 The Authors. Transfusion Medicine published by John Wiley & Sons Ltd on behalf of British Blood Transfusion Society.)

Background: Some jurisdictions require leukoreduction of cellular blood components. The only whole blood collection set with a platelet-saving filter uses citrate-phosphate-dextrose (CPD) as storage solution. Substituting CPD with citrate-phosphate-dextrose-adenine (CPDA-1) increases shelf life from 21 to 35 days. This would simplify prehospital and rural resupply and reduce wastage. We investigated in vitro quality and hemostatic properties of CPDA-1 whole blood leukoreduced with a platelet-saving filter., Study Design and Methods: CPDA-1 whole blood was leukoreduced using a platelet-saving filter and stored 35 days. EDQM requirements, hematology, metabolic parameters, thromboelastography, light transmission aggregometry, fibrinogen, factor VIII, and interleukin-6 were measured on Days 0, 1, 14, 21, and 35 and compared to non-leukoreduced blood., Results: All units met EDQM requirements. Leukoreduction yielded residual white blood cell count <1 × 10 6 and 87% platelet recovery on Day 1. It caused reduction in thromboelastography parameters, but not aggregometry response. No hemolysis >0.8% was observed. Factor VIII was higher on Day 35 in the leukoreduced group, 37.9 (95% CI: 26.0, 49.8) versus 13.8 (9.4, 18.2) IU/dL. In both groups, aggregation was significantly reduced by Day 14. Thromboelastography showed remaining platelet activity on Day 35, MA 46.9 (42.1, 51.7) in the leukoreduced and 44.3 (39.6, 49.0) mm in the non-leukoreduced group. Fibrinogen was within reference ranges at Day 35 (>2 g/dL). Interleukin-6 was not detectable., Conclusion: Leukoreducing CPDA-1 whole blood with a platelet-saving filter did not compromise hemostatic properties. We encourage development of a single bag CPDA-1 whole blood collection set with in-line platelet-saving filter., (© 2020 The Authors. Transfusion published by Wiley Periodicals, Inc. on behalf of AABB.)

Objectives: Tonsillectomy (TE) or adenotonsillectomy (ATE) may have a beneficial effect on the clinical course in children with the periodic fever, aphthous stomatitis, pharyngitis and cervical adenitis (PFAPA) syndrome. However, an immunological reason for this effect remains unknown. This literature review summarizes the current knowledge regarding the immunological role of the tonsils in the PFAPA syndrome., Methods: We searched PubMed, Medline, EMBASE and Cochrane for papers written in English dated from 1 January 1987 to 30 April 2019. The search included all studies reporting histological, immunological or microbiological workup of tonsil specimens from children aged 0-18 years with PFAPA., Results: Thirteen articles reported histological, immunological or microbiological workup of tonsil specimens in children with PFAPA. The histology of tonsil specimens from children with PFAPA displayed chronic tonsillar inflammation with lymphoid hyperplasia. No uniform immunological pattern was identified, but some studies found fewer B-lymphocytes and smaller germinal centers in PFAPA compared to controls. A difference in tonsillar microbiota between PFAPA and controls was found in one study., Conclusion: A uniform immunological or microbiological pattern explaining the clinical effect of TE in children with PFAPA has not been revealed. Future targeted immunological studies of tonsils in PFAPA patients could possibly illuminate the understanding of the immunology in this disease., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Cold storage of platelets may extend shelf life compared to room temperature storage. This study aimed to investigate in vitro platelet quality and function in cold-stored and delayed-cold-stored nonagitated apheresis platelets in platelet additive solution during storage for 21 days., Study Design and Methods: Ten double apheresis platelet concentrates in 37% plasma/63% PAS-IIIM were split into two groups; nonagitated 2 to 6°C storage (CSPs) and delayed cold storage (DCSPs) with 7 days agitated storage at 20-24°C followed by nonagitated cold storage for 14 additional days. Platelet count, metabolism, viscoelastic properties, and aggregation ability were measured on Days 1, 7, 14, and 21., Results: All platelet units, both CSPs and DCSPs, complied with the EU guidelines throughout storage for 21 days. Swirling was not detectable after cold storage. Cold storage improved platelet function; however, DCSP on Day 7 showed poorer results compared to CSP. Cold storage slowed down metabolism, with lower lactate and higher glucose concentrations in the CSP compared to the DCSP throughout storage for 21 days., Conclusion: Cold storage of platelets improved platelet function in in vitro assays, even though delayed cold storage on Day 7 showed poorer results compared to continuous cold storage. This difference could be explained by accelerated metabolism and higher glucose consumption during the period of room temperature storage. Cold storage and delayed cold storage could ease inventory management. Further studies investigating the in vitro and clinical effects of cold-stored and delayed-cold-stored platelets are encouraged., (© 2019 AABB.)

Background: Natural killer (NK) cells are potential effectors in anti-cancer immunotherapy; however only a subset potently kills cancer cells. Here, we examined whether pretreatment of glioblastoma (GBM) with the proteasome inhibitor, bortezomib (BTZ), might sensitize tumour cells to NK cell lysis by inducing stress antigens recognized by NK-activating receptors. Methods: Combination immunotherapy of NK cells with BTZ was studied in vitro against GBM cells and in a GBM-bearing mouse model. Tumour cells were derived from primary GBMs and NK cells from donors or patients. Flow cytometry was used for viability/cytotoxicity evaluation as well as in vitro and ex vivo phenotyping. We performed a Seahorse assay to assess oxygen consumption rates and mitochondrial function, Luminex ELISA to determine NK cell secretion, protein chemistry and LC-MS/MS to detect BTZ in brain tissue. MRI was used to monitor therapeutic efficacy in mice orthotopically implanted with GBM spheroids. Results: NK cells released IFNγ, perforin and granzyme A cytolytic granules upon recognition of stress-ligand expressing GBM cells, disrupted mitochondrial function and killed 24-46% of cells by apoptosis. Pretreatment with BTZ further increased stress-ligands, induced TRAIL-R2 expression and enhanced GBM lysis to 33-76% through augmented IFNγ release ( p < 0.05). Blocking NKG2D, TRAIL and TRAIL-R2 rescued GBM cells treated with BTZ from NK cells, p = 0.01. Adoptively transferred autologous NK-cells persisted in vivo ( p < 0.05), diminished tumour proliferation and prolonged survival alone (Log Rank 10.19 , p = 0.0014, 95%CI 0.252-0.523) or when combined with BTZ (Log Rank 5.25 , p = 0.0219, 95%CI 0.295-0.408), or either compared to vehicle controls (median 98 vs. 68 days and 80 vs. 68 days, respectively). BTZ crossed the blood-brain barrier, attenuated proteasomal activity in vivo ( p < 0.0001; p < 0.01 compared to vehicle control or NK cells only, respectively) and diminished tumour angiogenesis to promote survival compared to vehicle-treated controls (Log Rank 6.57 , p = 0.0104, 95%CI 0.284-0.424, median 83 vs. 68 days). However, NK ablation with anti-asialo-GM1 abrogated the therapeutic efficacy. Conclusions: NK cells alone or in combination with BTZ inhibit tumour growth, but the scheduling of BTZ in vivo requires further investigation to maximize its contribution to the efficacy of the combination regimen.

