CD137 is a Useful Marker for Identifying CD4+ T Cell Responses to Mycobacterium tuberculosis

Upregulation of CD137 on recently activated CD8⁺ T cells has been used to identify rare viral and tumour antigen‐specific T cells from the peripheral blood. We aimed to evaluate the accuracy of CD137 for identifying Mycobacterium tuberculosis (Mtb)‐reactive CD4⁺ T cells in the peripheral blood of infected individuals by flow cytometry and to investigate the characteristics of these CD137⁺CD4⁺ T cells. We initially enrolled 31 active tuberculosis (TB) patients, 31 individuals with latent TB infection (LTBI) and 25 healthy donors. The intracellular CD137 and interferon‐γ (IFN‐γ) production by CD4⁺ T cells was simultaneously detected under unstimulated and CFP10‐stimulated (culture filtrate protein 10, a Mtb‐specific antigen) conditions. In unstimulated CD4⁺ T cells, we found that the CD137 expression in the TB group was significantly higher than that in the LTBI group. Stimulation with CFP10 largely increased the CD4⁺ T cell CD137 expression in both the TB and LTBI groups. After CFP10 stimulation, the frequency of CD137⁺CD4⁺ T cells was higher than that of IFN‐γ⁺CD4⁺ T cells in both the TB and LTBI groups. Most of the CFP10‐activated IFN‐γ‐secreting cells were CD137‐positive, but only a small fraction of the CD137‐positive cells expressed IFN‐γ. An additional 20 patients with TB were enrolled to characterize the CD45RO⁺CCR7⁺, CD45RO⁺CCR7⁻ and CD45RO⁻ subsets in the CD137⁺CD4⁺ T cell populations. The Mtb‐specific CD137⁺CD4⁺ T cells were mainly identified as having an effector memory phenotype. In conclusion, CD137 is a useful marker that can be used for identifying Mtb‐reactive CD4⁺ T cells by flow cytometry.     

Donor‐specific alloreactive T cells can be quantified from whole blood,and may predict cellular rejection after renal transplantation

Preformed cellular alloreactivity can exist prior to transplantation and may contribute to rejection. Here, we used a rapid flow‐cytometric whole‐blood assay to characterize the extent of alloreactive T cells among 1491 stimulatory reactions from 61 renal transplant candidates and 75 controls. The role of preformed donor‐specific alloreactive T cells in cellular rejection was prospectively analyzed in 21 renal transplant recipients. Alloreactive CD8⁺ T cells were more frequent than respective CD4⁺ T cells, and these levels were stable over time. CD8⁺ T cells were effector‐memory T cells largely negative for expression of CD27, CD62L, and CCR7, and were susceptible to steroid and calcineurin inhibitor inhibition. Alloreactivity was more frequent in samples with higher number of HLA mismatches. Moreover, the percentage of individuals with alloreactive T cells was higher in transplant candidates than in controls. Among transplant candidates, 5/61 exhibited alloreactive CD8⁺ T cells against most stimulators, 23/61 toward a limited number of stimulators, and 33/61 did not show any alloreactivity. Among 21 renal transplant recipients followed prospectively, one had donor‐specific preformed T‐cell alloreactivity. She was the only patient who developed cellular rejection posttransplantation. In conclusion, donor‐specific alloreactive T cells may be rapidly quantified from whole blood, and may predict cellular rejection after transplantation.     

The effect of allogeneic in vitro stimulation and in vivo immunization on memory CD4+ T‐cell APOBEC3G expression and HIV‐1 infectivity

