HLA-G expression defines a novel regulatory T-cell subset present in human peripheral blood and sites of inflammation

Blood ◽  
2007 ◽  
Vol 110 (2) ◽  
pp. 568-577 ◽  
Author(s):  
Ute Feger ◽  
Eva Tolosa ◽  
Yu-Hwa Huang ◽  
Anne Waschbisch ◽  
Tilo Biedermann ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
T Cells ◽  
Cerebrospinal Fluid ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Inflammatory Responses ◽  
Human Peripheral Blood ◽  
Cell Subset ◽  
Single Positive

Abstract Regulatory T cells can inhibit harmful immunopathologic responses directed against self and foreign antigens and play a major role in controlling autoimmunity. Here we have identified and characterized a subpopulation of CD4 and CD8 T cells in human peripheral blood expressing the immune tolerizing molecule HLA-G. HLA-G–expressing T cells are hypoproliferative, are CD25- and FOXP3-negative, and exhibit potent suppressive properties that are partially mediated by HLA-G. HLA-G–positive (HLA-Gpos) T cells are found at low percentages among CD4 and CD8 single-positive thymocytes, suggesting a thymic origin. The presence of HLA-Gpos T cells at sites of inflammation such as inflamed skeletal muscle in myositis or the cerebrospinal fluid of patients with acute neuroinflammatory disorders suggests an important function in modulating parenchymal inflammatory responses in vivo.

scholarly journals The CD8+ Dendritic Cell Subset Selectively Endocytoses Dying Cells in Culture and In Vivo

2002 ◽  
Vol 195 (10) ◽  
pp. 1289-1302 ◽  
Author(s):  
Tomonori Iyoda ◽  
Susumu Shimoyama ◽  
Kang Liu ◽  
Yoshiki Omatsu ◽  
Yuji Akiyama ◽  
...  
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
Ex Vivo ◽  
Osmotic Shock ◽  
Cell Subset ◽  
Fluorescent Labeling ◽  
Dendritic Cell Subset ◽  
Dying Cells

Dendritic cells (DCs) are able in tissue culture to phagocytose and present antigens derived from infected, malignant, and allogeneic cells. Here we show directly that DCs in situ take up these types of cells after fluorescent labeling with carboxyfluorescein succinimidyl ester (CFSE) and injection into mice. The injected cells include syngeneic splenocytes and tumor cell lines, induced to undergo apoptosis ex vivo by exposure to osmotic shock, and allogeneic B cells killed by NK cells in situ. The CFSE-labeled cells in each case are actively endocytosed by DCs in vivo, but only the CD8+ subset. After uptake, all of the phagocytic CD8+ DCs can form major histocompatibility complex class II–peptide complexes, as detected with a monoclonal antibody specific for these complexes. The CD8+ DCs also selectively present cell-associated antigens to both CD4+ and CD8+ T cells. Similar events take place with cultured DCs; CD8+ DCs again selectively take up and present dying cells. In contrast, both CD8+ and CD8− DCs phagocytose latex particles in culture, and both DC subsets present soluble ovalbumin captured in vivo. Therefore CD8+ DCs are specialized to capture dying cells, and this helps to explain their selective ability to cross present cellular antigens to both CD4+ and CD8+ T cells.

Autoproliferation Contributes to Clonal Expansion and Changes In Cellular Topography In T Cell Large Granular Lymphocyte (LGL) Leukemia

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1373-1373
Author(s):  
JianXiang Zou ◽  
Jeffrey S Painter ◽  
Fanqi Bai ◽  
Lubomir Sokol ◽  
Thomas P. Loughran ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Cd8 T Cell ◽  
Effector Memory ◽  
Ki67 Expression ◽  
Terminal Effector ◽  
Lgl Leukemia

