Sequential monitoring of lymphocyte subsets and of T-and-B cell neogenesis indexes to identify time-varying immunologic profiles in relation to graft-versus-host disease and relapse after allogeneic stem cell transplantation

T and B lymphocyte subsets have been not univocally associated to Graft-versus-host disease (GVHD) and relapse of hematological alignancies after stem cell transplantation (SCT). Their sequential assessment together with B and T cell neogenesis indexes has been not thoroughly analysed in relation to these changing and interrelated immunologic/clinic events yet. Lymphocyte subsets in peripheral blood (PB) and B and T cell neogenesis indexes were analysed together at different time points in a prospective study of 50 patients. Principal component analysis (PCA) was used as first step of multivariate analysis to address issues related to a high number of variables versus a relatively low number of patients. Multivariate analysis was completed by Fine-Gray proportional hazard regression model. PCA identified 3 clusters of variables (PC1-3), which correlated with acute GVHD: PC1 (pre-SCT: KRECs 656608/ml, unswitched memory B 44%, CD8+TCM cells>4%; HR 1.9, p = 0.01), and PC3 (at aGVHD onset: CD4+TEMRA69%, switched memory CD19+ = 0 cells and KRECs<6614/ml at +90; HR 0.1, p = 0.008). All these immunologic parameters were independent indicators of chronic GVHD and relapse, also considering the possible effect of previous steroid-therapy for acute GVHD. Specific time-varying immunologic profiles were associated to GVHD and relapse. Pre-SCT host immune-microenvironment and changes of B cell homeostasis could influence GVH- and Graft-versus-Tumor reactions. The paradoxical increase of EM Treg in PB of patients with GVHD could be explained by their compartmentalization outside lymphoid tissues, which are of critical relevance for regulation of GVH reactions

Postremission sequential monitoring of minimal residual disease by WT1 Q-PCR and multiparametric flow cytometry assessment predicts relapse and may help to address risk-adapted therapy in acute myeloid leukemia patients

Risk stratification in acute myeloid leukemia (AML) patients using prognostic parameters at diagnosis is effective, but may be significantly improved by the use of on treatment parameters which better define the actual sensitivity to therapy in the single patient. Minimal residual disease (MRD) monitoring has been demonstrated crucial for the identification of AML patients at high risk of relapse, but the best method and timing of MRD detection are still discussed. Thus, we retrospectively analyzed 104 newly diagnosed AML patients, consecutively treated and monitored by quantitative polymerase chain reactions (Q-PCR) on WT1 and by multiparametric flow cytometry (MFC) on leukemia-associated immunophenotypes (LAIPs) at baseline, after induction, after 1st consolidation and after 1st intensification. By multivariate analysis, the factors independently associated with adverse relapse-free survival (RFS) were: bone marrow (BM)-WT1 ≥ 121/10(4) ABL copies (P = 0.02) and LAIP ≥ 0.2% (P = 0.0001) (after 1st consolidation) (RFS at the median follow up of 12.5 months: 51% vs. 82% [P < 0.0001] and 57% vs. 81%, respectively [P = 0.0003]) and PB-WT1 ≥ 16/10(4) ABL copies (P = 0.0001) (after 1st intensification) (RFS 43% vs. 95% [P < 0.0001]) Our data confirm the benefits of sequential MRD monitoring with both Q-PCR and MFC. If confirmed by further prospective trials, they may significantly improve the possibility of a risk-adapted, postinduction therapy of AML

NEUROLOGICAL COMPLICATIONS IN ADULT ALLOGENEIC HEMATOPOIETIC STEM CELL TRANSPLANT PATIENTS: RESULTS FROM A RETROSPECTIVE MULTICENTRE STUDY

Patients undergoing allogeneic hematopoietic stem cell transplant (HSCT) are exposed to a number of neurological complications that may be related to drugs, infections, metabolic alterations, cerebrovascular events and immune-\uadmediated disorders including myositis, myasthenia gravis, Guillain-\uadBarr\ue8-\uadlike demyelinating polyneuropathy and central nervous system (CNS) manifestations of graft versus host disease (GVHD). The multifactorial etiology of neurological complications in HSCT patients makes diagnosis difficult. However a timely and rigorous characterization of such complications should be obtained in the attempt to avoid fatal outcomes or long-\uadterm effects. Data regarding neurological complications in HSCT patients derives from small series and varies largely in respect to incidence and severity. Aim of this study is to describe incidence, characteristics and outcome of neurological complications in a large series of consecutive HSCT patients

Which sample tube should be used for routine glucose determination ?

tBackground: Glucose is one of the most frequently requested analytes in clinical laboratory.Blood glucose analysis is affected from in vitro glycolysis. In order to determine the mostsuitable blood collection tube for this purpose we have compared different tubes: sodiumfluoride, lithium heparin, sodium fluoride/citrate buffer containing tubes and serum withclot activator tube for the measurement of glucose when the tube has been kept at roomtemperature (RT) for up to 4 h.Methods: Venous blood was collected from 49 healthy volunteers into Sarstedt S-Monovettesfor glucose analysis. Reference plasma glucose was determined in a lithium heparin tubeand immediately placed in an ice/water slurry. Within 10 min it was centrifuged at 4◦C andplasma was separated from the blood cells. Samples have been preserved at RT for 1, 2 and4 h after drawing. Glucose has been determined using a hexokinase method.Results: Glucose levels tested in a serum with clot activator tube, in lithium heparin andin sodium fluoride/sodium EDTA tubes when compared with lithium-heparin referenceplasma did not meet the desirable bias for glucose (±1.8%) when kept at RT for up to 4 h.GlucoEXACT tubes, when corrected by the Sarsted recommended factor of 1.16, showed amean (95% CI) bias of +0.96% (0.45–1.47) at 1 h, +1.40% (0.88–1.93) at 2 h and +0.95% (0.44–1.46)at 4 h, reaching the analytical goal for the desirable bias.Conclusions: Samples collected into GlucoEXACT tubes containing sodium fluoride/citratebuffer liquid mixture are equivalent to those collected in reference plasma tubes avoidingglycolysis completely and within a 4 h delay in plasma separation