Blood donation for iron removal in individuals with HFE mutations: study of efficacy and safety and short review on hemochromatosis and blood donation.

Background: Elevated serum ferritin with/without HFE variants in asymptomatic persons leads frequently to referral for blood donation. Hemochromatosis (p.C282Y/p.C282Y) only requires treatment. We evaluated safety and feasibility of iron removal in healthy persons with elevated ferritin and HFE variants using blood donation procedures. Materials and methods: Thirty subjects with ferritin >200 ng/mL (women) or >300 ng/mL (men) with p.C282Y/p.C282Y, p.C282Y/p.H63D or p.H63D/p.H63D were randomized to weekly phlebotomy (removal of 450 mL whole blood) or erythrapheresis (removal of 360 mL red blood cells) every 14 days. The ferritin target was <100 ng/mL. A full blood count and ferritin were measured at each visit. Hemoglobin (Hb) ≥140 g/L was required at inclusion. If Hb dropped to <120 g/L (women) or <130 g/L (men), procedures were postponed (7 or 14 days). Primary endpoint was the number of procedures needed to the ferritin target; secondary objectives were duration of treatment and compliance. The treatment effect was tested with Poisson regression; number of procedures and treatment duration were compared between study arms with the Kruskal-Wallis test. Results: Twenty-five of 30 participants were men (83%); mean age was 47 years (SD 10.5), mean BMI 26.6 kg/m2 (SD 3.6); 17 had p.C282Y/p.C282Y, nine p.C282Y/p.H63D, four p.H63D/p.H63D. Median baseline Hb was 150 g/L (IQR 144, 1,559), median ferritin 504 ng/mL (IQR 406,620). Twenty-seven subjects completed the study. Treatment arm (p < 0.001) and HFE variant (p = 0.007) influenced the primary endpoint significantly. To ferritin levels <100 ng/mL, a median number of 7.5 (IQR 6.2, 9.8) phlebotomies and 4.0 (IQR 3.0, 5.8) erythraphereses (p = 0.001) was needed during a median of 66.5 days (IQR 49,103) and 78.5 days (IQR 46139), respectively (p = 0.448). Low Hb was the principal reason for protocol violation; anemia occurred in 13 participants (48%). Immediate complications were infrequent; fatigue was reported after 25% of phlebotomies and 45% of erythraphereses. Thirty-five procedures were postponed because of low Hb and 15 for non-medical reasons. The median interval was 7.0 (IQR 7.7) and 14.0 (IQR 14, 20) days between phlebotomies and erythraphereses, respectively. Conclusion: Blood donation procedures remove iron effectively in HC, but frequent treatments cause Hb decrease and fatigue that can impair feasibility.

Red blood cell transfusion-related dynamics of extracellular vesicles in intensive care patients: a prospective subanalysis.

Extracellular vesicles (EVs) accumulate during packed red blood cell (PRBC) storage. To date, the involvement of EVs in transfusion-related immunomodulation (TRIM) has not been prospectively evaluated in intensive care unit (ICU) patients. This was a prospective subanalysis of a recent observational feasibility study in postoperative ICU patients after: (1) open aortic surgery (Aorta), (2) bilateral lung transplantation (LuTx), and (3) other types of surgery (Comparison). Patient plasma was collected three times each before and after leukoreduced PRBC transfusion at 30-min intervals. The total number of EVs and EVs derived from erythrocytes (EryEVs), total platelets (total PEVs), activated platelets, granulocytes (GEVs), monocytes, and myeloid cells in PRBC samples and patient plasma were analyzed by flow cytometry. Statistical analysis was performed by Spearman's correlation test, linear mixed models and pairwise comparisons by Wilcoxon matched-pairs test. Twenty-three patients (Aorta n = 5, LuTx n = 9, Comparison n = 9) were included in the final analysis. All EV subgroups analyzed were detectable in all PRBCs samples (n = 23), but concentrations did not correlate with storage time. Moreover, all EVs analyzed were detectable in all plasma samples (n = 138), and EV counts were consistent before transfusion. Concentrations of total EVs, EryEVs, total PEVs, and GEVs increased after transfusion compared with baseline in the entire cohort but not in specific study groups. Furthermore, the change in plasma EV counts (total EVs and EryEVs) after transfusion correlated with PRBC storage time in the entire cohort. Extracellular vesicles were detectable in all PRBC and plasma samples. Individual EV subtypes increased after transfusion in the entire cohort, and in part correlated with storage duration. Future clinical studies to investigate the role of EVs in TRIM are warranted and should anticipate a larger sample size.Trial registration: Clinicaltrials.gov: NCT03782623.

Red blood cell transfusion-related eicosanoid profiles in intensive care patients-A prospective, observational feasibility study.

