Preoperative platelet transfusions to reverse antiplatelet therapy for urgent non-cardiac surgery: an observational cohort study.

Essentials An increasing number of patients requiring surgery receive antiplatelet therapy (APT). We analyzed 181 patients receiving presurgery platelet transfusions to reverse APT. No coronary thrombosis occurred after platelet transfusion. This justifies a prospective trial to test preoperative platelet transfusions to reverse APT. SUMMARY: Background Patients receiving antiplatelet therapy (APT) have an increased risk of perioperative bleeding and cardiac adverse events (CAE). Preoperative platelet transfusions may reduce the bleeding risk but may also increase the risk of CAE, particularly coronary thrombosis in patients after recent stent implantation. Objectives To analyze the incidence of perioperative CAE and bleeding in patients undergoing non-cardiac surgery using a standardized management of transfusing two platelet concentrates preoperatively and restart of APT within 24-72 h after surgery. Methods A cohort of consecutive patients on APT treated with two platelet concentrates before non-cardiac surgery between January 2012 and December 2014 was retrospectively identified. Patients were stratified by the risk of major adverse cardiac and cerebrovascular events (MACCE). The primary objective was the incidence of CAE (myocardial infarction, acute heart failure and cardiac troponine T increase). Secondary objectives were incidences of other thromboembolic events, bleedings, transfusions and mortality. Results Among 181 patients, 88 received aspirin, 21 clopidogrel and 72 dual APT. MACCE risk was high in 63, moderate in 103 and low in 15 patients; 67 had cardiac stents. Ten patients (5.5%; 95% CI, 3.0-9.9%) developed a CAE (three myocardial infarctions, four cardiac failures and three troponin T increases). None was caused by coronary thrombosis. Surgery-related bleeding occurred in 22 patients (12.2%; 95% CI, 8.2-17.7%), making 12 re-interventions necessary (6.6%; 95% CI, 3.8-11.2%). Conclusion Preoperative platelet transfusions and early restart of APT allowed urgent surgery and did not cause coronary thromboses, but non-thrombotic CAEs and re-bleeding occurred. Randomized trials are warranted to test platelet transfusion against other management strategies.

Platelets kill bacteria by bridging innate and adaptive immunity via platelet factor 4 and FcγRIIA.

Essentials Human platelets specifically interact with IgG opsonized bacteria through FcγRIIA. Platelet factor 4 (PF4) binds to polyanions (P) and undergoes a conformational change. Anti-PF4/P IgG opsonizes PF4-coated Gram-positive and Gram-negative bacteria. Platelets specifically kill E.coli opsonized with PF4 and human anti-PF4/P IgG. SUMMARY: Background Activated platelets release the chemokine platelet factor 4 (PF4) stored in their granules. PF4 binds to polyanions (P) on bacteria, undergoes a conformational change and exposes neoepitopes. These neoepitopes induce production of anti-PF4/P antibodies. As PF4 binds to a variety of bacteria, anti-PF4/P IgG can bind and opsonize several bacterial species. Objective Here we investigated whether platelets are able to kill bacteria directly after recognizing anti-PF4/P IgG opsonized bacteria in the presence of PF4 via their FcγRIIA. Methods Using platelet-bacteria suspension co-culture experiments and micropatterns with immobilized viable bacteria, in combination with pharmacological inhibitors and human anti- PF4/P IgG we analyzed the role of platelet-mediated killing of bacteria. Results In the presence of PF4, human anti-PF4/P IgG and platelets, E. coli killing (> 50%) with colony forming units (CFU mL-1 ) 0.71 × 104 ± 0.19 was observed compared with controls incubated only with anti-PF4/P IgG (CFU mL-1 3.4 × 104 ± 0.38). Blocking of platelet FcγRIIA using mAb IV.3 (CFU mL-1 2.5 × 104 ± 0.45), or integrin αIIbß3 (CFU mL-1 2.26 × 104 ± 0.31), or disruption of cytoskeletal functions (CFU mL-1 2.7 × 104 ± 0.4) markedly reduced E. coli killing by this mechanism. Our observation of E. coli killing by platelets on micropatterned arrays is compatible with the model that platelets kill bacteria by covering them, actively concentrating them into the area under their granulomere and then releasing antimicrobial substances of platelet α-granules site directed towards bacteria. Conclusion These findings collectively indicate that by bridging of innate and adaptive immune mechanisms, platelets and anti-PF4/polyanion antibodies cooperate in an antibacterial host response.

