Andrew W. Shih

Haptoglobin testing in hemolysis: Measurement and interpretation

Haptoglobin is primarily produced in the liver and is functionally important for binding free hemoglobin from lysed red cells in vivo, preventing its toxic effects. Because haptoglobin levels become depleted in the presence of large amounts of free hemoglobin, decreased haptoglobin is a marker of hemolysis. Despite its ubiquity and importance, a paucity of literature makes testing difficult to interpret. This review highlights the many physiological roles that have been recently elucidated in the literature. Different methodologies have been developed for testing, including spectrophotometry, immunoreactive methods, and gel electrophoresis. These are covered along with their respective advantages and disadvantages. As there is no single gold standard for hemolysis, validation studies must rely on a combination of factors, which are reviewed in this article. Pitfalls and limitations of testing are also addressed. False positives can occur in improper specimen preparations, cirrhosis, elevated estrogen states, and hemodilution. False negatives can occur in hypersplenism and medications such as androgens and corticosteroids. Haptoglobin testing in the setting of inflammation is additionally discussed as interpretation can be difficult in this setting. Given the widespread use of haptoglobin testing, it is vital that clinicians and laboratory staff understand the principles and correct interpretation of this test. Am. J. Hematol. 89:443–447, 2014.

Novel treatments for immune thrombocytopenia

Primary immune thrombocytopenia (ITP) is caused by platelet autoantibodies and T-cell dysregulation. Both platelets and their precursor megakaryocytes may be targeted leading to platelet destruction and underproduction. Current treatments for ITP are inadequate since they do not reverse the disease process and generally do not result in durable remissions. In addition, many treatments are limited by side effects including infection and potentially thrombosis. Novel agents that are currently in development target certain key steps in the disease process, including: (1) the interaction between T-cell and antigen presenting cells (CD40-CD154 interaction); (2) the binding of the Fc portion of platelet autoantibodies to Fc-receptors on macrophages (soluble Fc-RIIb); and (3) the signaling pathways leading to platelet phagocytosis by macrophages (Syk inhibition). Other strategies have been to augment platelet production by simulating thrombopoiesis or by neutralizing physiological inhibitors of megakaryopoiesis. Targeted therapies in ITP have the potential to improve disease morbidity and mortality while limiting systemic side effects. Before these agents can be used in practice, additional clinical studies are needed with rational study outcomes including platelet count, bleeding and quality of life. An individualized treatment strategy is needed for patients since ITP is a distinctly heterogeneous disease.

Blood-Borne Pathogens: A Canadian Blood Services Centre for Innovation Symposium

Abstract Testing donations for pathogens and deferring selected blood donors have reduced the risk of transmission of known pathogens by transfusion to extremely low levels in most developed countries. Protecting the blood supply from emerging infectious threats remains a serious concern in the transfusion medicine community. Transfusion services can employ indirect measures such as surveillance, hemovigilance, and donor questioning (defense), protein-, or nucleic acid based direct testing (detection), or pathogen inactivation of blood products (destruction) as strategies to mitigate the risk of transmission-transmitted infection. In the North American context, emerging threats currently include dengue, chikungunya, and hepatitis E viruses, and Babesia protozoan parasites. The 2003 SARS and 2014 Ebola outbreaks illustrate the potential of epidemics unlikely to be transmitted by blood transfusion but disruptive to blood systems. Donor-free blood products such as ex vivo generated red blood cells offer a theoretical way to avoid transmission-transmitted infection risk, although biological, engineering, and manufacturing challenges must be overcome before this approach becomes practical. Similarly, next generation sequencing of all nucleic acid in a blood sample is currently possible but impractical for generalized screening. Pathogen inactivation systems are in use in different jurisdictions around the world, and are starting to gain regulatory approval in North America. Cost concerns make it likely that pathogen inactivation will be contemplated by blood operators through the lens of health economics and risk-based decision making, rather than in zero-risk paradigms previously embraced for transfusable products. Defense of the blood supply from infectious disease risk will continue to require innovative combinations of surveillance, detection, and pathogen avoidance or inactivation.

Quantification of Cell-Free DNA in Red Blood Cell Units in Different Whole Blood Processing Methods.