Abstract Abstract 1373 Introduction: LGL leukemia is associated with cytopenias and expansion of clonally-derived mature cytotoxic CD8+ lymphocytes. The etiology of LGL leukemia is currently unknown, however, T cell activation, loss of lymph node homing receptor L-selectin (CD62L), and increased accumulation of T cells in the bone marrow may lead to suppressed blood cell production. The broad resistance to Fas (CD95) apoptotic signals has lead to the hypothesis that amplification of clonal cells occurs through apoptosis resistance. However, the proliferative history has not been carefully studied. To define possible mechanism of LGL leukemia expansion, T cell phenotype, proliferative history, and functional-related surface marker expression were analyzed. Methods: Peripheral blood mononuclear cells (PBMCs) were obtained from 16 LGL leukemia patients that met diagnostic criteria based on the presence of clonal aβ T cells and >300 cells/ml CD3+/CD57+ T cells in the peripheral blood. Samples were obtained from 10 age-matched healthy individuals from the Southwest Florida Blood Services for comparisons. Multi-analyte flow cytometry was conducted for expression of CD3, CD4/8, CD45RA, CD62L, CD27, CD28, CD25, CD127, IL15Ra, IL21a, CCR7 (all antibodies from BD Biosciences). The proliferative index was determined by Ki67 expression in fixed and permeabilized cells (BD Biosciences) and the proliferative history in vivo was assessed by T-cell-receptor excision circle (TREC) measurement using real-time quantitative PCR (qRT-PCR) in sorted CD4+ and CD8+ T cells. TRECs are episomal fragments generated during TCR gene rearrangements that fail to transfer to daughter cells and thus diminish with each population doubling that reflects the in vivo proliferative history. Results: Compared to healthy controls, significantly fewer CD8+ naïve cells (CD45RA+/CD62L+, 8.4 ± 10.8 vs 24.48 ± 11.99, p=0.003) and higher CD8+ terminal effector memory (TEM) T cells (CD45RA+/CD62L-, 67.74 ± 28.75 vs 39.33 ± 11.32, p=0.007) were observed in the peripheral blood. In contrast, the percentage of CD4+ naïve and memory cells (naïve, central memory, effector memory, and terminal effector memory based on CD45RA and CD62L expression) was similar in patients as compared to controls. The expression of CD27 (31.32 ± 34.64 vs 71.73 ± 20.63, p=0.003) and CD28 (31.38 ± 31.91 vs 70.02 ± 22.93, p=0.002) were lower in CD8+ T cell from patients with LGL leukemia and this reduction predominated within the TEM population (17.63±24.5 vs 70.98±22.5 for CD27, p<0.0001 and 13±20.5 vs 69.43± 21.59 for CD28, p<0.0001). Loss of these markers is consistent with prior antigen activation. There was no difference in CD25 (IL2Ra, p=0.2) expression on CD4+ or CD8+ T cells, but CD127 (IL7Ra, p=0.001), IL15Ra, and IL21Ra (p=0.15) were overexpressed in TEM CD8+ T cell in patients vs controls. All of these cytokine receptors belong to the IL2Rβg-common cytokine receptor superfamily that mediates homeostatic proliferation. In CD8+ T cells in patients, the IL-21Ra was also overexpressed in naïve, central and effector memory T cells. The topography of the expanded CD8+ T cell population was therefore consistent with overexpression of activation markers and proliferation-associated cytokine receptors. Therefore, we next analyzed Ki67 expression and TREC DNA copy number to quantify actively dividing cells and determine the proliferative history, respectively. We found that LGL leukemia patients have more actively dividing CD8+ TEM T cells compared to controls (3.2 ± 3.12 in patients vs 0.44 ± 0.44 in controls, p=0.001). Moreover, the TREC copy number in CD8+ T cells was statistically higher in healthy individuals after adjusting for age (177.54 ± 232 in patients vs 1015 ± 951 in controls, p=0.019). These results show that CD8+ cells in the peripheral compartment have undergone more population doublings in vivo compared to healthy donors. In contrast, the TREC copies in CD4+ T-cells were similar between LGL patients and controls (534.4 ± 644 in patients vs 348.78 ± 248.16 in controls, p>0.05) demonstrating selective cellular proliferation within the CD8 compartment. Conclusions: CD8+ T- cells are undergoing robust cellular activation, contraction in repertoire diversity, and enhanced endogenous proliferation in patients with LGL leukemia. Collectively, these results suggest that clonal expansion is at least partially mediated through autoproliferation in T-LGL leukemia. Disclosures: No relevant conflicts of interest to declare.

