Induction and inhibition of human lymphocyte transformation by the lectin from the red marine alga Amansia multifida

Lectins are powerful stimulants of quiescent peripheral blood lymphocytes. They can induce blast transformation leading to mitosis of these cells in vitro. We report here the dose-dependent proliferative curve for human peripheral blood monouclear cells (PBMC) stimulated by the lectin amansin, from Amansia multifida. Amansin stimulated proliferation of (PBMC) at relatively low concentrations (3.12 to 12.5 μg mL^-1). We observed also a gradual reduction in mitogenic capacity with progressive increase in the lectin concentration above 12.5 μg mL^-1. This decrease in the mitogenic potential did not result from a toxic effect on the cells, and was predominant at a lectin concentration above 50 μg mL^-1. This decrease in lymphocyte proliferation could be blocked by avidin and could not be overcome by IL-2 or another lectin (Con Br) at stimulatory concentrations. Additionally, we observed that cells incubated at stimulatory concentrations of amansin produced IFN-γ. Analysis of the culture supernatants established a direct correlation between the IFN-γ and the mitogenic and anti-mitogenic capacity of amansin. This revised version was published online in June 2006 with corrections to the Cover Date.     

T lymphocyte cytokine profile at a single cell level in alveolar Echinococcosis.

The metacestode Echinococcus multilocularis causes a life-threatening disease in humans, named alveolar echinococcosis (AE). A comparative analysis of the early activation marker CD69 on peripheral blood mononuclear cells (PBMC) of patients with AE and healthy controls after in vitro culture with crude E. multilocularis antigen revealed that specific expression of CD69 was induced in CD4(+)T lymphocytes as well as in CD8(+)T lymphocytes. Using a protocol for intracellular staining of cytokines followed by fluorescence activating cell sorting (FACS) analysis, production of interleukin (IL)-2, IL-5 and IL-10 was detected in CD4(+)as well as in CD8(+)lymphocytes. Most notably, there was a definite increase in the expression of IL-10 in CD8(+) lymphocytes from patients with alveolar echinococcosis. The data support an important role of CD8(+) lymphocytes in the long persistence of the metacestode.     

Antibodies against the CD44 p80, lymphocyte homing receptor molecule augment human peripheral blood T cell activation

The CD44 inhibitor Lutheran [In(Lu)]-related p80 molecule has recently been shown to be identical to the Hermes-1 lymphocyte homing receptor and to the human Pgp-1 molecule. We have determined the effect of addition of CD44 antibodies to in vitro activation assays of PBMC. CD44 antibodies did not induce PBMC proliferation alone, but markedly enhanced PBMC proliferation induced by a mitogenic CD2 antibody pair or by CD3 antibody. CD44 antibody addition had no effect upon PBMC activation induced by PHA or tetanus toxoid. CD44 antibody enhancement of CD2 antibody-induced T cell activation was specific for mature T cells as thymocytes could not be activated in the presence of combinations of CD2 and CD44 antibodies. CD44 antibody enhancement of CD2-mediated T cell triggering occurred if CD44 antibody was placed either on monocytes or on T cells. In experiments with purified monocyte and T cell suspensions, CD44 antibodies A3D8 and A1G3 augmented CD2-mediated T cell activation by three mechanisms. First, CD44 antibody binding to monocytes induced monocyte IL-1 release, second, CD44 antibodies enhanced the adhesion of T cells and monocytes in CD2 antibody-stimulated cultures, and third, CD44 antibodies augmented T cell IL-2 production in response to CD2 antibodies. Thus, ligand binding to CD44 molecules on T cells and monocytes may regulate numerous events on both cell types that are important for T cell activation. Given that recent data suggest that the CD44 molecule may bind to specific ligands on endothelial cells (vascular addressin) and within the extracellular matrix (collagen, fibronectin), these data raise the possibility that binding of T cells to endothelial cells or extracellular matrix proteins may induce or up-regulate T cell activation in inflammatory sites.     