Background. Whole blood donations in Canada are processed by either the red cell filtration (RCF) or whole blood filtration (WBF) methods, where leukoreduction is potentially delayed in WBF. Fresh WBF red blood cells (RBCs) have been associated with increased in-hospital mortality after transfusion. Cell-free DNA (cfDNA) is released by neutrophils prior to leukoreduction, degraded during RBC storage, and is associated with adverse patient outcomes. We explored cfDNA levels in RBCs prepared by RCF and WBF and different storage durations. Methods. Equal numbers of fresh (stored ≤14 days) and older RBCs were sampled. cfDNA was quantified by spectrophotometry and PicoGreen. Separate regression models determined the association with processing method and storage duration and their interaction on cfDNA. Results. cfDNA in 120 RBC units (73 RCF, 47 WBF) were measured. Using PicoGreen, WBF units overall had higher cfDNA than RCF units (p = 0.0010); fresh WBF units had higher cfDNA than fresh RCF units (p = 0.0093). Using spectrophotometry, fresh RBC units overall had higher cfDNA than older units (p = 0.0031); fresh WBF RBCs had higher cfDNA than older RCF RBCs (p = 0.024). Conclusion. Higher cfDNA in fresh WBF was observed compared to older RCF blood. Further study is required for association with patient outcomes.

Evaluation of the appropriateness of frozen plasma usage after introduction of prothrombin complex concentrates: a retrospective study.

Prothrombin complex concentrates (PCCs) can be used instead of frozen plasma (FP) transfusion to reverse the effect of warfarin. Audits have demonstrated over usage of FP transfusions even before the introduction of PCC. The objective of this study was to determine the appropriateness of current FP transfusion practice in the current era since the introduction of PCCs.

Audit of provincial IVIG Request Forms and efficacy documentation in four Ontario tertiary care centres

Retrospective audit of IVIG Request Forms in four Ontario tertiary care centres: to determine the case mix of new IVIG requests, to authenticate information provided, and to determine documentation of clinical efficacy.

Best practices in the differential diagnosis and reporting of acute transfusion reactions

License. The full terms of the License are available at http://creativecommons.org/licenses/by-nc/3.0/. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. Permissions beyond the scope of the License are administered by Dove Medical Press Limited. Information on how to request permission may be found at: http://www.dovepress.com/permissions.php International Journal of Clinical Transfusion Medicine 2016:4 1–14 International Journal of Clinical Transfusion Medicine Dovepress

Impact of organizational interventions on reducing inappropriate intravenous immunoglobulin (IVIG) usage: A systematic review and meta-analysis

BACKGROUND With increasing global use of intravenous immunoglobulin (IVIG), there is interest in its appropriate usage. Efforts to regulate IVIG usage have primarily taken the form of organizational interventions implemented in hospitals to monitor and improve physician prescribing. Similar interventions have proven effective in reducing the inappropriate and total hospital usage of other blood products, but their efficacy on IVIG use is less understood. Thus, we performed a systematic review of studies reporting the change in inappropriate IVIG use following such interventions in hospitals or regions. METHODS A systematic search was carried out using MEDLINE and EMBASE (1966-June 2016) for English language studies if they 1) were primary research, 2) described an organizational intervention to target plasma, IVIG, or albumin, and 3) reported appropriateness of usage and total usage preand post-intervention. Review Manager v5.0 was utilized to perform a random-effects meta-analysis on eligible IVIG studies, where the risk ratio (RR) of inappropriate IVIG transfusion comparing pre- and postintervention periods was calculated with 95% confidence intervals (CI). RESULTS Our search retrieved three retrospective cohort studies, where metaanalysis encompassing 2100 episodes of IVIG transfusion demonstrated no decrease in inappropriate IVIG use (RR 1.55, 95% CI 0.78-3.07). Heterogeneity between studies was considerable (I2 = 89%). CONCLUSION Organizational interventions were ineffective at changing inappropriate IVIG use, but more high-quality studies describing the effects of these interventions are required before any conclusions can be drawn. Futureresearch efforts should also be directed at evolving evidence-based IVIGguidelines to improve patient safety and burdens on healthcare systems.

Intersecting Worlds of Transfusion and Transplantation Medicine: An International Symposium Organized by the Canadian Blood Services Centre for Innovation

The principal theme of the symposium was centered on how the world of regenerative medicine intersects with that of transfusion medicine, with a particular focus on hematopoietic stem cells (HSCs) and stem cell therapies. The symposium highlighted several exciting developments and identified areas where additional research is needed. A revised map of human hematopoietic hierarchy was presented based on the functional and phenotypic analysis of thousands of single stem and progenitor cells from adult bone marrow and fetal liver. These analyses revealed that multipotency is largely restricted to the HSC and multipotent progenitor compartments in adult bone marrow where most progenitors are unipotent, whereas fetal liver contains a large number of distinct oligopotent progenitors. Furthermore, unlike adult bone marrow, multipotency is extended in the downstream progenitors in the hierarchy in the fetal liver stage. Production of platelets ex vivo from HSCs is emerging as a potentially viable option because of advances in culture techniques that combine cytokine mixtures, small molecules, and shear stress. However, limited HSC expansion and low platelet yield from culture-derived megakaryocytes remain problematic. Evidence was presented to support stricter guidelines for transfusion of platelets and red blood cells practices in allogeneic HSC transplant patients, although evidence is often extrapolated from general indications. Basic principles of human leukocyte antigen testing in HSC transplant were described, emphasizing the need for a national (and global) stem cell donor registry. Ongoing research is aimed at improving cellular cryopreservation including the establishment of a new thawing protocol that improves viability of umbilical cord blood CD34+ cells. Umbilical cord blood transplantation practices have also been improved; recent studies suggest noninferior outcomes when patients are transplanted with umbilical cord blood vs a matched adult donor. Finally, mesenchymal stem cell infusion is an example of a cellular therapy useful for immunomodulation. Preclinical trials suggest that mesenchymal stem cells may be effective in managing sepsis. In conclusion, practices and research surrounding HSCs are continuing to evolve rapidly as new information is obtained.

