Flow Cytometric Analysis of Mechanically Disaggregated Bone Marrow Trephine Biopsies.

BACKGROUND: Bone marrow (BM) aspirate analysis by flow cytometry (FC) is a key hematopathologic technique but dry taps hinder diagnosis. We describe the utility and limitations of a method of mechanically disaggregating BM trephine biopsies for FC. METHOD: Trephine biopsies mechanically disaggregated for FC between 2010 and 2016 were reviewed. We recorded cell yield, pathological findings, and turnaround time. Where available, results of trephine FC were compared with aspirate FC. RESULTS: Eighty BM trephine samples were processed, representing 4.3% of BM biopsies. Mechanical disaggregation yielded cellular samples in 98% of cases (median yield 3 × 106 nucleated cells). The most frequent diagnoses were B cell lymphoproliferative disorders (n = 26) and acute leukemia (n = 18). Trephine FC correlated with histochemistry findings in 94% of cases, but two Hodgkin lymphoma infiltrates were missed by FC, and trephine FC underestimated B cell infiltrate burden compared to immunohistochemistry (IHC). Fluorescence intensity of CD34 and CD45 on disease populations was similar in processed trephine and contemporaneous aspirate samples. Trephine FC results were available 2 days earlier than stained IHC slides. CONCLUSION: Mechanical disaggregation of BM trephine samples provided a cellular suspension suitable for diagnostic FC in most cases. Limitations were similar to aspirate FC: disease burden was underestimated and some infiltrates were missed. Trephine FC results were available earlier than trephine IHC. We conclude that trephine FC is a useful technique to complement trephine IHC in the event of a failed aspirate, providing rapid diagnostic immunophenotyping. © 2018 International Clinical Cytometry Society.

Glycolipid-peptide conjugate vaccines enhance CD8+ T cell responses against human viral proteins.

An important goal of vaccination against viruses and virus-driven cancers is to elicit cytotoxic CD8+ T cells specific for virus-derived peptides. CD8+ T cell responses can be enhanced by engaging help from natural killer T (NKT) cells. We have produced synthetic vaccines that induce strong peptide-specific CD8+ T cell responses in vivo by incorporating an NKT cell-activating glycolipid. Here we examine the effect of a glycolipid-peptide conjugate vaccine incorporating an NKT cell-activating glycolipid linked to an MHC class I-restricted peptide from a viral antigen in human peripheral blood mononuclear cells. The vaccine induces CD1d-dependent activation of human NKT cells following enzymatic cleavage, activates human dendritic cells in an NKT-cell dependent manner, and generates a pool of activated antigen-specific CD8+ T cells with cytotoxic potential. Compared to unconjugated peptide, the vaccine upregulates expression of genes encoding interferon-γ, CD137 and granzyme B. A similar vaccine incorporating a peptide from the clinically-relevant human papilloma virus (HPV) 16 E7 oncoprotein induces cytotoxicity against peptide-expressing targets in vivo, and elicits a better antitumor response in a model of E7-expressing lung cancer than its unconjugated components. Glycolipid-peptide conjugate vaccines may prove useful for the prevention or treatment of viral infections and tumors that express viral antigens.

Consensus guidelines for antifungal prophylaxis in haematological malignancy and haemopoietic stem cell transplantation, 2014.

There is a strong argument for the use of antifungal prophylaxis in high-risk patients given the significant mortality associated with invasive fungal disease, the late identification of these infections, and the availability of safe and well-tolerated prophylactic medications. Clinical decisions about which patients should receive prophylaxis and choice of antifungal agent should be guided by risk stratification, knowledge of local fungal epidemiology, the efficacy and tolerability profile of available agents, and estimates such as number needed to treat and number needed to harm. There have been substantial changes in practice since the 2008 guidelines were published. These include the availability of new medications and/or formulations, and a focus on refining and simplifying patient risk stratification. Used in context, these guidelines aim to assist clinicians in providing optimal preventive care to this vulnerable patient demographic.

Imipenem versus piperacillin/tazobactam for empiric treatment of neutropenic fever in adults.

Australian guidelines for neutropenic fever recommend piperacillin/tazobactam (PIP-TAZ) or cefepime for first-line empiric treatment of neutropenic fever. We compared outcomes among haematology patients before and after changing our first-line neutropenic fever treatment from imipenem to PIP-TAZ. Forty-five patients received imipenem and 60 PIP-TAZ. Despite a higher rate of antibiotic modification in the PIP-TAZ cohort, treatment success and time to defervescence were similar, with a trend towards fewer Clostridium difficile infections in the PIP-TAZ cohort.

Fibrinogen concentrate for acquired hypofibrinogenaemic states.

This study aims to assess the efficacy and safety of fibrinogen concentrate in acquired hypofibrinogenaemic states. Cryoprecipitate is standard treatment for replacement of fibrinogen in acquired hypofibrinogenaemia. A virally inactivated fibrinogen concentrate (Haemocomplettan((R)); CSL Behring, Marburg, Germany) is licensed in some European countries. Clinical data for its use in acquired hypofibrinogenaemic states are scarce. Demographic and pretreatment clinical data of patients treated with fibrinogen concentrate for acquired hypofibrinogenaemia were retrospectively reviewed. Pretreatment and posttreatment fibrinogen levels, transfusion requirements, outcomes and adverse events were recorded. Thirty adult patients who received fibrinogen concentrate for acquired hypofibrinogenaemia (fibrinogen <1.5 g L(-1)) were included in the study. Causes of hypofibrinogenaemia included placental abruption, disseminated intravascular coagulation as a result of massive blood loss and transfusion, liver failure and cardiac surgery. Following a median dose of 4 g fibrinogen concentrate, median Clauss fibrinogen level rose from 0.65 to 2.01 g L(-1), with a median fibrinogen increment of 0.25 g L(-1) per 1 g fibrinogen concentrate administered. Forty-six per cent of patients stopped bleeding with blood components and fibrinogen concentrate alone, and a further 29% stopped bleeding with surgical or endoscopic intervention. Inpatient mortality was 40%. No venous thromboses were observed. Four patients with massive perioperative haemorrhage and hypotension (including three postcardiothoracic surgery) had arterial ischaemic events, none of which was attributable to fibrinogen overreplacement. The cost of fibrinogen concentrate was comparable with that of cryoprecipitate. Purified, virally inactivated fibrinogen concentrate appears effective in the management of acquired hypofibrinogenaemic states and enables rapid administration of a known fibrinogen dose in an emergency setting.

Techniques for predicting a favourable response to renal angioplasty in patients with renovascular disease.

Renovascular disease is present in some 10-40% of patients with end-stage renal disease, and constitutes the fastest-growing group of end-stage renal disease patients. The unselective correction of renal artery stenosis has led to disappointing results. Most studies that compared conservative treatment with angioplasty found only modest or no beneficial effects of angioplasty on renal function and blood pressure. It is therefore mandatory to evaluate the functional significance of a stenosis before intervention. Patients with a high likelihood of a favourable response should be identified. Factors that affect outcome include the severity of renal artery stenosis, the procedure used to treat renal artery stenosis (antihypertensive drugs, angioplasty with or without stenting, or surgery), radiocontrast nephrotoxicity, atheroembolism and, most importantly, underlying renal disease, forestalling a favourable response of renal function or blood pressure even after the successful correction of renal artery stenosis. Evaluation of the renal resistance index using Doppler ultrasound or captopril scintigraphy are the best methods by which to classify patients as responders or non-responders to intervention. Each factor has to be considered before the correction of renal artery stenosis to achieve satisfactory results with regard to an improvement in renal function and blood pressure.