Challenging the dogma: Red blood cell-directed autoimmunity as risk factor for red blood cell alloimmunisation after blood transfusion.

Red blood cell autoimmunity and alloimmunity are potentially linked. Quantification of this association can tailor extensively matched red blood cell transfusions in patients with autoimmunity. Using an incident new-user cohort comprising 47 285 previously non-transfused, non-alloimmunised patients, we compared transfusion-induced red blood cell alloimmunisation incidences in direct antiglobulin test (DAT)-positive and control patients. Additionally, we performed case-control analyses to handle potential confounding by clinical immunomodulators. Among (IgG and/or C3d) DAT-positive patients (N = 380), cumulative red blood cell alloimmunisation incidences after 10 units transfused reached 4.5% (95% confidence interval [CI] 2.5-8.2) versus 4.2% (CI 3.9-4.5, p = 0.88) in controls. In case-control analyses, alloimmunisation relative risks among DAT-positive patients increased to 1.7 (CI 1.1-2.8). Additional adjustments for pre-DAT transfusion exposure or the extent of Rh/K mismatching did not impact results. In conclusion, while patients with DAT positivity show an intrinsically increased alloimmune red blood cell response, their absolute risk is comparable to control patients due to counteracting co-existing immunosuppressive conditions. Consequently, isolated DAT positivity in patients lacking overt haemolysis or complicated alloantibody testing does not seem to warrant extended matching strategies.

Transfusion of ever-pregnant donor red blood cells and mortality of male patients.

Previous studies found exposure to red blood cell transfusions from female donors who have been pregnant reduces survival in male patients compared to exposure to male donor products, but evidence is not consistent. We postulate the previously observed association is modified by offspring sex, with an expected increased mortality risk for male patients receiving units from female donors with sons. Here, marginal structural models were used to assess the association between exposure to units from ever-pregnant donors, ever-pregnant donors with sons and ever-pregnant donors with daughters, and mortality. Clinical data were collected on first-ever transfusion recipients in the Netherlands and donor data were supplemented with information about offspring sex and date of birth. In this analysis, 56,825 patients were included, of whom 8,288 died during follow-up. Exposure to red blood cell units from everpregnant donors with sons was not associated with increased all-cause mortality risk among male transfusion recipients (hazard ratio [HR] 0.91, 95% confidence interval 0.83-1.01). Exposure to ever-pregnant donors, irrespective of offspring sex, was associated with mortality in male patients aged between 18 and 50 years (ever-pregnant donors: HR 1.81, 95% CI 1.31-2.51) compared to male donor units, but was protective in female patients. This study suggests that the observed increased mortality risk for exposure to red blood cell units from parous female donors does not depend on offspring sex. The increased risk of mortality seen in younger adult male patients is consistent with previous observations, but the underlying biological mechanism could not be identified in this study.

Donor pregnancies and transfusion recipient mortality: A role for red blood cell storage?

BACKGROUND AND OBJECTIVES: Donor characteristics have been implicated in transfusion-related adverse events. Uncertainty remains about whether sex, and specifically pregnancy history of the blood donor, could affect patient outcomes. Whether storage duration of the blood product could be important for patient outcomes has also been investigated, and a small detrimental effect of fresh products remains a possibility. Here, we hypothesize that fresh red blood cell products donated by ever-pregnant donors are associated with mortality in male patients. MATERIALS AND METHODS: We used data from a cohort study of adult patients receiving a first transfusion between 2005 and 2015 in the Netherlands. The risk of death after receiving a transfusion from one of five exposure categories (female never-pregnant stored ≤10 days, female never-pregnant stored >10 days, female ever-pregnant stored ≤10 days, female ever-pregnant stored >10 days and male stored for ≤10 days), compared to receiving a unit donated by a male donor, which was stored for >10 days (reference), was calculated using a Cox proportional hazards model. RESULTS: The study included 42,456 patients who contributed 88,538 person-years in total, of whom 13,948 died during the follow-up of the study (33%). Fresh units (stored for ≤10 days) from ever-pregnant donors were associated with mortality in male patients, but the association was not statistically significant (hazard ratio 1.39, 95% confidence interval 0.97-1.99). Sensitivity analyses did not corroborate this finding. CONCLUSION: These findings do not consistently support the notion that the observed association between ever-pregnant donor units and mortality is mediated by blood product storage.