Anti-Thymocyte Globulin (ATG) Differentially Depletes naïve and Memory T Cells and Permits Memory-Type Regulatory T Cells

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4670-4670
Author(s):  
Chang-Qing Xia ◽  
Anna Chernatynskaya ◽  
Clive Wasserfall ◽  
Benjamin Looney ◽  
Suigui Wan ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Memory T Cells ◽  
Conflicts Of Interest ◽  
T Cell Subsets ◽  
Hematopoietic Stem

Abstract Abstract 4670 Anti-thymocyte globulin (ATG) has been used in clinic for the treatment of allograft rejection and autoimmune diseases. However, its mechanism of action is not fully understood. To our knowledge, how ATG therapy affects naïve and memory T cells has not been well investigated. In this study, we have employed nonobese diabetic mouse model to investigate how administration of anti-thymocyte globulin (ATG) affects memory and naïve T cells as well as CD4+CD25+Foxp3+ regulatory T cells in peripheral blood and lymphoid organs; We also investigate how ATG therapy affects antigen-experienced T cells. Kinetic studies of peripheral blood CD4+ and CD8+ T cells post-ATG therapy shows that both populations decline to their lowest levels at day 3, while CD4+ T cells return to normal levels more rapidly than CD8+ T cells. We find that ATG therapy fails to eliminate antigen-primed T cells, which is consistent with the results that ATG therapy preferentially depletes naïve T cells relative to memory T cells. CD4+ T cell responses post-ATG therapy skew to T helper type 2 (Th2) and IL-10-producing T regulatory type 1 (Tr1) cells. Intriguingly, Foxp3+ regulatory T cells (Tregs) are less sensitive to ATG depletion and remain at higher levels following in vivo recovery compared to controls. Of note, the frequency of Foxp3+ Tregs with memory-like immunophenotype is significantly increased in ATG-treated animals, which might play an important role in controlling effector T cells post ATG therapy. In summary, ATG therapy may modulate antigen-specific immune responses through modulation of naïve and memory T cell pools and more importantly through driving T cell subsets with regulatory activities. This study provides important data for guiding ATG therapy in allogenieic hematopoietic stem cell transplantation and other immune-mediated disorders. Disclosures: No relevant conflicts of interest to declare.

A Mouse Model for XLP-2 Disease Uncovers a Critical Function for IL-1beta and TNF in Driving Hyper-Inflammation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 1403-1403
Author(s):  
Philipp J. Jost ◽  
Monica Yabal ◽  
Heiko Adler ◽  
Nathalie Knies ◽  
Christina Groß ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
T Lymphocytes ◽  
Viral Infections ◽  
Inflammatory Responses ◽  
Conflicts Of Interest ◽  
Human Peripheral Blood ◽  
Critical Function ◽  
Deficient Mice