CD11b expression identifies CD8+CD28+ T lymphocytes with phenotype and function of both naive/memory and effector cells

A previously unreported CD8(+)CD28(+)CD11b(+) T cell subset occurs in healthy individuals and expands in patients suffering from primary viral infections. In functional terms, these cells share the features of naive/memory CD8(+)CD28(+)CD11b(-) and terminally differentiated effector CD8(+)CD28(-)CD11b(+) subpopulations. Like CD28(-) cells, CD28(+)CD11b(+) lymphocytes have the ability to produce IFN-gamma, to express perforin granules in vivo, and to exert a potent cytolytic activity. Moreover, these cells can respond to chemotactic stimuli and can efficiently cross the endothelial barrier. In contrast, like their CD11b(-) counterpart, they still produce IL-2 and retain the ability to proliferate following mitogenic stimuli. The same CD28(+)CD11b(+) subpopulation detected in vivo could be generated by culturing naive CD28(+)CD11b(-) cells in the presence of mitogenic stimuli following the acquisition of a CD45RO(+) memory phenotype. Considering both phenotypic and functional properties, we argue that this subset may therefore constitute an intermediate phenotype in the process of CD8(+) T cell differentiation and that the CD11b marker expression can distinguish between memory- and effector-type T cells in the human CD8(+)CD28(+) T cell subset.     

The capsular polysaccharides of Cryptococcus neoformans activate normal CD4(+) T cells in a dominant Th2 pattern.

Capsular components of Cryptococcus neoformans induce several deleterious effects on T cells. However, it is unknown how the capsular components act on these lymphocytes. The present study characterized cellular and molecular events involved in immunoregulation of splenic CD4(+) T cells by C. neoformans capsular polysaccharides (CPSs). The results showed that CPSs induce proliferation of normal splenic CD4(+) T cells, but not of normal CD8(+) T or B lymphocytes. Such proliferation depended on physical contact between CPSs and viable splenic adherent cells (SAC) and CD40 ligand-induced intracellular signal transduction. The absence of lymphoproliferation after fixation of SAC with paraformaldehyde has discarded the hypothesis of a superantigen-like activation. The evaluation of a cytokine pattern produced by the responding CD4(+) T lymphocytes revealed that CPSs induce a dominant Th2 pattern, with high levels of IL-4 and IL-10 production and undetectable inflammatory cytokines, such as TNF-alpha and IFN-gamma. Blockade of CD40 ligand by relevant mAb down-regulated the CPS-induced anti-inflammatory cytokine production and abolished the enhancement of fungus growth in cocultures of SAC and CD4(+) T lymphocytes. Our findings suggest that CPSs induce proliferation and differentiation of normal CD4(+) T cells into a Th2 phenotype, which could favor parasite growth and thus important deleterious effects to the host.     

Primary human lymphocytes transduced with NY-ESO-1 antigen-specific TCR genes recognize and kill diverse human tumor cell lines

cDNAs encoding TCR alpha- and beta-chains specific for HLA-A2-restricted cancer-testis Ag NY-ESO-1 were cloned using a 5'RACE method from RNA isolated from a CTL generated by in vitro stimulation of PBMC with modified NY-ESO-1-specific peptide (p157-165, 9V). Functionality of the cloned TCR was confirmed by RNA electroporation of primary PBL. cDNA for these alpha- and beta-chains were used to construct a murine stem cell virus-based retroviral vector, and high titer packaging cell lines were generated. Gene transfer efficiency in primary T lymphocytes of up to 60% was obtained without selection using a method of precoating retroviral vectors onto culture plates. Both CD4(+) and CD8(+) T cells could be transduced at the same efficiency. High avidity Ag recognition was demonstrated by coculture of transduced lymphocytes with target cells pulsed with low levels of peptide (<20 pM). TCR-transduced CD4 T cells, when cocultured with NY-ESO-1 peptide pulsed T2 cells, could produce IFN-gamma, GM-CSF, IL-4, and IL-10, suggesting CD8-independent, HLA-A2-restricted TCR activation. The transduced lymphocytes could efficiently recognize and kill HLA-A2- and NY-ESO-1-positive melanoma cell lines in a 4-h (51)Cr release assay. Finally, transduced T cells could efficiently recognize NY-ESO-1-positive nonmelanoma tumor cell lines. These results strongly support the idea that redirection of normal T cell specificity by TCR gene transfer can have potential applications in tumor adoptive immunotherapy.     