Allogeneic immunity is one of the most potent natural immune responses. APOBEC3G (A3G) is an intracellular anti‐viral factor that deaminates cytidine to uridine. Allogeneic stimulation of human CD4⁺ T cells in vitro upregulated A3G mRNA and a significant correlation was found between the mixed leukocyte reaction and A3G mRNA. The mechanism of upregulation of A3G mRNA involves interaction between HLA on DC and TCR of CD4⁺ T cells, which is ZAP70 and downstream ERK phosphokinase signalling dependent and induces CD40L and A3G mRNA expression in CD4⁺ T cells. Alloimmune‐induced A3G was found to be significantly increased in CD45RA^?, CCR5⁺ and CD45RA^?CCR7^? subsets of effector memory T cells. In vivo studies of women alloimmunized with their partners' PBMC also showed a significant increase in A3G protein in CD4⁺ T cells, CD45RO⁺ memory and CCR7^? effector memory T cells. The functional effect of allostimulation upregulating A3G mRNA was demonstrated by a significant decrease in in vitro infectivity, using GFP‐labelled pseudovirus and confirmed by a decrease in HIV‐1 (BaL) infection of primary CD4⁺ T cells. The results suggest that alloimmunization offers an alternative or complementary strategy in inducing an innate anti‐viral factor that inhibits HIV‐1 infection.     

Differential expression of CCR7 defines two distinct subsets of human memory CD4+CD25+ Tregs

Natural Tregs play an essential role in controlling self-tolerance but the in vivo sites of Treg-mediated suppression remain controversial. We have previously reported the identification of distinct naïve and memory Treg populations in human circulating lymphocytes. Here we show that memory Tregs contain high proportions of inflammatory chemokine-expressing cells and comprise two populations that differ in the expression of the lymphoid chemokine receptor CCR7 and represent the counterparts of conventional CCR7⁺ central memory (CM) and CCR7⁻ effector memory (EM) T cells. CM and EM Tregs exert comparable ex vivo suppressor functions but the EM population is more prominent among Tregs as compared to conventional CD4⁺ T cells, and is the main Treg subset found in ovarian tumors. Our data suggest that a division of labor between CM and EM Tregs ensures tolerance at lymphoid and peripheral locations including tumor sites.     

Interleukin 10 and dendritic cells are the main suppression mediators of regulatory T cells in human neurocysticercosis

Neurocysticercosis is caused by the establishment of Taenia solium cysticerci in the central nervous system. It is considered that, during co‐evolution, the parasite developed strategies to modulate the host's immune response. The action mechanisms of regulatory T cells in controlling the immune response in neurocysticercosis are studied in this work. Higher blood levels of regulatory T cells with CD4⁺CD45RO⁺forkhead box protein 3 (FoxP3)^high and CD4⁺CD25^highFoxP3⁺CD95^high phenotype and of non‐regulatory CD4⁺CD45RO⁺FoxP3^med T cells were found in neurocysticercosis patients with respect to controls. Interestingly, regulatory T cells express higher levels of cytotoxic T lymphocyte antigen 4 (CTLA‐4), lymphocyte‐activation gene 3 (LAG‐3), programmed death 1 (PD‐1) and glucocorticoid‐induced tumour necrosis factor receptor (GITR), suggesting a cell‐to‐cell contact mechanism with dendritic cells. Furthermore, higher IL‐10 and regulatory T cell type 1 (Tr1) levels were found in neurocysticercosis patients’ peripheral blood, suggesting that the action mechanism of regulatory T cells involves the release of immunomodulatory cytokines. No evidence was found of the regulatory T cell role in inhibiting the proliferative response. Suppressive regulatory T cells from neurocysticercosis patients correlated negatively with late activated lymphocytes (CD4⁺CD38⁺). Our results suggest that, during neurocysticercosis, regulatory T cells could control the immune response, probably by a cell‐to‐cell contact with dendritic cells and interleukin (IL)‐10 release by Tr1, to create an immunomodulatory environment that may favour the development of T. solium cysticerci and their permanence in the central nervous system.     