Introduction: Eicosanoids are bioactive lipids present in packed red blood cells (PRBCs), and might play a role in transfusion-related immunomodulation (TRIM). We tested the feasibility of analyzing eicosanoid profiles in PRBC supernatant and in plasma samples of postoperative intensive care unit (ICU) patients transfused with one unit of PRBCs. Methods: We conducted a prospective, observational feasibility study enrolling postoperative ICU patients: 1) patients treated with acetylsalicylic acid following abdominal aortic surgery (Aorta); 2) patients on immunosuppressants after bilateral lung transplantation (LuTx); and 3) patients undergoing other types of major surgery (Comparison). Abundances of arachidonic acid (AA) and seven pre-defined eicosanoids were assessed by liquid chromatography and tandem mass spectrometry. PRBC supernatant was sampled directly from the unit immediately prior to transfusion. Spearman's correlations between eicosanoid abundance in PRBCs and storage duration were assessed. Patient plasma was collected at 30-min intervals: Three times each before and after transfusion. To investigate temporal changes in eicosanoid abundances, we fitted linear mixed models. Results: Of 128 patients screened, 21 were included in the final analysis (Aorta n = 4, LuTx n = 8, Comparison n = 9). In total, 21 PRBC and 125 plasma samples were analyzed. Except for 20-hydroxyeicosatetraenoic acid (HETE), all analyzed eicosanoids were detectable in PRBCs, and their abundance positively correlated with storage duration of PRBCs. While 5-HETE, 12-HETE/8-HETE, 15-HETE, 20-HETE, and AA were detectable in virtually all plasma samples, 9-HETE and 11-HETE were detectable in only 57% and 23% of plasma samples, respectively. Conclusions: Recruitment of ICU patients into this transfusion study was challenging but feasible. Eicosanoid abundances increased in PRBC supernatants during storage. In plasma of ICU patients, eicosanoid abundances were ubiquitously detectable and showed limited fluctuations over time prior to transfusion. Taken together, larger clinical studies seem warranted and feasible to further investigate the role of PRBC-derived eicosanoids in TRIM.

Increasing test specificity without impairing sensitivity: lessons learned from SARS-CoV-2 serology.

BACKGROUND: Serological tests are widely used in various medical disciplines for diagnostic and monitoring purposes. Unfortunately, the sensitivity and specificity of test systems are often poor, leaving room for false-positive and false-negative results. However, conventional methods were used to increase specificity and decrease sensitivity and vice versa. Using SARS-CoV-2 serology as an example, we propose here a novel testing strategy: the 'sensitivity improved two-test' or 'SIT²' algorithm. METHODS: SIT² involves confirmatory retesting of samples with results falling in a predefined retesting zone of an initial screening test, with adjusted cut-offs to increase sensitivity. We verified and compared the performance of SIT² to single tests and orthogonal testing (OTA) in an Austrian cohort (1117 negative, 64 post-COVID-positive samples) and validated the algorithm in an independent British cohort (976 negatives and 536 positives). RESULTS: The specificity of SIT² was superior to single tests and non-inferior to OTA. The sensitivity was maintained or even improved using SIT² when compared with single tests or OTA. SIT² allowed correct identification of infected individuals even when a live virus neutralisation assay could not detect antibodies. Compared with single testing or OTA, SIT² significantly reduced total test errors to 0.46% (0.24-0.65) or 1.60% (0.94-2.38) at both 5% or 20% seroprevalence. CONCLUSION: For SARS-CoV-2 serology, SIT² proved to be the best diagnostic choice at both 5% and 20% seroprevalence in all tested scenarios. It is an easy to apply algorithm and can potentially be helpful for the serology of other infectious diseases.

INTERCEPT Pathogen Reduction in Platelet Concentrates, in Contrast to Gamma Irradiation, Induces the Formation of trans-Arachidonic Acids and Affects Eicosanoid Release during Storage.

Pathogen inactivation techniques for blood products have been implemented to optimize clinically safe blood components supply. The INTERCEPT system uses amotosalen together with ultraviolet light wavelength A (UVA) irradiation. Irradiation-induced inactivation of nucleic acids may actually be accompanied by modifications of chemically reactive polyunsaturated fatty acids known to be important mediators of platelet functions. Thus, here, we investigated eicosanoids and the related fatty acids released upon treatment and during storage of platelet concentrates for 7 days, complemented by the analysis of functional and metabolic consequences of these treatments. Metabolic and functional issues like glucose consumption, lactate formation, platelet aggregation, and clot firmness hardly differed between the two treatment groups. In contrast to gamma irradiation, here, we demonstrated that INTERCEPT treatment immediately caused new formation of trans-arachidonic acid isoforms, while 11-hydroxyeicosatetraenoic acid (11-HETE) and 15-HETE were increased and two hydroperoxyoctadecadienoic acid (HpODE) isoforms decreased. During further storage, these alterations remained stable, while the release of 12-lipoxygenase (12-LOX) products such as 12-HETE and 12-hydroxyeicosapentaenoic acid (12-HEPE) was further attenuated. In vitro synthesis of trans-arachidonic acid isoforms suggested that thiol radicals formed by UVA treatment may be responsible for the INTERCEPT-specific effects observed in platelet concentrates. It is reasonable to assume that UVA-induced molecules may have specific biological effects which need to be further investigated.