The shift toward using a transfusion strategy in a ratio to mimic whole blood (WB) functionality has revitalized WB as a viable option to replace severe blood loss in civilian health care. A military-civilian collaboration has contributed to the reintroduction of WB at Haukeland University Hospital in Bergen, Norway. WB has logistical and hemostatic advantages in both the pre- and in-hospital settings where the goal is a perfectly timed balanced transfusion strategy. In this paper, we describe an event leading to activation of our emergency WB collection strategy for the first time. We evaluate the feasibility of our civilian walking blood bank (WBB) to cover the need of a massive amount of blood in an emergency situation. The challenges are discussed in relation to the different stages of the event with the recommendations for improvement in practice. We conclude that the use of pre-screened donors as a WBB in a civilian setting is feasible. The WBB can provide platelet containing blood components for balanced blood resuscitation in a clinically relevant time frame., (© 2019 AABB.)

Background: Damage control resuscitation principles advocate the use of blood to treat traumatic hemorrhage. Hemorrhage is a leading cause of preventable death on the battlefield, but making blood components available far forward presents logistical challenges due to shelf life and storage requirements. Whole blood simplifies logistics and enables collection in the field but can cause leukocyte-related transfusion reactions. A field-adapted leukoreduction system must be fast and safe, and storage of whole blood should preserve hemostatic function., Methods: Blood was collected using Imuflex WB-SP and leukoreduced at 0, 150, or 300 mm Hg. Additional bags were stored at 4°C for 21 days unagitated, mixed daily, agitated or head-over-heel rotated, at 22°C for 3 days, or 32°C for 2 hours. Hematology, coagulation, CD62P/CD42b, thromboelastography (TEG)/thromboelastometry (ROTEM), and Multiplate was performed., Results: Filtration time was 35 ± 1, 14 ± 0, and 9 ± 0 minutes at 0, 150, and 300 mm Hg, respectively. One of 10 units at 150 mm Hg and 4 of 11 at 300 mm Hg had residual whole blood cells greater than 5.0 × 10 per unit. One of 11 at 300 mm Hg had platelet recovery of less than 80%. Hemolysis was less than 0.2%. Filtration decreased thromboelastography/thromboelastometry and Multiplate aggregation response. Stored at 4°C, α and MA/MCF moderately decreased regardless of mixing. Significant loss of aggregation response and increased CD62P expression was seen by Day 10. By Day 3, storage at 22°C caused loss of most aggregation. Two-hour storage at 32°C did not significantly affect hemostatic capacity., Conclusion: Forced filtration reduced leukoreduction time, but increased residual whole blood cells reduced hemostatic function. Aggregation response deteriorated early in storage, while viscoelastic assays decreased more gradually. Mixing showed no benefits., Level of Evidence: Diagnostic study, level IV.

Background: Allogeneic hematopoietic stem cell transplantation is associated with a high risk of immune-mediated post-transplant complications. Graft depletion of immunocompetent cell subsets is regarded as a possible strategy to reduce this risk without reducing antileukemic immune reactivity., Study Design and Methods: We investigated the effect of hematopoietic stem cell mobilization with granulocyte colony-stimulating factor (G-CSF) on peripheral blood and stem cell graft levels of various T, B, and NK cell subsets in healthy donors. The results from flow cytometric cell quantification were examined by bioinformatics analyses., Results: The G-CSF-induced mobilization of lymphocytes was a non-random process with preferential mobilization of naïve CD4 + and CD8 + T cells together with T cell receptor αβ + T cells, naïve T regulatory cells, type 1 T regulatory cells, mature and memory B cells, and cytokine-producing NK cells. Analysis of circulating lymphoid cell capacity to release various cytokines (IFNγ, IL10, TGFβ, IL4, IL9, IL17, and IL22) showed preferential mobilization of IL10 releasing CD4 + T cells and CD3 - 19 - cells. During G-CSF treatment, the healthy donors formed two subsets with generally strong and weaker mobilization of immunocompetent cells, respectively; hence the donors differed in their G-CSF responsiveness with regard to mobilization of immunocompetent cells. The different responsiveness was not reflected in the graft levels of various immunocompetent cell subsets. Furthermore, differences in donor G-CSF responsiveness were associated with time until platelet engraftment. Finally, strong G-CSF-induced mobilization of various T cell subsets seemed to increase the risk of recipient acute graft versus host disease, and this was independent of the graft T cell levels., Conclusion: Healthy donors differ in their G-CSF responsiveness and preferential mobilization of immunocompetent cells. This difference seems to influence post-transplant recipient outcomes.