Effector memory CD4+ T cells differentially express activation associated molecules depending on the duration of American cutaneous leishmaniasis lesions

A high number of Leishmania‐responder T cells is found in cutaneous leishmaniasis lesions, suggesting that important immunological events occur at the site of infection. Although activated, cytotoxic and regulatory T cells infiltrating into lesions may influence disease pathogenesis, the role of the T cell differentiation pattern of lymphocytes in lesions is unknown. Our aim was to investigate whether the phase of lesion development (early or late) is influenced by the functional status of cells present in inflammatory infiltrate. Activation, cytotoxity and T cell differentiation molecules were evaluated in lesion mononuclear cells by flow cytometry. The frequency of T cells was correlated with the lesion area (r = 0·68; P = 0·020). CD4⁺CD25⁺ T cells predominated over CD4⁺CD69⁺ T cells in early lesions (less than 30 days), whereas late lesions (more than 60 days) exhibited more CD4⁺CD69⁺ T cells than CD4⁺CD25⁺ T cells. The duration of illness was correlated positively with CD4⁺CD69⁺ (r = 0·68; P = 0·005) and negatively with CD4⁺CD25⁺ T cells (r = ?0·45; P = 0·046). Most CD8⁺ T cells expressed cytotoxic‐associated molecules (CD244⁺), and the percentages were correlated with the lesion area (r = 0·52; P = 0·04). Both CD4⁺ and CD8⁺ effector memory T cells (T_EM‐CD45RO⁺CCR7^–) predominated in CL lesions and were significantly higher than central memory (T_CM‐CD45RO⁺CCR7⁺) or naive T cells (CD45RO^–CCR7⁺). An enrichment of T_EM cells and contraction of naive T cells were observed in lesions in comparison to blood (P = 0·006) for both CD4⁺ and CD8⁺ T cells. Lesion chronicity is associated with a shift in activation phenotype. The enrichment of T_EM and activated cytotoxic cells can contribute to immune‐mediated tissue damage.     

Peripherally Circulating CD4(+) FOXP3(+) CXCR3(+) T Regulatory Cells Correlate with Renal Allograft Function

Peripheral immunoregulation depends on T regulatory cell trafficking into the allograft to modulate the local alloresponse. Little is known about the relevance of trafficking receptors for Tregs after solid organ transplantation in humans. In this study, expression of the peripheral chemokine receptors CXCR3 and CCR5 on CD4⁺ FOXP3⁺ Treg cells was analysed and correlated with allograft function in renal transplant recipients. Flow cytometry analysis of peripheral blood mononuclear cells of 54 renal transplant recipients receiving a calcineurin inhibitor‐based immunosuppression was performed for CD4, CD25, FOXP3, CXCR3 and CCR5 within the first 18 months post‐transplantation. Correlation analysis of chemokine receptor expression and glomerular filtration rate as calculated by MDRD (eGFR) was performed. Expression of the peripheral homing receptors CXCR3 (r = 0.44, P < 0.05) and CCR5 (r = 0.45, P < 0.05) on FOXP3⁺ Tregs correlated with renal allograft function (eGFR) in patients receiving tacrolimus (n = 28), but not cyclosporine A (CsA) (n = 26). CsA but not tacrolimus reduced surface expression of CXCR3 on FOXP3⁺ Tregs in renal transplant recipients as correlated to trough levels (r = ?0.42, P < 0.05). In contrast to CD4⁺ CXCR3⁺ CD25^lo T cells, flow‐sorted CD4⁺ CXCR3⁺ CD25^hi Tregs isolated from healthy individuals did not produce IFNγ or IL‐17 ex vivo and expressed high levels of GARP mRNA both at baseline as well as after TCR activation indicating functional regulatory activity. Expression of the peripheral trafficking receptors CXCR3 and CCR5 on FOXP3⁺ Tregs is associated with renal allograft function. These results suggest that Treg trafficking may also depend on the interaction of CXCR3 or CCR5 and their respective ligands.     

Ex vivo monitoring of human cytomegalovirus‐specific CD8+ T‐Cell responses using the QuantiFERON®‐CMV assay in allogeneic hematopoietic stem cell transplant recipients attending an Irish hospital