Indices of iron homeostasis in asymptomatic subjects with HFE mutations and moderate ferritin elevation during iron removal treatment.

We analysed iron biomarkers and their relationships in 30 subjects with HFE mutations and moderate hyperferritinaemia undergoing iron removal at our blood donation centre. Body mass index (BMI) and liver enzymes were assessed. Serum iron (SI), ferritin, transferrin saturation (TSAT), hepcidin and non-transferrin bound iron (NTBI) were measured serially. Seventeen subjects had p.C282Y/p.C282Y, nine p.C282Y/p.H63D, four p.H63D/p.H63D. Median age (p = 0.582), BMI (p = 0.500) and ferritin (p = 0.089) were comparable. At baseline, 12/17 p.C282Y/p.C282Y and 2/9 p.C282Y/p.H63D had measurable NTBI (p = 0.003). The p.C282Y/p.C282Y had higher TSAT (p < 0.001), lower hepcidin (p = 0.031) and hepcidin/ferritin ratio (p = 0.073). After treatment, iron indices were similar among groups, except TSAT (higher in p.C282Y/p.C282Y; p = 0.06). Strong relationships were observed between ferritin and TSAT (R = 0.71), NTBI and TSAT (R = 0.61), NTBI and SI (R = 0.54) in p.C282Y/p.C282Y. Hepcidin correlated weakly with ferritin in p.C282Y/p.C282Y (R = 0.37) but strongly in p.C282Y/p.H63D (R = 0.66) and p.H63D/p.H63D (R = 0.72), while relationships with TSAT were weak (R = 0.27), moderate (R = 0.55) and strong (R = 0.61), respectively. Low penetrance p.C282Y/p.C282Y phenotype displays hepcidin dysregulation and biochemical risk for iron toxicity.

Extracorporeal Photopheresis With Low-Dose Immunosuppression in High-Risk Heart Transplant Patients-A Pilot Study.

In severely ill patients undergoing urgent heart transplant (HTX), immunosuppression carries high risks of infection, malignancy, and death. Low-dose immunosuppressive protocols have higher rejection rates. We combined extracorporeal photopheresis (ECP), an established therapy for acute rejection, with reduced-intensity immunosuppression. Twenty-eight high-risk patients (13 with high risk of infection due to infection at the time of transplant, 7 bridging to transplant via extracorporeal membrane oxygenation, 8 with high risk of malignancy) were treated, without induction therapy. Prophylactic ECP for 6 months (24 procedures) was initiated immediately postoperatively. Immunosuppression consisted of low-dose tacrolimus (8-10 ng/ml, months 1-6; 5-8 ng/ml, >6 months) with delayed start; mycophenolate mofetil (MMF); and low maintenance steroid with delayed start (POD 7) and tapering in the first year. One-year survival was 88.5%. Three patients died from infection (POD 12, 51, 351), and one from recurrence of cancer (POD 400). Incidence of severe infection was 17.9% (n = 5, respiratory tract). Within the first year, antibody-mediated rejection was detected in one patient (3.6%) and acute cellular rejection in four (14.3%). ECP with reduced-intensity immunosuppression is safe and effective in avoiding allograft rejection in HTX recipients with risk of severe infection or cancer recurrence.

Using the World Apheresis Association Registry Helps to Improve the Treatment Quality of Therapeutic Apheresis.

Therapeutic apheresis (TA) is prescribed to patients that suffer from a severe progressive disease that is not sufficiently treated by conventional medications. A way to gain more knowledge about this treatment is usually by the local analysis of data. However, the use of large quality assessment registries enables analyses of even rare findings. Here, we report some of the recent data from the World Apheresis Association (WAA) registry. Data from >104,000 procedures were documented, and TA was performed on >15,000 patients. The main indication for TA was the collection of autologous stem cells (45% of patients) as part of therapy for therapy. Collection of stem cells from donors for allogeneic transplantation was performed in 11% of patients. Patients with indications such as neurological diseases underwent plasma exchange (28%). Extracorporeal photochemotherapy, lipid apheresis, and antibody removal were other indications. Side effects recorded in the registry have decreased significantly over the years, with approximately only 10/10,000 procedures being interrupted for medical reasons. CONCLUSION: Collection of data from TA procedures within a multinational and multicenter concept facilitates the improvement of treatment by enabling the analysis of and feedback on indications, procedures, effects, and side effects.