Abstract The hyper-inflammatory syndrome X-linked lymphoproliferative syndrome type 2 (XLP-2) is defined by mutations in BIRC4 (XIAP). XLP-2 is often diagnosed in paediatric patients and is characterized by hyper-inflammation triggered by common viral infections. Symptoms include splenomegaly, HLH, fevers, and chronic haemorrhagic colitis among others. Recent work has also shown that mutations in BIRC4 predispose to the development of early-onset IBD, which is not necessarily associated with symptoms of systemic hyper-inflammation. Symptoms of XLP-2 are mostly attributed to the aberrant activation of macrophages and dendritic cells (DC) and the subsequent accumulation of activated T-lymphocytes. We have characterized the inflammatory response of mice deficient for BIRC4 (XIAP) to viral infections with the murine herpes virus 68 (MHV-68) as the closest murine model for human EBV-driven mononucleosis. Xiap-/- mice were capable of clearing the virus normally during early infection (day 6, 16), but failed to do so during the course of the infection measured as elevated viral genomic loads during late (day 43) and very late (day 84) latency. Xiap-/- mice responded to intranasal application of the virus with systemic hyper-inflammation exemplified by elevated IL-1beta levels, splenomegaly and increased activation of peripheral T lymphocytes such as CD4+ effector T cells, regulatory T cells (Treg), and IFNg+ T cells. In previous work, we have shown that TNF is critically required to drive the hyper-inflammatory phenotype of macrophages and dendritic cells of XIAP-deficient mice. Indeed, genetic deletion of TNF in vivo or, alternatively, anti-TNF treatment in vivo using Eternacept (Enbrel) ameliorated the symptoms of XIAP-deficient mice in response to viral infection. Elevated IL-1beta levels were also observed in human peripheral blood-derived monocytes from XLP-2 patients (7 patients from 5 different families) when compared to healthy controls. In conclusion, this data supports the notion that anti-TNF treatment might be able to ameliorate the hyper-inflammatory responses in XLP-2 patients, when used early during an infection. Disclosures No relevant conflicts of interest to declare.

scholarly journals Interleukin 2 induction of pore-forming protein gene expression in human peripheral blood CD8+ T cells.

1990 ◽  
Vol 171 (4) ◽  
pp. 1269-1281 ◽  
Author(s):  
M J Smyth ◽  
J R Ortaldo ◽  
Y Shinkai ◽  
H Yagita ◽  
M Nakata ◽  
...  
Keyword(s):  
T Cells ◽  
Mrna Expression ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Interleukin 2 ◽  
Cytotoxic Lymphocytes ◽  
Mrna Levels ◽  
Cytotoxic Potential

Our studies have analyzed pore-forming protein (PFP) mRNA expression in resting and stimulated human peripheral blood CD3- large granular lymphocytes (LGL), CD3+ T cells, and their CD4+ or CD8+ subsets. Signals that stimulate T cells to develop cytotoxic activity (i.e., IL-2 or OKT-3 mAb) led to the induction of PFP mRNA in T cells. The data indicated that IL-2 directly increased PFP mRNA in the CD8+ subset of T cells, in the absence of new DNA or protein synthesis. Abrogation of IL-2-induced PFP mRNA expression and cytotoxic potential of T cells by the anti-p75 IL-2 receptor mAb suggested that low numbers of p75 IL-2 receptors on CD8+ T cells were capable of transducing signals responsible for these IL-2-induced effects. The induction of T cell PFP mRNA via CD3, using OKT-3 mAb, was less rapid but greater than that caused by IL-2; however, a combination of PMA and ionomycin, which bypasses crosslinking of the TCR/CD3 complex, could not mimic this increase in PFP mRNA levels in T cells. The role of second messenger systems in regulating PFP mRNA expression remains to be determined. In contrast, high constitutive PFP mRNA expression was observed in CD3- LGL and these mRNA levels could not be enhanced by stimulation with IL-2. The cytotoxic potential of peripheral blood T cells and LGL induced in response to IL-2 correlated with IL-2-induced PFP mRNA levels in these cells and was consistent with PFP being one of several important molecules involved in the effector function of cytotoxic lymphocytes.

scholarly journals Identification of Cytolytic CD161−CD56+ Regulatory CD8 T Cells in Human Peripheral Blood

PLoS ONE ◽  
2013 ◽  
Vol 8 (3) ◽  
pp. e59545 ◽  
Author(s):  
Dan Hu ◽  
Howard L. Weiner ◽  
Jerome Ritz
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Human Peripheral Blood

scholarly journals Constitutive expression of pore-forming protein in peripheral blood gamma/delta T cells: implication for their cytotoxic role in vivo.

