The major population of PHA-stimulated PBMC infected by R5 or X4 HIV variants after a single cycle of infection is predominantly composed of CD45RO<Superscript>+</Superscript>CD4<Superscript>+</Superscript> T lymphocytes

Summary. PHA-stimulated peripheral blood mononuclear cells (PBMCs) are widely used for investigating replication and neutralization of HIV primary isolates in vitro. The objective of this study was to identify the T lymphocyte subset(s) that are found infected after one replication cycle by either R5- or X4-HIV-1 variants in PHA-stimulated PBMCs from healthy donors. Infected T lymphocytes were detected by intracellular p24 staining and characterized by cell surface immunophenotyping using flow cytometry. The predominant lymphocyte subset expressing p24 after 24 h of infection with either R5 or X4 HIV-1 strains was found to exhibit mainly the memory CD45RO phenotype, a greater percentage of CD62L⁺CD45RO⁺ central memory T lymphocytes was infected with X4 HIV strains. Although some CD45RA⁺ lymphocytes were also infected, these cells co-expressed CD45RO⁺. The proportion of lymphocytes expressing CD4 and CD4/CD45RO decreased by 20% after 24 h of infection. A 2-fold decrease of CD4⁺CD8⁺ T lymphocytes could also be recorded, even though this subset accounted for less than 5% of total lymphocytes in control cultures. Moreover, CD4⁺CD8⁺ T cells further decreased by 90% after 4 days of infection, a time at which they scored p24⁺. Therefore, our results indicate that the in vitro infection system of PHA-stimulated PBMC utilized in neutralization assays provides an appropriate model for the study of infected CD45RO⁺ lymphocytes but not CD45RA⁺ lymphocytes.     

Analysis of FOXP3<Superscript>+</Superscript> regulatory T cell subpopulations in peripheral blood and tissue of patients with systemic lupus erythematosus

Regulatory T cells (Tregs) are critical mediators of immune tolerance, yet their involvement in the autoimmune disease systemic lupus erythematosus (SLE) is incompletely understood. We analyzed CD4⁺ T cell subpopulations with Treg-related phenotypes and their association with disease activity in peripheral blood (PB) and tissues of patients with SLE. In detail, we quantified subpopulations regarding CD25, FOXP3, CD62L, CCR6, CD27, CD45RA, and CD45RO expression in PB from 31 patients with SLE divided into two disease activity groups and 32 healthy controls using flow cytometry. CD4⁺ and FOXP3⁺ T cells in skin and kidney biopsies of patients with SLE were quantified by immunohistochemistry. CD4⁺CD25^+/++FOXP3⁺ and CD4⁺CD25⁺CD45RA^?/CD45RO⁺ T cell frequencies were significantly higher in PB from patients with active compared to inactive SLE. The fraction of CD4⁺CD25⁺⁺FOXP3⁺ Tregs and CD4⁺CD25⁺CD45RA⁺/CD45RO^? naïve Tregs was not significantly different between these groups. CD4⁺CD25⁺⁺ Tregs from active SLE patients comprised significantly less CD27⁺ cells and more CCR6⁺ cells compared to patients with inactive SLE. The percentage of CD4⁺FOXP3⁺ T cells among inflammatory infiltrates in skin and kidney biopsies of SLE patients was not different from other inflammatory skin/kidney diseases. In conclusion, although CD4⁺FOXP3⁺ T cell frequencies in the inflamed tissues of SLE patients were comparable to other inflammatory diseases, distinct T cell subpopulations appeared misbalanced in PB of patients with active SLE. Here, cells phenotypically resembling activated T cells, but not Tregs, were increased compared to patients with inactive SLE. Within Tregs of patients with active SLE, markers related to Treg function and homing were altered.     