A comprehensive antigen production and characterisation study for easy-to-implement, specific and quantitative SARS-CoV-2 serotests.

BACKGROUND: Antibody tests are essential tools to investigate humoral immunity following SARS-CoV-2 infection or vaccination. While first-generation antibody tests have primarily provided qualitative results, accurate seroprevalence studies and tracking of antibody levels over time require highly specific, sensitive and quantitative test setups. METHODS: We have developed two quantitative, easy-to-implement SARS-CoV-2 antibody tests, based on the spike receptor binding domain and the nucleocapsid protein. Comprehensive evaluation of antigens from several biotechnological platforms enabled the identification of superior antigen designs for reliable serodiagnostic. Cut-off modelling based on unprecedented large and heterogeneous multicentric validation cohorts allowed us to define optimal thresholds for the tests' broad applications in different aspects of clinical use, such as seroprevalence studies and convalescent plasma donor qualification. FINDINGS: Both developed serotests individually performed similarly-well as fully-automated CE-marked test systems. Our described sensitivity-improved orthogonal test approach assures highest specificity (99.8%); thereby enabling robust serodiagnosis in low-prevalence settings with simple test formats. The inclusion of a calibrator permits accurate quantitative monitoring of antibody concentrations in samples collected at different time points during the acute and convalescent phase of COVID-19 and disclosed antibody level thresholds that correlate well with robust neutralization of authentic SARS-CoV-2 virus. INTERPRETATION: We demonstrate that antigen source and purity strongly impact serotest performance. Comprehensive biotechnology-assisted selection of antigens and in-depth characterisation of the assays allowed us to overcome limitations of simple ELISA-based antibody test formats based on chromometric reporters, to yield comparable assay performance as fully-automated platforms. FUNDING: WWTF, Project No. COV20-016; BOKU, LBI/LBG.

De novo Vessel Formation Through Cross-Talk of Blood-Derived Cells and Mesenchymal Stromal Cells in the Absence of Pre-existing Vascular Structures.

BACKGROUND: The generation of functional blood vessels remains a key challenge for regenerative medicine. Optimized in vitro culture set-ups mimicking the in vivo perivascular niche environment during tissue repair may provide information about the biological function and contribution of progenitor cells to postnatal vasculogenesis, thereby enhancing their therapeutic potential. AIM: We established a fibrin-based xeno-free human 3D in vitro vascular niche model to study the interaction of mesenchymal stromal cells (MSC) with peripheral blood mononuclear cells (PBMC) including circulating progenitor cells in the absence of endothelial cells (EC), and to investigate the contribution of this cross-talk to neo-vessel formation. MATERIALS AND METHODS: Bone marrow-derived MSC were co-cultured with whole PBMC, enriched monocytes (Mo), enriched T cells, and Mo together with T cells, respectively, obtained from leukocyte reduction chambers generated during the process of single-donor platelet apheresis. Cells were embedded in 3D fibrin matrices, using exclusively human-derived culture components without external growth factors. Cytokine secretion was analyzed in supernatants of 3D cultures by cytokine array, vascular endothelial growth factor (VEGF) secretion was quantified by ELISA. Cellular and structural re-arrangements were characterized by immunofluorescence and confocal laser-scanning microscopy of topographically intact 3D fibrin gels. RESULTS: 3D co-cultures of MSC with PBMC, and enriched Mo together with enriched T cells, respectively, generated, within 2 weeks, complex CD31+/CD34+ vascular structures, surrounded by basement membrane collagen type-IV+ cells and matrix, in association with increased VEGF secretion. PBMC contained CD31+CD34+CD45dimCD14- progenitor-type cells, and EC of neo-vessels were PBMC-derived. Vascular structures showed intraluminal CD45+ cells that underwent apoptosis thereby creating a lumen. Cross-talk of MSC with enriched Mo provided a pro-angiogenic paracrine environment. MSC co-cultured with enriched T cells formed "cell-in-cell" structures generated through internalization of T cells by CD31+CD45 dim⁣/ - cells. No vascular structures were detected in co-cultures of MSC with either Mo or T cells. CONCLUSION: Our xeno-free 3D in vitro vascular niche model demonstrates that a complex synergistic network of cellular, extracellular and paracrine cross-talk can contribute to de novo vascular development through self-organization via co-operation of immune cells with blood-derived progenitor cells and MSC, and thereby may open a new perspective for advanced vascular tissue engineering in regenerative medicine.