Routine Lupus Anticoagulant Sensitive aPTT Testing Can Prevent Unnecessary LA Testing.

Even though routine screening of the general hospital population is discouraged, medical laboratories may use a "lupus sensitive" activated partial thromboplastin time test (aPTT) with phospholipid concentrations that are susceptible to inhibition by lupus anticoagulant (LA), to screen for the presence of LA. If deemed necessary, follow-up testing according to ISTH guidelines may be performed. However, LA testing is a laborious and time-consuming effort that is often not readily available due to a lack of automation and/or temporary unavailability of experienced staff. In contrast, the aPTT is a fully automated test that is available 24/7 in almost all medical laboratories and is easily interpreted with the use of reference ranges. In addition to clinical signs, the result of an LA sensitive aPTT may thus be used to lower the suspicion of the presence of LA and reduce costly follow-up testing. In this study, we show that a normal LA sensitive aPTT result may be safely used to refrain from LA testing in the absence of strong clinical suspicion.

The effect of extended c, E and K matching in females under 45 years of age on the incidence of transfusion-induced red blood cell alloimmunisation.

Maternal alloantibodies directed against fetal red blood cell (RBC) antigens may cause potentially life-threatening haemolytic disease of the fetus and newborn (HDFN). Dutch transfusion guidelines therefore prescribe preventive cEK matching for all (pre-)fertile females. To quantify the impact of cEK matching, we compared overall and antigen-specific cumulative RBC alloimmunisation incidences in females and males aged <45 years. Among a multicentre cohort comprised of patients who received their first and subsequent RBC unit between 2005 and 2019, first-formed RBC alloantibodies were detected in 47 of 2998 (1·6%) females and 49 of 2507 (2·0%) males. Comparing females and males, overall alloimmunisation incidences were comparable (3·1% [95% confidence interval (CI) 2·1-4·4] versus 3·5% (95% CI 2·4-4·9, P = 0·853) after 10 units transfused). However, cEK alloimmunisation incidences were significantly lower among females (0·6% (95% CI 0·3-1.5) versus 2·2% (95% CI 1·5-3·4, P = 0·001) after 10 units transfused). Yet, despite cEK-matching guidelines being in effect, 6·5%, 3·6% and 0·2% of all RBC units remained mismatched for c, E or K antigens respectively. Most of these mismatches were almost always due to emergency settings. Even though cEK alloimmunisation was not prevented completely, implementation of cEK matching resulted in an alloantigen-exposure risk reduction of up to 98%.

Association between renal failure and red blood cell alloimmunization among newly transfused patients.

BACKGROUND: Renal failure and renal replacement therapy (RRT) affect the immune system and could therefore modulate red blood cell (RBC) alloimmunization after transfusion. STUDY DESIGN AND METHODS: We performed a nationwide multicenter case-control study within a source population of newly transfused patients between 2005 and 2015. Using conditional multivariate logistic regression, we compared first-time transfusion-induced RBC alloantibody formers (N = 505) with two nonalloimmunized recipients with similar transfusion burden (N = 1010). RESULTS: Renal failure was observed in 17% of the control and 13% of the case patients. A total of 41% of the control patients and 34% of case patients underwent acute RRT. Renal failure without RRT was associated with lower alloimmunization risks after blood transfusion (moderate renal failure: adjusted relative rate [RR], 0.82 [95% confidence interval (CI), 0.67-1.01]); severe renal failure, adjusted RR, 0.76 [95% CI, 0.55-1.05]). With severe renal failure patients mainly receiving RRT, the lowest alloimmunization risk was found in particularly these patients [adjusted RR 0.48 (95% CI 0.39-0.58)]. This was similar for patients receiving RRT for acute or chronic renal failure (adjusted RR, 0.59 [95% CI, 0.46-0.75]); and adjusted RR, 0.62 [95% CI 0.45-0.88], respectively). CONCLUSION: These findings are indicative of a weakened humoral response in acute as well as chronic renal failure, which appeared to be most pronounced when treated with RRT. Future research should focus on how renal failure and RRT mechanistically modulate RBC alloimmunization.

