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Roles of CX3CR1+ macrophages and CD8+ T cells for gut IgA production and immune regulation

Roles of CX3CR1+ macrophages and CD8+ T cells for gut IgA production and immune regulation

  • 김영인 (Young-In Kim, 아주대학교 대학원 약학대학)

초록/요약

Mucosal tissues such as the gastrointestinal, respiratory and genital tract are exposed to large numbers of commensal microbiota and pathogens. Therefore, the mucosal immune system has various lymphoid cells to protect the host from the harsh environment. The current study is about the intestinal CX3CR1+ macrophages and CD8+ T cells and how they contribute for immune regulation in the gut. Various innate immune cells in the intestine contribute to support the production of secretory IgA. IgA is the most abundant immunoglobulin in the gut and is crucial for intestinal homeostasis. This research shows that intestinal CX3CR1+ macrophages can support B cells to produce IgA in the gut. Intestinal CX3CR1+ macrophages express BAFF, APRIL and TNF-α which was responsible for inducing IgA by B cells. Unlike classical dendritic cells, support of IgA production from intestinal CX3CR1+ macrophages were independent of retinoic acid and microbial signals. CD8+ T cells can further enhance CX3CR1+ macrophages to induce IgA production by secreting IL-9 and IL-13. The IgA production support of intestinal CX3CR1+ macrophages occurred in the small intestine lamina propria and did not require migration to the mesenteric lymph nodes. CD8+ T cells primed by CX3CR1+ macrophages express IL-10 which is crucial for the control of inflammatory disorders in the intestine. However, the master regulatory cells which produce IL-10 are the Foxp3+CD4+ regulatory T cells. For functional activation of Foxp3+CD4+ regulatory T cells, they need to migrate from the periphery to lymph nodes via expression of CCR7. In this study, CCR7-/- mice were fed with dextran sulfate sodium to induce colitis. Since CCR7-/- Foxp3+CD4+ regulatory T cells are reported to have abnormal functionality due to aberrant localization, dextran sulfate sodium induced colitis was expected to worsen in CCR7-/- mice compared to wild type mice. However, there was no significant difference in the pathological grade of colon tissues. The body weight of CCR7-/- mice did not decrease as much and survived longer than wild type mice. The suppressive cytokine IL-10 expression was increased in CD8+ T cells of CCR7-/- mice. Also, plasmacytoid dendritic cells expanded in CCR7-/- mice. Depletion of CD8+ T cells and plasmacytoid dendritic cells in CCR7-/- mice showed severe colitis after DSS treatment, which shows their role of immune suppression in CCR7-/- mice. Taken together, intestinal CX3CR1+ macrophages help B cells secrete IgA after circulatory antigen delivery, which can be enhanced in the presence of CD8+ T cells. Also, when CCR7-/- mice were induced colitis, IL-10 expressing CD8+ T cells and plasmacytoid dendritic cells compensated for the suppressive effect upon inflammation. This research reveals how intestinal CX3CR1+ macrophages and CD8+ T cells function for gut immune regulation.

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목차

Abstract ‧‧‧ i
Contents ‧‧‧ iii
List of Table ‧‧‧ vii
List of Figures ‧‧‧ viii
Abbreviations ‧‧‧ xi

Introduction ‧‧‧ 1
1. Intestinal dendritic cells and macrophages ‧‧‧ 1
2. Secretory IgA production in the intestine ‧‧‧ 2
3. Inflammatory bowel disease ‧‧‧ 5
4. Regulatory CD4+ and CD8+ T cells ‧‧‧ 6
5. CCR7 ‧‧‧ 7
6. Purpose of the study ‧‧‧ 7

Results ‧‧‧ 9
Part I. CX3CR1+ macrophages and CD8+ T cells support IgA production in the gut ‧‧‧ 9
1. Intestinal CX3CR1+ macrophages support B cells produce IgA ‧‧‧ 9
2. Intestinal CX3CR1+ macrophages express BAFF, APRIL and TNF-α for support of IgA production ‧‧‧ 17
3. Intestinal CX3CR1+ macrophages are less dependent from signals of commensal microbiota for IgA induction ‧‧‧ 20
4. Circulatory antigen delivery promotes antigen specific production of Ig ‧‧‧ 24
5. Intestinal CX3CR1+ macrophages are responsible for the increase of antigen specific IgA after circulatory antigen delivery ‧‧‧ 29
6. CD8+ T cells further enhance IgA production of CX3CR1+ macrophages via IL-9 and IL-13 ‧‧‧ 34
7. Migration to the MLN is not required for intestinal CX3CR1+ macrophages to induce IgA production after circulatory antigen delivery ‧‧‧ 40
8. Antigen-specific Ig generated after circulatory antigen delivery contributes to host protection ‧‧‧ 46
9. CX3CR1+ macrophages contribute to antigen-specific Ig generation in other immunization routes ‧‧‧ 52

Part II. CD8+ T cells and plasmacytoid dendritic cells maintain gut immune homeostasis in CCR7-/- mice ‧‧‧ 55
1. DSS-induced colitis in mice did not show more susceptibility for the deficiency of CCR7 ‧‧‧ 55
2. Infiltration of innate immune cells to the inflamed colon reduced in CCR7-/- mice ‧‧‧ 59
3. CD4+ Tregs accumulated in the intestine of CCR7-/- mice ‧‧‧ 62
4. CD8+ T cells express high levels of IL-10 in the colon of CCR7-/- mice ‧‧‧ 70
5. CCR7-/- CD8+ T cells have immune suppressive effects ‧‧‧ 73
6. DSS-induced colitis worsened after depletion of CD8+ T cells in CCR7-/- mice ‧‧‧ 76
7. Expansion of plasmacytiod DCs suppressed DSS-induced colitis in CCR7-/- mice ‧‧‧ 80

Discussion ‧‧‧ 85
Conclusion ‧‧‧ 89
Materials and Methods ‧‧‧ 91
1. Mice ‧‧‧ 91
2. Immunization and cell depletion ‧‧‧ 91
3. Cell isolation ‧‧‧ 92
4. In vitro cell culture for IgA production ‧‧‧ 93
5. T cell proliferation and inhibition ‧‧‧ 93
6. Fluorescent conjugated antibody staining for flow cytometry ‧‧‧ 94
7. Gene expression ‧‧‧ 94
8. Determination of Ig concentration by ELISA ‧‧‧ 95
9. Determination of OVA-specific antibody-secreting cells ‧‧‧ 96
10. Depletion of commensal microbiota in mice ‧‧‧ 96
11. Salmonella Typhimurium infection ‧‧‧ 96
12. DSS induced colitis and cell depletion ‧‧‧ 97
13. Histology of colon tissues ‧‧‧ 97
14. Statistics ‧‧‧ 97
References ‧‧‧ 98
Abstract in Korean (초록) ‧‧‧ 107

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