Vorinostat in the acute neuroinflammatory form of X-linked adrenoleukodystrophy.

OBJECTIVE: To identify a pharmacological compound targeting macrophages, the most affected immune cells in inflammatory X-linked adrenoleukodystrophy (cerebral X-ALD) caused by ABCD1 mutations and involved in the success of hematopoietic stem cell transplantation and gene therapy. METHODS: A comparative database analysis elucidated the epigenetic repressing mechanism of the related ABCD2 gene in macrophages and identified the histone deacetylase (HDAC) inhibitor Vorinostat as a compound to induce ABCD2 in these cells to compensate for ABCD1 deficiency. In these cells, we investigated ABCD2 and pro-inflammatory gene expression, restoration of defective peroxisomal ß-oxidation activity, accumulation of very long-chain fatty acids (VLCFAs) and their differentiation status. We investigated ABCD2 and pro-inflammatory gene expression, restoration of defective peroxisomal ß-oxidation activity, accumulation of very long-chain fatty acids (VLCFA) and differentiation status. Three advanced cerebral X-ALD patients received Vorinostat and CSF and MRI diagnostics was carried out in one patient after 80 days of treatment. RESULTS: Vorinostat improved the metabolic defects in X-ALD macrophages by stimulating ABCD2 expression, peroxisomal ß-oxidation, and ameliorating VLCFA accumulation. Vorinostat interfered with pro-inflammatory skewing of X-ALD macrophages by correcting IL12B expression and further reducing monocyte differentiation. Vorinostat normalized the albumin and immunoglobulin CSF-serum ratios, but not gadolinium enhancement upon 80 days of treatment. INTERPRETATION: The beneficial effects of HDAC inhibitors on macrophages in X-ALD and the improvement of the blood-CSF/blood-brain barrier are encouraging for future investigations. In contrast with Vorinostat, less toxic macrophage-specific HDAC inhibitors might improve also the clinical state of X-ALD patients with advanced inflammatory demyelination.

The assessment of platelet function by thromboelastometry as a point-of-care test to guide Intercept-treated platelet support in hemato-oncological patients and hematopoietic stem cell transplantation recipients.

BACKGROUND: Pathogen inactivation (PI) techniques for platelet concentrates (PCs) are one of the latest innovations to improve blood safety and reduce the risk of transfusion-transmitted infections (TTIs). An impaired function and in vivo recovery of platelets as well as an increased PC demand are concerns regarding these techniques. The intent of this study was to evaluate the hemostatic effect of PCs treated with the Intercept™ System by thromboelastometry (TEM) and to assess the clinical validity of its results in comparison to post-transfusion increase (PTI) and corrected count increment (CCI). STUDY-DESIGN AND METHODS: This prospective-observational study included 47 patients (m:f = 25:22; median age: 54 years [21-70]) of our Bone Marrow Transplantation unit with hemato-oncological malignancies transfused with Intercept™-treated PCs. Serial TEM measurements were performed just before and 1 hour after PC transfusion and were analyzed for their correlation with PTI and CCI as well as for clinical variables. RESULTS: The majority of our patients had received a hematopoietic stem cell transplantation (HSCT) (n = 41; 87%). In median 9 (1-50) PCs were transfused. Serial TEM, PTI, and CCI measurements were available for 150 transfusion episodes. The median platelet dose transfused was 2.65 × 1011 /unit (1.8-6). The median CCI was 9.250 (0-28.000). We observed a significant improvement in TEM parameters (p < 0.05) after transfusion of PI PCs, which did not mandatory correlate with the 1-hour PTI and CCI. CONCLUSION: Serial TEM measurements indicate the hemostatic effect of Intercept™-treated PCs. The 1-hour PTI and CCI may not appropriately reflect the in vivo function of platelets after PI PC transfusion.

The impact of ABO blood type on the prevalence of portal vein thrombosis in patients with advanced chronic liver disease.

