Edwin Leeansyah

Acquisition of innate-like microbial reactivity in mucosal tissues during human fetal MAIT-cell development

Innate-like, evolutionarily conserved MR1-restricted mucosa-associated invariant T (MAIT) cells represent a large antimicrobial T-cell subset in humans. Here, we investigate the development of these cells in second trimester human fetal tissues. MAIT cells are rare and immature in the fetal thymus, spleen and mesenteric lymph nodes. In contrast, mature IL-18Rα+ CD8αα MAIT cells are enriched in the fetal small intestine, liver and lung. Independently of localization, MAIT cells express CD127 and Ki67 in vivo and readily proliferate in response to Escherichia coli in vitro. Maturation is accompanied by the gradual post-thymic acquisition of the PLZF transcription factor and the ability to produce IFNγ and IL-22 in response to bacteria in mucosa. Thus, MAIT cells acquire innate-like antimicrobial responsiveness in mucosa before exposure to environmental microbes and the commensal microflora. Establishment of this arm of immunity before birth may help protect the newborn from a range of pathogenic microbes.

Arming of MAIT Cell Cytolytic Antimicrobial Activity Is Induced by IL-7 and Defective in HIV-1 Infection

Mucosa-associated invariant T (MAIT) cells represent a large innate-like evolutionarily conserved antimicrobial T-cell subset in humans. MAIT cells recognize microbial riboflavin metabolites from a range of microbes presented by MR1 molecules. MAIT cells are impaired in several chronic diseases including HIV-1 infection, where they show signs of exhaustion and decline numerically. Here, we examined the broader effector functions of MAIT cells in this context and strategies to rescue their functions. Residual MAIT cells from HIV-infected patients displayed aberrant baseline levels of cytolytic proteins, and failed to mobilize cytolytic molecules in response to bacterial antigen. In particular, the induction of granzyme B (GrzB) expression was profoundly defective. The functionally impaired MAIT cell population exhibited abnormal T-bet and Eomes expression patterns that correlated with the deficiency in cytotoxic capacity and cytokine production. Effective antiretroviral therapy (ART) did not fully restore these aberrations. Interestingly, IL-7 was capable of arming resting MAIT cells from healthy donors into cytotoxic GrzB+ effector T cells capable of killing bacteria-infected cells and producing high levels of pro-inflammatory cytokines in an MR1-dependent fashion. Furthermore, IL-7 treatment enhanced the sensitivity of MAIT cells to detect low levels of bacteria. In HIV-infected patients, plasma IL-7 levels were positively correlated with MAIT cell numbers and function, and IL-7 treatment in vitro significantly restored MAIT cell effector functions even in the absence of ART. These results indicate that the cytolytic capacity in MAIT cells is severely defective in HIV-1 infected patients, and that the broad-based functional defect in these cells is associated with deficiency in critical transcription factors. Furthermore, IL-7 induces the arming of effector functions and enhances the sensitivity of MAIT cells, and may be considered in immunotherapeutic approaches to restore MAIT cells.

Nonreversible MAIT cell‐dysfunction in chronic hepatitis C virus infection despite successful interferon‐free therapy

Immune exhaustion is a hallmark of chronic viral infections. However, pathogen eradication can result in reinvigorated immune responses. Indeed, this was recently suggested for antigen‐specific CD8+ T cells and NK cells in HCV‐infected patients receiving an interferon‐free treatment regimen. Mucosal‐associated invariant T (MAIT) cells are evolutionarily conserved innate‐like effector T cells. Here, we show that MAIT cells are severely diminished in frequency in chronic HCV‐infection, and in this regard the most affected immune cell type in peripheral blood of humans with this disease. Residual MAIT cells show an activated phenotype with high expression of granzyme B, HLA‐DR, PD‐1, and CD69 as well as altered transcription factor expression and suppressed responsiveness to MR1‐dependent antigen stimulation. In contrast to other immune cells, MAIT cells are not reinvigorated after successful HCV‐clearance using interferon‐free therapy. The present results hence demonstrate persistent immune cell‐dysfunction in humans despite successful elimination of a chronic pathogen.

Multiple layers of heterogeneity and subset diversity in human MAIT cell responses to distinct microorganisms and to innate cytokines