Background: Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and thereafter harvested by leukapheresis are commonly used for allogeneic stem cell transplantation., Methods: Plasma levels of 38 soluble mediators (cytokines, soluble adhesion molecules, proteases, protease inhibitors) were analyzed in samples derived from healthy stem cell donors before G-CSF treatment and after 4 days, both immediately before and after leukapheresis., Results: Donors could be classified into two main subsets based on their plasma mediator profile before G-CSF treatment. Seventeen of 36 detectable mediators were significantly altered by G-CSF; generally an increase in mediator levels was seen, including pro-inflammatory cytokines, soluble adhesion molecules and proteases. Several leukocyte- and platelet-released mediators were increased during apheresis. Both plasma and graft mediator profiles were thus altered and showed correlations to graft concentrations of leukocytes and platelets; these concentrations were influenced by the apheresis device used. Finally, the mediator profile of the allotransplant recipients was altered by graft infusion, and based on their day +1 post-transplantation plasma profile our recipients could be divided into two major subsets that differed in overall survival., Discussion: G-CSF alters the short-term plasma mediator profile of healthy stem cell donors. These effects together with the leukocyte and platelet levels in the graft determine the mediator profile of the stem cell grafts. Graft infusion also alters the systemic mediator profile of the recipients, but further studies are required to clarify whether such graft-induced alterations have a prognostic impact., (Copyright © 2018. Published by Elsevier Inc.)

Titanium is used in a wide variety of materials ranging from medical devices to materials used in everyday life. Adverse biological reactions that could occur in patients, consumers, and workers should be monitored and prevented. There is a lack of available agents to test and predict titanium-related hypersensitivity. The aim of this study was to develop two bioavailable titanium substances in ionic and nanoparticulate form to serve as antigens for hypersensitivity testing in vitro. Peripheral blood mononuclear cells from 20 test subjects were stimulated with the antigens and secretion of monocytic and lymphatic cytokines and chemokines were measured by a multiplex bead assay. Lymphocyte stimulation indices were also determined in a subset of test subjects by measuring CD69 and HLA-DR expression by flow cytometry. Cytokine profiling revealed that both antigens increased production of typical monocyte and macrophage secreted cytokines after 24 h, with significant increases in IL-1β, IL-7, IL-10, IL-12, IL-2R, IL-6, GM-CSF, TNF-α, IL-1RA, MIP-1α, MIP-1β, IFN-α, and IL-15. Lymphatic cytokines and chemokines were not significantly induced by activation. After seven days of stimulation, ionic-Ti (2.5 μg/mL) caused proliferation (stimulation index > 2) of CD4+ cells and CD8+ cells in all persons tested ( N = 6), while titanium dioxide nanoparticles (50 μg/mL) only caused significant proliferation of CD4+ cells. Our preliminary results show that the experimental titanium antigens, especially the ionic form, induce a general inflammatory response in vitro. A relevant cohort of test subjects is required to further elucidate their potential for predictive hypersensitivity testing., Competing Interests: The authors declare no conflict of interest.

Background: Endogenous antibodies to signaling molecules and receptors (Abs) are associated with Alzheimer's disease (AD)., Objectives: To investigate the association of 33 Abs to dopaminergic, serotoninergic, muscarinic, adrenergic, vascular, and immune receptors with cognitive, neuropsychiatric, and mortality outcomes., Methods: Ninety-one patients with mild AD were followed annually for 5 years with the Mini-Mental State Examination (MMSE) and the Neuropsychiatric Inventory (NPI; composite outcomes: "psychosis" (item 1 + 2), "mood" (item 4 + 5 + 7), and "agitation" (item 3 + 8 + 9)). Abs were quantified in sera obtained at baseline by ELISA and reduced to principal components (PCs). Associations between Abs and outcomes were assessed by a mixed model (MMSE decline), zero-inflated fixed effects count models (composite NPI scores), and Cox regression (mortality). The resulting p-values were adjusted for multiple testing according to a false discovery rate of 0.05 (Benjamini-Hochberg)., Results: The measured levels of the 33 Abs formed four PCs. PC1 (dopaminergic and serotonergic Abs) was associated with increased mortality (Hazard ratio 2.57, p < 0.001), PC2 (serotonergic, immune, and vascular Abs) with decreased agitation symptoms (β - 0.19, p < 0.001), and PC3 (cholinergic receptor Abs) with increased mood symptoms (β 0.04, p = 0.002), over time. There were no associations between Abs and MMSE decline., Conclusion: The associations between Abs, mortality, and neuropsychiatric symptoms reported in this cohort are intriguing. They cannot, however, be generalized. Validation in independent sample sets is required.