An open conformation of ADAMTS-13 is a hallmark of acute acquired thrombotic thrombocytopenic purpura.

Essentials Conformational changes in ADAMTS-13 are part of its mode-of-action. The murine anti-ADAMTS-13 antibody 1C4 discriminates between folded and open ADAMTS-13. ADAMTS-13 conformation is open in acute acquired thrombotic thrombocytopenic purpura (TTP). Our study forms an important basis to fully elucidate the pathophysiology of TTP. SUMMARY: Background Acquired thrombotic thrombocytopenic purpura (aTTP) is an autoimmune disorder characterized by absent ADAMTS-13 activity and the presence of anti-ADAMTS-13 autoantibodies. Recently, it was shown that ADAMTS-13 adopts a folded or an open conformation. Objectives As conformational changes in self-antigens play a role in the pathophysiology of different autoimmune diseases, we hypothesized that the conformation of ADAMTS-13 changes during acute aTTP. Methods Antibodies recognizing cryptic epitopes in the spacer domain were generated. Next, the conformation of ADAMTS-13 in 40 healthy donors (HDs), 99 aTTP patients (63 in the acute phase versus 36 in remission), 12 hemolytic-uremic syndrome (HUS) patients and 63 sepsis patients was determined with ELISA. Results The antibody 1C4 recognizes a cryptic epitope in ADAMTS-13. Therefore, we were able to discriminate between a folded and an open ADAMTS-13 conformation. We showed that ADAMTS-13 in HDs does not bind to 1C4, indicating that ADAMTS-13 circulates in a folded conformation. Similar results were obtained for HUS and sepsis patients. In contrast, ADAMTS-13 of acute aTTP patients bound to 1C4 in 92% of the cases, whereas, in most cases, this binding was abolished during remission, showing that the conformation of ADAMTS-13 is open during an acute aTTP episode. Conclusions Our study shows that, besides absent ADAMTS-13 activity and the presence of anti-ADAMTS-13 autoantibodies, an open ADAMTS-13 conformation is also a hallmark of acute aTTP. Demonstrating this altered ADAMTS-13 conformation in acute aTTP will help to further unravel the pathophysiology of aTTP and lead to improved therapy and diagnosis.

Motivational factors for blood donation in first-time donors and repeat donors: a cross-sectional study in West Pomerania.

OBJECTIVES: This study aimed to analyse motivational factors for blood donation in different donor groups. BACKGROUND: As the demographic change will result in a decrease of the population in age groups of blood donors, the risk of blood product shortage increases. METHODS: During a 12-month period, every sixth blood donor presenting at the blood donation centre of the University Hospital was asked to complete a self-administered questionnaire assessing motivational factors for blood donation. Despite the formalised enrolment protocol, frequent donors were over-represented in the study cohort, which was adjusted by weighting donors with different numbers of donations per year in such a way that the distribution of numbers of donations per year was the same in the sample as in the donor population. RESULTS: Of 2443 participants, 14·3% were first-time and 85·3% repeat donors. To "help other people" (>90%) and receiving "medical assessment of my blood values" (63-69%) were the strongest motivational factors in all donor groups. Receiving remuneration (49·2% vs 38·1%) was more important for repeat donors than for first-time donors, whereas it was the opposite for "being taken by a friend to the donor clinic" (47·0% vs 15·5%). A potentially important observation is that 33·9% of frequent donors reported feeling physically better after blood donation compared to infrequent donors (29·5%). CONCLUSION: Identification of motivational factors can lead to the design of targeted motivation campaigns for blood donation. The underlying cause of the perceived well-being after blood donation requires further studies.

Platelet transfusion to reverse antiplatelet therapy before decompressive surgery in patients with intracranial haemorrhage.