Significance Mucosa-associated invariant T (MAIT) cells are a large subset of unconventional T cells in humans, recognizing microbial riboflavin metabolites presented by the monomorphic MR1 molecule. The extraordinary level of conservation of MR1 and the limited diversity of riboflavin-derived antigens have suggested that MAIT cells are homogeneous, and their functional specialization has not been thoroughly investigated. Here, we show that MAIT cell responses against two distinct riboflavin biosynthesis-competent microorganisms display microbe-specific response patterns with multiple layers of heterogeneity. Furthermore, a set of natural killer cell-associated receptors define a subset with enhanced capacity to respond to innate cytokine stimulus. Thus, MAIT cells harbor multiple layers of functional heterogeneity and can adapt their antimicrobial responses to the type of microbial stimuli. Mucosa-associated invariant T (MAIT) cells are a large innate-like T-cell subset in humans defined by invariant TCR Vα7.2 use and expression of CD161. MAIT cells recognize microbial riboflavin metabolites of bacterial or fungal origin presented by the monomorphic MR1 molecule. The extraordinary level of evolutionary conservation of MR1 and the limited known diversity of riboflavin metabolite antigens have suggested that MAIT cells are relatively homogeneous and uniform in responses against diverse microbes carrying the riboflavin biosynthesis pathway. The ability of MAIT cells to exhibit microbe-specific functional specialization has not been thoroughly investigated. Here, we found that MAIT cell responses against Escherichia coli and Candida albicans displayed microbe-specific polyfunctional response profiles, antigen sensitivity, and response magnitudes. MAIT cell effector responses against E. coli and C. albicans displayed differential MR1 dependency and TCR β-chain bias, consistent with possible divergent antigen subspecificities between these bacterial and fungal organisms. Finally, although the MAIT cell immunoproteome was overall relatively homogenous and consistent with an effector memory-like profile, it still revealed diversity in a set of natural killer cell-associated receptors. Among these, CD56, CD84, and CD94 defined a subset with higher expression of the transcription factors promyelocytic leukemia zinc finger (PLZF), eomesodermin, and T-bet and enhanced capacity to respond to IL-12 and IL-18 stimulation. Thus, the conserved and innate-like MAIT cells harbor multiple layers of functional heterogeneity as they respond to bacterial or fungal organisms or innate cytokines and adapt their antimicrobial response patterns in a stimulus-specific manner.

MAIT cells reside in the female genital mucosa and are biased towards IL-17 and IL-22 production in response to bacterial stimulation

The female genital tract (FGT) mucosa is a critically important site for immune defense against microbes. Mucosal-associated invariant T (MAIT) cells are an innate-like T-cell population that recognizes microbial riboflavin metabolite antigens in an MR1-dependent manner. The role of MAIT cells in the FGT mucosa is unknown. Here, we found that MAIT cells and MR1+ antigen-presenting cells were present in the upper and lower FGT, with distinct tissue localization of MAIT cells in endometrium vs. cervix. The MAIT cells from the FGT and blood displayed a distinct phenotype with expression of interleukin (IL)-18Rα, CD127, α4β7, PD-1, as well as the transcription factors promyelocytic leukemia zinc finger (PLZF), RORγt, Helios, Eomes, and T-bet. Their expression levels of PLZF and Eomes were lower in the FGT compared with blood. When stimulated with Escherichia coli, MAIT cells from the FGT displayed a bias towards IL-17 and IL-22 expression, whereas blood MAIT cells produced primarily IFN-γ, TNF, and Granzyme B. Furthermore, both FGT- and blood-derived MAIT cells were polyfunctional and contributed to the T-cell-mediated response to E. coli. Thus, MAIT cells in the genital mucosa have a distinct IL-17/IL-22 profile and may have an important role in the immunological homeostasis and control of microbes at this site.

Human MAIT-cell responses to Escherichia coli: activation, cytokine production, proliferation, and cytotoxicity

Mucosa‐associated invariant T cells are a large and relatively recently described innate‐like antimicrobial T‐cell subset in humans. These cells recognize riboflavin metabolites from a range of microbes presented by evolutionarily conserved major histocompatibility complex, class I‐related molecules. Given the innate‐like characteristics of mucosa‐associated invariant T cells and the novel type of antigens they recognize, new methodology must be developed and existing methods refined to allow comprehensive studies of their role in human immune defense against microbial infection. In this study, we established protocols to examine a range of mucosa‐associated invariant T‐cell functions as they respond to antigen produced by Escherichia coli. These improved and dose‐ and time‐optimized experimental protocols allow detailed studies of MR1‐dependent mucosa‐associated invariant T‐cell responses to Escherichia coli pulsed antigen‐presenting cells, as assessed by expression of activation markers and cytokines, by proliferation, and by induction of apoptosis and death in major histocompatibility complex, class I‐related–expressing target cells. The novel and optimized protocols establish a framework of methods and open new possibilities to study mucosa‐associated invariant T‐cell immunobiology, using Escherichia coli as a model antigen. Furthermore, we propose that these robust experimental systems can also be adapted to study mucosa‐associated invariant T‐cell responses to other microbes and types of antigen‐presenting cells.

Contact-dependent interference with invariant NKT cell activation by herpes simplex virus-infected cells.