Plasma transfusion trials and tribulations

F rozen plasma (FP) transfusions are commonly used to correct clotting factor deficiencies especially among critically ill patients. In such “coagulopathic” patients with bleeding, FP transfusions are justified to restore hemostasis and avoid excess morbidity or mortality. However, most FP transfusions are administered to prevent, rather than to treat, bleeding, typically in patients with an upcoming invasive procedure. In that setting, it remains unclear whether FP transfusions make any difference. Complicating matters further is the way in which intensive care unit (ICU)-acquired factor deficiencies and, in turn, response to FP transfusions, are measured. The international normalized ratio (INR) has been misused for so long that its interpretation has become obscure. Developed as a measure of the anticoagulant effect of warfarin, the INR is a standardized way of reporting the prothrombin time (PT) normalized for different types of PT reagents used across laboratories. The INR has excellent correlation with effective warfarin dose, but it may not accurately assess global impairments in hemostasis due to liver disease, sepsis, or coagulation factor consumption, which are so commonly encountered in critically ill patients. Equipoise exists around the use of prophylactic FP transfusion to correct a prolonged INR in preparation for an invasive procedure: On the one hand, it may reduce bleeding and improve clinical outcomes; on the other hand, it is associated with a risk of infection, allergic reactions, transfusion-associated circulatory overload, transfusion-associated lung injury, and multiorgan failure. The pressing need for randomized controlled trials (RCTs) in this area has been recognized for some time; however, logistic and methodologic challenges including difficulties with blinding, patient selection, and buy-in have hampered progress. The RCT by Müller and colleagues, published in this issue of TRANSFUSION was a brave attempt to address this enduring and important clinical question. In this trial, ICU patients with a mild INR elevation (1.5-3.0) were randomly assigned to receive or not receive FP transfusion before a planned invasive procedure (central venous catheter placement, percutaneous tracheostomy, chest tube, or abscess drainage procedures). The primary outcome was procedure-related bleeding. The trial faced several challenges that ultimately resulted in its early termination. Enrollment was slow because of strict eligibility criteria. The screening process was complex and involved identifying potential participants with an INR that was within study range, applying exclusion criteria, and capturing those patients for whom a procedure was planned. On top of that, a significant proportion of patients were missed, refused to participate, or were denied participation by their treating physician. Thus, 81 of 263 eligible patients (30.8%) with a planned procedure, or 81 of 1478 patients (5.5%) with a suitable INR, were ultimately randomized over 3 years across four centers. Because of a low event rate, the investigators changed the primary outcome from major procedure-related bleeding to procedure-related bleeding of any severity. The initial sample size was calculated based on an estimated event rate of major procedure-related bleeding of 1%; however, estimates of bleeding rates in patients with an elevated INR who undergo invasive procedures have not been well established due to heterogeneous definitions of coagulopathy and bleeding and diverse patient populations. After 81 patients were enrolled (of the planned 400), one major bleed had occurred. While this event was consistent with their prediction, this low event rate was too small to conduct the planned inferiority analysis. In their final conclusion, the authors reported that there was no difference in bleeding complications with or without FP; however, the true meaning of these results is uncertain since most bleeds were minor and the post hoc analysis was underpowered. ICU trials are no easy task, because of the high acuity of illness among the participants, frequent reliance on substitute decision makers for consent, and rotating staff caring for the patients. Moreover, FP transfusion practice is habitual, often rooted in tradition and associated with strong beliefs, making its omission potentially unpalatable for clinicians even in the context of an RCT. A previous RCT using prophylactic FP transfusion to prevent hepatobiliary procedure-related bleeding was also stopped early because of difficulties with accrual. Thus, many of these challenges could have been anticipated by the investigators, but the extent to which these and other barriers would impact the trial’s success was underestimated. Ultimately, the trial was truncated after only one-fifth of the target sample was enrolled thus leaving us still uncertain as to whether withholding FP transfusion is no worse than giving it. Incomplete trials also present ethical dilemmas since they expose patients to potential risks without the