搜索结果: 'EasySep Release'

Transcripts expressed in cytotoxic T lymphocytes (CTL) have mechanistic and diagnostic importance in transplantation. We used microarrays to select CTL-associated transcripts (CATs) expressed in human CD4(+) CTL,CD8(+) CTL and NK cells,excluding transcripts expressed in B cells,monocytes and kidney. This generated three transcript sets: CD4(+)-associated,CD8(+)-associated and NK-associated. Surprisingly,many CATs were expressed in effector memory cells e.g. granzyme B/GZMB,interferon-gamma/IFNG. Transcript expression was very similar between CD4(+) and CD8(+) CTL. There were no transcripts highly selective for CD4(+) CTL or CD8(+) CTL: for example,cytotoxic molecule transcripts (perforin,granzymes,granulysin) were shared between CD8(+) CTL and CD4(+) CTL although expression remained higher in CD8(+) CTL. Transcripts that differentiated between CD8(+) CTL and CD4(+) CTL were primarily those shared between CD8(+) CTL and NK cells (e.g. NK receptors KLRC1,KLRC3,KLRD1,KLRK1). No transcripts could differentiate CD4(+) CTL from CD8(+) CTL but NK cell-associated transcripts could differentiate NK cells from CTL. This study serves as a foundation for the interpretation of CATs in rejecting allografts and highlights the extensive sharing of CATs among CD4(+) CTL,CD8(+) CTL and effector memory T cells. View Publication

The CD31(+) subset of human naive CD4(+) T cells is thought to contain the population of cells that have recently emigrated from the thymus,while their CD31(-) counterparts have been proposed to originate from CD31(+) cells after homeostatic cell division. Naive T-cell maintenance is known to involve homeostatic cytokines such as interleukin-7 (IL-7). It remains to be investigated what role this cytokine has in the homeostasis of naive CD4(+) T-cell subsets defined by CD31 expression. We provide evidence that IL-7 exerts a preferential proliferative effect on CD31(+) naive CD4(+) T cells from adult peripheral blood compared with the CD31(-) subset. IL-7-driven proliferation did not result in loss of CD31 expression,suggesting that CD31(+) naive CD4(+) T cells can undergo cytokine-driven homeostatic proliferation while preserving CD31. Furthermore,IL-7 sustained or increased CD31 expression even in nonproliferating cells. Both proliferation and CD31 maintenance were dependent on the activation of phosphoinositide 3-kinase (PI3K) signaling. Taken together,our data suggest that during adulthood CD31(+) naive CD4(+) T cells are maintained by IL-7 and that IL-7-based therapies may exert a preferential effect on this population. View Publication

Identification of rational therapeutic targets is an important strategy to improve the cure rate of diffuse large B-cell lymphoma (DLBCL). We previously showed that inhibition of the phosphodiesterase 4B (PDE4B) unleashes cyclic-AMP (cAMP) inhibitory effects toward the PI3K/AKT pathway and induces apoptosis. These data raised important considerations as to which upstream regulators mediate cAMP inhibition of PI3K/AKT,and how identifying this signaling route could be translated into clinical initiatives. We found that in normal and malignant B cells,cAMP potently inhibit the phosphorylation and activity of the tyrosine kinase SYK. Using genetic models of gain- and loss-of-function,we demonstrated the essential role for PDE4B in controlling these effects in DLBCL. Furthermore,we used a constitutively active SYK mutant to confirm its central role in transducing cAMP effects to PI3K/AKT. Importantly,given SYK credentials as a therapeutic target in B-cell tumors,we explored the role of PDE4B in these responses. In multiple DLBCL models,we found that genetically,hence specifically,inhibiting PDE4B expression significantly improved the efficacy of SYK inhibitors. Our data defined a hitherto unknown role for cAMP in negatively regulating SYK and indicate that combined inhibition of PDE4B and SYK should be actively pursued. View Publication

The lack of natural killer (NK) cell-specific markers,as well as the overlap among several common surface antigens and functional properties,has obscured the delineation between NK cells and dendritic cells. Here,novel subsets of peripheral blood CD3/14/19(neg) NK cells and monocyte/dendritic cell (DC)-like cells were identified on the basis of CD7 and CD4 expression. Coexpression of CD7 and CD56 differentiates NK cells from CD56+ monocyte/DC-like cells,which lack CD7. In contrast to CD7+CD56+ NK cells,CD7(neg)CD56+ cells lack expression of NK cell-associated markers,but share commonalities in their expression of various monocyte/DC-associated markers. Using CD7,we observed approximately 60% of CD4+CD56+ cells were CD7(neg) cells,indicating the actual frequency of activated CD4+ NK cells is much lower in the blood than previously recognized. Functionally,only CD7+ NK cells secrete gamma interferon (IFNgamma) and degranulate after interleukin-12 (IL-12) plus IL-18 or K562 target cell stimulation. Furthermore,using CD7 to separate CD56+ NK cells and CD56+ myeloid cells,we demonstrate that unlike resting CD7+CD56+ NK cells,the CD7(neg)CD56+ myeloid cells stimulate a potent allogeneic response. Our data indicate that CD7 and CD56 coexpression discriminates NK cells from CD7(neg)CD56+ monocyte/DC-like cells,thereby improving our ability to study the intricacies of NK-cell subset phenotypes and functions in vivo. View Publication