Accumulation of CD5<Superscript>+</Superscript>CD19<Superscript>+</Superscript> B lymphocytes expressing PD-1 and PD-1L in hypertrophied pharyngeal tonsils

Programmed death-1 (PD-1) is one of the most important inhibitory co-receptors expressed predominantly on activated T and B lymphocytes whose expression could be sustained by permanent antigenic stimulation accompanying chronic or recurrent tonsillitis. The expression of PD-1 and PD-1L was analyzed using flow cytometry on hypertrophied tonsils collected from 57 children. We observed high expression of PD-1 and PD-1L on certain lymphocytes subpopulations of hypertrophied tonsils; among T cells, the expression of PD-1 on protein level was higher on CD4⁺ cells (70.3 %) than on CD8⁺ cells (35 %). Interestingly, a limited expression of PD-1 was observed on CD19⁺ B lymphocytes (6.5 %), while CD5⁺CD19⁺ B cells overexpressed PD-1 (52.5 %). Moreover, the expression of PD-1L was also higher on CD5⁺CD19⁺ B cells (16.5 %) than on CD19⁺ B cells (3.5 %) and on CD4⁺ T cells (20 %) than on CD8⁺ T cells (10 %). PD-1 and PD-1L expressions correlated only on CD5⁺CD19⁺ cells. In conclusion, high expression of PD-1 and PD-1L on T and B cells could represent hallmark of immune system adaptation to chronic antigenic exposition in patients with tonsillitis.     

Aerobic training increases the stimulated percentage of CD4<Superscript>+</Superscript>CD25<Superscript>+</Superscript> in older men but not older women

The purpose of the present study was to determine whether 12 months of moderate intensity cycling would increase the expression of IL-2 (CD25⁺) receptors in T helper (CD4⁺) lymphocytes in men and women aged 65–75 years. Fourteen men and 10 women completed 52 weeks of moderate intensity cycling (60% VO₂peak). Subjects trained (TR) three times per week for 45 min per session. Eight age-matched untrained (UT) male and eight UT female subjects acted as controls. Resting blood samples were taken from TR and UT subjects every 4 weeks. Leukocyte concentration was measured using a full blood count. PHA-stimulated CD4⁺ lymphocytes were analysed for changes in the expression of CD25⁺, by flow cytometry. Training significantly increased VO_2peak (l min⁻¹, ml kg⁻¹ min⁻¹) in male (+14.3, +16%) and female (+16.7, +27.8%) groups. The TR male group showed a significantly lower percentage of CD4⁺CD25⁺ than the male UT in January but the TR male percentage was significantly higher than the UT male group during February, March, April, May, June, September B and December. The female TR group showed a significantly higher percentage CD4⁺CD25⁺ than the female UT only during July. There were also significant sequential monthly changes in the percentage of CD4⁺CD25⁺ for male and female UT and TR groups. Significant increases in the percentage of CD4⁺CD25⁺ in the male TR group suggest training-enhanced lymphocyte mitogenic responsiveness. Moderate intensity long-term training may increase the recruitment of active memory CD4⁺CD25⁺ in men rather than women.     

Expression of CD62L on Donor CD4<Superscript>+</Superscript> T Cells in Allografts: Correlation with Graft-Versus-Host Disease after Unmanipulated Allogeneic Blood and Marrow Transplantation

Introduction  The aim of this study was to investigate the association of donor CD4⁺ T cells expressing CD62L with transplant outcomes. Materials and Methods  We report a prospective analysis of 31 patients who were treated with a Bu/Cy regimen, followed by unmanipulated blood and marrow transplantation. Results  Median number (range) of CD4⁺CD62L⁺, CD4⁺CD45RA⁺CD62L⁺, and CD4⁺CD45RO⁺CD62L⁺ cells infused were 0.31(0.05–1.10)×10⁸/kg, 0.22(0.03–0.95)× 10⁸/kg, and 0.17(0.01–0.81)×10⁸/kg, respectively. The incidence of grades II to IV aGVHD was 36%. In a multivariate analysis, infusion of >0.22 × 10⁸ CD4⁺CD45RA⁺CD62L⁺ cells infused/kg increased the risk of grades II to IV aGVHD (HR = 4.741, 95% CI = 1.037–21.662, P = 0.045). Thirteen of 31 patients experienced cGVHD, the risk of cGVHD was increased in patients receiving >0.45 × 10⁸ CD4⁺CD45RA⁺ cells infused/kg (HR = 4.614, 95% CI = 1.265–16.829, P = 0.021). Conclusion  Our results suggest that a high cell dose of CD4⁺CD45RA⁺CD62L⁺ cells increase the incidence of grades II–IV aGVHD. A high number of CD4⁺CD45RA⁺ cells infused were associated with increased risk of cGVHD in our transplant settings. Ying-Jun Chang: performed research, analysis and interpretation of data, and drafting of the article, and gave final approval of the version to be published; Xiang-Yu Zhao: performed research, analysis and interpretation of data, and drafting of the article and gave final approval of the version to be published; Ming-Rui Huo: performed research, analysis and interpretation of data, and drafting of the article and gave final approval of the version to be published; Xiao-Jun Huang: involved in conception and design, revising the article critically, and final approval of the version to be published.     