Reconstitution of human cytomegalovirus (HCMV) T‐cell immunity is crucial in hematopoietic stem cell transplant (HSCT) recipients. The QuantiFERON®‐CMV assay for cellular HCMV‐specific immunity was evaluated in allogeneic HSCT recipients (n = 43) and patients with hematological malignancies (n = 29) attending a tertiary‐care Irish hospital. An intracellular cytokine (ICC) assay correlated with the QuantiFERON®‐CMV assay. Although there was agreement between HCMV seropositivity and QuantiFERON®‐CMV assay, six HCMV seropositive immunosuppressed patients with hematological malignancy had negative QuantiFERON®‐CMV results. The 43 HSCT recipients were classified as high risk (D^?/R⁺) (n = 18), intermediate risk (D⁺/R⁺ and D⁺/R^?) (n = 17), and low risk (D^?/R^?) (n = 8). During episodes of HCMV DNAemia no evidence of HCMV‐specific immunity was found using the QuantiFERON®‐CMV assay. Furthermore, the recovery of HCMV‐specific CD8⁺ T‐cell responses in high‐risk seropositive recipients of matched unrelated donors was severely delayed, a mean of 200 (SD = 117) days compared to 58 (SD = 23) days for sibling donors (P ≤ 0.028). In addition, three patients with late HCMV infection (infection >100 days post‐transplant) had delayed reconstitution of HCMV‐specific CD8⁺ T cells. Interestingly, two recipients (R⁺/D^?) developed rapid immune reconstitution by days 15 and 36 post‐HSCT, suggesting HCMV‐specific T‐cell lymphopoiesis of recipient origin. Levels of CD8⁺ T‐cell immunity in HCMV seropositive HSCT recipients were lowest following HSCT. A high number (33%) of indeterminate results was observed immediately after transplantation. Patients with indeterminate QuantiFERON®‐CMV results had low levels of HCMV‐specific CD8⁺ T cells. J. Med. Virol. 82:433–440, 2010. © 2010 Wiley‐Liss, Inc.     

Importance of B cell co-stimulation in CD4(+) T cell differentiation: X-linked agammaglobulinaemia, a human model

We were interested in the question of whether the congenital lack of B cells actually had any influence on the development of the T cell compartment in patients with agammaglobulinaemia. Sixteen patients with X‐linked agammaglobulinaemia (XLA) due to mutations in Btk, nine patients affected by common variable immune deficiency (CVID) with <2% of peripheral B cells and 20 healthy volunteers were enrolled. The T cell phenotype was determined with FACSCalibur and CellQuest Pro software. Mann–Whitney two‐tailed analysis was used for statistical analysis. The CD4 T cell memory compartment was reduced in patients with XLA of all ages. This T cell subset encompasses both CD4⁺CD45RO⁺ and CD4⁺CD45RO⁺CXCR5⁺ cells and both subsets were decreased significantly when compared to healthy controls: P = 0·001 and P < 0·0001, respectively. This observation was confirmed in patients with CVID who had <2% B cells, suggesting that not the lack of Bruton's tyrosine kinase but the lack of B cells is most probably the cause of the impaired CD4 T cell maturation. We postulate that this defect is a correlate of the observed paucity of germinal centres in XLA. Our results support the importance of the interplay between B and T cells in the germinal centre for the activation of CD4 T cells in humans.     

Differential expression of type I insulin-like growth factor receptors in different stages of human T cells

Insulin-like growth factor I (IGF-I) has been implicated to play a regulatory role in T cell development and in T cell function. We investigated the expression of type I IGF receptors on human peripheral T cells related to the maturation and activation stage using the type I IGF receptor-specific monoclonal antibody αIR3. It appeared that 87% of the CD4⁺CD45RA⁺ cells and 66% of the CD8⁺CD45RA⁺ cells were αIR3⁺, whereas only 37% of the CD4⁺CD45RO⁺ cells and 38% of the CD8⁺CD45RO⁺ cells bound αIR3. We also found that the fraction of αIR3⁺ cells within in vivo or in vitro activated (HLA-DR⁺) T cells is markedly lower than in nonactivated (HLA-DR^?) cells. In vitro phytohemagglutinin-activated T cells and CD4⁺CD45RO⁺ cells activated with recall antigens also contained less αIR3⁺ cells (1–6%) than nonactivated cells (30–54%).     