Differential expression of a 70 kDa O-glycoprotein on T cells: a possible marker for naive and early activated murine T cells

We purified a 70 kDa O-glycoprotein that binds to the GalNAc specific lectin from Amaranthus leucocarpus (ALLr) and determined its expression pattern on T lymphocytes from different murine lymphoid organs. High level of ALLr expression was demonstrated in 95-98% of both CD4(+)8(+) and CD4(-)8(+) thymocytes, and in 80-95% of CD8(+) T cells from peripheral blood, lymph nodes, and spleen, whereas a minor fraction of CD4(+)8(-) thymocytes (46-67%) and peripheral CD4(+) T cells (9-40%) showed low ALLr expression. Peripheral CD19(+) B cells were ALLr negative and most of the peripheral ALL(+) T cells showed a CD62L(hi)CD45RB(hi)CD44(lo/-) phenotype, indicating features of naive cells. Mitogenic activation of peripheral T cells increased 3-fold the number of ALL(+)CD4(+) T cells 24 h after stimulation, as opposed to a >80% decrease in CD8(+) T cells 72 h after stimulation. Our results suggest that ALL detects a non-described surface O-glycoprotein selectively expressed by naive CD8(+) T cells and by early activated CD4(+) T cells.     

Selective expansion and partial activation of human NK cells and NK receptor-positive T cells by IL-2 and IL-15.

IL-2 and IL-15 are lymphocyte growth factors produced by different cell types with overlapping functions in immune responses. Both cytokines costimulate lymphocyte proliferation and activation, while IL-15 additionally promotes the development and survival of NK cells, NKT cells, and intraepithelial lymphocytes. We have investigated the effects of IL-2 and IL-15 on proliferation, cytotoxicity, and cytokine secretion by human PBMC subpopulations in vitro. Both cytokines selectively induced the proliferation of NK cells and CD56(+) T cells, but not CD56(-) lymphocytes. All NK and CD56(+) T cell subpopulations tested (CD4(+), CD8(+), CD4(-)CD8(-), alphabetaTCR(+), gammadeltaTCR(+), CD16(+), CD161(+), CD158a(+), CD158b(+), KIR3DL1(+), and CD94(+)) expanded in response to both cytokines, whereas all CD56(-) cell subpopulations did not. Therefore, previously reported IL-15-induced gammadelta and CD8(+) T cell expansions reflect proliferations of NK and CD56(+) T cells that most frequently express these phenotypes. IL-15 also expanded CD8alpha(+)beta(-) and Valpha24Vbeta11 TCR(+) T cells. Both cytokines stimulated cytotoxicity by NK and CD56(+) T cells against K562 targets, but not the production of IFN-gamma, TNF-alpha, IL-2, or IL-4. However, they augmented cytokine production in response to phorbol ester stimulation or CD3 cross-linking by inducing the proliferation of NK cells and CD56(+) T cells that produce these cytokines at greater frequencies than other T cells. These results indicate that IL-2 and IL-15 act at different stages of the immune response by expanding and partially activating NK receptor-positive lymphocytes, but, on their own, do not influence the Th1/Th2 balance of adaptive immune responses.     

