A subset of follicular helper-like MAIT cells can provide B cell help and support antibody production in the mucosa

We identify a MAIT cell subset expressing T follicular helper markers and show the ability of MAIT cells to support B cell responses in the mucosa.

Multi-Dimensionality Immunophenotyping Analyses of MAIT Cells Expressing Th1/Th17 Cytokines and Cytotoxic Markers in Latent Tuberculosis Diabetes Comorbidity

Mucosal-associated invariant T (MAIT) cells are innate like, and play a major role in restricting disease caused by Mycobacterium tuberculosis (Mtb) disease before the activation of antigen-specific T cells. Additionally, the potential link and synergistic function between diabetes mellitus (DM) and tuberculosis (TB) has been recognized for a long time. However, the role of MAIT cells in latent TB (LTB) DM or pre-DM (PDM) and non-DM (NDM) comorbidities is not known. Hence, we examined the frequencies (represented as geometric means, GM) of unstimulated (UNS), mycobacterial (purified protein derivative (PPD) and whole-cell lysate (WCL)), and positive control (phorbol myristate acetate (P)/ionomycin (I)) antigen stimulated MAIT cells expressing Th1 (IFNγ, TNFα, and IL-2), Th17 (IL-17A, IL-17F, and IL-22), and cytotoxic (perforin (PFN), granzyme (GZE B), and granulysin (GNLSN)) markers in LTB comorbidities by uniform manifold approximation (UMAP) and flow cytometry. We also performed a correlation analysis of Th1/Th17 cytokines and cytotoxic markers with HbA1c, TST, and BMI, and diverse hematological and biochemical parameters. The UMAP analysis demonstrated that the percentage of MAIT cells was higher; T helper (Th)1 cytokine and cytotoxic (PFN) markers expressions were different in LTB-DM and PDM individuals in comparison to the LTB-NDM group on UMAP. Similarly, no significant difference was observed in the geometric means (GM) of MAIT cells expressing Th1, Th17, and cytotoxic markers between the study population under UNS conditions. In mycobacterial antigen stimulation, the GM of Th1 (IFNγ (PPD and WCL), TNFα (PPD and WCL), and IL-2 (PPD)), and Th17 (IL-17A, IL-17F, and IL-22 (PPD and/or WCL)) cytokines were significantly elevated and cytotoxic markers (PFN, GZE B, and GNLSN (PPD and WCL)) were significantly reduced in the LTB-DM and/or PDM group compared to the LTB-NDM group. Some of the Th1/Th17 cytokines and cytotoxic markers were significantly correlated with the parameters analyzed. Overall, we found that different Th1 cytokines and cytotoxic marker population clusters and increased Th1 and Th17 (IL-17A, IL-22) cytokines and diminished cytotoxic markers expressing MAIT cells are associated with LTB-PDM and DM comorbidities.

Human MAIT Cells Respond to Staphylococcus aureus with Enhanced Anti-Bacterial Activity

Mucosal-Associated Invariant T (MAIT) cells have been shown to play protective roles during infection with diverse pathogens through their propensity for rapid innate-like cytokine production and cytotoxicity. Among the potential applications for MAIT cells is to defend against Staphylococcus aureus, a pathogen of serious clinical significance. However, it is unknown how MAIT cell responses to S. aureus are elicited, nor has it been investigated whether MAIT cell cytotoxicity is mobilized against intracellular S. aureus. In this study, we investigate the capacity of human MAIT cells to respond directly to S. aureus. MAIT cells co-cultured with dendritic cells (DCs) infected with S. aureus rapidly upregulate CD69, express IFNγ and Granzyme B and degranulate. DC secretion of IL-12, but not IL-18, was implicated in this immune response, while TCR binding of MR1 is required to commence cytokine production. MAIT cell cytotoxicity resulted in apoptosis of S. aureus-infected cells, and reduced intracellular persistence of S. aureus. These findings implicate these unconventional T cells in important, rapid anti-S. aureus responses that may be of great relevance to the ongoing development of novel anti-S. aureus treatments.