Expression of P2X Receptor Subtypes on CD34<Superscript>+</Superscript> Cells and c-kit<Superscript>+</Superscript> Cells of Human Umbilical Blood

The presence of several subtypes of P2X receptors on early hemopoietic precursors (CD34⁺) from human umbilical blood was detected by flow cytometry. The expression of P2X receptors on umbilical blood lymphocytes was an order of magnitude higher than that on adult human blood cells. Our results attest to early involvement of P2X receptors in differentiation of human hemopoietic cells.     

Highly enriched CD133<Superscript>+</Superscript>CD44<Superscript>+</Superscript> stem-like cells with CD133<Superscript>+</Superscript>CD44<Superscript>high</Superscript> metastatic subset in HCT116 colon cancer cells

Stem-like cancer cells (SLCCs) are distinct cellular subpopulation in colon cancer that is essential for tumor maintenance. Previous studies indicated that SLCCs accounted for only a minor subset in a given cancer model. However, we found that SLCCs frequency varied among a panel of colon cancer cell lines, with HCT116 cells composed mainly of SLCCs, as demonstrated by colonosphere forming capability and CD133 expression. Indeed, flow cytometric analysis revealed more than 60% HCT116 cells co-expressed the putative SLCCs markers CD133 and CD44. Compared with non-CD133⁺CD44⁺ cells, FACS sorted CD133⁺CD44⁺ cells were undifferentiated, endowed with extensive self-renewal and epithelial lineage differentiation capacity in vitro. CD133⁺CD44⁺ exhibited enhanced tumorigeneicity in NOD/SCID mice. One thousand CD133⁺CD44⁺ cells initiated xenograft tumors efficiently (3/6) while 1 × 10⁵ non-CD133⁺CD44⁺ cells could only form palpable nodule with much slower growth rate (1/6). More interestingly, long-term cultured self-renewing CD133⁺CD44⁺ cells enriched CD133⁺CD44^high subset, which expressed epithelial to mesenchymal transition marker, were more invasive in vitro and responsible solely for liver metastasis in vivo. In conclusion, these data demonstrated for the first time that CD133⁺CD44⁺ SLCCs were highly enriched in HCT116 cells and that metastatic SLCCs resided exclusively in a CD133⁺CD44^high subpopulation.     

Quantitative differences in lipid raft components between murine CD4<Superscript>+</Superscript> and CD8<Superscript>+</Superscript> T cells

Background  

Lipid rafts have been shown to play a role in T cell maturation, activation as well as in the formation of immunological synapses in CD4⁺ helper and CD8⁺ cytotoxic T cells. However, the differential expression of lipid raft components between CD4⁺ and CD8⁺ T cells is still poorly defined. To examine this question, we analyzed the expression of GM1 in T cells from young and aged mice as well as the expression of the glycosylphosphatidylinositol (GPI)-linked protein Thy-1 and cholesterol in murine CD4⁺ and CD8⁺ T cell subpopulations.     