BACKGROUND: Platelet concentrates (PC) are transfused to improve primary haemostasis before urgent neurosurgery in patients with intracranial haemorrhage (ICH) receiving antiplatelet therapy (APT). It is unresolved, whether PCs increase the risk for major cardio- and cerebrovascular adverse events. We evaluated a standardized transfusion regimen to reverse APT in patients with ICH who required decompressive neurosurgery. METHODS: Analysed were consecutive patients between 2012 and 2014. The primary outcome was the frequency of new arterial thrombotic complications. The secondary outcome was the frequency of recurrent ICH. RESULTS: Of 72 patients, 14 received acetylsalicylic acid and a P2Y12 inhibitor, 53 received acetylsalicylic acid and five clopidogrel. No acute coronary syndrome (95% CI: 0-5·07) and one ischaemic stroke occurred (1·4%; 95% CI: 0·25-7·46). In contrast, 26·4% of patients developed recurrent ICH (95% CI: 17·59-37·58). The risk of bleeding was significantly higher compared to the risk of arterial thrombosis (P < 0·00001) and was increased for patients with chronic ICH (OR: 4·78; 95% CI: 1·57-14·55) and those receiving clopidogrel (OR: 2·78; 95% CI: 0·90-8·57). CONCLUSION: Platelet concentrate transfusion before cranial decompressive surgery in patients with ICH complicating APT showed a low risk for cardio-cerebral thrombotic complications. However, the risk of rebleeding remains high, especially in patients with chronic ICH and those pretreated with clopidogrel.

Autoimmune heparin-induced thrombocytopenia.

Autoimmune heparin-induced thrombocytopenia (aHIT) indicates the presence in patients of anti-platelet factor 4 (PF4)-polyanion antibodies that are able to activate platelets strongly even in the absence of heparin (heparin-independent platelet activation). Nevertheless, as seen with serum obtained from patients with otherwise typical heparin-induced thrombocytopenia (HIT), serum-induced platelet activation is inhibited at high heparin concentrations (10-100 IU mL-1 heparin). Furthermore, upon serial dilution, aHIT serum will usually show heparin-dependent platelet activation. Clinical syndromes associated with aHIT include: delayed-onset HIT, persisting HIT, spontaneous HIT syndrome, fondaparinux-associated HIT, heparin 'flush'-induced HIT, and severe HIT (platelet count of < 20 × 109  L-1 ) with associated disseminated intravascular coagulation (DIC). Recent studies have implicated anti-PF4 antibodies that are able to bridge two PF4 tetramers even in the absence of heparin, probably facilitated by non-heparin platelet-associated polyanions (chondroitin sulfate and polyphosphates); nascent PF4-aHIT-IgG complexes recruit additional heparin-dependent HIT antibodies, leading to the formation of large multimolecular immune complexes and marked platelet activation. aHIT can persist for several weeks, and serial fibrin, D-dimer, and fibrinogen levels, rather than the platelet count, may be helpful for monitoring treatment response. Although standard anticoagulant therapy for HIT ought to be effective, published experience indicates frequent failure of activated partial thromboplastin time (APTT)-adjusted anticoagulants (argatroban, bivalirudin), probably because of underdosing in the setting of HIT-associated DIC, known as 'APTT confounding'. Thus, non-APTT-adjusted therapies with drugs such as danaparoid and fondaparinux, or even direct oral anticoagulants, such as rivaroxaban or apixaban, are suggested therapies, especially for long-term management of persisting HIT. In addition, emerging data indicate that high-dose intravenous immunoglobulin can interrupt HIT antibody-induced platelet activation, leading to rapid platelet count recovery.

Diagnosis of inherited platelet disorders on a blood smear: a tool to facilitate worldwide diagnosis of platelet disorders.