Patients with Alzheimer's disease (AD) have blood-brain barrier (BBB) dysfunction. Methods to study cells of the BBB in vivo would facilitate analyses of neurovascular damage in early AD. Thus, we conducted a pilot study to investigate if brain-derived endothelial cells (BDCECs) could be identified from a cell population of circulating endothelial cells (CECs). Peripheral blood was sampled from early AD patients (n = 9), patients with vascular diseases (myocardial infarction (n = 8) and ischemic stroke (n = 8)), and healthy controls (n = 8). We enumerated CD34 + /CD146 + /CD45 - cells (CECs) and Glucose transporter-1 (Glut1 + CECs (BDCECs)) by flow cytometry. We found that BDCECs formed a separate, aggregate cell population. Glut1 expression on BDCECs, measured by the median fluorescence intensity, was significantly decreased in patients with AD compared to both the healthy controls and patients with myocardial infarction ((p < .05, Kruskal-Wallis, Dunn's post hoc test). We found no significant differences in cell numbers. Our study shows that isolation of BDCECs offers a promising non-invasive tool to investigate cells derived from the BBB. Downregulation of Glut1 at the mild stages of AD suggests that agents that increase Glut1 levels may be therapeutic candidates to improve energy availability to the brain., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Human cytomegalovirus (HCMV) antigens in glioblastoma (GBM) present opportunities for personalised immunotherapy. However, their presence in GBM tissue is still under debate, and evidence of their impact on functional immune responses and prognosis is sparse. Here, we investigated the presence of pp65 (UL83) and immediate early 1 (IE-1) HCMV antigens in a cohort of Norwegian GBM patients ( n = 177), using qPCR, immunohistochemistry, and serology. HCMV status was then used to investigate whether viral antigens influenced immune cell phenotype, infiltration, activation and patient survival. Pp65 and IE-1 were detected by qPCR in 23% and 43% of GBM patients, respectively. Furthermore, there was increased seropositivity in GBM patients relative to donors (79% vs . 48%, respectively; Logistic regression, OR = 4.05, 95%CI [1.807-9.114], P = 0.001, also when adjusted for age (OR = 2.84, 95%CI [1.110-7.275], P = 0.029). Tissue IE-1-positivity correlated with increased CD3 + CD8 + T-cell infiltration ( P < 0.0001), where CD8 + effector memory T (T EM ) cells accounted for the majority of CD8 + T cells compared with peripheral blood of HCMV + patients ( P < 0.0001), and HCMV + ( P < 0.001) and HCMV - ( P < 0.001) donors. HLA-A2/B8-restricted HCMV-specific CD8 + T cells were more frequent in blood and tumor of HCMV + GBM patients compared with seronegative patients, and donors irrespective of their serostatus. In biopsies, the HCMV-specific CD8 + T EM cells highly expressed CTLA-4 and PD-1 immune checkpoint protein markers compared with populations in peripheral blood ( P < 0.001 and P < 0.0001), which expressed 3-fold greater levels of CD28 ( P < 0.001 and P < 0.0001). These peripheral blood T cells correspondingly secreted higher levels of IFNγ in response to pp65 and IE-1 peptide stimulation ( P < 0.001). Thus, despite apparent increased immunogenicity of HCMV compared with tumor antigens, the T cells were tolerised, and HCMV status did not impact patient survival (Log Rank 3.53 HR = 0.85 95%CI [0.564-1.290], P = 0.45). Enhancing immune functionality in the tumor microenvironment thus may improve patient outcome.

Background: Alzheimer's disease (AD) is associated with several antibodies as well as signaling molecules and receptors. These may be detrimental in the presence of a disrupted blood-brain barrier (BBB)., Objective: To investigate whether the levels of antibodies toward 33 signaling molecules involved in neurotransmitter, vascular, and immune functions were associated with AD and, within the AD group; cognitive function and mood., Methods: Antibodies in sera from patients with mild AD [(n = 91) defined as a Mini-Mental State Examination ≥ 20 or a Clinical Dementia Rating Scale≤1] and healthy controls (n = 102) were measured with enzyme-linked immunosorbent assays. Levels in AD and controls were compared by Mann-Whitney test. In the AD group, associations between antibodies and psychometric test scores were analyzed by robust regression. The false discovery threshold was set to 0.05., Results: Antibodies to serotonin receptors [5-HT2AR (effect size (r) = 0.21, p = 0.004), 5-HT2CR (r = 0.25, p = 0.0005) and 5-HT7R (r = 0.21, p = 0.003)], vascular endothelial growth factor receptor 1 [VEGFR1 (r = 0.29, p < 0.001)] and immune-receptors (Stabilin-1 (r = 0.23, p = 0.001) and C5aR1 (r = 0.21, p = 0.004) were higher in AD. Psychomotor speed was associated with D1R-abs (β 0.49, p < 0.001), depression with ETAR-abs (β 0.31, p < 0.001), and visuospatial function with 5-HT1AR-abs (β 0.27, p = 0.004) despite similar antibody levels compared to controls., Conclusions: Antibody levels to VEGFR1, serotonergic receptors, and receptors in the immune system were increased in AD. Antibodies at similar levels as in controls were associated cognitive dysfunction and depression in AD.

Myalgic Encephalopathy/Chronic Fatigue Syndrome (ME/CFS) is a disease of unknown etiology. We have previously suggested clinical benefit from B-cell depletion using the monoclonal anti-CD20 antibody rituximab in a randomized and placebo-controlled study. Prolonged responses were then demonstrated in an open-label phase-II study with maintenance rituximab treatment. Using blood samples from patients in the previous two clinical trials, we investigated quantitative changes in T-lymphocyte subsets, in immunoglobulins, and in serum levels of two B-cell regulating cytokines during follow-up. B-lymphocyte activating factor of the tumor necrosis family (BAFF) in baseline serum samples was elevated in 70 ME/CFS patients as compared to 56 healthy controls (p = 0.011). There were no significant differences in baseline serum BAFF levels between patients with mild, moderate, or severe ME/CFS, or between responders and non-responders to rituximab. A proliferation-inducing ligand (APRIL) serum levels were not significantly different in ME/CFS patients compared to healthy controls at baseline, and no changes in serum levels were seen during follow-up. Immunophenotyping of peripheral blood T-lymphocyte subsets and T-cell activation markers at multiple time points during follow-up showed no significant differences over time, between rituximab and placebo groups, or between responders and non-responders to rituximab. Baseline serum IgG levels were significantly lower in patients with subsequent response after rituximab therapy compared to non-responders (p = 0.03). In the maintenance study, slight but significant reductions in mean serum immunoglobulin levels were observed at 24 months compared to baseline; IgG 10.6-9.5 g/L, IgA 1.8-1.5 g/L, and IgM 0.97-0.70 g/L. Although no functional assays were performed, the lack of significant associations of T- and NK-cell subset numbers with B-cell depletion, as well as the lack of associations to clinical responses, suggest that B-cell regulatory effects on T-cell or NK-cell subsets are not the main mechanisms for the observed improvements in ME/CFS symptoms observed in the two previous trials. The modest increase in serum BAFF levels at baseline may indicate an activated B-lymphocyte system in a subgroup of ME/CFS patients.