BACKGROUND AND AIMS: Non-O blood type (BT) is a risk factor for thromboses, which has been attributed to its effects on von Willebrand factor (VWF)/factor VIII (FVIII) levels. Although high VWF/FVIII may be risk factors for portal vein thrombosis (PVT) in patients with advanced chronic liver disease (ACLD), the impact of BT on PVT is unknown. We aimed to assess (I) whether non-O-BT is a risk factor for PVT and (II) whether non-O-BT impacts VWF/factor VIII in patients with ACLD. METHODS: Retrospective analysis comprising two cohorts: (I) "US" including all adult liver transplantations in the US in the MELD era and (II) "Vienna" comprising patients with a hepatic venous pressure gradient (HVPG) ≥6 mmHg. RESULTS: (I) The "US cohort" included 84 947 patients (non-O: 55.43%). The prevalence of PVT at the time of listing (4.37% vs 4.56%; P = .1762) and at liver transplantation (9.56% vs 9.33%; P = .2546) was similar in patients with O- and non-O-BT. (II) 411 patients were included in the "Vienna cohort" (non-O: 64%). Mean HVPG was 18(9) mmHg and 90% had an HVPG ≥10 mmHg. Patients with non-O-BT had slightly increased VWF levels (318(164)% vs 309(176)%; P = .048; increase of 23.8%-23.9% in adjusted analyses), but this difference was driven by patients with less advanced disease. However, non-O-BT explained only 1% of the variation in VWF and had no effect on FVIII. CONCLUSIONS: Although non-O-BT impacts VWF in patients with early stage ACLD, its contribution to VWF variation is considerably smaller than in the general population. Moreover, non-O-BT had no impact on FVIII. These findings may explain the absence of an association between non-O-BT and PVT in patients with advanced cirrhosis.

Transfusion of standard-issue packed red blood cells induces pulmonary vasoconstriction in critically ill patients after cardiac surgery-A randomized, double-blinded, clinical trial.

BACKGROUND: Experimental and volunteer studies have reported pulmonary vasoconstriction during transfusion of packed red blood cells (PRBCs) stored for prolonged periods. The primary aim of this study was to evaluate whether transfusion of PRBCs stored over 21 days (standard-issue, siPRBCs) increases pulmonary artery pressure (PAP) to a greater extent than transfusion of PRBCs stored for less then 14 days (fresh, fPRBCs) in critically ill patients following cardiac surgery. The key secondary aim was to assess whether the pulmonary vascular resistance index (PVRI) increases after transfusion of siPRBCs to a greater extent than after transfusion of fPRBCs. METHODS: The study was performed as a single-center, double-blinded, parallel-group, randomized clinical trial. Leukoreduced PRBCs were transfused while continuously measuring hemodynamic parameters. Systemic concentrations of syndecan-1 were measured to assess glycocalyx injury. After randomizing 19 patients between January 2014 and June 2016, the study was stopped due to protracted patient recruitment. RESULTS: Of 19 randomized patients, 11 patients were transfused and included in statistical analyses. Eight patients were excluded prior to transfusion, 6 patients received fPRBCs (10±3 storage days), whereas 5 patients received siPRBCs (33±4 storage days). The increase in PAP (7±3 vs. 2±2 mmHg, P = 0.012) was greater during transfusion of siPRBCs than during transfusion of fPRBCs. In addition, the change in PVRI (150±89 vs. -4±37 dyn·s·cm-5·m2, P = 0.018) was greater after transfusion of siPRBCs than after transfusion of fPRBCs. The increase in PAP correlated with the change of systemic syndecan-1 concentrations at the end of transfusion (R = 0.64,P = 0.034). CONCLUSION: Although this study is underpowered and results require verification in larger clinical trials, our findings suggest that transfusion of siPRBCs increases PAP and PVRI to a greater extent than transfusion of fPRBCs in critically ill patients following cardiac surgery. Glycocalyx injury might contribute to pulmonary vasoconstriction associated with transfusion of stored blood.

Time from venipuncture to cell isolation: Impact on granulocyte-reactive antibody testing.

BACKGROUND AND OBJECTIVES: Classical neutrophil-reactive antibody testing depends on the quick isolation of neutrophils from freshly taken whole blood. To allow a better logistic preparation before testing, the influence of time interval between venipuncture and cell isolation has been evaluated in this study. MATERIALS AND METHODS: Neutrophils and whole leukocytes were isolated from EDTA whole blood immediately (T0) as well as 4, 8 and 24 h after blood donation (T4, T8 and T24). These cells were tested against reference sera containing antibodies against HNA-1b, -2, -3a and HLA class I using granulocyte aggregation test (GAT), microscopic granulocyte immunofluorescence test (GIFT) and flow-cytometric white blood cell immunofluorescence test (Flow-GIFT/WIFT). RESULTS: GAT was the most error-prone test displaying overall weaker aggregation strengths already at T4 (overall accuracy OA = 0.72, κ = 0.58). GIFT results showed good agreement at T4 (OA = 0.86, κ = 0.79) and remained stable until T8, while test results were slightly impaired at T24 (OA = 0.71, κ = 0.55). Flow-GIFT/WIFT was identified as the most robust screening method, remaining stable even at T24. Calculated ratios (sample/negative control) decreased non-significantly and remained highly above the cut-off in all samples. CONCLUSION: Acceptable time limits for cell isolation are different for each screening method investigated. For GAT, cell isolation should be performed within 4 h, while GIFT tolerates a neutrophil isolation delay of 8 h. Flow-GIFT/WIFT isolation can be performed even after 24 h without impairment of the results. Using the latter test as a stand-alone pre-screening test, whole blood can be used from donors who are not directly accessible.