CD4<Superscript>+</Superscript>FOXP3<Superscript>+</Superscript> T regulatory cells decrease and CD3<Superscript>+</Superscript>CD8<Superscript>+</Superscript> T cells recruitment in TILs from melanoma metastases after electrochemotherapy

Electrochemotherapy (ECT) represents an effective local treatment for skin unresectable melanoma metastases with high overall objective response rate. ECT is based on the combination of anti-neoplastic drugs administration and cancer cells electroporation. Whether ECT can also activate the immune system is a matter of debate, however a significant recruitment of dendritic cells in melanoma treated metastases has been described. Herein we investigated immediate and late effects of ECT treatment on T cell subsets in ECT-treated lesions by fluorescent immunohistochemistry. Biopsies from melanoma patients (n = 10) were taken before ECT (t0), at d1 and d14 from treatment. At t0, CD3⁺CD4⁺ T cells were the most represented T cells, well detected in the perilesional dermis, particularly at tumour margin, while CD3⁺CD8⁺ T cells were less represented. CD4⁺FOXP3⁺ T regulatory (Treg) cells were present in the perilesional dermis and within the lesion. ECT induced a significant decrease of CD4⁺FOXP3⁺ Treg cells percentage in the perilesional dermis, observed at d1 and at d14 (p < 0.001). CD3⁺CD8⁺ T cells frequency significantly increased at d14 from treatment in the perilesional dermis (p < 0.001). Furthermore calreticulin translocation to the plasma membrane, a hallmark of immunogenic cell death, was observed in metastatic cells after ECT. The data reported here confirm that ECT induces a local response, with a lymphoid infiltrate characterized by CD4⁺FOXP3⁺ Treg cells decrease and CD3⁺CD8⁺ T cells recruitment in the treated lesions. These results might contribute to design novel combinational therapeutic approaches with ECT and immunotherapy in order to generate a systemic long-lasting anti-melanoma immunity.     

Evaluation of the expression of early activation marker CD69 by CD8<Superscript>+</Superscript> lymphocytes in HIV-infected patients

We studied the expression of CD69 antigen on CD8⁺ lymphocytes in response to nonspecific activator phytohemagglutinin in 202 patients with HIV infection. It was found that the number of CD8⁺CD69⁺ lymphocytes in HIV-infected patients increased; this parameter negatively correlated with relative and absolute content of CD4⁺ cells and positively correlated with IgG concentration. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 144, No. 12, pp. 661–663, December, 2007     

Lenalidomide potentiates CD4<Superscript>+</Superscript>CD25<Superscript>+</Superscript>Treg-related suppression of lymphoma B-cell proliferation

We have previously found that ex vivo expanded human CD4⁺CD25⁺Treg cells suppress proliferation of lymphoma B-cell lines. Here we demonstrate that the immunomodulatory drug lenalidomide potentiates suppression of lymphoma B-cell proliferation by freshly isolated CD4⁺CD25⁺Tregs, as well as suppression by Tregs expanded polyclonally in the presence of rapamycin from CD4⁺CD25⁺T cells or CD4⁺CD25⁺CD127^loT cells. The regulation of lymphoma cell proliferation by Tregs pre-expanded with “third-party” allogeneic MoDCs in the presence of rapamycin was also potentiated by lenalidomide. Lenalidomide contributed to the suppression exerted by Tregs despite concomitant downregulation of Treg proliferation. Lenalidomide did not reduce the suppression of conventional T cells by expanded Tregs. The exposure of polyclonally expanded Tregs to lenalidomide did not significantly alter their phenotype. There was no uniform pattern of lenalidomide effect on Treg-mediated regulation of lymphoma B cells freshly isolated from patients. Freshly isolated lymphoma cells activated with multimeric CD40L and IL-4 to support their survival in vitro varied in their sensitivity to lenalidomide, and the regulatory effect of Tregs on such lymphoma cells ranged from suppression to help in individual patients. Lenalidomide potentiated or attenuated Treg effects on the survival of freshly isolated lymphoma cells. A combination of lenalidomide treatment with adoptive transfer of CD4⁺CD25⁺Tregs or CD4⁺CD25⁺CD127^loTregs expanded ex vivo could be used to suppress proliferation of residual lymphoma in select patients with lymphoma responsive to the regulation by Tregs and sensitive to lenalidomide.     