CMV drives the expansion of highly functional memory T cells expressing NK‐cell receptors in renal transplant recipients

Cytomegalovirus (CMV) is a common opportunistic infection encountered in renal transplant recipients (RTRs) and may be reactivated without symptoms at any time post‐transplant. We describe how active and latent CMV affect T‐cell subsets in RTRs who are stable on maintenance therapy. T‐cell responses to CMV were assessed in RTRs (n = 54) >2 years post‐transplant, and healthy controls (n = 38). Seven RTRs had CMV DNA detectable in plasma. CMV antibody and DNA aligned with increased proportions of CD8⁺ T cells and reduced CD4/CD8 ratios. This paralleled an expansion of effector memory T‐cell (T_EM), terminally differentiated T‐cell (T_EMRA) and CD57⁺ T_EMRA cell populations. Expression of NK‐cell receptors, LIR‐1 and KLRG1 on CD4⁺ and CD8⁺ CD57⁺ T_EM and T_EMRA cells correlated with elevated interferon‐γ and cytotoxic responses to anti‐CD3 and increased cytotoxic responses to CMV phosphoprotein (pp) 65 in RTRs who carried CMV DNA. CD8⁺ T cells from all CMV seropositive RTRs responded efficiently to CMV immediate early (IE) ‐1 peptides. The data show that latent and active CMV infection can alter T‐cell subsets in RTRs many years after transplantation, and up‐regulate T‐cell expression of NK‐cell receptors. This may enhance effector responses of CD4⁺ and CD8⁺ T cells against CMV.     

Human and mouse perforin are processed in part through cleavage by the lysosomal cysteine proteinase cathepsin L

Summary Infection of mice with the gastrointestinal nematode Trichuris muris represents a valuable tool to investigate and dissect intestinal immune responses. Resistant mouse strains respond to T. muris infection by mounting a T helper type 2 response. Previous results have shown that CD4⁺ T cells play a critical role in protective immunity, and that CD4⁺ T cells localize to the infected large intestinal mucosa to confer protection. Further, transfer of CD4⁺ T cells from immune mice to immunodeficient SCID mice can prevent the development of a chronic infection. In the current study, we characterize the protective CD4⁺ T cells, describe their chemokine receptor expression and explore the functional significance of these receptors in recruitment to the large intestinal mucosa post‐T. muris infection. We show that the ability to mediate expulsion resides within a subpopulation of CD4⁺ T cells marked by down‐regulation of CD62L. These cells can be isolated from intestine‐draining mesenteric lymph nodes (MLN) from day 14 post‐infection, but are rare or absent in MLN before this and in spleen at all times post‐infection. Among CD4⁺ CD62L^low MLN cells, the two most abundantly expressed chemokine receptors were CCR6 and CXCR3. We demonstrate for the first time that CD4⁺ CD62L^low T‐cell migration to the large intestinal mucosa is dependent on the family of Gα_i‐coupled receptors, to which chemokine receptors belong. CCR6 and CXCR3 were however dispensable for this process because neutralization of CCR6 and CXCR3 did not prevent CD4⁺ CD62L^low cell migration to the large intestinal mucosa during T. muris infection.     

Regulatory T‐cells in acute dengue viral infection

Although regulatory T‐cells (T_regs) have been shown to be expanded in acute dengue, their role in pathogenesis and their relationship to clinical disease severity and extent of viraemia have not been fully evaluated. The frequency of T_regs was assessed in 56 adult patients with acute dengue by determining the proportion of forkhead box protein 3 (FoxP3) expressing CD4⁺ CD25⁺T‐cells (FoxP3⁺ cells). Dengue virus (DENV) viral loads were measured by quantitative real‐time polymerase chain reaction (PCR) and DENV‐specific T‐cell responses were measured by ex‐vivo interferon (IFN)‐γ enzyme‐linked immunospot (ELISPOT) assays to overlapping peptide pools of DENV‐NS3, NS1 and NS5. CD45RA and CCR4 were used to phenotype different subsets of T‐cells and their suppressive potential was assessed by their expression of cytotoxic T lymphocyte‐antigen 4 (CTLA‐4) and Fas. While the frequency of FoxP3⁺ cells in patients was significantly higher (P < 0·0001) when compared to healthy individuals, they did not show any relationship with clinical disease severity or the degree of viraemia. The frequency of FoxP3⁺ cells did not correlate with either ex‐vivo IFN‐γ DENV‐NS3‐, NS5‐ or NS1‐specific T‐cell responses. FoxP3⁺ cells of patients with acute dengue were predominantly CD45RA⁺ FoxP3^low, followed by CD45RA‐FoxP3^low, with only a small proportion of FoxP3⁺ cells being of the highly suppressive effector Treg subtype. Expression of CCR4 was also low in the majority of T‐cells, with only CCR4 only being expressed at high levels in the effector T_reg population. Therefore, although FoxP3⁺ cells are expanded in acute dengue, they predominantly consist of naive T_regs, with poor suppressive capacity.     

