Epithelial Ikkβ deletion modulates immune responses and the IFNγ/CXCL9 axis during early esophageal carcinogenesis.

Abstract

Esophageal cancer is a major cause of cancer-related death, often preceded with chronic inflammation and injuries. The NFκB/IKKβ pathway plays a central role in inflammation, yet its role in early esophageal carcinogenesis remains unclear. This study investigated the role of epithelial IKKβ in early esophageal carcinogenesis. Mice were treated with the carcinogen 4-nitroquinoline-1-oxide (4-NQO) or a vehicle for one month to induce precancerous lesions. Esophagi were harvested and examined through histological, protein, flow cytometry, and RNA analyses. Histological analysis revealed that 4-NQO treatment led to increased inflammation, intraepithelial CD45+ immune cells, and elevated IKKβ phosphorylation levels. Mice with esophageal epithelial-specific Ikkβ deletion (4-NQO/IkkβEEC-KO) showed delayed progression to a precancerous state, with reduced immune cell recruitment compared to 4-NQO/controls. Immunophenotyping showed decreased recruitment of T cells, including CD4+, CD8+ and regulatory (Tregs) T cells, and increased recruitment of macrophages in 4-NQO/IkkβEEC-KO mice compared to 4-NQO/controls. RNA sequencing data identified 262 differentially expressed genes in 4-NQO/IkkβEEC-KO mice, implicating pathways related to inflammation and wound healing. Notably, the chemokine CXCL9, a T cell chemoattractant, was significantly upregulated in 4-NQO control mice, but not in 4-NQO/IkkβEEC-KO mice. Further analysis identified IFNγ as an upstream regulator of Cxcl9 expression, and neutralization of IFNγ reduced Cxcl9 expression levels in 4-NQO treated mice. Additionally, in vitro studies demonstrated that IFNγ upregulates Cxcl9 in an NF-ĸB dependent manner in esophageal keratinocytes. These findings suggest that epithelial IKKβ regulates the immune microenvironment in early esophageal carcinogenesis through the IFNγ/CXCL9 axis and influencing T cell recruitment and inflammatory responses.