Peripheral blood stem cells from healthy donors mobilized by granulocyte colony-stimulating factor (G-CSF) and harvested by leukapheresis are commonly used for allogeneic stem cell transplantation. The frequency of severe graft versus host disease is similar for patients receiving peripheral blood and bone marrow allografts, even though the blood grafts contain more T cells, indicating mobilization-related immunoregulatory effects. The regulatory phosphoprotein osteopontin was quantified in plasma samples from healthy donors before G-CSF treatment, after four days of treatment immediately before and after leukapheresis, and 18-24 h after apheresis. Myeloma patients received chemotherapy, combined with G-CSF, for stem cell mobilization and plasma samples were prepared immediately before, immediately after, and 18-24 h after leukapheresis. G-CSF treatment of healthy stem cell donors increased plasma osteopontin levels, and a further increase was seen immediately after leukapheresis. The pre-apheresis levels were also increased in myeloma patients compared to healthy individuals. Finally, in vivo G-CSF exposure did not alter T cell expression of osteopontin ligand CD44, and in vitro osteopontin exposure induced only small increases in anti-CD3- and anti-CD28-stimulated T cell proliferation. G-CSF treatment, followed by leukapheresis, can increase systemic osteopontin levels, and this effect may contribute to the immunomodulatory effects of G-CSF treatment.

Background: Allergic transfusion reactions (ATRs) present with a broad range of symptoms probably caused by mediators released from mast cells and basophil granulocytes upon activation. Passive immunoglobulin (Ig)E sensitization may yield clinical symptoms and positive allergy tests. Unexpected findings of IgE antibodies in pooled solvent/detergent (S/D)-treated plasma (Octaplas, Octapharma) during routine analysis initiated an investigation of serum proteins., Study Design and Methods: Consecutive batches of S/D-plasma transfused during September 2014 through March 2015 were investigated for IgE, IgG, IgA IgM, C3, C4, haptoglobin, anti-nuclear antibodies (ANAs), and red blood cell (RBC) antibodies., Results: During the study period, 4203 S/D-plasma units were transfused. Nineteen (14 Octaplas A and five Octaplas AB) of 20 batches of S/D-plasma were included, representing 99.9% of total number of plasma units. A total of 0.4% of units and five batches reported ATRs. Concentrations of total IgE higher than expected values in adults (<120 kU/L) were observed in 18 of the 19 (95%) batches investigated (median concentration [quartiles], 161 [133-183]). Specific IgE antibodies (expected < 0.35 kilounits antigen [kUA]/L) against house dust mite (2.52 [1.01-5.09]), timothy (2.83 [2.48-3.24]), cat (1.13 [0.58-1.52]), dog (0.83 [0.50-1.05]), mugwort (0.69 [0.53-0.97]), birch (0.62 [0.28-0.92]), peanut (0.52 [0.29-075]), wheat (0.46 [0.33-0.69]), and latex (0.32 [0.21-0.53]) were also detected. IgG, IgA, IgM, C3, C4, and haptoglobin were within or below normal ranges. No RBC antibodies were observed, but 18% of batches showed low levels of ANA (anti-RNP)., Conclusion: Specific IgE antibodies against airborne allergens, food allergens, and latex were detected in S/D-treated pooled plasma., (© 2016 AABB.)

Background: Myalgic Encephalopathy/Chronic Fatigue Syndrome (ME/CFS) is a disease of unknown etiology. We previously reported a pilot case series followed by a small, randomized, placebo-controlled phase II study, suggesting that B-cell depletion using the monoclonal anti-CD20 antibody rituximab can yield clinical benefit in ME/CFS., Methods: In this single-center, open-label, one-armed phase II study (NCT01156909), 29 patients were included for treatment with rituximab (500 mg/m2) two infusions two weeks apart, followed by maintenance rituximab infusions after 3, 6, 10 and 15 months, and with follow-up for 36 months., Findings: Major or moderate responses, predefined as lasting improvements in self-reported Fatigue score, were detected in 18 out of 29 patients (intention to treat). Clinically significant responses were seen in 18 out of 28 patients (64%) receiving rituximab maintenance treatment. For these 18 patients, the mean response durations within the 156 weeks study period were 105 weeks in 14 major responders, and 69 weeks in four moderate responders. At end of follow-up (36 months), 11 out of 18 responding patients were still in ongoing clinical remission. For major responders, the mean lag time from first rituximab infusion until start of clinical response was 23 weeks (range 8-66). Among the nine patients from the placebo group in the previous randomized study with no significant improvement during 12 months follow-up after saline infusions, six achieved a clinical response before 12 months after rituximab maintenance infusions in the present study. Two patients had an allergic reaction to rituximab and two had an episode of uncomplicated late-onset neutropenia. Eight patients experienced one or more transient symptom flares after rituximab infusions. There was no unexpected toxicity., Conclusion: In a subgroup of ME/CFS patients, prolonged B-cell depletion with rituximab maintenance infusions was associated with sustained clinical responses. The observed patterns of delayed responses and relapse after B-cell depletion and regeneration, a three times higher disease prevalence in women than in men, and a previously demonstrated increase in B-cell lymphoma risk for elderly ME/CFS patients, suggest that ME/CFS may be a variant of an autoimmune disease., Trial Registration: ClinicalTrials.gov NCT01156909.

The syndrome of periodic fever, aphthous stomatitis, pharyngitis and cervical adenitis (PFAPA) is an autoinflammatory disorder of unknown aetiology. Tonsillectomy may cause a prompt resolution of the syndrome. The aim was to study the histologic and immunological aspects of the palatine tonsils in PFAPA, to help understand the pathophysiology of the syndrome. Tonsils from children with PFAPA (n = 11) and children with tonsillar hypertrophy (n = 16) were evaluated histologically after haematoxylin and eosin staining. The number of different cell types was identified immunohistochemically by cluster of differentiation (CD) markers: CD3 (T cells), CD4 (T helper cells), CD8 (cytotoxic T cells), CD15 (neutrophils), CD20 (B cells), CD45 (all leucocytes), CD57 (NK cells) and CD163 (monocytes and macrophages). Tonsils from children with PFAPA showed reactive lymphoid hyperplasia dominated by well-developed germinal centres with many tingible body macrophages. The histologic findings were unspecific, and a similar morphologic appearance was also found in the tonsils from controls. The number of CD8+ cells in germinal centres differed between children with PFAPA [median 9 cells (quartiles: 5, 15)] and controls [18 cells (12, 33) (P = 0.001)] and between children with PFAPA with (median 14 cells; 9, 16) and without (4 cells; 3, 8) aphthous stomatitis (P = 0.015). For the other cell types, no differences in germinal centres were found between children with PFAPA and controls. In conclusion, a lower number of CD8+ cells were found in germinal centres of tonsils in children with PFAPA compared to controls, which may be a feature linked to the aetiology of the syndrome., (© 2015 John Wiley & Sons Ltd.)