Donor Safety in Haemapheresis: Development of an Internet-Based Registry for Comprehensive Assessment of Adverse Events from Healthy Donors.

BACKGROUND: Currently, there is an extensive but highly inconsistent body of literature regarding donor adverse events (AEs) in haemapheresis. As the reports diverge with respect to types and grading of AEs, apheresis procedures and machines, the range of haemapheresis-related AEs varies widely from about 0.03% to 6.6%. METHODS: The German Society for Transfusion Medicine and Immunohaematology (DGTI) formed a 'Haemapheresis Vigilance Working Party' (Arbeitsgemeinschaft Hämapheresevigilanz; AGHV) to create an on-line registry for comprehensive and comparable AE assessment with all available apheresis devices in all types of preparative haemapheresis: plasmapheresis (PLS), plateletpheresis (PLT), red blood cell apheresis, all kind of leukaphereses (autologous/allogeneic blood stem cell apheresis, granulocyte apheresis, lymphocyte/monocyte apheresis) and all possible types of multi-component apheresis. To ensure the comparability of the data, the AGHV adopted the 'Standard for Surveillance of Complications Related to Blood Donation' from the International Society for Blood Transfusion in cooperation with the International Haemovigilance Network (IHN) and the American Association of Blood Banks for AE acquisition and automated evaluation. The registry is embedded in a prospective observational multi-centre study with a study period of 7 years. RESULTS: A preliminary evaluation encompassed the time period from January, 2012 to December, 2015. During this time, the system proved to be safe and stable. Out of approximately 345,000 haemaphereses 16,477 AEs were reported (4.9%) from 20 participating centres. The majority of AEs occurred in PLSs (63%), followed by PLT (34.5%) and SC (2.2%). Blood access injuries (BAI) accounted for about 55% of the supplied AEs, whereas citrate toxicity symptoms, vasovagal reactions and technical events (e.g. disposable leakages, software failures) rather equally affected haemaphereses at 8-15%. Out of 12,348 finalized AEs, 8,759 (70.1%) were associated with a procedure-related break-off, with BAI being the prevailing cause (5,463/8,759; 62.4%). An automated centre- and procedure-specific AE evaluation according to the latest IHN standard and AGHV pre-settings is available within a few minutes. CONCLUSIONS: An on-line electronic platform for comprehensive assessment and centre-specific automated evaluation of AEs in haemaphereses has been developed and proved to be stable and safe over a period of 4 years.

Microparticle detection to guide platelet management for the reduction of platelet refractoriness in children - A study proposal.

Microparticles have been shown to shed from a variety of viable cells as a consequence of inflammatory processes, activation or physical stress. Seventy to 90% of circulating microparticles are thought to be platelet-derived. The content of microparticles in blood collected from normal blood donors is highly variable and transfers into the final blood component. Elevated microparticle content (MPC) in donor blood might indicate an asymptomatic clinical condition of the donor which might affect the transfusion recipient, particularly pediatric patients. ThromboLUX is a new technology designed to routinely test biological samples for microparticle content. We compared MPC in platelet-rich plasma (PRP) of apheresis donors and the corresponding INTERCEPT-treated apheresis products (N=24). The MPCs in donor and product samples were correlated (r=0.74, P<0.001). Microparticles were significantly reduced after plasma replacement and INTERCEPT treatment. These findings are supported by phase contrast microscopy. Platelet transfusions given to patients with fever or systemic inflammation are less efficacious. In addition, transfusing heterogeneous platelets - concentrates with high MPC and activated platelets - to patients whose immune systems are activated might tip them over a threshold and cause platelet refractoriness. Restricting prophylactic platelet transfusions to homogeneous products - concentrates with resting platelets and therefore low MPC - may reduce the risk of refractoriness in cancer patients, especially children with immature immunity. To test this hypothesis we introduce an evaluation protocol for platelet management, i.e., keeping a split inventory of homogeneous and heterogeneous platelets, and using only homogeneous platelets for prophylaxis as a strategy to reduce refractoriness.

Plerixafor as preemptive strategy results in high success rates in autologous stem cell mobilization failure.