Essentials There are many hereditary platelet disorders (HPD) but diagnosing these is challenging. We provide a method to diagnose several HPDs using standard blood smears requiring < 100 µL blood. By this approach, the underlying cause of HPD was characterized in ~25-30% of referred individuals. The method facilitates diagnosis of HPD for patients of all ages around the world. SUMMARY: Background Many hereditary thrombocytopenias and/or platelet function disorders have been identified, but diagnosis of these conditions remains challenging. Diagnostic laboratory techniques are available only in a few specialized centers and, using fresh blood, often require the patient to travel long distances. For the same reasons, patients living in developing countries usually have limited access to diagnosis. Further, the required amount of blood is often prohibitive for pediatric patients. Objectives By a collaborative international approach of four centers, we aimed to overcome these limitations by developing a method using blood smears prepared from less than 100 µL blood, for a systematic diagnostic approach to characterize the platelet phenotype. Methods We applied immunofluorescence labelling (performed centrally) to standard air-dried peripheral blood smears (prepared locally, shipped by regular mail), using antibodies specific for proteins known to be affected in specific hereditary platelet disorders. Results By immunofluorescence labelling of blood smears we characterized the underlying cause in 877/3217 (27%) patients with suspected hereditary platelet disorders (HPD). Currently about 50 genetic causes for HPD are identified. Among those, the blood smear method was especially helpful to identify MYH9 disorders/MYH9-related disease, biallelic Bernard-Soulier syndrome, Glanzmann thrombasthenia and gray platelet syndrome. Diagnosis could be established for GATA1 macrothrombocytopenia, GFI1B macrothrombocytopenia, ß1-tubulin macrothrombocytopenia, filamin A-related thrombocytopenia and Wiskott-Aldrich syndrome. Conclusion Combining basic and widely available preanalytical methods with the immunomorphological techniques presented here, allows detailed characterization of the platelet phenotype. This supports genetic testing and facilitates diagnosis of hereditary platelet disorders for patients of all ages around the world.

[Recurrent thromboembolisms despite full anticoagulation in a patient with antiphospholipid syndrome]. / Rezidivierende Thrombembolien trotz Vollantikoagulation bei Patientin mit Antiphospholipidsyndrom.

We report on a female patient with confirmed secondary antiphospholipid syndrome (APS) due to underlying systemic lupus erythematosus (SLE). Despite a thromboplastin time within the normal range (international normalized ratio, INR) under treatment with a vitamin K antagonist (VKA), a recurrent thrombotic event occurred, this time as pulmonary embolism due to bilateral deep vein thrombosis. Despite an INR value in the therapeutic range, clotting factors II, VII, IX and X were found to be insufficiently decreased suggesting inefficient anticoagulation. Thus, the anticoagulation regimen was changed to the direct oral anticoagulant dabigatran. This case demonstrates that the INR in APS patients may be artificially prolonged in rare cases, despite a normal activated partial thromboplastin time (aPTT) and cannot be used for monitoring VKA anticoagulant therapy. Suspicion of ineffective anticoagulation despite VKA therapy should prompt measurement of the individual clotting factors.

Patients with Bernard-Soulier syndrome and different severity of the bleeding phenotype.

Bernard-Soulier syndrome is a rare (1:1million), hereditary bleeding disorder caused by defects of the platelet GPIb-IX-V complex. Patients suffer from mucocutaneous bleedings. Typical are thrombocytopenia, giant platelets and impaired agglutination after stimulation with ristocetin. In populations in which consanguineous marriages are common the frequency of the disorder is increased because Bernard-Soulier syndrome is mostly inherited autosomal recessively. Genetic analyses of the disease-related genes may help to gain more insights regarding the phenotype/genotype correlation. Here, we investigated several patients with Bernard-Soulier syndrome from different families. We analyzed two patients with severe bleeding symptoms from one family of middle east origin and confirmed the diagnosis by identifying a pathogenic variant in GP1BB. We compared phenotype/genotype correlation of this GP1BB mutation with the GP9 (p.Asn61Ser) European founder mutation present in 9 patients out of 4 families for whom we also performed molecular genetic analysis.

Platelet activation in the presence of neutral protamine Hagedorn insulin: a new feature of antibodies against protamine/heparin complexes.