Purification of Ag-specific T lymphocytes after direct peripheral blood mononuclear cell stimulation followed by CD25 selection. I. Application to CD4(+) or CD8(+) cytomegalovirus phosphoprotein pp65 epitope determination

The two main constraints that currently limit a broader usage of T cell therapy against viruses are the delay required to obtain specific T cells and the safety of the selection procedure. In the present work we developed a generally applicable strategy that eliminates the need for APC for timing reasons, and the need for infectious viral strains for safety concerns. As a model, we used the selection of T lymphocytes specific for the immunodominant CMV phosphoprotein pp65. PBMC from healthy seropositive donors were first depleted of IL-2R alpha-chain CD25(+) cells and were then stimulated for 24-96 h with previously defined peptide Ags or with autologous PBMC infected with a canarypox viral vector encoding the total pp65 protein (ALVAC-pp65). Subsequent immunomagnetic purification of newly CD25-expressing cells allowed efficient recovery of T lymphocytes specific for the initial stimuli, i.e., for the already known immunodominant epitope corresponding to the peptides used as a model or for newly defined epitopes corresponding to peptides encoded by the transfected pp65 protein. Importantly, we demonstrated that direct PBMC stimulation allowed recovery not only of CD8(+) memory T lymphocytes, but also of the CD4(+) memory T cells, which are known to be crucial to ensure persistence of adoptively transferred immune memory. Finally, our analysis of pp65-specific T cells led to the identification of several new helper and cytotoxic epitopes. This work thus demonstrates the feasibility of isolating memory T lymphocytes specific for a clinically relevant protein without the need to prepare APC, to use infectious viral strains, or to identify immunodominant epitopes.     

Lectin ligands on human dendritic cells and identification of a peanut agglutinin positive subset in blood

As only a few cell surface markers for dendritic cells (DC) have been identified to date, this study examined the expression of ligands for lectin on different human DC populations. The ability of Concanavalin A (Con A), Wheat Germ Agglutinin (WGA), peanut agglutinin (PNA), and Helix pomatia (HPA) to bind to cell lines and PBMC and DC populations was analyzed by flow cytometry and specificity of binding confirmed using inhibitory and noninhibitory sugars. The cell lines showed non-lineage-restricted binding with Con A and WGA, independent of sialidase treatment. HPA and PNA bound to a restricted number of lines, but showed broad reactivity after sialidase treatment. The peripheral blood mononuclear cells (PBMC) and directly isolated blood DC, activated CD83(+) blood DC, epidermal Langerhans cells (LC), and monocyte-derived DC (Mo-DC) showed strong binding of Con A and WGA, both before and after sialidase treatment. No HPA binding ligands were detected on PBMC populations, including directly isolated blood DC. Following sialidase treatment CD3(+), CD16(+), and a subset of CD19(+) lymphocytes bound HPA. The lectin PNA bound weakly to CD14(+) monocytes and a subpopulation of circulating DC that were HLA-DR(hi)CDw123 Dr(hi)CDw123(dim)/(neg)CD11c(+). The HLA-DR(mod)CDw123(hi)CD11c(neg) subpopulation did not bind PNA. Without sialidase treatment LC expressed both HPA and PNA ligands, but these were either absent on activated CD83(+) blood DC or weakly expressed on Mo-DC. Following sialidase treatment PBMC populations, activated CD83(+) blood DC, and Mo-DC became PNA positive. Thus human DC express several lectin ligands and PNA binding identifies a subset of blood DC. That may reflect discrete changes associated with stages of DC development or functional maturation.     

Intracellular events involved in CD5-induced human T cell activation and proliferation.

In this report we describe a novel pathway of human T cell activation and proliferation involving the CD5 surface Ag. The CD5-specific Cris1 mAb induces by itself monocyte-dependent proliferation of PBMC. Among a panel of CD5-specific mAb (Leu1, OKT1, LO-CD5a, F101-1C5, and F145GF3), only the F145GF3 mAb shared this property with Cris1. The analysis of the biochemical pathway involved in this activation showed the lack of detectable hydrolysis of inositol phosphates or early increments in the intracellular cytosolic calcium concentration, after triggering cells with the mitogenic CD5 mAb. However, stimulation with CD5 induces activation of protein kinase C, as measured by phosphorylation of a specific peptide substrate (peptide GS), which can be inhibited by a pseudosubstrate peptide inhibitor. Stimulation with CD5 mAb induces also tyrosine kinase activity, with a substrate pattern that differs from that induced after triggering lymphocytes through the TCR-CD3 complex. On the other hand the IL-2/IL-2R pathway seems involved in the CD5-mediated proliferation of PBMC because anti-IL-2R-specific mAb inhibits CD5-induced proliferation, and stimulation with mitogenic CD5 mAb induces production of IL-2 and expression of IL-2R alpha and beta chains. Therefore, the triggering of the CD5 Ag can induce IL-2- and monocyte-dependent human T cell proliferation by a biochemical pathway that differs, at least in the first stages, from the one that mediates TCR-CD3 complex-induced T cell activation.     