Plerixafor in combination with granulocyte-colony stimulating factor (G-CSF) is approved for autologous stem cell mobilization in poor mobilizing patients with multiple myeloma or malignant lymphoma. The purpose of this study was to evaluate efficacy and safety of plerixafor in an immediate rescue approach, administrated subsequently to G-CSF alone or chemotherapy and G-CSF in patients at risk for mobilization failure. Eighty-five patients mobilized with G-CSF alone or chemotherapy were included. Primary endpoint was the efficacy of the immediate rescue approach of plerixafor to achieve ≥2.0 × 106 CD34+ cells/kg for a single or ≥5 × 106 CD34+ cells/kg for a double transplantation and potential differences between G-CSF and chemotherapy-based mobilization. Secondary objectives included comparison of stem cell graft composition including CD34+ cell and lymphocyte subsets with regard to the mobilization regimen applied. No significant adverse events were recorded. A median 3.9-fold increase in CD34+ cells following plerixafor was observed, resulting in 97% patients achieving at least ≥2 × 106 CD34+ cells/kg. Significantly more differentiated granulocyte and monocyte forming myeloid progenitors were collected after chemomobilization whereas more CD19+ and natural killer cells were collected after G-CSF. Fifty-two patients underwent transplantation showing rapid and durable engraftment, irrespectively of the stem cell mobilization regimen used. The addition of plerixafor in an immediate rescue model is efficient and safe after both, G-CSF and chemomobilization and results in extremely high success rates. Whether the differences in graft composition have a clinical impact on engraftment kinetics, immunologic recovery, and graft durability have to be analysed in larger prospective studies.

Complete neurologic and cognitive recovery after plasmapheresis in a patient with chronic inflammatory demyelinating polyneuropathy after allogeneic hematopoietic stem cell transplantation.

UNLABELLED: Neurologic complications after allogeneic hematopoietic stem cell transplantation (HSCT) are rare but poorly understood. We present a case report of a 57-year-old-male patient who was diagnosed in 2009 with acute myeloid leukemia (AML). He received two standard induction chemotherapies, as well as a following consolidation. Six months later, an allogeneic HSCT was performed. Shortly after HSCT the patient developed progressive polyneuropathy of the lower legs and hypoesthesia. Five months later a severe dementia followed. All images of the brain and spine showed no specific pathologies. High dose corticosteroids and immunoglobulins did not improve the neurologic symptoms. Due to severe worsening of the neuropsychiatric status and the clinical presentation, chronic inflammatory demyelinating polyneuropathy (CIDP) was suspected. Therefore, the patient received ten cycles of plasmapheresis. The patient showed a significant improvement of the neuropsychiatric symptoms and cognitive status. CONCLUSIONS: Immune mediated neuropathies after allogeneic HSCT, such as CIDP, have great variability in symptoms and presentation and are challenging to diagnose and treat. Plasmapheresis is a safe and efficient treatment for patients with unclear persisting autoimmune neuropathy after HSCT.

Pharmacological- and non-pharmacological therapeutic approaches in inflammatory bowel disease in adults.

Inflammatory bowel diseases (IBDs) are a group of chronic inflammatory conditions mainly of the colon and small intestine. Crohn's disease (CD) and ulcerative colitis (UC) are the most frequent types of IBD. IBD is a complex disease which arises as a result of the interaction of environmental, genetic and immunological factors. It is increasingly thought that alterations of immunological reactions of the patients to their own enterable bacteria (microfilm) may contribute to inflammation. It is characterized by mucosal and sub mucosal inflammation, perpetuated by infiltration of activated leukocytes. CD may affect the whole gastrointestinal tract while UC only attacks the large intestine. The therapeutic goal is to achieve a steroid-free long lasting remission in both entities. UC has the possibility to be cured by a total colectomy, while CD never can be cured by any operation. A lifelong intake of drugs is mostly necessary and essential. Medical treatment of IBD has to be individualized to each patient and usually starts with anti-inflammatory drugs. The choice what kind of drugs and what route administered (oral, rectal, intravenous) depends on factors including the type, the localization, and severity of the patient's disease. IBD may require immune-suppression to control symptoms such as prednisolone, thiopurines, calcineurin or sometimes folic acid inhibitors or biologics like TNF-α inhibitors or anti-integrin antibodies. For both types of disease (CD, UC) the same drugs are available but they differ in their preference in efficacy between CD and UC as 5-aminosalicylic acid for UC or budesonide for ileocecal CD. As therapeutic alternative the main mediators of the disease, namely the activated pro-inflammatory cytokine producing leukocytes can be selectively removed via two apheresis systems (Adacolumn and Cellsorba) in steroid-refractory or dependent cases. Extracorporeal photopheresis results in an increase of regulatory B cells, regulatory CD8(+) T cells and T-regs Type 1. Both types of apheresis were able to induce clinical remission and mucosal healing accompanied by tapering of steroids.