1990 ◽  
Vol 172 (6) ◽  
pp. 1877-1880 ◽  
Author(s):  
M Nakata ◽  
M J Smyth ◽  
Y Norihisa ◽  
A Kawasaki ◽  
Y Shinkai ◽  
...  
Keyword(s):  
T Cells ◽  
Cytotoxic Activity ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Constitutive Expression ◽  
Immunocytochemical Staining ◽  
Target Cells ◽  
Gamma Delta T Cells ◽  
Gamma Delta

The cytotoxic activity and pore-forming protein (PFP) expression of human peripheral blood (PB) gamma/delta T cells were examined. Fresh gamma/delta T cells isolated from PB lymphocytes by fluorescence-activated cell sorting exhibited a substantial natural killer-like cytotoxic activity against K562 target cells and had a high cytotoxic potential triggered by anti-CD3 monoclonal antibody (mAb) against P815 target cells bearing Fc gamma R. Immunocytochemical staining with an anti-PFP mAb revealed that virtually all PB gamma/delta T cells are granular lymphocytes with abundant PFP in their cytoplasmic granules. Constitutive expression of PFP in PB gamma/delta T cells was also demonstrated by Northern blot analysis. These observations support the proposed role of gamma/delta T cells in cytolytic immune surveillance in vivo.

Apoptotic Pathways of Human Regulatory T Cells in Chronic Graft Versus Host Disease (cGVHD) After Allogeneic Hematopoietic Stem Cell Transplantation (HSCT)

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 225-225
Author(s):  
Kazuyuki Murase ◽  
Yutaka Kawano ◽  
Jeremy Ryan ◽  
Ken-ichi Matsuoka ◽  
Gregory Bascug ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Low Dose ◽  
Cell Subset ◽  
Mitochondrial Pathway ◽  
T Cell Subsets ◽  
T Cell Subset ◽  
Mitochondrial Membrane Depolarization