Preferential infiltration of interleukin‐4‐producing CXCR4+ T cells in the lesional muscle but not skin of patients with dermatomyositis

Dermatomyositis (DM) and polymyositis (PM) are collectively termed autoimmune myopathy. To investigate the difference between muscle‐ and skin‐infiltrating T cells and to address their role for myopathy, we characterized T cells that were directly expanded from the tissues. Enrolled into this study were 25 patients with DM and three patients with PM. Muscle and skin biopsied specimens were immersed in cRPMI medium supplemented with interleukin (IL)‐2 and anti‐CD3/CD28 antibody‐conjugated microbeads. The expanded cells were subjected to flow cytometry to examine their phenotypes. We analysed the cytokine concentration in the culture supernatants from the expanded T cells and the frequencies of cytokine‐bearing cells by intracellular staining. There was non‐biased in‐vitro expansion of tissue‐infiltrating CD4⁺ and CD8⁺ T cells from the muscle and skin specimens. The majority of expanded T cells were chemokine receptor (CCR) type 7^–CD45RO⁺ effecter memory cells with various T cell receptor (TCR) Vβs. The skin‐derived but not muscle‐derived T cells expressed cutaneous lymphocyte antigen (CLA) and CCR10 and secreted large amounts of IL‐17A, suggesting that T helper type 17 (Th17) cells may have a crucial role in the development of skin lesions. Notably, the frequency of IL‐4‐producing chemokine (C‐X‐C motif) receptor (CXCR)4⁺ Th2 cells was significantly higher in the muscle‐derived cells and correlated inversely with the serum creatine phosphokinase (CPK) and lactate dehydrogenase (LDH) levels. stromal‐derived factor (SDF)‐1/CXCL12, a ligand for CXCR4, was expressed at a high level in the vascular endothelial cells between muscular fasciculi. Our study suggests that T cell populations in the muscle and skin are different, and the Th2 cell infiltrate in the muscle is associated with the low severity of myositis in DM.     

Inhibition of R5‐tropic HIV type‐1 replication in CD4+ natural killer T cells by γδ T lymphocytes

After the development of highly active anti‐retroviral therapy, it became clear that the majority of emergent HIV‐1 is macrophage‐tropic and infects CD4⁺, CCR5‐expressing cells (R5‐tropic). There are three distinct cell populations, R5‐tropic, HIV‐1‐susceptible CD4⁺ cells: (i) natural killer T (NKT) cells, (ii) dendritic cells and macrophages, and (iii) tissue‐associated T cells residing primarily at mucosal surfaces. We have confirmed that CD4⁺ NKT cells derived from peripheral blood mononuclear cells (PBMCs) predominantly express CCR5 rather than CXCR4, whereas the reverse is true for CD4⁺ T cells derived from circulating PBMCs, and that R5‐tropic HIV‐1 expands efficiently in the CD4⁺ NKT cells. Moreover, when PBMCs depleted of CD8α⁺ cells were stimulated in the presence of α‐galactosylceramide (α‐GalCer) and R5‐tropic HIV‐1 [NL(AD8)], the production of HIV‐1 virions was not suppressed, whereas, similar to the untreated PBMCs, depletion of CD8β⁺ cells from PBMCs significantly inhibited virion production. These findings suggest that CD8αα⁺ but not CD8αβ⁺ cells may have the ability to inhibit R5‐tropic HIV‐1 replication in CD4⁺ NKT cells. Here, we show that co‐culturing R5‐tropic HIV‐1‐infected CD4⁺ NKT cells with CD8αα⁺ γδ T cells, in particular Vγ1Vδ1 cells, but not with CD8αα⁺ NKT cells or CD8αα⁺ dendritic cells, inhibits HIV‐1 replication mainly by secreting chemokines, such as macrophage inflammatory proteins 1α and 1β and RANTES. Collectively, these results indicate the importance of CD8αα⁺ γδ T cells in the control of R5‐tropic HIV‐1 replication and persistence in CD4⁺ NKT cells.     