Background: Formulation of a medical preparedness plan for treating severely bleeding casualties during naval deployment is a significant challenge because of territory covered during most missions. The aim of this study was to evaluate the concept of "walking blood bank" as a supportable plan for supplying safe blood and blood products., Methods: In 2013, the Royal Norwegian Navy conducted antipiracy operations from a frigate, beginning in the Gulf of Aden and ending in the Indian Ocean. Crews were on 24-hour emergency alert in preparation for an enemy assault on the frigate. Under an approved command protocol, a "walking blood bank," using crew blood donations, was established for use on board and on missions conducted in rigid-hulled inflatable boats, during which freeze-dried plasma and leukoreduced, group O low anti-A/anti-B titer, cold-stored whole blood were stored in Golden Hour Boxes. Data demonstrating the ability to collect, store, and provide whole blood were collected to establish feasibility of implementing a whole blood-focused remote damage-control resuscitation program aboard a naval vessel. In addition, ROTEM data were collected to demonstrate feasibility of performing this analysis on a large naval vessel and to also measure hemostatic efficacy of cold-stored leukoreduced whole blood (CWB) stored during a period of 14 days. ROTEM data on CWB was compared with reconstituted whole blood., Results: Drills simulating massive transfusion activation were conducted, in which 2 U of warm fresh whole blood with platelet sparing leukoreduction were produced in 40 minutes, followed by collection of two additional units at 15-minute increments. The ROTEM machine performed well during ship-rolling, as shown by the overlapping calculated and measured mechanical piston movements measured by the ROTEM device. Error messages were recorded in 4 (1.5%) of 267 tests. CWB yielded reproducible ROTEM results demonstrating preserved fibrinogen function and platelet function for at least 3.5 weeks and 2 weeks, respectively. The frequency of ROTEM tests were as follows: EXTEM (n = 88), INTEM (n = 85), FIBTEM (n = 82), and APTEM (n = 12). CWB results were grouped. Compared with Days 0 to 2, EXTEM maximum clot firmness was significantly reduced, beginning on Days 10 to 14; however, results through that date remained within reference ranges and were comparable with the EXTEM maximum clot firmness for the reconstituted whole blood samples containing Day 5 room temperature-stored platelets., Conclusion: A "walking blood bank" can provide a balanced transfusion product to support damage-control resuscitation/remote damage-control resuscitation aboard a frigate in the absence of conventional blood bank products. ROTEM analysis is feasible to monitor damage-control resuscitation and blood product quality. ROTEM analysis was possible in challenging operational conditions., Level of Evidence: Therapeutic study, level V.

Background: The last decade of military trauma care has emphasized the role of blood products in the resuscitation of hemorrhaging patients. Damage-control resuscitation advocates decreased crystalloid use and reintroduces blood components as primary resuscitative fluids. The systematic use of blood products have been described in military settings, but reports describing the use of freeze dried plasma (FDP) or red blood cells (RBCs) in civilian prehospital care are few. We describe our preliminary results after implementing RBCs and FDP into our Helicopter Emergency Medical Service (HEMS)., Methods: We collected data on the use of FDP (LyoPlas N-w (AB)) during a 12-month period from May 31, 2013, to May 30, 2014, before RBC (0Rh (D) negative) introduction in June 2014. FDP and RBCs were indicated in trauma and medical patients presenting with clinical significant hemorrhage on scene. Data were obtained from HEMS registry and patient records., Results: Our preliminary results show that FDP was used in 16 patients (88% males) during the first year. Main patient categories were blunt trauma (n = 5), penetrating trauma (n = 4), and nontrauma (n = 7). Ten patients (62%) were hypotensive with systolic blood pressures less than 90 mm Hg on scene. The majority (75%) received tranexamic acid. Of 14 patients admitted to the hospital, 11 received emergency surgery and 8 needed additional transfusions within the first 24 hours. No transfusion-related complications were recorded. Two of the FDP patients died on scene, and the remaining 14 patients were alive after 30 days. Early results from the recent introduction of RBC show that RBCs were given to four patients. Two patients (one penetrating trauma and one blunt trauma patient) died on scene because of exsanguination, while additional two patients (one blunt trauma patient and one with ruptured aortic aneurism) survived to hospital discharge., Conclusion: Our small study indicates that introduction of FDP into civilian HEMS seems feasible and may be safe and that logistical and safety issues for the implementation of RBCs are solvable. FDP ensures both coagulation factors and volume replacement, has a potentially favorable safety profile, and may be superior to other types of plasma for prehospital use. Further prospective studies are needed to clarify the role of FDP (and RBCs) in civilian prehospital hemorrhagic shock resuscitation and to aid the development of standardized protocols for prehospital use of blood products., Level of Evidence: Therapeutic study, level V.

Human babesiosis is a rare but potentially life-threatening parasitic disease transmitted by ixodid ticks, and has not previously been reported in Norway. We report a case of severe babesiosis that occurred in Norway in 2007. The patient had previously undergone a splenectomy. He was frequently exposed to tick bites in an area endemic for bovine babesiosis in the west of Norway. The patient presented with severe haemolysis and multiorgan failure. Giemsa-stained blood smears revealed 30% parasitaemia with Babesia spp. He was treated with quinine in combination with clindamycin, apheresis, and supportive treatment with ventilatory support and haemofiltration, and made a complete recovery. This is the first case reported in Norway; however Babesia divergens seroprevalence in cattle in Norway is high, as is the risk of Ixodes ricinus tick bite in the general population. Babesiosis should be considered in the differential diagnosis of unexplained febrile haemolytic disease., (Copyright © 2015. Published by Elsevier Ltd.)