Lifetime Transfusion Burden and Transfusion-Related Iron Overload in Adult Survivors of Solid Malignancies.

BACKGROUND: Limited data exist on transfusion burden and transfusion-related iron overload in adult survivors of solid malignancies. METHODS: Hospital-specific cancer registry data of patients with solid tumor receiving systemic anticancer treatment between January 2008 and September 2009 at the Oncology Department of the Leiden University Medical Center (The Netherlands) were retrieved and cross-referenced with red blood cell (RBC) transfusion records. Individual lifetime transfusion burden was captured in April 2015. Multitransfused long-term survivors with serum ferritin >500 µg/L were subsequently screened for hepatic and cardiac iron overload using 1.5 Tesla magnetic resonance imaging. RESULTS: The study population consisted of 775 adult patients with solid cancer (45.2% male; median age, 58 years; >75% chemotherapy-treated), 423 (54.6%) of whom were transfused with a median of 6.0 RBC units (range 1-67). Transfusion triggers were symptomatic anemia or hemoglobin <8.1-8.9 g/dL prior to each myelosuppressive chemotherapy cycle. We identified 123 (15.9%) patients across all tumor types with a lifetime transfusion burden of ≥10 RBC units. In the absence of a hemovigilance program, none of these multitransfused patients was screened for iron overload despite a median survival of 4.6 years. In 2015 at disclosure of transfusion burden, 26 multitransfused patients were alive. Six (23.1%) had hepatic iron overload: 3.9-11.2 mg Fe/g dry weight. No cardiac iron depositions were found. CONCLUSION: Patients with solid malignancies are at risk for multitransfusion and iron overload even when adhering to restrictive RBC transfusion policies. With improved long-term cancer survivorship, increased awareness of iatrogenic side effects of supportive therapy and development of evidence-based guidelines are essential. IMPLICATIONS FOR PRACTICE: In the presence of a restrictive transfusion policy, ∼30% of transfused adult patients with solid cancer are multitransfused and ∼50% become long-term survivors, underscoring the need for evidence-based guidelines for the detection and management of transfusion-related iron overload in this group of patients. In each institution, a hemovigilance program should be implemented that captures the lifetime cumulative transfusion burden in all patients with cancer, irrespective of tumor type. This instrument will allow timely assessment and treatment of iron overload in cancer survivors, thus preventing organ dysfunction and decreased quality of life.

Age of platelet concentrates and time to the next transfusion.

BACKGROUND: Storage time of platelet (PLT) concentrates has been negatively associated with clinical efficacy outcomes. The aim of this study was to quantify the association between storage time of PLT concentrates and interval to the next PLT transfusion for different types of PLT components, stored for up to 7 days and transfused to transfusion-dependent hematooncology patients with thrombocytopenia. STUDY DESIGN AND METHODS: From a cohort of patients from 10 major Dutch hospitals, patients were selected whose transfusion patterns were compatible with PLT transfusion dependency due to hematooncologic disease. Mean time to the next transfusion and mean differences in time to the next transfusion for different storage time categories (i.e., fresh, <4 days; intermediate, 4-5 days; and old, >5 days) were estimated, per component type, using multilevel mixed-effects linear models. RESULTS: Among a cohort of 29,761 patients who received 140,896 PLT transfusions we selected 4441 hematooncology patients who had received 12,724 PLT transfusions during periods of PLT transfusion dependency. Transfusion of fresh, compared to old, buffy coat-derived PLTs in plasma was associated with a delay to the next transfusion of 6.2 hours (95% confidence interval [CI], 4.5-8.0 hr). For buffy coat-derived PLTs in PAS-B and -C this difference was 7.7 hours (95% CI, 2.2-13.3 hr) and 3.9 hours (95% CI, -2.1 to 9.9 hr) while for apheresis PLTs in plasma it was only 1.8 hours (95% CI, -3.5 to 7.1 hr). CONCLUSION: Our results indicate that the time to the next transfusion shortens with increasing age of transfused buffy coat-derived PLT concentrates. This association was not observed for apheresis PLTs.