CD8+ alpha beta+ T cells that lack surface CD5 antigen expression are a major lymphotactin (XCL1) source in peripheral blood lymphocytes

To better characterize the cellular source of lymphotactin (XCL1), we compared XCL1 expression in different lymphocyte subsets by real-time PCR. XCL1 was constitutively expressed in both PBMC and CD4(+) cells, but its expression was almost 2 log higher in CD8(+) cells. In vitro activation was associated with a substantial increase in XCL1 expression in both PBMC and CD8(+) cells, but not in CD4(+) lymphocytes. The preferential expression of XCL1 in CD8(+) cells was confirmed by measuring XCL1 production in culture supernatants, and a good correlation was found between figures obtained by real-time PCR and XCL1 contents. XCL1 expression was mostly confined to a CD3(+)CD8(+) subset not expressing CD5, where XCL1 expression equaled that shown by gammadelta(+) T cells. Compared with the CD5(+) counterpart, CD3(+)CD8(+)CD5(-) cells, which did not express CD5 following in vitro activation, showed preferential expression of the alphaalpha form of CD8 and a lower expression of molecules associated with a noncommitted/naive phenotype, such as CD62L. CD3(+)CD8(+)CD5(-) cells also expressed higher levels of the XCL1 receptor; in addition, although not differing from CD3(+)CD8(+)CD5(+) cells in terms of the expression of most alpha- and beta-chemokines, they showed higher expression of CCL3/macrophage inflammatory protein-1alpha. These data show that TCR alphabeta-expressing lymphocytes that lack CD5 expression are a major XCL1 source, and that the contribution to its synthesis by different TCR alphabeta-expressing T cell subsets, namely CD4(+) lymphocytes, is negligible. In addition, they point to the CD3(+)CD8(+)CD5(-) population as a particular T cell subset within the CD8(+) compartment, whose functional properties deserve further attention.     

MELOE-1 Antigen Contains Multiple HLA Class II T Cell Epitopes Recognized by Th1 CD4+ T Cells from Melanoma Patients

MELOE-1 is an overexpressed melanoma antigen containing a HLA-A2 restricted epitope, involved in melanoma immunosurveillance of patients adoptively transferred with tumour infiltrating lymphocytes (TIL). The use of the full-length antigen (46 aa) for anti-melanoma vaccination could be considered, subject to the presence of Th epitopes all along MELOE-1 sequence. Thus, in this study we evaluated in vitro the immunoprevalence of the different regions of MELOE-1 (i.e. their ability to induce CD4 T cell responses in vitro from PBMC). Stimulation of PBMC from healthy subjects with MELOE-1 induced the amplification of CD4 T cells specific for various regions of the protein in multiple HLA contexts, for each tested donor. We confirmed these results in a panel of melanoma patients, and documented that MELOE-1 specific CD4 T cells, were mainly Th1 cells, presumably favourable to the amplification of CD8 specific T cells. Using autologous DC, we further showed that these class II epitopes could be naturally processed from MELOE-1 whole protein and identified minimal epitopes derived from each region of MELOE-1, and presented in four distinct HLA contexts. In conclusion, vaccination with MELOE-1 whole polypeptide should induce specific Th1 CD4 responses in a majority of melanoma patients, stimulating the amplification of CD8 effector cells, reactive against melanoma cells.     