Essentials Protamine (PRT) is used to stabilize insulin in neutral protamine Hagedorn (NPH) insulin. The interaction between NPH-insulin, anti-PRT/heparin antibodies and platelets was investigated. Anti-PRT/heparin antibodies activate platelets in presence of NPH-insulin dependent on heparin. Cross-reactivity seems to have no major effect on the clinical outcome of medical patients. SUMMARY: Background Protamine (PRT) is used to stabilize insulin in neutral protamine Hagedorn (NPH) insulin, a commonly used therapeutic agent for diabetes mellitus. Immunization against PRT/heparin complexes is common in diabetic patients. Objectives To investigate the impact of NPH-insulin on the interaction between anti-PRT/heparin antibodies and platelets. Methods The interaction between NPH-insulin and anti-PRT/heparin antibodies was tested using in-house enzyme immunoassays. The ability of anti-PRT/heparin antibodies to activate platelets in the presence of NPH-insulin (and heparin) was investigated using flow cytometry. Results Twenty-one out of 80 sera containing anti-PRT/heparin IgG showed binding to NPH-insulin. Anti-PRT/heparin IgG from immunized patients bound to platelets in the presence of NPH-insulin, but not in the presence of native insulin. Anti-PRT/heparin antibodies induced P-selectin expression in the presence of NPH-insulin in a heparin-dependent way (median mean fluorescence intensity in the presence of NPH-insulin: 55, 95% confidence interval [CI] 18.7-100.5 vs. NPH-insulin and heparin: 204, 95% CI 106.5-372.8). The clinical relevance of platelet-activating anti-PRT/heparin antibodies was assessed by investigating a multicenter study cohort of 332 acutely ill medical patients who received heparin. None of the 21 patients with anti-PRT/heparin IgG developed thrombocytopenia or thromboembolic complications. Conclusions Anti-PRT/heparin antibodies activate platelets in the presence of NPH-insulin in a heparin-dependent way. However, results from our preliminary study indicate no major impact of these antibodies on the clinical outcome in medical patients receiving heparin, particularly on thromboembolic complications.

Implications of a switch to a 100% apheresis platelet supply for patients and for blood donors: a risk benefit analysis.

BACKGROUND: A 100% apheresis platelet supply is considered to increase transfusion safety by lowering donor exposures for transfusion recipients. We performed a risk benefit analysis to contrast the reduction of donor exposures and the risk of contaminated blood products in the nation-wide inventory with the donor risks associated with a switch to a 100% apheresis platelet supply in Germany. METHODS: Donor exposures and the number of contaminated blood products resulting from HIV-like, HBV-like, HCV-like pathogens and two theoretical agents with infection rates of 10 and 1000 in 100 000, respectively, were calculated for a 100% apheresis platelet supply in Germany based on the 2006-2012 hemovigilance reports. These numbers were compared with the current mixed platelet supply of pooled and apheresis platelets. Moreover, additional donation time and apheresis donor complications resulting from a 100% apheresis platelet supply were estimated. RESULTS: Per million total blood products (red cells, platelets, fresh frozen plasma), a 100% apheresis platelet supply would reduce donor exposures by 87 100 and the number of contaminated blood products ranging from 0·8 to 871·1. On the other hand, this requires additional 29 478 apheresis donations, 3·4 years additional donor time, and would be associated with 58 additional donor complications, respectively. CONCLUSIONS: A 100% apheresis platelet supply would reduce donor exposures and the number of contaminated blood products in the inventory, but would increase apheresis complications in donors. Potential risks for patients must be carefully weighed against the risks for donors, dependent on the specific pathogen scenario.

Predonation finger lancet punctures: a potential risk factor for interdonor pathogen transmission in the blood donor clinic.

BACKGROUND AND OBJECTIVES: Point-of-care testing using capillary blood from a finger prick is widely used for predonation haemoglobin testing of blood donors. It is common practice to cover the finger prick with a cotton swab and to instruct the donor to press for few minutes. The finger prick can cause blood contamination of surfaces in contact with the lanced finger, especially door handles, risking infectious disease transmission, particularly if another person touching the contaminated door handle also has a punctured fingertip. MATERIALS AND METHODS: First, we investigated contamination by blood (benzidine assay) of the door handles of our blood donor clinic, taking 175 samples 3 h after opening of the donation centre (baseline). We then introduced band-aids to cover the finger prick and started an information campaign using educational flyers to sensitize blood donors and staff to this problem (period-1). Thereafter, the staff was instructed to use the non-dominant hand for blood sampling and mandated to replace any discarded band-aids immediately (period-2). RESULTS: At baseline, 82% of the nurse room door handles showed contamination with blood. This decreased somewhat (10-40%) after period-1, but only after immediate mandatory band-aid replacement on any donor finger without a band-aid (period-2), no further blood contaminations were detected. CONCLUSION: Blood contamination of shared surfaces can occur after finger prick for capillary blood sampling. Application of a band-aid and use of the non-dominant hand for fingertip incision are easy to apply and effective in reducing this iatrogenic health hazard.