Association of Blood Transfusion From Female Donors With and Without a History of Pregnancy With Mortality Among Male and Female Transfusion Recipients.

Importance: Transfusion of red blood cells from female donors has been associated with increased mortality in male recipients. Objective: To quantify the association between red blood cell transfusion from female donors with and without a history of pregnancy and mortality of red blood cell recipients. Design, Setting, and Participants: Retrospective cohort study of first-time transfusion recipients at 6 major Dutch hospitals enrolled from May 30, 2005, to September 1, 2015; the final follow-up date was September 1, 2015. The primary analysis was the no-donor-mixture cohort (ie, either all red blood cell transfusions exclusively from male donors, or all exclusively from female donors without a history of pregnancy, or all exclusively from female donors with a history of pregnancy). The association between mortality and exposure to transfusions from ever-pregnant or never-pregnant female donors was analyzed using life tables and time-varying Cox proportional hazards models. Exposures: Red blood cell transfusions from ever-pregnant or never-pregnant female donors, compared with red blood cell transfusions from male donors. Main Outcomes and Measures: All-cause mortality during follow-up. Results: The cohort for the primary analyses consisted of 31 118 patients (median age, 65 [interquartile range, 42-77] years; 52% female) who received 59 320 red blood cell transfusions exclusively from 1 of 3 types of donors (88% male; 6% ever-pregnant female; and 6% never-pregnant female). The number of deaths in this cohort was 3969 (13% mortality). For male recipients of red blood cell transfusions, all-cause mortality rates after a red blood cell transfusion from an ever-pregnant female donor vs male donor were 101 vs 80 deaths per 1000 person-years (time-dependent "per transfusion" hazard ratio [HR] for death, 1.13 [95% CI, 1.01-1.26]). For receipt of transfusion from a never-pregnant female donor vs male donor, mortality rates were 78 vs 80 deaths per 1000 person-years (HR, 0.93 [95% CI, 0.81-1.06]). Among female recipients of red blood cell transfusions, mortality rates for an ever-pregnant female donor vs male donor were 74 vs 62 per 1000 person-years (HR, 0.99 [95% CI, 0.87 to 1.13]); for a never-pregnant female donor vs male donor, mortality rates were 74 vs 62 per 1000 person-years (HR, 1.01 [95% CI, 0.88-1.15]). Conclusions and Relevance: Among patients who received red blood cell transfusions, receipt of a transfusion from an ever-pregnant female donor, compared with a male donor, was associated with increased all-cause mortality among male recipients but not among female recipients. Transfusions from never-pregnant female donors were not associated with increased mortality among male or female recipients. Further research is needed to replicate these findings, determine their clinical significance, and identify the underlying mechanism.

Fibrinogen determination according to Clauss: commutability assessment of International and commercial standards and quality control samples.

BACKGROUND: Many clinical laboratories use a clotting rate assay according to Clauss for the determination of fibrinogen in citrated plasma. The aim of the present study was to assess the commutability of the current International Standard for fibrinogen (coded 09/264), three commercial fibrinogen standards, and 10 freeze-dried plasma quality control samples from various sources. METHODS: Clotting rate assays according to Clauss were performed on three automated instruments (Sysmex CA1500, STA-Rack Evolution and ACL-Top 700), using three commercial thrombin reagents (Siemens, Stago, and Instrumentation Laboratory). Relationships between the results obtained with the three instruments were determined with 25 fresh-frozen plasma samples obtained from patients. The deviations of the assay results obtained with the freeze-dried samples were compared with the deviations obtained with the fresh-frozen samples, according to approved CLSI guideline C53A. RESULTS: Freezing and thawing had no influence on the assay results. There were significant differences in the mean assay results (fibrinogen, g/L) for the fresh-frozen plasma samples between the three automated instruments: 2.51 (STA-Rack Evolution), 2.25 (ACL-Top 700) and 2.20 (Sysmex CA1500). Similar differences were observed for several freeze-dried plasma samples. Some freeze-dried plasma samples, including the International Standard, were out of the 95% confidence interval for the relationship between STA-Rack Evolution and Sysmex CA1500. CONCLUSIONS: Some freeze-dried plasmas including the international standard for fibrinogen are not commutable among automated instruments for fibrinogen clotting rate assays according to Clauss. Our results have consequences for all interested parties in the traceability chain (WHO, industry, external quality assessment schemes, clinical laboratories).