Background: Autoantibodies with agonist function are described in cardiovascular disorders. Since vascular risk factors are associated with an increased risk for Alzheimer's disease (AD), we investigated a potential association between antibodies to the angiotensin 2 type 1 receptor (anti-AT1R) and AD., Objective: The primary objective of this study was to investigate the association between anti-AT1R and AD. The secondary objective was to investigate the association between clinical or biomarker features of AD and anti-AT1R., Methods: Samples from patients with mild AD participating in a longitudinal study in Western Norway (n = 92, 65 [71%] females, mean age 74.8 [range 50-89]) and age- and gender-matched healthy controls (n = 102) were included. Cerebrospinal fluid (CSF) AD biomarkers were assessed in a subgroup of patients. Patients were examined annually, including Mini-Mental State Examination. ELISA was used to measure anti-AT1R in serum. Non-parametric tests were used for statistical calculations and a p <  0.05 was considered significant., Results: AD patients had significantly higher levels of anti-AT1R compared with healthy controls (10.2 U/mL versus 8.1 U/mL, p = 0.04). This difference was found only in patients without hypertension and diabetes. Anti-AT1R levels correlated with CSF total tau (p = 0.03) and phosphorylated tau (p = 0.03) levels, and inversely with blood pressure in AD (Spearman R -0.277, p = 0.008)., Discussion: AD is associated with increased levels of anti-AT1R, and the antibodies correlated with CSF total, and phosphorylated tau levels. Further research is needed to understand the blood pressure response in AD without hypertension and a potential link between tau and anti-AT1R in AD.

In past and ongoing military conflicts, the use of whole blood (WB) as a resuscitative product to treat trauma-induced shock and coagulopathy has been widely accepted as an alternative when availability of a balanced component-based transfusion strategy is restricted or lacking. In previous military conflicts, ABO group O blood from donors with low titers of anti-A/B blood group antibodies was favored. Now, several policies demand the exclusive use of ABO group-specific WB. In this short review, we argue that the overall risks, dangers, and consequences of "the ABO group-specific approach," in emergencies, make the use of universal group O WB from donors with low titers of anti-A/B safer. Generally, risks with ABO group-specific transfusions are associated with in vivo destruction of the red blood cells transfused. The risk with group O WB is from the plasma transfused to ABO-incompatible patients. In the civilian setting, the risk of clinical hemolytic transfusion reactions (HTRs) due to ABO group-specific red blood cell transfusions is relatively low (approximately 1:80,000), but the consequences are frequently severe. Civilian risk of HTRs due to plasma incompatible transfusions, using titered donors, is approximately 1:120,000 but usually of mild to moderate severity. Emergency settings are often chaotic and resource limited, factors well known to increase the potential for human errors. Using ABO group-specific WB in emergencies may delay treatment because of needed ABO typing, increase the risk of clinical HTRs, and increase the severity of these reactions as well as increase the danger of underresuscitation due to lack of some ABO groups. When the clinical decision has been made to transfuse WB in patients with life-threatening hemorrhagic shock, we recommend the use of group O WB from donors with low anti-A/B titers when logistical constraints preclude the rapid availability of ABO group-specific WB and reliable group matching between donor and recipient is not feasible.

Military experience and recent in vitro laboratory data provide a biological rationale for whole-blood use in the treatment of exsanguinating hemorrhage and have renewed interest in the reintroduction of fresh whole blood and cold-stored whole blood to patient care in austere environments. There is scant evidence to support, in a field environment, that a whole blood-based resuscitation strategy is superior to a crystalloid/colloid approach even when augmented by a limited number of red blood cell (RBC) and plasma units. Recent retrospective evidence suggests that, in this setting, resuscitation with a full compliment of RBCs, plasma, and platelets may offer an advantage, especially under conditions where evacuation is delayed. No current evacuation system, military or civilian, is capable of providing RBC, plasma, and platelet units in a prehospital environment, especially in austere settings. As a result, for the vast minority of casualties, in austere settings, with life-threatening hemorrhage, it is appropriate to consider a whole blood-based resuscitation approach to provide a balanced response to altered hemostasis and oxygen debt, with the goal of reducing the risk of death from hemorrhagic shock. To optimize the successful use of fresh whole blood/cold-stored whole blood in combat field environments, proper planning and frequent training to maximize efficiency and safety will be required. Combat medics will need proper protocol-based guidance and education if whole-blood collection and transfusion are to be successfully and safely performed in austere environments. In this article, we present the Norwegian Naval Special Operation Commando unit-specific remote damage control resuscitation protocol, which includes field collection and transfusion of whole blood. This protocol can serve as a template for others to use and adjust for their own military or civilian unit-specific needs and capabilities for care in austere environments.

Background: The Periodic Fever, Aphthous stomatitis, Pharyngitis and cervical Adenitis syndrome (PFAPA) is the most common periodic fever syndrome in childhood. Clinically, PFAPA may resemble autoinflammatory diseases, but the etiology is not fully understood., Methods: We measured inflammatory proteins in plasma and hematologic parameters in children with PFAPA during and between febrile episodes, and in a control group with suspected bacterial pneumonia. In children with PFAPA, a first blood sample was taken within 24 hours of a febrile episode and a second sample between episodes. In children with pneumonia, the first sample was taken shortly after admission and a second sample after full recovery., Results: A total of 22 children with PFAPA and 14 children with pneumonia were included. In children with PFAPA, levels of interleukin (IL) 6, CXCL10 and CCL4 were significantly increased during febrile episodes. The levels of IL-6 and CXCL10 were higher in children with PFAPA during febrile episodes than in children with pneumonia. The levels of CXCL10 remained higher in children with PFAPA between febrile episodes compared to children with pneumonia after recovery. Children with PFAPA had a relative eosinopenia and lymphocytopenia with reduced numbers of both CD4+ and CD8+ T cells during febrile episodes. This pattern was not observed in the children with pneumonia., Conclusions: The results indicate an innate immune response as the initial step in PFAPA, and a subsequent adaptive response with activation and redistribution of T cells. Moreover, an activation of the innate immune system involving CXCL10 may persist between febrile episodes. CXCL10 may be a possibly clinical marker in children with PFAPA.