CC chemokine receptor 9 expression defines a subset of peripheral blood lymphocytes with mucosal T cell phenotype and Th1 or T-regulatory 1 cytokine profile

The chemokine receptor CCR9 is expressed on most small intestinal lamina propria and intraepithelial lymphocytes and on a small subset of peripheral blood lymphocytes. CCR9-expressing lymphocytes may play an important role in small bowel immunity and inflammation. We studied the phenotype and functional characteristics of CCR9(+) lymphocytes in blood from normal donors. A subset of CCR9(+) T cells have a phenotype of activated cells and constitutively express the costimulatory molecules CD40L and OX-40. In contrast to CCR9(-), CCR9(+)CD4(+) peripheral blood T cells proliferate to anti-CD3 or anti-CD2 stimulation and produce high levels of IFN-gamma and IL-10. IL-10-producing cells were exclusively detected within the CCR9(+) subset of CD4(+) T cells by intracellular staining and were distinct from IL-2- and IFN-gamma-producing cells. Moreover, memory CCR9(+)CD4(+) lymphocytes respond to CD2 stimulation with proliferation and IFN-gamma/IL-10 production, whereas memory CCR9(-)CD4(+) cells were unresponsive. In addition, memory CCR9(+)CD4(+) T cells support Ig production by cocultured CD19(+) B cells in the absence of prior T cell activation or addition of exogenous cytokines. Our data show that the memory subset of circulating CCR9(+)CD4(+) T cells has characteristics of mucosal T lymphocytes and contains cells with either Th1 or T-regulatory 1 cytokine profiles. Studies on the cytokine profile and Ag specificity of this cell subset could provide important insight into small intestinal immune-mediated diseases and oral tolerance in humans.     

CD4+CD25+ and CD4+CD25- T cells act respectively as inducer and effector T suppressor cells in superantigen-induced tolerance

The repeated injection of low doses of bacterial superantigens (SAg) is known to induce specific T cell unresponsiveness. We show in this study that the spleen of BALB/c mice receiving chronically, staphylococcal enterotoxin B (SEB) contains SEB-specific CD4(+) TCRBV8(+) T cells exerting an immune regulatory function on SEB-specific primary T cell responses. Suppression affects IL-2 and IFN-gamma secretion as well as proliferation of T cells. However, the suppressor cells differ from the natural CD4(+) T regulatory cells, described recently in human and mouse, because they do not express cell surface CD25. They are CD152 (CTLA-4)-negative and their regulatory activity is not associated with expression of the NF Foxp3. By contrast, after repeated SEB injection, CD4(+)CD25(+) splenocytes were heterogenous and contained both effector as well as regulatory cells. In vivo, CD4(+)CD25(-) T regulatory cells prevented SEB-induced death independently of CD4(+)CD25(+) T cells. Nevertheless, SEB-induced tolerance could not be achieved in thymectomized CD25(+) cell-depleted mice because repeated injection of SEB did not avert lethal toxic shock in these animals. Collectively, these data demonstrate that, whereas CD4(+)CD25(+) T regulatory cells are required for the induction of SAg-induced tolerance, CD4(+)CD25(-) T cells exert their regulatory activity at the maintenance stage of SAg-specific unresponsiveness.     

Novel pathway for induction of latent virus from resting CD4(+) T cells in the simian immunodeficiency virus/macaque model of human immunodeficiency virus type 1 latency

Although combination therapy allows the suppression of human immunodeficiency virus type 1 (HIV-1) viremia to undetectable levels, eradication has not been achieved because the virus persists in cellular reservoirs, particularly the latent reservoir in resting CD4(+) T lymphocytes. We previously established a simian immunodeficiency virus (SIV)/macaque model to study latency. We describe here a novel mechanism for the induction of SIV from latently infected resting CD4(+) T cells. Several human cell lines including CEMx174 and Epstein-Barr virus-transformed human B-lymphoblastoid cell lines mediated contact-dependent activation of resting macaque T cells and induction of latent SIV. Antibody-blocking assays showed that interactions between the costimulatory molecule CD2 and its ligand CD58 were involved, whereas soluble factors and interactions between T-cell receptors and major histocompatibility complex class II were not. Combinations of specific antibodies to CD2 also induced T-cell activation and virus induction in human resting CD4(+) T cells carrying latent HIV-1. This is the first demonstration that costimulatory signals can induce latent virus without the coengagement of the T-cell receptor, and this study might provide insights into potential pathways to target latent HIV-1.     

