Evaluation of a potassium removal filter on irradiated red cells stored in SAGM.

BACKGROUND AND OBJECTIVES: Irradiation of red cell concentrates RCCs) can lead to well-documented elevated extracellular potassium concentrations. Transfusion of these products has the potential, if given as a massive/rapid transfusion, to lead to transient hyperkalemia. A potassium absorption filter (PAF) has recently been developed and has been proven to effectively remove excess K(+) . However, data are lacking on the red cell quality parameters over storage after irradiation. METHODS: Thirty RCCs were pooled and split into 3 groups of 10. All RCCs were irradiated on day 14 and filtered on day 28 (group 1 control), day 15 (group 2) or day 17 (group 3). Pre-irradiation, pre- and post-filtration and day 28 samples were taken for each study. Standard red cell quality parameters were measured over storage at the above time points. RESULTS: Losses for haemoglobin, haematocrit and volume were minimal after filtration with all units containing >40 g Hgb unit(-1). Statistically, significant differences were observed for K(+) and Na(+) levels in groups filtered at either 24 or 72 h post-irradiation, and this was observed directly after filtration and remained by day 28. Filtration had no significant impact on any other parameters measured. CONCLUSIONS: PAF effectively removed supernatant potassium (93%) from all RCC units. Early removal of K(+) at either day 15 or 17 on RCCs subsequently stored to day 28 had no measurable effect on red cell quality, suggesting this may be a useful device to ensure further safety for at-risk immunocompromised patient groups requiring irradiated RCCs.

Immunology in the clinic review series; focus on cancer: double trouble for tumours: bi-functional and redirected T cells as effective cancer immunotherapies.

Cancer is one of the most important pathological conditions facing mankind in the 21st century, and is likely to become the most important cause of death as improvements continue in health, diet and life expectancy. The immune response is responsible for controlling nascent cancer through immunosurveillance. If tumours escape this control, they can develop into clinical cancer. Although surgery and chemo- or radiotherapy have improved survival rates significantly, there is a drive to reharness immune responses to treat disease. As T cells are one of the key immune cells in controlling cancer, research is under way to enhance their function and improve tumour targeting. This can be achieved by transduction with tumour-specific T cell receptor (TCR) or chimaeric antigen receptors (CAR) to generate redirected T cells. Virus-specific cells can also be transduced with TCR or CAR to create bi-functional T cells with specificity for both virus and tumour. In this review we outline the development and optimization of redirected and bi-functional T cells, and outline the results from current clinical trials using these cells. From this we discuss the challenges involved in generating effective anti-tumour responses while avoiding concomitant damage to normal tissues and organs.

Rapid detection, enrichment and propagation of specific T cell subsets based on cytokine secretion.

T cell lines with defined cytokine profiles are an invaluable tool for assessing the control of immune responses both in vitro and in vivo. Production of such cell lines can be complex and time-consuming. Here we present a powerful technique to assay the cytokines produced by T cells activated polyclonally or with specific antigens. This paper presents a detailed methodology for the identification and isolation of cytokine-producing T cells activated with the artificial superantigen, CytoStim, or viral and fungal antigens. These cells can be analysed for different cytokines simultaneously, or cultured further to rapidly establish T cell lines making known cytokine types. We highlight the enumeration, isolation and phenotype of interleukin-17-producing T cells, and the rapid generation of virus-specific Th1 T cell lines.

Immature monocytes from G-CSF-mobilized peripheral blood stem cell collections carry surface-bound IL-10 and have the potential to modulate alloreactivity.

Production of the anti-inflammatory cytokine IL-10 by monocytes has been implicated as a probable negative regulator of graft-versus-host disease (GvHD) in patients undergoing allogeneic stem cell transplants (SCT). Monocytes from G-CSF-mobilized peripheral blood stem cell (gmPBSC) collections have been reported to produce more IL-10 than unmobilized monocytes in response to proinflammatory factors such as LPS. Why this should occur is unclear. In this study, monocyte phenotype and IL-10 localization and release were investigated in PB mononuclear cells (MNC) from 27 healthy donors mobilized for allogeneic SCT and from 13 patients with hematological malignancies mobilized for autologous SCT. All isolates contained elevated total percentages of monocytes in comparison with unmobilized PB, a high proportion of which displayed an immature phenotype. Stimulation of gmPB MNC with an inflammatory stimulus [fixed Staphylococcus aureus cells (SAC)] induced rapid up-regulation of CD14, indicating conversion to mature status. Localization studies indicated that IL-10 was predominantly present, bound on the surface of CD64(+)/CD14(low/neg) immature monocytes. Inflammatory stimuli (LPS, polyinosinic:polycytidylic acid, or SAC) induced release of variable quantities of IL-10 from the cell surface. MNC, separated into surface IL-10-positive or -negative fractions, differed in their ability to stimulate alloreactivity in MLR, and IL-10(+) MNC induced significantly lower levels of proliferation than IL-10(-) MNC. Thus, the subset of immature monocytes carrying surface-bound IL-10 in gmPB has the potential to modulate alloreactivity and GvHD after allogeneic SCT through cell-to-cell contact and released IL-10.

Characterization of efferent lymph cells and their function following immunization of cattle with an allogenic Theileria annulata infected cell line.

Immunization of cattle with in vitro propagated bovine mononuclear cells infected with Theileria annulata induces a protective immune response. Activation and effector function of T cells exiting the lymph node draining the site of cell line immunization were investigated to understand the mechanisms involved in the generation of immunity. Immunized animals exhibited a biphasic immune response in efferent lymph as well as peripheral blood. The first phase corresponded to allogenic responses against MHC antigens of the immunizing cell line and the second was associated with parasite specific responses. An increase in the output of CD2(+) cells and MHC class II(+) cells in efferent lymph was observed after cell line immunization with a corresponding decrease in WC1(+) cells. Although the percentage of CD4(+) T cells did not change significantly over the course of the experiment, they became activated. Both CD25 and MHC class II expressing CD4(+) T cells were detected from day 7 onwards, peaking around day 13. Efferent lymph leukocytes (ELL) exhibited sustained responses to IL-2 in vitro following cell line immunization. Antigen specific proliferation was also detected first to the immunizing cell line and then to parasite antigens. The two peaks of CD2(+) cells were observed, which corresponded to similar peaks of CD8(+) cells. The increase in CD8(+) cells was more pronounced during the second parasite specific phase than the first allogenic phase. Activated CD8(+) T cells mainly expressed MHC class II and some expressed CD25. Significantly the peak of activated CD4(+) T cells preceded the peak of activated CD8(+) T cells, highlighting the role of T. annulata specific CD4(+) T cells in inducing parasite specific CD8(+) cytotoxic responses. A biphasic cytotoxic response also appeared in efferent lymph and peripheral blood, the first directed against MHC antigens of the immunizing cell line followed by MHC class I restricted parasite specific cytotoxicity. The cytotoxic responses in efferent lymph appeared earlier than peripheral blood, suggesting that activated CD8(+) cells exiting the draining lymph node following immunization with T. annulata infected schizonts play an important role in the development of protective immune responses.