CCR7+ Central and CCR7− Effector Memory CD4+ T Cells in Human Cutaneous Leishmaniasis

Purpose

The profile of central (=T_CM) and effector (=T_EM) memory CD4⁺ T cell subsets and the possible role as surrogate markers of protection is studied in the volunteers with history of cutaneous leishmaniasis (HCL).

Methods

Profile of T cell subsets based on CCR7/CD45RA expressions and phenotypic changes after soluble Leishmania antigen (SLA) stimulation were analyzed. Then, sorted CD4⁺CD45RO^?CD45RA⁺ naïve T, CD4⁺CD45RO⁺CD45RA^?CCR7^? T_EM, CD4⁺CD45RO⁺CD45RA^?CCR7⁺ T_CM subsets were cultured with SLA for proliferation, cytokine production and intracellular cytokine assays.

Results

In the HCL and control volunteers, the mean frequencies of CD4⁺CD45RA⁺CCR7⁺ naïve T cells and CD4⁺CD45RA^?CCR7^? T_EM cells were higher than the other subsets before culture. Frequency of naïve T cells and CD4⁺CD45RA^?CCR7⁺ T_CM cells was significantly decreased (P?=?0.01 for naïve T and P?<?0.05 for T_CM cells) and frequency of T_EM cells was significantly increased after SLA stimulation compared to before culture (P?<?0.001). By CFSE labeling, CD4⁺CD45RO⁺CD45RA^?CCR7⁺ T_CM cells showed more proliferation potential than CD4⁺CD45RO⁺CD45RA^?CCR7^? T_EM cells. Stimulation of the T_EM cells in HCL volunteers induced a significantly higher IFN-γ production (P?=?0.04) with higher number of intracellular IFN-γ positive cells (P?=?0.032) than the same cells from controls. A significantly higher number of T_CM cells produced IL-2 in HCL volunteers compared with controls (P?<?0.05). Most of the intracellular IFN-γ positive T_EM cells were proliferating CFSE-dim populations (P?<?0.05).

Conclusions

A combination of Leishmania-reactive IFN-γ producing CD4⁺CD45RO⁺CD45RA^?CCR7^? T_EM and Leishmania-reactive IL-2 producing CD4⁺CD45RO⁺CD45RA^?CCR7⁺ T_CM are identified in individuals with history of CL which might play a role in protective recall immune response against Leishmania infection.     

Critical role of CCL22/CCR4 axis in the maintenance of immune homeostasis during apoptotic cell clearance by splenic CD8α+ CD103+ dendritic cells

Macrophages and dendritic cells (DCs) in murine spleen are essential for the maintenance of immune homeostasis by elimination of blood‐borne foreign particles and organisms. It has been reported that splenic DCs, especially CD8α⁺ CD103⁺ DCs, are responsible for tolerance to apoptosis‐associated antigens. However, the molecular mechanism by which these DCs maintain immune homeostasis by blood‐borne apoptotic cell clearance remains elusive. Here, we found that the CCL22/CCR4 axis played a critical role in the maintenance of immune homeostasis during apoptotic cell clearance by splenic CD8α⁺ CD103⁺ DCs. The present results revealed that systemic administration of apoptotic cells rapidly induced a large number of CCL22 and CCR4⁺ regulatory T (Treg) cells in the spleen of C57BL/6J mice. Further study demonstrated that CD8α⁺ CD103⁺ DCs dominantly produce much higher CCL22 than CD8α⁺ CD103^? DCs. Moreover, the transient deletion of CD8α⁺ CD103⁺ DCs caused a decrease in CCL22 levels together with CCR4⁺ Treg cell percentage. Subsequently, the levels of some pro‐inflammatory cytokines, such as interleukin‐17 and interferon‐γ in the spleen with the absence of CD8α⁺ CD103⁺ DCs increased in response to the administration of apoptotic cells. Hence, intravenous injection of apoptotic cells induced a subsequent increase in CCL22 expression and CCR4⁺ Treg cells, which contribute to the maintenance of immune homeostasis at least partially by splenic CD8α⁺ CD103⁺ DCs.     

