Circulating subpopulations of non-cytotoxic ILCs in diffuse large B-cell lymphoma.

Non-cytotoxic innate lymphoid cells (ILCs) have been added to the list of immune cells that may contribute to the tumor microenvironment. Elevated levels of total ILCs and their subgroups have been reported in peripheral blood and tissue samples from patients with solid tumors, but their frequency in non-Hodgkin lymphomas, particularly diffuse large B-cell lymphoma (DLBCL), has not been clearly established. This study examined frequency and subset distribution in newly diagnosed DLBCL patients (nodal and extra-nodal) and compared it with blood specimens from healthy donors. The percentage of total ILCs (Lin - CD127+) was assessed by flow cytometry, as well as the four ILC subsets, defined as ILC1 (Lin - CD127 + cKit - CRTH2-), ILC2 (Lin - CD127 + cKit+/- CRTH2+), ILCp NCR- (Lin - CD127 + cKit + CRTH2- NKp46-) and NCR + ILC3 (Lin - CD127 + cKit + NKp46+). In the studied group of patients (n = 54), significantly lower levels of circulating total ILCs, ILC1, and ILCp NCR- were observed compared to the control group (n = 43). Similarly, there was a statistically significant decrease in the median frequency of NKp46 + ILC3 cells in lymphoma patients. Analysis of the ILC2 subpopulation showed no significant differences. The correlation of the distribution of individual subpopulations of ILCs with the stage and location of the tumor was also demonstrated. Our results suggest that circulating ILCs are activated and differentiated and/or differentially recruited to the lymph nodes or tumor microenvironment where they may be involved in antitumor defense. However, our observations require confirmation in functional studies.

Comparison of the cryoprotective solutions based on human albumin vs. autologous plasma: its effect on cell recovery, clonogenic potential of peripheral blood hematopoietic progenitor cells and engraftment after autologous transplantation.

BACKGROUND AND OBJECTIVES: Cryopreservation of peripheral blood hematopoietic progenitor/stem cells (PBPCs) requires the addition of cryoprotectant such as DMSO, often prediluted using human serum albumin solution (HSAS). The goal of our study was to verify whether the HSAS may be replaced by autologous plasma (AP) without negative impact on PBPCs quality and engraftment. AP usage is less expensive and allows performing cell preparation in a 'closed system', and hence to reduce the risk of product contamination. MATERIALS AND METHODS: Peripheral blood progenitor cells from 18 patients were divided into two aliquots (500 µl) placed in separate vials, each containing 7·5% DMSO prediluted with 5% HSAS or AP. Post-thaw cell recovery and clonogenic potential was evaluated. During clinical part of the study, the impact of both cryoprotective solution on hematopoietic engraftment was evaluated in two cohorts (n = 26) matched for diagnosis, age and the number of transplanted CD34+ cells. RESULTS: The median recovery of nucleated cells and the number of colony-forming units did not differ between tested cryoprotective mixtures. For AP mixture, neither total protein nor albumin concentration of plasma correlated with cell recovery and clonogenic potential of the PBPCs after cryopreservation. In clinical evaluation, the median time to leucocyte recovery and reconstitution of neutrophils was comparable in both groups: 10 days. We did not observe either significant difference with regard to the time of platelet recovery (median: 15 days for AP vs. 16 for HSAS; P = 0·79). CONCLUSIONS: HSA in cryoprotective mixture may be replaced by AP without negative impact on cell recovery, clonogenic potential or engraftment.

Intermediate-dose Ara-C plus G-CSF for stem cell mobilization in patients with lymphoid malignancies, including predicted poor mobilizers.

The optimal protocol for mobilization of hematopoietic stem cells in patients with lymphoid malignancies has not been determined so far. We retrospectively analyzed the efficacy and safety of Ara-C at a dose of 1.6 g/m(2) compared with CY at a dose of 4.0 g/m(2), both combined with filgrastim. Seventy and forty-five patients, respectively, were included, among whom 60% were defined as 'predicted poor mobilizers'. The use of Ara-C was associated with significantly higher peak number of circulating CD34(+) cells compared with CY (P<0.0001). In the Ara-C group, 95% of patients with multiple myeloma (MM) collected at least 5 × 10(6) CD34(+) cells/kg required for tandem transplantation, and 97% of lymphoma patients collected at least 2 × 10(6) CD34(+) cells/kg, needed for a single autologous hematopoietic SCT (autoHSCT), which was achieved with a single leukapheresis in 91% of cases. Results for the CY group were significantly inferior (P<0.0001). No patient mobilized with Ara-C experienced febrile neutropenia, whereas 35% required platelet transfusions. Among patients who proceeded to autoHSCT, the time of both neutrophil and platelet recovery was significantly shorter for those mobilized with Ara-C than CY. We conclude that intermediate-dose Ara-C+filgrastim is a very effective and relatively safe mobilization protocol for patients with lymphoid malignancies.

Impact of different dimethyl sulphoxide concentrations on cell recovery, viability and clonogenic potential of cryopreserved peripheral blood hematopoietic stem and progenitor cells.

BACKGROUND AND OBJECTIVES: The procedure of autologous hematopoietic stem/progenitor cell transplantation requires cryopreservation. Addition of DMSO is necessary to secure the viability of such cells, but this solvent is potentially toxic to stem cells' recipient. 10% DMSO solution is used by the majority of transplant centres. The aim of our study was to test if DMSO concentration might be reduced without negative impact on cell recovery and clonogenicity. MATERIALS AND METHODS: Samples were prospectively collected from 20 patients. Small volumes of leukapheresis products were frozen with different cryoprotective mixtures, containing 10%, 7·5%, 5% and 2·5% DMSO, respectively. The quality of cryoprotective mixtures was evaluated based on recovery, viability and clonogenic potential of hematopoietic stem cells after defreezing. RESULTS: Reduction in DMSO concentration to 7·5% or lower was associated with decreased recovery of nucleated cells. In contrast, the number of colonies was highest for 7·5% DMSO with significant differences when compared to 10% DMSO solution. CONCLUSION: Reduction in DMSO concentration from 10% to 7·5% may have favourable impact on hematopoietic recovery after autologous transplantation. The findings require confirmation in a clinical setting.

A new cholesterol derivative suitable for transfecting certain type of cells in the presence of 10% serum.

We developed a new cationic lipid suitable for use as a DNA carrier in the presence of 10% sera. The novel compound (abbreviated as Arg-Chol) contains cholesterol and a dipeptide consisting of glycine and sterically protected arginine. The efficiency of reporter gene transfection using liposomes based on this new reagent was compared with that of liposomes made with other cationic derivatives of cholesterol. Lipoplexes formulated with the newly synthesized lipid mediate in vitro transfection of B16(F10) murine melanoma cells in the presence of 10% sera more efficiently than in other cell lines and compared with other cholesterol derivatives studied.