Leucocyte function-associated antigen-1 (LFA-1) is known to be involved in immune reactions leading to allograft rejection. The role of deactivating LFA-1 in this context has not been investigated yet,although it is accepted that regulating LFA-1 activity is essential for T-cell function. Expressing LFA-1 locked in an active state in mice (LFA-1(d/d)) allowed us to investigate the in vivo function of LFA-1 deactivation for allograft rejection in a model of heterotopic cardiac transplantation. We provide in vivo evidence that regulating LFA-1 activity from an active to an inactive state controls antigen-specific priming and proliferation of T cells in response to allogeneic stimuli. Consequently,defective LFA-1 deactivation significantly prolonged cardiac allograft survival. Furthermore,reduced numbers of alloantigen-specific T cells and non-allo-specific innate immune cells within allografts of LFA-1(d/d) recipients indicate that expression of active LFA-1 impairs inflammatory responses involving all major leucocyte subpopulations. Taken together,our in vivo data suggest that LFA-1 deactivation is important for the formation of inflammatory lesions and rejection of cardiac allografts. Thus,the dynamic regulation of LFA-1 activity,rather than the mere presence of LFA-1,appears to contribute to the control of immune reactions inducing allogeneic transplant rejection. View Publication

View Publication

Progressive multifocal leukoencephalopathy (PML) is an often fatal demyelinating disease caused by lytic infection of oligodendrocytes with JC virus (JCV). The development of PML in non-immunosuppressed individuals is a growing concern with reports of mortality in patients treated with mAb therapies. JCV can persist in the kidneys,lymphoid tissue and bone marrow. JCV gene expression is restricted by non-coding viral regulatory region sequence variation and cellular transcription factors. Because JCV latency has been associated with cells undergoing haematopoietic development,transcription factors previously reported as lymphoid specific may regulate JCV gene expression. This study demonstrates that one such transcription factor,Spi-B,binds to sequences present in the JCV promoter/enhancer and may affect early virus gene expression in cells obtained from human brain tissue. We identified four potential Spi-B-binding sites present in the promoter/enhancer elements of JCV sequences from PML variants and the non-pathogenic archetype. Spi-B sites present in the promoter/enhancers of PML variants alone bound protein expressed in JCV susceptible brain and lymphoid-derived cell lines by electromobility shift assays. Expression of exogenous Spi-B in semi- and non-permissive cells increased early viral gene expression. Strikingly,mutation of the Spi-B core in a binding site unique to the Mad-4 variant was sufficient to abrogate viral activity in progenitor-derived astrocytes. These results suggest that Spi-B could regulate JCV gene expression in susceptible cells,and may play an important role in JCV activity in the immune and nervous systems. View Publication

Greater than 75% of all hematologic malignancies derive from germinal center (GC) or post-GC B cells,suggesting that the GC reaction predisposes B cells to tumorigenesis. Because GC B cells acquire expression of the highly mutagenic enzyme activation-induced cytidine deaminase (AID),GC B cells may require additional DNA repair capacity. The goal of this study was to investigate whether normal human B cells acquire enhanced expression of DNA repair factors upon AID induction. We first demonstrated that several DNA mismatch repair,homologous recombination,base excision repair,and ATR signaling genes were overexpressed in GC B cells relative to naïve and memory B cells,reflecting activation of a process we have termed somatic hyperrepair (SHR). Using an in vitro system,we next characterized activation signals required to induce AID expression and SHR. Although AID expression was induced by a variety of polyclonal activators,SHR induction strictly required signals provided by contact with activated CD4+ T cells,and B cells activated in this manner displayed reduced levels of DNA damage-induced apoptosis. We further show the induction of SHR is independent of AID expression,as GC B cells from AID-/-mice retained heightened expression of SHR proteins. In consideration of the critical role that CD4+ T cells play in inducing the SHR process,our data suggest a novel role for CD4+ T cells in the tumor suppression of GC/post-GC B cells. View Publication