Invariant CD1d-restricted NKT (iNKT) cells play important roles in generating protective immune responses against infections. In this study, we have investigated the role of human iNKT cells in HSV-1 infection and their interaction with epidermal keratinocytes. These cells express CD1d and are the primary target of the virus. Keratinocytes loaded with α-galactosyl ceramide (α-GalCer) could stimulate IFN-γ production and CD25 upregulation by iNKT cells. However, both α-GalCer–dependent and cytokine-dependent activation of iNKT cells was impaired after coculture with HSV-1–infected cells. Notably, CD1d downregulation was not observed on infected keratinocytes, which were also found to inhibit TCR-independent iNKT cell activation. Further examination of the cytokine profile of iNKT–keratinocyte cocultures showed inhibition of IFN-γ, IL-5, IL-10, IL-13, and IL-17 secretion but upregulation of IL-4 and TNF-α after the infection. Moreover, cell-to-cell contact between infected keratinocytes and iNKT cells was required for the inhibition of activation, as the cell-free supernatants containing virus did not affect activation. Productive infection of iNKT cells was however not required for the inhibitory effect. After coculture with infected cells, iNKT cells were no longer responsive to further stimulation with α-GalCer–loaded CD1d-expressing cells. We found that exposure to HSV-1–infected cells resulted in impaired TCR signaling downstream of ZAP70. Additionally, infected cells upregulated the expression of the negative T cell regulator, galectin-9; however, blocking experiments indicated that the impairment of iNKT cell responses was independent of galectin-9. Thus, interference with activation of human iNKT cells by HSV-1 may represent a novel immunoevasive strategy used by the virus to avoid immune clearance.

Innate Invariant NKT Cell Recognition of HIV-1–Infected Dendritic Cells Is an Early Detection Mechanism Targeted by Viral Immune Evasion

Invariant NKT (iNKT) cells are innate-like T cells that respond rapidly with a broad range of effector functions upon recognition of glycolipid Ags presented by CD1d. HIV-1 carries Nef- and Vpu-dependent mechanisms to interfere with CD1d surface expression, indirectly suggesting a role for iNKT cells in control of HIV-1 infection. In this study, we investigated whether iNKT cells can participate in the innate cell–mediated immune response to HIV-1. Infection of dendritic cells (DCs) with Nef- and Vpu-deficient HIV-1 induced upregulation of CD1d in a TLR7-dependent manner. Infection of DCs caused modulation of enzymes in the sphingolipid pathway and enhanced expression of the endogenous glucosylceramide Ag. Importantly, iNKT cells responded specifically to rare DCs productively infected with Nef- and Vpu-defective HIV-1. Transmitted founder viral isolates differed in their CD1d downregulation capacity, suggesting that diverse strains may be differentially successful in inhibiting this pathway. Furthermore, both iNKT cells and DCs expressing CD1d and HIV receptors resided in the female genital mucosa, a site where HIV-1 transmission occurs. Taken together, these findings suggest that innate iNKT cell sensing of HIV-1 infection in DCs is an early immune detection mechanism, which is independent of priming and adaptive recognition of viral Ag, and is actively targeted by Nef- and Vpu-dependent viral immune evasion mechanisms.

No difference in the rate of change in telomere length or telomerase activity in HIV-infected patients after three years of darunavir/ritonavir with and without nucleoside analogues in the MONET trial.

Objective To determine whether nucleos(t)ide reverse transcriptase inhibitors (NRTI) contribute to an accelerated loss in telomere length (TL) in HIV-infected patients on antiretroviral therapy (ART). Design Substudy of randomised controlled trial. Methods Patients with HIV RNA <50 copies/mL on combination ART (n = 256) were randomised to darunavir/ritonavir (DRV/r) 800/100 mg once daily, either as monotherapy (n = 127) or with 2 NRTIs (n = 129) for up to 144 weeks. TL and telomerase activity was quantified on stored peripheral blood mononuclear cells (PBMC; n = 124) using quantitative real time PCR. Results Patients in the sub-study had a mean age of 44 years and had received NRTI for a mean of 6.4 years (range 1–20 years). As expected, older patients have significantly shorter TL (p = 0.006), while women had significantly longer TL (p = 0.026). There was no significant association between TL and either the duration of prior NRTI treatment (p = 0.894) or the use of a PI versus NNRTI (p = 0.107). There was no significant difference between patients who continued or ceased NRTI in the mean change/year of TL or telomerase (p = 0.580 and 0.280 respectively). Conclusion Continuation versus cessation of NRTI treatment was not associated with an accelerated loss in TL or telomerase activity.

Will loss of your MAITs weaken your HAART

Mucosa-associated Invariant T (MAIT) cells are an evolutionarily conserved innate-like T cell subset that recognizes antigens presented by MR1 molecules. These antigens include vitamin B derivatives shared by many potentially pathogenic microbes, including Mycobacterium tuberculosis and Candida albicans. It was recently discovered that MAIT cells decay numerically and functionally in HIV-1 infection, and that they fail to recover despite several years of effective suppression of viral replication by antiretroviral therapy (ART). Here, we briefly discuss the roles of MAIT cells and their loss in HIV immunopathogenesis. We furthermore propose that the persistence of MAIT cell loss on ART needs to be taken into account when assessing the immunological response to treatment, and when treatment should commence. The importance of this T cell subset in HIV-1 infection needs further study, and interventions to restore the MAIT cell compartment should be considered.