Treatments for hematologic malignancies in contrast to those for solid cancers are associated with reduced red cell alloimmunization.

Red cell alloimmunization may induce severe hemolytic side effects. Identification of risk-modifying conditions will help tailor preventative strategies. This study aims to quantify the associations of hematologic malignancies and solid cancers with red cell alloimmunization in patients receiving red cell transfusions. We performed a nested multicenter case-control study in a source population of 24,063 patients receiving their first and subsequent red cell transfusions during an 8-year follow-up period. Cases (n=505), defined as patients developing a first transfusion-induced red cell alloantibody, were each compared with 2 non-alloimmunized controls (n=1010) who received a similar number of red cell units. Using multivariate logistic regression analyses, we evaluated the association of various malignancies and treatment regimens with alloimmunization during a delineated 5-week risk period. The incidence of alloimmunization among patients with acute (myeloid or lymphoid) leukemia and mature (B- or T-cell) lymphoma was significantly reduced compared to patients without these malignancies: adjusted relative risks (RR) with 95% confidence interval (CI) 0.36 (range 0.19-0.68) and 0.30 (range 0.12-0.81). Associations were primarily explained by immunosuppressive treatments [RR for (any type of) chemotherapy combined with immunotherapy 0.27 (95%CI: 0.09-0.83)]. Alloimmunization risks were similarly diminished in allogeneic or autologous stem cell transplanted patients (RR 0.34, 95%CI: 0.16-0.74), at least during the six months post transplant. Alloimmunization risks of patients with other hematologic diseases or solid cancers, and their associated treatment regimens were similar to risks in the general transfused population. Our findings suggest that, in contrast to malignancies in general, hemato-oncological patients treated with dose-intensive regimens have strongly diminished risk of red cell alloimmunization.

Red cell alloimmunisation in patients with different types of infections.

Red cell alloantigen exposure can cause alloantibody-associated morbidity. Murine models have suggested that inflammation modulates red cell alloimmunisation. This study quantifies alloimmunisation risks during infectious episodes in humans. We performed a multicentre case-control study within a source population of patients receiving their first and subsequent red cell transfusions during an 8-year follow-up period. Patients developing a first transfusion-induced red cell alloantibody (N = 505) were each compared with two similarly exposed, but non-alloimmunised controls (N = 1010) during a 5-week 'alloimmunisation risk period' using multivariate logistic regression analysis. Transfusions during 'severe' bacterial (tissue-invasive) infections were associated with increased risks of alloantibody development [adjusted relative risk (RR) 1·34, 95% confidence interval (95% CI) 0·97-1·85], especially when these infections were accompanied with long-standing fever (RR 3·06, 95% CI 1·57-5·96). Disseminated viral disorders demonstrated a trend towards increased risks (RR 2·41, 95% CI 0·89-6·53), in apparent contrast to a possible protection associated with Gram-negative bacteraemia (RR 0·58, 95% CI 0·13-1·14). 'Simple' bacterial infections, Gram-positive bacteraemia, fungal infections, maximum C-reactive protein values and leucocytosis were not associated with red cell alloimmunisation. These findings are consistent with murine models. Confirmatory research is needed before patients likely to develop alloantibodies may be identified based on their infectious conditions at time of transfusion.

Red-blood-cell alloimmunisation in relation to antigens' exposure and their immunogenicity: a cohort study.