Persistent low thymic activity and non‐cardiac mortality in children with chromosome 22q11·2 microdeletion and partial DiGeorge syndrome

A subgroup of patients with 22q11·2 microdeletion and partial DiGeorge syndrome (pDGS) appears to be susceptible to non‐cardiac mortality (NCM) despite sufficient overall CD4⁺ T cells. To detect these patients, 20 newborns with 22q11·2 microdeletion and congenital heart disease were followed prospectively for 6 years. Besides detailed clinical assessment, longitudinal monitoring of naive CD4⁺ and cytotoxic CD3⁺CD8⁺ T cells (CTL) was performed. To monitor thymic activity, we analysed naive platelet endothelial cell adhesion molecule‐1 (CD31⁺) expressing CD45RA⁺RO^‐CD4⁺ cells containing high numbers of T cell receptor excision circle (T_REC)‐bearing lymphocytes and compared them with normal values of healthy children (n = 75). Comparing two age periods, low overall CD4⁺ and naive CD4⁺ T cell numbers were observed in 65%/75%, respectively, of patients in period A (< 1 year) declining to 22%/50%, respectively, of patients in period B (> 1/< 7 years). The percentage of patients with low CTLs (< P10) remained robust until school age (period A: 60%; period B: 50%). Low numbers of CTLs were associated with abnormally low naive CD45RA⁺RO^‐CD4⁺ T cells. A high‐risk (HR) group (n = 11) and a standard‐risk (SR) (n = 9) group were identified. HR patients were characterized by low numbers of both naive CD4⁺ and CTLs and were prone to lethal infectious and lymphoproliferative complications (NCM: four of 11; cardiac mortality: one of 11) while SR patients were not (NCM: none of nine; cardiac mortality: two of nine). Naive CD31⁺CD45RA⁺RO^‐CD4⁺, naive CD45RA⁺RO^‐CD4⁺ T cells as well as T_RECs/10⁶ mononuclear cells were abnormally low in HR and normal in SR patients. Longitudinal monitoring of naive CD4⁺ and cytotoxic T cells may help to discriminate pDGS patients at increased risk for NCM.     

Robust T cell responses to aspergillosis in chronic granulomatous disease: implications for immunotherapy

Chronic granulomatous disease (CGD) patients are highly susceptible to invasive aspergillosis and might benefit from aspergillus‐specific T cell immunotherapy, which has shown promise in treating those with known T cell defects such as haematopoietic stem cell transplant (HSCT) recipients. But whether such T cell defects contribute to increased risks for aspergillus infection in CGD is unclear. Hence, we set out to characterize the aspergillus‐specific T cell response in CGD. In murine CGD models and in patients with CGD we showed that the CD4⁺ T cell responses to aspergillus were unimpaired: aspergillus‐specific T cell frequencies were even elevated in CGD mice (P < 0·01) and humans (P = 0·02), compared to their healthy counterparts. CD4‐depleted murine models suggested that the role of T cells might be redundant because resistance to aspergillus infection was conserved in CD4⁺ T cell‐depleted mice, similar to wild‐type animals. In contrast, mice depleted of neutrophils alone or neutrophils and CD4⁺ T cells developed clinical and pathological evidence of pulmonary aspergillosis and increased mortality (P < 0·05 compared to non‐depleted animals). Our findings that T cells in CGD have a robust aspergillus CD4⁺ T cell response suggest that CD4⁺ T cell‐based immunotherapy for this disease is unlikely to be beneficial.