Intestinal Epithelial TBK1 Prevents Differentiation of T-helper 17 Cells and Tumorigenesis in Mice.

Abstract

Intestinal epithelial cells (IECs) regulate intestinal immune cells, particularly development of T-helper 17 (Th17) cells. Deregulation of this process leads to intestinal inflammation and tumorigenesis, via unknown mechanisms. TANK-binding kinase 1 (TBK1) is expressed by IECs and cells in the innate immune system. We studied the functions of TBK1 in the intestinal immune response and tumorigenesis in mice. We performed studies of wild-type mice, mice with conditional disruption of Tbk1 (Tbk1^IEC-KO), Tbk1^IEC-KO mice crossed with Apc^Min/+ mice, and Mt^-/- mice crossed with Apc^Min/+ mice. Some mice were given intraperitoneal injections of a neutralizing antibody against interleukin 17 (IL17) or IL1β. Intestine tissues were collected from mice and analyzed by histology, for numbers of adenomas and Th17 cells, and expression of inflammatory cytokines by real-time PCR. IECs were isolated from wild-type and Tbk1^IEC-KO mice, stimulated with lipopolysaccharide, co-cultured for with bone marrow-derived macrophages, and analyzed by RNA sequencing and biochemical analyses. Compared to Apc^Min/+Tbk1^WT mice, Apc^Min/+Tbk1^IEC-KO mice had significant increases in number and size of intestinal polyps, and significantly more Th17 cells in lamina propria. Administration of an antibody against IL17 reduced the number of intestinal polyps in Apc^Min/+Tbk1^IEC-KO mice to that observed in Apc^Min/+Tbk1^WT mice. In culture, TBK1-deficient IECs promoted expression of IL1β by macrophages, which induced differentiation of naïve CD4⁺ T cells into Th17 cells. RNA sequencing analysis revealed that the TBK1-deficient IECs had increased expression of metallothionein 1 (MT1), an immune regulator that promotes intestinal inflammation. Intestine tissues from Apc^Min/+Mt^-/- mice had significant fewer Th17 cells than Apc^Min/+Mt^+/+ mice, and a significantly lower number of polyps. Analyses of colorectal tumors in the Cancer Genome Atlas found colorectal tumors with high levels of MT1 and IL17 mRNAs to be associated with reduced survival times of patients. Expression of TBK1 by IECs suppresses expression of MT1 and prevents expression of IL1β by macrophages and differentiation of Th17 cells, to prevent inflammation and tumorigenesis. Strategies to block this pathway might be developed for colorectal tumorigenesis.