Requirement of cognate CD4+ T-cell recognition for the regulation of allospecific CTL by human CD4+ CD127- CD25+ FOXP3+ cells generated in MLR

Although immunoregulation of alloreactive human CTLs has been described, the direct influence of CD4(+) Tregs on CD8(+) cytotoxicity and the interactive mechanisms have not been well clarified. Therefore, human CD4(+)CD127(-)CD25(+)FOXP3(+) Tregs were generated in MLR, immunoselected and their allospecific regulatory functions and associated mechanisms were then tested using modified (51)Chromium release assays (Micro-CML), MLRs and CFSE-based multi-fluorochrome flow cytometry proliferation assays. It was observed that increased numbers of CD4(+)CD127(-)CD25(+)FOXP3(+) cells were generated after a 7 day MLR. After immunoselection for CD4(+)CD127(-)CD25(+) cells, they were designated as MLR-Tregs. When added as third component modulators, MLR-Tregs inhibited the alloreactive proliferation of autologous PBMC in a concentration dependent manner. The inhibition was quasi-antigen specific, in that the inhibition was non-specific at higher MLR-Treg modulator doses, but non-specificity disappeared with lower numbers at which specific inhibition was still significant. When tested in micro-CML assays CTL inhibition occurred with PBMC and purified CD8(+) responders. However, antigen specificity of CTL inhibition was observed only with unpurified PBMC responders and not with purified CD8(+) responders or even with CD8(+) responders plus Non-T "APC". However, allospecificity of CTL regulation was restored when autologous purified CD4(+) T cells were added to the CD8(+) responders. Proliferation of CD8(+) cells was suppressed by MLR-Tregs in the presence or absence of IL-2. Inhibition by MLR-Tregs was mediated through down-regulation of intracellular perforin, granzyme B and membrane-bound CD25 molecules on the responding CD8(+) cells. Therefore, it was concluded that human CD4(+)CD127(-)CD25(+)FOXP3(+) MLR-Tregs down-regulate alloreactive cytotoxic responses. Regulatory allospecificity, however, requires the presence of cognate responding CD4(+) T cells. CD8(+) CTL regulatory mechanisms include impaired proliferation, reduced expression of cytolytic molecules and CD25(+) activation epitopes.     

Telomerase activity is increased and telomere length shortened in T cells from blood of patients with atopic dermatitis and psoriasis

We studied telomerase activity and telomere length in PBMC and purified CD4(+) and CD8(+) T cells from blood obtained from a total of 32 patients with atopic dermatitis, 16 patients with psoriasis, and 30 normal controls. The telomerase activity was significantly increased in PBMC from the patients compared with PBMC from normal donors. This increase was most pronounced in the subpopulation of CD4(+) T cells, which were significantly above the activity of the CD8(+) T cells in atopic dermatitis, psoriasis patients, and control persons. The telomere length was significantly reduced in all T cell subsets from both atopic dermatitis and psoriasis patients compared with normal individuals. Furthermore, the telomere length was found to be significantly shorter in CD4(+) memory T cells compared with the CD4(+) naive T cells, and both of the cell subsets from diseases were shown to be of significantly shorter telomere length than the same cell subsets from normal controls. No significant difference was observed between CD8(+)CD28(-) and CD8(+)CD28(+) T cell populations in both diseases. However, the telomere length of CD8(+)CD28(+) T cells from both diseases was significantly shorter than CD8(+)CD28(+) T cell subsets from normal donors. In conclusion, the increased telomerase activity and shortened telomere length indicates that T lymphocytes in atopic dermatitis and psoriasis are chronically stimulated and have an increased cellular turnover in vivo.