Abstract Abstract 225 CD4+CD25+Foxp3+ regulatory T cells (Treg) are known to play a central role in the maintenance of self-tolerance and immune homeostasis. After allogeneic stem cell transplantation, impaired recovery of Treg is associated with the development of cGVHD. Interleukin-2 (IL-2) is a critical regulator of Treg development, expansion and survival and lack of IL-2 results in Treg deficiency. In patients with cGVHD, we previously demonstrated that Treg proliferate at high levels but this subset is also highly susceptible to apoptosis leading to inadequate Treg numbers (Matsuoka et al. JCI 2010). We also reported that low-dose IL-2 administration resulted in selective expansion of Treg in vivo and clinical improvement of cGVHD (Koreth et al. NEJM 2011). To identify mechanisms responsible for increased Treg susceptibility to apoptosis in cGVHD we used a new flow cytometry-based assay to measure mitochondrial membrane depolarization in response to a panel of pro-apoptotic BH3 peptides (BIM, BID, BAD, NOXA, PUMA, BMF, HRK). This assessment allowed us to compare BH3 peptide-induced mitochondrial membrane depolarization (“priming”) in different T cell subsets, including CD4 Treg, conventional CD4 T cells (CD4 Tcon), and CD8 T cells. Expression of Bcl-2, CD95 and Ki67 were also studied in each T cell subset. We studied peripheral blood samples from 36 patients with hematologic malignancies (median age 59 yr) who are > 2 years post HSCT (27 patients with cGVHD and 9 patients without cGVHD) and 15 patients who received daily subcutaneous IL-2 for 8 weeks for treatment of steroid-refractory cGvHD. Severity of cGVHD was classified according to NIH criteria. In patients without cGVHD, BH3 priming was similar in all 3 T cell subsets (CD4 Treg, CD4 Tcon and CD8). In patients with cGVHD, CD4 Treg were more primed than CD4 Tcon when challenged with BIM, BAD, PUMA, BMF and the combination of BAD + NOXA peptides (p<0.01 – 0.0001). Treg were more primed than CD8 T cells when challenged with PUMA peptide (p<0.0001), but priming in Treg and CD8 T cells was similar for other BH3 peptides in patients with cGVHD. We also compared BH3 priming of each T cell subset in patients with different grades of cGVHD. When challenged with BH3 peptides, Treg, Tcon and CD8 T cells were less primed in patients with severe cGVHD. In patients with cGVHD, Treg expressed higher levels of Ki-67, higher levels of CD95 and lower levels of Bcl-2 than Tcon. Expression of CD95 did not vary with severity of GVHD in any T cell subset, but expression of Bcl-2 was significantly increased in all subsets in patients with severe cGVHD. Increased BH3 priming and high expression of CD95 indicate that Treg are more susceptible to apoptosis than Tcon in cGVHD. However, both Treg and Tcon become less primed and Bcl-2 levels increase in severe cGVHD suggesting that these cells are less susceptible to mitochondrial pathway apoptosis. Since the total number of Treg and Tcon are significantly reduced in patients with cGVHD, these findings suggest that the remaining circulating cells are relatively resistant to mitochondrial pathway apoptosis. CD95 expression in Treg remains high indicating no change in death receptor pathway apoptosis. Daily treatment with low-dose IL-2 for 8 weeks selectively expands Treg in vivo in patients with severe cGVHD. As the number of Treg increase, BH3 profiling shows that these cells gradually become more primed and therefore more susceptible to mitochondrial pathway apoptosis. Taken together, these studies help define the complex and distinct pathways that regulate survival in different T cell subsets and changes in these pathways that occur in patients with chronic GVHD. These pathways play important roles in the maintenance of T cell homeostasis and targeting these complex pathways can provide new opportunities to promote immune tolerance after allogeneic HSCT. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Human plasmacytoid dendritic cells efficiently cross-present exogenous Ags to CD8+ T cells despite lower Ag uptake than myeloid dendritic cell subsets

Blood ◽  
2013 ◽  
Vol 121 (3) ◽  
pp. 459-467 ◽  
Author(s):  
Jurjen Tel ◽  
Gerty Schreibelt ◽  
Simone P. Sittig ◽  
Till S. M. Mathan ◽  
Sonja I. Buschow ◽  
...  
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Human Peripheral Blood ◽  
Adaptive Immune System ◽  
Plasmacytoid Dendritic Cells ◽  
Cell Responses ◽  
Adaptive Immune ◽  
Antitumor Immunotherapy

Abstract In human peripheral blood, 4 populations of dendritic cells (DCs) can be distinguished, plasmacytoid dendritic cells (pDCs) and CD16+, CD1c+, and BDCA-3+ myeloid DCs (mDCs), each with distinct functional characteristics. DCs have the unique capacity to cross-present exogenously encountered antigens (Ags) to CD8+ T cells. Here we studied the ability of all 4 blood DC subsets to take up, process, and present tumor Ags to T cells. Although pDCs take up less Ags than CD1c+ and BDCA3+ mDCs, pDCs induce potent Ag-specific CD4+ and CD8+ T-cell responses. We show that pDCs can preserve Ags for prolonged periods of time and on stimulation show strong induction of both MHC class I and II, which explains their efficient activation of both CD4+ and CD8+ T cells. Furthermore, pDCs cross-present soluble and cell-associated tumor Ags to cytotoxic T lymphocytes equally well as BDCA3+ mDCs. These findings, and the fact that pDCs outnumber BDCA3+ mDCs, both in peripheral blood and lymph nodes, together with their potent IFN-I production, known to activate both components of the innate and adaptive immune system, put human pDCs forward as potent activators of CD8+ T cells in antitumor responses. Our findings may therefore have important consequences for the development of antitumor immunotherapy.

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