Human MAIT cells respond to and suppress HIV-1

Human MAIT cells sit at the interface between innate and adaptive immunity, are polyfunctional and are capable of killing pathogen infected cells via recognition of the Class IB molecule MR1. MAIT cells have recently been shown to possess an antiviral protective role in vivo and we therefore sought to explore this in relation to HIV-1 infection. There was marked activation of MAIT cells in vivo in HIV-1 infected individuals, which decreased following ART. Stimulation of THP1 monocytes with R5 tropic HIVBAL potently activated MAIT cells in vitro. This activation was dependent on IL-12 and IL-18 but was independent of the TCR. Upon activation, MAIT cells were able to up-regulate granzyme B, IFNg and HIV-1 restriction factors CCL3, 4 and 5. Restriction factors produced by MAIT cells inhibited HIV-1 infection of primary PBMCs and immortalized target cells in vitro. These data reveal MAIT cells to be an additional T cell population responding to HIV-1, with a potentially important role in controlling viral replication at mucosal sites.

Activation and Functional Alteration of Mucosal-Associated Invariant T Cells in Adult Patients With Community-Acquired Pneumonia

Community-acquired pneumonia (CAP) remains the significant infectious cause of morbidity and mortality worldwide. Although mucosal-associated invariant T cells (MAIT) play roles in the pathogenesis of children CAP and ICU-associated pneumonia, their roles in adult CAP are largely unexplored. In this study, we investigated the frequency, phenotype, and function of MAIT cells in peripheral blood and bronchoalveolar lavage fluid (BALF) of adult CAP patients. Our data indicate that MAIT-cell frequency is profoundly lower in the peripheral blood of CAP patients compared to that in healthy individuals. Furthermore, the circulatory MAIT cells express higher levels of CD69 and PD-1 compared to those in healthy individuals. In BALF of CAP patients, MAIT-cell frequency is higher and MAIT cells express higher levels of CD69 and PD-1 compared to their matched blood counterparts. Levels of IL-17A and IFN-γ are increased in BALF of CAP patients compared to those in BALF of patients with pulmonary small nodules. The IL-17A/IFN-γ ratio is significantly positively correlated with MAIT frequency in BALF of CAP patients, suggesting a pathogenic role of MAIT-17 cells in CAP. Of note, blood MAIT-cell frequency in CAP patients is strongly negatively correlated with high-sensitivity C-reactive protein (hsCRP) and neutrophil count percentage in blood. The ability of circulating MAIT cells in CAP patients to produce IFN-γ is significantly impaired compared to those in healthy individuals. In summary, our findings suggest the possible involvement of MAIT cells in the immunopathogenesis of adult CAP.

Clostridioides difficile Toxin CDT Induces Cytotoxic Responses in Human Mucosal-Associated Invariant T (MAIT) Cells

Clostridioides difficile is the major cause of antibiotic-associated colitis (CDAC) with increasing prevalence in morbidity and mortality. Severity of CDAC has been attributed to hypervirulent C. difficile strains, which in addition to toxin A and B (TcdA, TcdB) produce the binary toxin C. difficile transferase (CDT). However, the link between these toxins and host immune responses as potential drivers of immunopathology are still incompletely understood. Here, we provide first experimental evidence that C. difficile toxins efficiently activate human mucosal-associated invariant T (MAIT) cells. Among the tested toxins, CDT and more specifically, the substrate binding and pore-forming subunit CDTb provoked significant MAIT cell activation resulting in selective MAIT cell degranulation of the lytic granule components perforin and granzyme B. CDT-induced MAIT cell responses required accessory immune cells, and we suggest monocytes as a potential CDT target cell population. Within the peripheral blood mononuclear cell fraction, we found increased IL-18 levels following CDT stimulation and MAIT cell response was indeed partly dependent on this cytokine. Surprisingly, CDT-induced MAIT cell activation was found to be partially MR1-dependent, although bacterial-derived metabolite antigens were absent. However, the role of antigen presentation in this process was not analyzed here and needs to be validated in future studies. Thus, MR1-dependent induction of MAIT cell cytotoxicity might be instrumental for hypervirulent C. difficile to overcome cellular barriers and may contribute to pathophysiology of CDAC.