Co-expression of the CD45RA and CD45RO antigens on T lymphocytes in chronic arthritis

The site of T lymphocyte activation in chronic arthritis is unknown. Peripheral blood (PB) lymphocytes from chronic arthritis patients are in a ‘naïve’ or non-activated state, as defined by expression of the CD45RA antigen and lack of HLA class II expression. In contrast, most synovial fluid (SF) T lymphocytes express a ‘memory’ or activated phenotype, as defined by the CD45RO antigen and high HLA class II expression. Following stimulation, naive cells lose CD45RA and gain CD45RO expression to become memory cells with a transitional stage of dual CD45RA, CD45RO antigen expression. To localize where this change in phenotype occurs we used dual colour immunofluorescence labelling to compare the percentage of dual CD45RA, CD45ROpositive T lymphocytes in PB and SF from chronic arthritic patients and from normal PB, assuming this population would be increased at the primary site of T lymphocyte activation. Expression of the intermediate and late activation marker. HLA-DR, was also analysed using dual colour immunofluorescence labelling. The percentage of dual positive T lymphocytes was similar between arthritic PB, SF. and normal PB, as was the density of both CD45RA and CD45RO antigens. Thus, CD45 isoform expression did not indicate where T lymphocytes were activated. However, we identified a previously unreported population of CD45RA⁺ CD45RO⁺ HLA-DR^- T lymphocytes in arthritic and normal PB. In SF, this population was absent, but a substantial number of dual CD45RA, CD45RO-positive HLA-DR⁺ T lymphocytes were identified. This population would not be predicted by the current model of T lymphocyte activation. Division of T lymphocytes into functional groups on the basis of CD45 isoform expression is likely to be more complicated than previously thought. Based on our findings we propose an alternative model of T lymphocyte differentiation.     

CD4<Superscript>+</Superscript> T Cells Downregulate Bcl-2 in Germinal Centers

Germinal centers (GCs) are the main site of T cell-dependent antibody responses. Upon antigen challenge, GCs comprise mostly B cells undergoing proliferation, somatic hypermutation and antigen-affinity selection. GC B cells down-modulate the expression of Bcl-2 protein and are highly sensitive to apoptosis to eliminate autoreactive or low-affinity cells. Bcl-2 is still expressed in a few GC cells, whose identity remains unclear. To address this issue, we examined by confocal microscopy the expression of Bcl-2 by different GC lymphocyte subsets in hyperplastic tonsils. We found that the vast majority of Bcl-2⁺ GC cells are T lymphocytes. Conversely, while in the mantle zone and in the interfollicular areas T cells are almost exclusively Bcl-2⁺, in the GC, most T lymphocytes are Bcl-2⁻. In addition, most of the CD4⁺ GC T cells are Bcl-2⁻, while nearly 100% of the CD8⁺ GC T cells are Bcl-2⁺. The Bcl-2 downregulation by both B and CD4⁺ T GC cells supports the concept that these two subsets may undergo a selection process in this microenvironment.     

CD3 antigen-mediated calcium signals and protein kinase C activation are higher in CD45R0+ than in CD45RA+ human T lymphocyte subsets