BACKGROUND: Matching donor red blood cells based on recipient antigens prevents alloimmunisation. Knowledge about the immunogenicity of red-blood-cell antigens can help optimise risk-adapted matching strategies. We set out to assess the immunogenicity of red-blood-cell antigens. METHODS: In an incident new-user cohort of previously non-transfused, non-alloimmunised white patients receiving non-extended matched red-blood-cell transfusions in six Dutch hospitals between 2006 and 2013, we determined the cumulative number of mismatched red-blood-cell units per patient. We used multiple imputation to address missing antigen data. Using Kaplan-Meier analysis, we estimated cumulative alloimmunisation incidences per mismatched antigen dose as a measure of immunogenicity. FINDINGS: Of 54 347 patients assessed, 21 512 were included in our study. Alloantibodies occurred in 474 (2·2%) of all transfused patients, with cumulative alloimmunisation incidences increasing up to 7·7% (95% CI 4·9-11·2) after 40 units received. The antigens C, c, E, K, and Jk(a) were responsible for 78% of all alloimmunisations in our cohort. K, E, and C(w) were the most immunogenic antigens (cumulative immunisation incidences after 2 mismatched units of 2·3% [95% CI 1·0-4·8] for K, 1·5% [0·6-3·0] for E, and 1·2% [0·0-10·8] for C(w)). These antigens were 8·7 times (for K), 5·4 times (for E), and 4·6 times (for C(w)) as immunogenic as Fy(a). The next most immunogenic antigens were, in order, e (1·9 times as immunogenic as Fy(a)), Jk(a) (1·9 times), and c (1·6 times). INTERPRETATION: Red-blood-cell antigens vary in their potency to evoke a humoral immune response. Our findings highlight that donor-recipient red-blood-cell matching strategies will be most efficient when primarily focusing on prevention of C, c, E, K, and Jk(a) alloimmunisation. Matching for Fy(a) is of lower clinical relevance. Variations of antigen frequencies determined by ethnic background prevent extrapolating these conclusions to non-white populations. FUNDING: None.

Incidence of alloantibody formation after ABO-D or extended matched red blood cell transfusions: a randomized trial (MATCH study).

BACKGROUND: Most incidentally transfused patients receive only ABO-D-compatible transfusions and antibodies are formed in up to 8%. The effect of extended (c, C, E, K, Fy(a) , Jk(a) , and S antigens) matched (EM) and ABO-D-matched red blood cell (RBC) transfusions on the incidence of new clinically relevant RBC antibody formation after a first elective transfusion event in surgical patients was studied. STUDY DESIGN AND METHODS: A multicenter randomized trial was performed in nontransfused patients who were scheduled to experience a single elective transfusion event of maximal 4 RBC units. The primary outcome was the incidence of newly formed warm reacting clinically relevant RBC alloantibodies measured in three follow-up (FU) samples taken at 7 to 10 days, 4 to 6 weeks, and 4 to 6 months posttransfusion. RESULTS: A total of 853 patients were randomized, and of these, 333 patients were transfused with a total of 1035 RBC units. At least one FU sample was available from 97% of transfused patients. In intention-to-treat analysis, new antibodies were detected in 10 of 155 ABO-D and seven of 178 EM patients, respectively. Per-protocol analysis including 190 patients showed a nonsignificant absolute risk difference (ARD) of 5.3% (95% confidence interval [CI], -1.4% to 12%) in alloimmunization between study arms. In a post hoc analysis of 138 patients who received RBCs but no platelet (PLT) transfusions the ARD increased to significance, 8.0% (95% CI, 0.4-16.0). CONCLUSION: Extended matching for selected antigens reduced the alloimmunization risk by 64% in surgical patients. Extended matching seems successful only if the patient did not receive accompanying nonmatched PLT transfusions.

Successful treatment of a noninhibitory antibody-mediated acquired factor X deficiency in a patient with marginal-zone lymphoma.

Prolonged clotting times were observed in a patient with spontaneous hemorrhage. Analysis showed severe factor X deficiency due to clearance by a noninhibitory antibody. Lymphadenopathy identified on imaging led to diagnosis of marginal B-cell lymphoma. Treatment of lymphoma with rituximab and chlorambucil resulted in complete disappearance of the bleeding disorder.