Aim: To describe the incidence, epidemiology, clinical presentation and clinical outcome of children with the syndrome of periodic fever, aphthous stomatitis, pharyngitis and cervical adenitis (PFAPA) in a population-based study., Methods: In a prospective population-based study, all children in South Rogaland, Norway, diagnosed with PFAPA during 2004-2010 were evaluated clinically, and parents were interviewed systematically. A follow-up interview was performed for all patients., Results: A total of 46 children (32 boys; p = 0.011) were diagnosed with PFAPA. We calculated an incidence of 2.3 per 10 000 children up to 5 years of age. The median age of onset was 11.0 months (quartiles: 5.0, 14.8). Nearly 37 children were followed until resolution. In 17 of these, a tonsillectomy was performed with prompt resolution of PFAPA in all. The median age of spontaneous resolution was 60.2 months (range 24-120) and in children with tonsillectomy 50.9 months (range 15-128)., Conclusion: The incidence of PFAPA was 2.3 per 10 000 children up to 5 years of age. In the majority of cases, onset of symptoms may be during the first year of life., (©2012 The Author(s)/Acta Paediatrica ©2012 Foundation Acta Paediatrica.)

Background: A Giardia outbreak was associated with development of post-infectious functional gastrointestinal disorders (PI-FGID) and chronic fatigue syndrome (PI-CFS). Markers of immune dysfunction have given conflicting results in CFS and FGID patient populations. The aim of this study was to evaluate a wide selection of markers of immune dysfunction in these two co-occurring post-infectious syndromes., Methods: 48 patients, reporting chronic fatigue in a questionnaire study, were clinically evaluated five years after the outbreak and grouped according to Fukuda criteria for CFS (n=19) and idiopathic chronic fatigue (n=5) and Rome II criteria for FGIDs (n=54). 22 Giardia exposed non-fatigued individuals and 10 healthy unexposed individuals were recruited as controls. Peripheral blood lymphocyte subsets were analyzed by flow cytometry., Results: In peripheral blood we found significantly higher CD8 T-cell levels in PI-FGID, and significantly lower NK-cell levels in PI-CFS patients. Severity of abdominal and fatigue symptoms correlated negatively with NK-cell levels. A tendency towards lower T-cell CD26 expression in FGID was seen., Conclusion: Patients with PI-CFS and/or PI-FGID 5 years after Giardia lamblia infection showed alterations in NK-cell and CD8-cell populations suggesting a possible immunological abnormality in these conditions. We found no significant changes in other markers examined in this well-defined group of PI-CFS and PI-FGID elicited by a gastrointestinal infection. Controlling for co-morbid conditions is important in evaluation of CFS-biomarkers.

In these case reports, we investigated pandemic influenza 2009 vaccination of primary hypogammaglobulinaemic patients. Three combined variable immunodeficiency (CVID) patients and one X-linked agammaglobulinaemia (XLA) patient were vaccinated with the pandemic vaccine A/California/7/2009 (H1N1)-like split virus (X179a) adjuvanted with the oil-in-water emulsion AS03. Subsequently, serum and peripheral blood mononuclear cells were sampled and used to measure the haemagglutination inhibition (HI) and antibody-secreting cell (ASC) responses. In addition, the IFN-γ, IL-2 and TNF-α producing CD4(+) Th1-cell response was determined as these cytokines are important indicators of cell-mediated immunity. Two of the CVID patients responded to vaccination as determined by a >4-fold rise in HI antibodies. These subjects also had influenza-specific ASC numbers, which, albeit low, were higher than prevaccination levels. In addition, vaccination induced CD4(+) Th1-cell responses in both the XLA patient and the CVID patients, although the frequency of influenza-responsive cells varied amongst the patients. These results suggest that hypogammaglobulinaemia patients can mount a CD4(+) Th1 cell-mediated response to influenza vaccination and, additionally, that influenza vaccination of some hypogammaglobulinaemia patients can produce an influenza-specific humoral immune response. The findings should be confirmed in larger clinical studies., (© 2011 The Authors. Scandinavian Journal of Immunology © 2011 Blackwell Publishing Ltd.)

Background: Chronic fatigue syndrome (CFS) is a disease of unknown aetiology. Major CFS symptom relief during cancer chemotherapy in a patient with synchronous CFS and lymphoma spurred a pilot study of B-lymphocyte depletion using the anti-CD20 antibody Rituximab, which demonstrated significant clinical response in three CFS patients., Methods and Findings: In this double-blind, placebo-controlled phase II study (NCT00848692), 30 CFS patients were randomised to either Rituximab 500 mg/m(2) or saline, given twice two weeks apart, with follow-up for 12 months. Xenotropic murine leukemia virus-related virus (XMRV) was not detected in any of the patients. The responses generally affected all CFS symptoms. Major or moderate overall response, defined as lasting improvements in self-reported Fatigue score during follow-up, was seen in 10 out of 15 patients (67%) in the Rituximab group and in two out of 15 patients (13%) in the Placebo group (p = 0.003). Mean response duration within the follow-up period for the 10 responders to Rituximab was 25 weeks (range 8-44). Four Rituximab patients had clinical response durations past the study period. General linear models for repeated measures of Fatigue scores during follow-up showed a significant interaction between time and intervention group (p = 0.018 for self-reported, and p = 0.024 for physician-assessed), with differences between the Rituximab and Placebo groups between 6-10 months after intervention. The primary end-point, defined as effect on self-reported Fatigue score 3 months after intervention, was negative. There were no serious adverse events. Two patients in the Rituximab group with pre-existing psoriasis experienced moderate psoriasis worsening., Conclusion: The delayed responses starting from 2-7 months after Rituximab treatment, in spite of rapid B-cell depletion, suggests that CFS is an autoimmune disease and may be consistent with the gradual elimination of autoantibodies preceding clinical responses. The present findings will impact future research efforts in CFS., Trial Registration: ClinicalTrials.gov NCT00848692.