T lymphocytes may be separated into subsets according to their expression of CD45 isoforms. The CD45R0⁺ T cell subset has been reported to proliferate in response to recall antigen and to mitogenic mAb to a much greater extent than the CD45RA⁺ subset. This difference could be due to more efficient coupling of the T cell antigen receptor complex to mitogenic signaling pathways. To investigate this possibility, CD3 antigen-induced calcium signals, diacylglycerol (DAG) production and protein kinase C (PKC) activation levels were compared in CD45RA⁺ and CD45R0⁺ human T lymphocyte subsets derived from peripheral blood. The mean CD3-induced rise in intracellular calcium was 80% greater in CD45R0⁺ than in CD45RA⁺ cells. Basal DAG levels in CD45R0⁺ cells were found to be, on average, 60% higher than in CD45RA⁺ cells (p = 0.002), but the CD3-induced production of DAG over background was not different in the two subsets (p = 0.4). Basal PKC activity, and CD3-induced PKC activation levels over background, were found to be 50% and 140% higher, respectively, in CD45R0⁺ cells than in CD45RA⁺ cells (p = 0.015 and 0.023). The CD45R0⁺ subset contained a higher proportion of cells expressing activation markers, such as CD25, CD71 and major histocompatibility complex class II, when compared to the CD45RA⁺ subset. Our results suggest that the elevated basal DAG levels observed in the CD45R0⁺ subset may reflect the recent activation of these cells. Both the higher basal DAG and CD3-induced elevation in intracellular calcium observed in the CD45R0⁺ cells may contribute to the greater PKC activation signals triggered by CD3 mAb in this subset. These findings elucidate the greater response of CD45R0⁺ T cells to mitogenic stimuli compared to CD45RA⁺ cells.     

Expression of Markers of Regulatory CD4<Superscript>+</Superscript>CD25<Superscript>+</Superscript>foxp3<Superscript>+</Superscript> Cells in Atherosclerotic Plaques of Human Coronary Arteries

The content of marker foxp3 of regulatory T cells and chemokines in atherosclerotic plaques of human coronary arteries was measured by the polymerase chain reaction. In vitro migration of regulatory CD4⁺CD25⁺foxp3⁺ cells in the CD4⁺ lymphocyte population from healthy donors was studied after treatment with chemokines I-309, IP-10, and SDF-1. mRNA for the factor foxp3 and chemokines SDF-1, I-309, and MIP-1β were found in the majority of samples from atherosclerotic plaques. SDF-1 induced maximum migratory response of CD4⁺CD25⁺foxp3⁺ cells.     

CD27<Superscript>+</Superscript>TIM-1<Superscript>+</Superscript> memory B cells promoted the development of Foxp3<Superscript>+</Superscript> Tregs and were associated with better survival in acute respiratory distress syndrome

Acute respiratory distress syndrome (ARDS) is a rapid onset life-threatening condition involving uncontrolled propagation of inflammatory responses. Here, we observed that ARDS patients that survived presented significantly higher frequencies of TIM-1⁺ B cells, especially the CD27⁺TIM-1⁺ B cells, than the ARDS patients who succumbed to the condition. We then found that using BCR/CD40 antigen-dependent stimulation or Staphylococcus aureus Cowan (SAC) antigen-independent stimulation, TIM-1⁺ B cells presented significantly higher IL-10 secretion and/or TGF-β1 secretion, with SAC stimulation being more effective. CD4⁺ T cells that incubated with TIM-1⁺ B cells presented significantly elevated IL-10 secretion, TGF-β1 secretion, and Foxp3 expression, than CD4⁺ T cells that incubated with TIM-1^? B cells, suggesting TIM-1⁺ B cells promoted the in vitro development of Foxp3⁺ Treg cells. Interestingly, this TIM-1⁺ B cell-mediated promotion of Foxp3 expression was mostly dependent on TGF-β1 but not IL-10, since neutralization of TGF-β1, but not IL-10, resulted in the suppression of Foxp3 expression. We further showed that in TIM-1⁺ B cells, the CD27⁺ classical memory B cell subset demonstrated more regulatory potency than the CD27^? subset. Together, our results suggested that the TIM-1⁺ B cells, especially those that expressed CD27, could promote Foxp3 expression. Their clinical efficacy in treating ARDS should be examined in in vivo experiments.     

