Functional Plasticity of Human MAIT Cells and Their Role in Immune Regulation

Academic Background

Mucosal-Associated Invariant T cells (MAIT cells) are a type of unconventional T lymphocyte found abundantly in healthy humans, especially in mucosal tissues. MAIT cells exhibit innate-like characteristics, rapidly recognizing antigens derived from microbial riboflavin metabolic pathways, which are presented by non-polymorphic MHC-Ib-related proteins (MR1). MAIT cells play a crucial role in antimicrobial immunity, but the mechanisms regulating their functional heterogeneity remain unclear. The functional plasticity of MAIT cells, i.e., their ability to exhibit different functional states under varying microenvironmental conditions, has been a focus of recent immunological research.

The aim of this study was to explore how MAIT cells adjust their functional responses during antigen recognition based on cytokine signals in the microenvironment. Specifically, the research team sought to reveal how MAIT cells regulate inflammatory or anti-inflammatory responses upon exposure to different cytokines (such as IL-12, IL-18, etc.), and to further elucidate the underlying molecular mechanisms. This research not only enhances our understanding of MAIT cells’ roles in infection and inflammation but also provides a theoretical foundation for developing MAIT cell-based immunotherapeutic strategies.

Paper Source

This paper was co-authored by Caroline Boulouis, Elli Mouchtaridi, Thomas R. Müller, and others, with the research team hailing from institutions such as Karolinska Institutet (Sweden) and The University of Queensland (Australia). It was published in the journal PNAS (Proceedings of the National Academy of Sciences) on February 4, 2025, titled “Human MAIT cell response profiles biased toward IL-17 or IL-10 are distinct effector states directed by the cytokine milieu.”

Research Process and Results

Research Process

The research team used a combination of functional analysis and single-cell transcriptomics (scRNA-seq) to systematically investigate MAIT cell functional responses under different cytokine environments. Below are the specific steps:

1. Isolation and Stimulation of MAIT Cells
   MAIT cells were isolated from peripheral blood samples of healthy volunteers. THP-1 cells were used as antigen-presenting cells, loaded with 5-OP-RU (an antigen recognized by MAIT cells), and stimulated in the presence of various cytokines (e.g., IL-12, IL-18, IL-23).

2. Functional Analysis
   Flow cytometry was used to detect cytokine secretion (e.g., IL-10, IL-17, IFNγ, TNF) and surface marker expression (e.g., CD69, CD107a) in MAIT cells after stimulation. UMAP (Uniform Manifold Approximation and Projection) was employed for unsupervised clustering analysis of MAIT cell functional states.

3. Single-Cell Transcriptomics Analysis
   Single-cell RNA sequencing (scRNA-seq) was performed on stimulated MAIT cells to analyze transcriptional features under different stimulation conditions. Gene Set Enrichment Analysis (GSEA) and regulatory network analysis (PySCENIC) were conducted to identify transcription factors associated with IL-10 and IL-17 expression.

4. CRISPR/Cas9 Gene Knockout Experiments
   To verify the role of transcription factors c-Maf and Aiolos in IL-10 expression, the research team used CRISPR/Cas9 technology to knock out these genes and observed their effects on MAIT cell function.

5. Reversibility of Functional States
   Secretion capture techniques were used to isolate IL-10+ and IL-17+ MAIT cells, which were then cultured for two weeks and re-stimulated to assess the reversibility of their functional states.

Main Results

1. Cytokine Environment Significantly Influences MAIT Cell Functional Responses
   The study found that when MAIT cells recognize antigens in the presence of IL-12 or IL-23, they tend to secrete the anti-inflammatory cytokine IL-10. In contrast, IL-18 stimulation induces a strong inflammatory response, secreting pro-inflammatory cytokines such as IL-17 and GM-CSF. Notably, IL-18 significantly inhibits IL-10 expression.

2. Transcriptomics Reveals Shifts in MAIT Cell Functional States
   scRNA-seq analysis showed that MAIT cells stimulated under different cytokine conditions have unique transcriptional profiles. For example, IL-12 and IL-23 induced immune-regulatory gene expression (such as IL10, LAG3, CTLA4), while IL-18 activated genes related to inflammation and chemotaxis (such as IFNG, CSF2, CCL20).

3. c-Maf is a Critical Transcription Factor for IL-10 Expression in MAIT Cells
   Through CRISPR/Cas9 knockout experiments, the research team confirmed that c-Maf is essential for IL-10 expression in MAIT cells, whereas Aiolos, though highly expressed in IL-10+ cells, is not necessary.

4. Reversible Functional States of MAIT Cells
   Experiments demonstrated that the functional states of IL-10+ and IL-17+ MAIT cells are not stably differentiated but reversible effectors that can adjust their functions according to changes in the microenvironment.

Conclusion and Significance

This study systematically revealed how MAIT cells adjust their functional responses during antigen recognition based on cytokine signals in the microenvironment. The findings show that MAIT cells tend to secrete IL-10 and exhibit immune-regulatory functions under IL-12 and IL-23 stimulation, while displaying strong inflammatory reactions under IL-18 stimulation. c-Maf was identified as a critical transcription factor for IL-10 expression in MAIT cells. These discoveries deepen our understanding of MAIT cell functional plasticity and provide new insights for developing MAIT cell-based immunotherapeutic strategies.

Research Highlights

1. New Discoveries in Functional Plasticity: This study systematically reveals the functional plasticity of MAIT cells in different cytokine environments and its underlying molecular mechanisms.
2. Application of Transcriptomics: scRNA-seq comprehensively parses the transcriptional characteristics of MAIT cells, offering new perspectives on their functional regulation.
3. Validation via CRISPR/Cas9: Gene knockout experiments confirm the critical role of c-Maf in IL-10 expression, supporting the study’s conclusions.
4. Discovery of Reversible Functional States: The study shows that MAIT cell functional states are reversible, providing new possibilities for their application in immune regulation.

Other Valuable Information

The study also found that IL-10 secreted by MAIT cells can regulate monocyte immune responses through paracrine action, promoting the differentiation of anti-inflammatory macrophages. This finding further highlights the important role of MAIT cells in immune regulation.