Pseudo-Pelger-Huët anomaly (PPHA) has been documented in association with transplant medications and other drugs. This iatrogenic neutrophilic dysplasia is reversible with cessation or adjustment of medications but is frequently confused with myelodysplastic syndrome (MDS) based on the conventional concept that PPHA is a marker for dysplasia. We investigated the clinicopathologic features in iatrogenic PPHA and compared them with MDS-related PPHA. The 13 cases studied included 5 bone marrow/stem cell transplantations,3 solid organ transplantations,1 autoimmune disease,3 chronic lymphocytic leukemias,and 1 breast carcinoma. For 12 cases,there was follow-up evaluation,and all demonstrated at least transient normalization of neutrophilic segmentation. All 9 cases of MDS demonstrated at least 2 of the following pathologic abnormalities on bone marrow biopsy: hypercellularity (8/9),morphologic dysplasia (8/9),clonal cytogenetic abnormality (7/9),and increased blasts (3/9),whereas these abnormalities were typically absent in iatrogenic PPHA. Iatrogenic PPHA displayed a higher proportion of circulating PPHA cells than in MDS (mean,47.4%; SD,31.6% vs mean,12.3%; SD,9.8; P textless .01). A diagnostic algorithm is proposed in which isolated PPHA is indicative of transient or benign PPHA unless proven otherwise. View Publication

Although most novel tuberculosis (TB) vaccines are designed for delivery via the muscle or skin for enhanced protection in the lung,it has remained poorly understood whether systemic vaccine-induced memory T cells can readily home to the lung mucosa prior to and shortly after pathogen exposure. We have investigated this issue by using a model of parenteral TB immunization and intravascular immunostaining. We find that systemically induced memory T cells are restricted to the blood vessels in the lung,unable to populate either the lung parenchymal tissue or the airway under homeostatic conditions. We further find that after pulmonary TB infection,it still takes many days before such T cells can enter the lung parenchymal tissue and airway. We have identified the acquisition of CXCR3 expression by circulating T cells to be critical for their entry to these lung mucosal compartments. Our findings offer new insights into mucosal T cell biology and have important implications in vaccine strategies against pulmonary TB and other intracellular infections in the lung. View Publication

Efforts to improve the efficacy of adoptive T-cell therapies and immune checkpoint therapies in myelogenous leukemia are desired. In this study,we evaluated the antileukemia activity of adoptively transferred polyclonal cancer antigen-reactive T cells deficient in the regulator diacylglycerol kinase zeta (DGKζ) with or without PD-1/PD-L1 blockade. In the C1498 mouse model of myeloid leukemia,we showed that leukemia was eradicated more effectively in DGKζ-deficient (DGKζ-/-) mice than wild-type mice. T cells transferred from DGKζ-deficient mice to wild-type tumor-bearing recipients conferred this benefit. Leukemia clearance was similar to mice treated with anti-PD-L1. Strikingly,we found that the activity of adoptively transferred DGKζ-/- T cells relied partly on induction of sustainable host T-cell immunity. Transferring DGKζ-deficient T cells increased the levels of IFNγ and other cytokines in recipient mice,especially with coadministration of anti-PD-L1. Overall,our results offered evidence that targeting DGKζ may leverage the efficacy of adoptive T-cell and immune checkpoint therapies in leukemia treatment. Furthermore,they suggest that DGKζ targeting might decrease risks of antigen escape or resistance to immune checkpoint blockade. Cancer Res; 77(20); 5676-86. textcopyright2017 AACR. View Publication

IL2 is an immunostimulatory cytokine for key immune cells including T cells and natural killer (NK) cells. Systemic IL2 supplementation could enhance NK-mediated immunity in a variety of diseases ranging from neoplasms to viral infection. However,its systemic use is restricted by its serious side effects and limited efficacy due to activation of T regulatory cells (Tregs). IL2 signaling is mediated through interactions with a multi-subunit receptor complex containing IL2Rα,IL2Rβ,and IL2Rγ. Adult natural killer (NK) cells express only IL2Rβ and IL2Rγ subunits and are therefore relatively insensitive to IL2. To overcome these limitations,we created a novel chimeric IL2-IL2Rβ fusion protein of IL2 and its receptor IL2Rβ joined via a peptide linker (CIRB). NK92 cells expressing CIRB (NK92CIRB) were highly activated and expanded indefinitely without exogenous IL2. When compared with an IL2-secreting NK92 cell line,NK92CIRB were more activated,cytotoxic,and resistant to growth inhibition. Direct contact with cancer cells enhanced the cytotoxic character of NK92CIRB cells,which displayed superior in vivo antitumor effects in mice. Overall,our results showed how tethering IL2 to its receptor IL2Rβ eliminates the need for IL2Rα and IL2Rβ,offering a new tool to selectively activate and empower immune therapy. Cancer Res; 77(21); 5938-51. textcopyright2017 AACR. View Publication