Effects of simvastatin on the function of splenic CD4<Superscript>+</Superscript> and CD8<Superscript>+</Superscript> T cells in sepsis mice

Simvastatin may be beneficial for treating sepsis due to its immune-regulating properties, although the mechanisms remain elusive. Herein, we hypothesized simvastatin may attenuate T cell dysfunction induced by sepsis. To test this hypothesis, we used a model based on cecal ligation and puncture (CLP) to induce sepsis in mice. Male C57BL/6 mice were pre-treated with simvastatin (0.2 μg/g of body weight) before CLP. The expression of B and T lymphocyte attenuator (BTLA) on splenic CD4⁺ T cells and T cell apoptosis, CD4⁺ and CD8⁺ T cells were quantified by flow cytometry. Immunohistochemical staining was performed to evaluate the loss of immune effector cells. Formation of TNF-α and interleukin 10 (IL-10) in the spleen and plasma levels of presepsin, IL-1β, and IL-6 were determined using enzyme-linked immunosorbent assay. Simvastatin markedly inhibited the reduction in cytokine secretion from lipopolysaccharide (LPS)-stimulated splenocytes. Simvastatin-treated mice had significantly decreased the percentages of negative costimulatory receptor BTLA on CD4 T cell expression. Simvastatin markedly reduced T cell apoptosis through downregulating the Fas/FasL expression and decrease the percentage of caspase-3 activity in spleen tissue. There was significantly less depletion of splenic CD4⁺ and CD8⁺ T cells in simvastatin-treated mice. Simvastatin reduced plasma levels of presepsin, IL-1β, and IL-6. Simvastatin can be a powerful regulator of immune function under sepsis conditions by improving T cell function in sepsis.     

CD8+ and CD45RA+ human peripheral blood lymphocytes are potent sources of macrophage inflammatory protein 1α, interleukin-8 and RANTES

The chemokines macrophage inflammatory protein 1α (MIP 1α), interleukin-8 (IL-8) and RANTES are potent regulators of leukocyte trafficking. Examination of chemokine secretion by human peripheral blood lymphocytes after stimulation with anti-CD3 or phorbol 12, 13 myristate acetate and ionomycin showed CD8⁺ cells were the dominant source of MIP 1α and RANTES. Although production of MIP 1α and IL-8 were similar in pharmacologically stimulated CD4⁺ CD45RA⁺, CD4⁺ CD45RO⁺, and CD8⁺ CD45RA⁺ cells, the largest amounts of MIP 1α and RANTES were secreted by CD8⁺ CD45RO⁺ lymphocytes. A parallel pattern of prolonged chemokine mRNA expression for at least 18 h after activation was observed in the T cell subsets. These results confirm that human T lymphocytes have a unique capacity for secretion of these three chemokines. In addition, CD8⁺ cells have an unrecognized role in recruiting cells to sites of inflammation, and adult human CD45RA⁺ cells have a physiologically significant secretory capacity.     

Immunological role of CD4<Superscript>+</Superscript>CD28<Superscript>null</Superscript> T lymphocytes,natural killer cells,and interferon-gamma in pediatric patients with sickle cell disease: relation to disease severity and response to therapy

Sickle cell disease (SCD) is associated with alterations in immune phenotypes. CD4⁺CD28^null T lymphocytes have pro-inflammatory functions and are linked to vascular diseases. To assess the percentage of CD4⁺CD28^null T lymphocytes, natural killer cells (NK), and IFN-gamma levels, we compared 40 children and adolescents with SCD with 40 healthy controls and evaluated their relation to disease severity and response to therapy. Patients with SCD steady state were studied, focusing on history of frequent vaso-occlusive crisis, hydroxyurea therapy, and IFN-gamma levels. Analysis of CD4⁺CD28^null T lymphocytes and NK cells was done by flow cytometry. Liver and cardiac iron overload were assessed. CD4⁺CD28^null T lymphocytes, NK cells, and IFN-gamma levels were significantly higher in patients than controls. Patients with history of frequent vaso-occlusive crisis and those with vascular complications had higher percentage of CD4⁺CD28^null T lymphocytes and IFN-gamma while levels were significantly lower among hydroxyurea-treated patients. CD4⁺CD28^null T lymphocytes were positively correlated to transfusional iron input while these cells and IFN-gamma were negatively correlated to cardiac T2* and duration of hydroxyurea therapy. NK cells were correlated to HbS and indirect bilirubin. Increased expression of CD4⁺CD28^null T lymphocytes highlights their role in immune dysfunction and pathophysiology of SCD complications.