Transcriptomes and metabolism define mouse and human MAIT cell heterogeneity

Mucosal-associated invariant T (MAIT) cells are a subpopulation of T lymphocytes that respond to microbial metabolites. We performed single-cell RNA sequencing and metabolic analyses of MAIT cell subsets in thymus and peripheral tissues from mice and humans to define the heterogeneity and developmental pathway of these innate-like lymphocytes. We show that the predominant mouse subset, which produces IL-17 (MAIT17), and the subset that produces IFNγ (MAIT1), have greatly different transcriptomes and metabolic states in the thymus and periphery. A splenic MAIT subset has a transcriptome similar to circulating lymphocytes, and in mice these also are found in recent thymic emigrants, suggesting partially mature cells emigrate from the thymus. Human MAIT cells are predominantly MAIT1 cells, but have a different metabolism from their mouse counterparts with increased fatty acid uptake and storage. Although mouse and human subsets are similar in thymus, in the periphery they diverge, likely reflecting environmental influences.

Single-cell Sequencing Resolves Landscape of Immune cell and Regulatory Mechanisms in HIV-infected immune non-responders

Immune non-responder (INR) after highly active antiretroviral therapy (HAART) is the main cause of opportunistic infection and high mortality in AIDS patients, but the mechanism underlying immune reconstitution failure is poorly understood. Here we performed scRNA-seq and scATAC-seq analysis of peripheral blood mononuclear cells (PBMCs) derived from Immune nonresponder (INR) and responder (IR) HIV-1-infected subjects. We then describe the mitochondrial function of MAIT by flow cytometry in INRs and IRs. We founded there were diminished frequencies and numbers of MAIT cells in INRs, and MAIT cells from INRs displayed transcriptional profiles associated with the impairment of mitochondrial function and apoptosis signaling. ScATAC-seq and flow cytometry revealed diminished mitochondrial fitness in MAIT cells from INRs, and MAIT had low expression of transcription factor A for mitochondria (TFAM) and PPARA. These findings demonstrate the restoring mitochondrial function could modulate the immune dysfunction characteristic of MAIT, which against bacterial co-infections in INRs subjects.

Harnessing the Potential of MAIT Cells as Cellular Adjuvants in Mucosal Vaccines

<p>The development of vaccines is considered one of the most successful medical interventions to date, preventing millions of deaths every year. However, the majority of vaccines are administered peritoneally, despite the vast majority of pathogens invade the human host at mucosal sites. By vaccinating at distal sites, little to no protection is developed at the mucosa where the initial invasion occurs. There are however, a handful of licenced mucosally administered vaccines against infections such as poliovirus, influenza and Salmonella Typhi that are able to induce both a systemic and mucosal protective immune response. All but one of the current licenced mucosal vaccines are live attenuated due in part to the difficulty of developing new mucosal adjuvants. Recombinant cholera toxin subunit B is the only adjuvant used in the current licenced mucosal vaccines. While inactivated and subunit vaccines are considered safer as they are unable to revert back to virulent pathogens, adjuvants are required to boost their immunogenicity. This thesis therefore explores whether mucosal-associated invariant T (MAIT) cells which are found in mucosal tissues, are invariant in nature and have rapid activation, could be exploited as cellular adjuvants in mucosal vaccines. This thesis was able to show that intranasally administered MAIT cell agonist components, 5-A-RU and methylglyoxal (MG), are able to induce both MAIT cell and conventional dendritic cell (cDC) activation in the lung tissue and mediastinal lymph node (mLN). In this model CD40L and RANKL co-stimulatory interactions are involved in ICOSL expression on cDCs in the lung and associated with cDC activation. The MAIT cells within this model also maintained a RORyT and GATA3 phenotype after both one and three doses of the 5-A-RU + MG vaccine. Furthermore, a prime-boost intranasal vaccine scheme of 5-A-RU + MG and the model antigen OVA, was able to induce MR1-dependent accumulation of TFH cells and antigen-specific germinal center B cells in the mLN along with systemic antigen-specific IgG antibody production. This humoral response was also dependent on the presence of both cDC1 and cDC2 populations. Together, this thesis suggests MAIT cells have the potential to be utilised as cellular adjuvants in mucosal vaccines.</p>