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Revealing immune responses in the Mycobacterium avium subsp. paratuberculosis-infected THP-1 cells using single cell
Hong-Tae Park1, Woo Bin Park1, Suji Kim1
1Department of Infectious Disease, College of Veterinary Medicine, Seoul National University, Seoul, Korea.
Abstract:
Mycobacterium avium subsp. paratuberculosis (MAP) is a causative agent of Johne's disease, which is a chronic and debilitating disease in ruminants. MAP is also considered to be a possible cause of Crohn's disease in humans. However, few studies have focused on the interactions between MAP and human macrophages to elucidate the pathogenesis of Crohn's disease. We sought to determine the initial responses of human THP-1 cells against MAP infection using single-cell RNA-seq analysis. Clustering analysis showed that THP-1 cells were divided into seven different clusters in response to phorbol-12-myristate-13-acetate (PMA) treatment. The characteristics of each cluster were investigated by identifying cluster-specific marker genes. From the results, we found that classically differentiated cells express CD14, CD36, and TLR2, and that this cell type showed the most active responses against MAP infection. The responses included the expression of proinflammatory cytokines and chemokines such as CCL4, CCL3, IL1B, IL8, and CCL20. In addition, the Mreg cell type, a novel cell type differentiated from THP-1 cells, was discovered. Thus, it is suggested that different cell types arise even when the same cell line is treated under the same conditions. Overall, analyzing gene expression patterns via scRNA-seq classification allows a more detailed observation of the response to infection by each cell type.
Insights
Mycobacterium avium subsp. paratuberculosis (MAP) infection triggers diverse responses in human THP-1 cells. Single-cell RNA sequencing identified distinct cell types, including a novel Mreg cell, and revealed classical cells mounting a strong inflammatory response.
Area of Science:
- Immunology
- Microbiology
- Genomics
Background:
- Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne's disease in ruminants and is implicated in human Crohn's disease.
- Limited research exists on MAP interactions with human macrophages, hindering understanding of Crohn's disease pathogenesis.
- THP-1 cells, a human monocytic cell line, serve as a model for studying macrophage responses to infection.
Purpose of the Study:
- To investigate the initial cellular and molecular responses of human THP-1 cells upon infection with MAP.
- To utilize single-cell RNA sequencing (scRNA-seq) to analyze heterogeneity in THP-1 cell responses.
- To identify specific cell types and their gene expression profiles during MAP infection.
Main Methods:
- THP-1 cells were differentiated using phorbol-12-myristate-13-acetate (PMA).
- Single-cell RNA sequencing (scRNA-seq) was performed on differentiated THP-1 cells after MAP infection.
- Clustering analysis and marker gene identification were used to characterize distinct cell populations and their responses.
Main Results:
- scRNA-seq revealed seven distinct clusters of THP-1 cells following PMA treatment.
- Classically differentiated cells (expressing CD14, CD36, TLR2) exhibited the most robust response to MAP.
- This response included upregulated expression of pro-inflammatory cytokines and chemokines (e.g., CCL4, IL1B, IL8).
- A novel cell type, termed Mreg cells, was identified, highlighting cellular heterogeneity even under uniform conditions.
Conclusions:
- MAP infection induces diverse responses within the THP-1 cell population, indicating cellular heterogeneity.
- Classically differentiated macrophages are key responders, initiating inflammatory pathways relevant to MAP infection.
- The discovery of Mreg cells suggests previously unrecognized cellular differentiation pathways in response to stimuli.
- scRNA-seq is a powerful tool for dissecting complex cellular responses to pathogens at a single-cell level.
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