Eosinophils Exert Antitumorigenic Effects in the Development of Esophageal Squamous Cell Carcinoma.

BACKGROUND AND AIMS: Eosinophils are present in several solid tumors and have context-dependent function. Our aim is to define the contribution of eosinophils in esophageal squamous cell carcinoma (ESCC), as their role in ESCC is unknown. METHODS: Eosinophils were enumerated in tissues from 2 ESCC cohorts. Mice were treated with 4-NQO for 8 weeks to induce precancer or 16 weeks to induce carcinoma. The eosinophil number was modified by a monoclonal antibody to interleukin-5 (IL5mAb), recombinant IL-5 (rIL-5), or genetically with eosinophil-deficient (ΔdblGATA) mice or mice deficient in eosinophil chemoattractant eotaxin-1 (Ccl11-/-). Esophageal tissue and eosinophil-specific RNA sequencing was performed to understand eosinophil function. Three-dimensional coculturing of eosinophils with precancer or cancer cells was done to ascertain direct effects of eosinophils. RESULTS: Activated eosinophils are present in higher numbers in early-stage vs late-stage ESCC. Mice treated with 4-NQO exhibit more esophageal eosinophils in precancer vs cancer. Correspondingly, epithelial cell Ccl11 expression is higher in mice with precancer. Eosinophil depletion using 3 mouse models (Ccl11-/- mice, ΔdblGATA mice, IL5mAb treatment) all display exacerbated 4-NQO tumorigenesis. Conversely, treatment with rIL-5 increases esophageal eosinophilia and protects against precancer and carcinoma. Tissue and eosinophil RNA sequencing revealed eosinophils drive oxidative stress in precancer. In vitro coculturing of eosinophils with precancer or cancer cells resulted in increased apoptosis in the presence of a degranulating agent, which is reversed with NAC, a reactive oxygen species scavenger. ΔdblGATA mice exhibited increased CD4 T cell infiltration, IL-17, and enrichment of IL-17 protumorigenic pathways. CONCLUSION: Eosinophils likely protect against ESCC through reactive oxygen species release during degranulation and suppression of IL-17.

Eosinophils exert direct and indirect anti-tumorigenic effects in the development of esophageal squamous cell carcinoma.

Background/Aims: Eosinophils are present in several solid tumors and have context-dependent function. Our aim is to define the contribution of eosinophils in esophageal squamous cell carcinoma (ESCC), since their role in ESCC is unknown. Methods: Eosinophils were enumerated in tissues from two ESCC cohorts. Mice were treated with 4-nitroquinolone-1-oxide (4-NQO) for 8 weeks to induce pre-cancer or 16 weeks to induce carcinoma. Eosinophil number was modified by monoclonal antibody to IL-5 (IL5mAb), recombinant IL-5 (rIL-5), or genetically with eosinophil-deficient (ΔdblGATA) mice or mice deficient in eosinophil chemoattractant eotaxin-1 ( Ccl11 -/- ). Esophageal tissue and eosinophil specific RNA-sequencing was performed to understand eosinophil function. 3-D co-culturing of eosinophils with pre-cancer or cancer cells was done to ascertain direct effects of eosinophils. Results: Activated eosinophils are present in higher numbers in early stage versus late stage ESCC. Mice treated with 4-NQO exhibit more esophageal eosinophils in pre-cancer versus cancer. Correspondingly, epithelial cell Ccl11 expression is higher in mice with pre-cancer. Eosinophil depletion using three mouse models ( Ccl11 -/- mice, ΔdblGATA mice, IL5mAb treatment) all display exacerbated 4-NQO tumorigenesis. Conversely, treatment with rIL-5 increases esophageal eosinophilia and protects against pre-cancer and carcinoma. Tissue and eosinophil RNA-sequencing revealed eosinophils drive oxidative stress in pre-cancer. In vitro co-culturing of eosinophils with pre-cancer or cancer cells resulted in increased apoptosis in the presence of a degranulating agent, which is reversed with N-acetylcysteine, a reactive oxygen species (ROS) scavenger. ΔdblGATA mice exhibited increased CD4 T cell infiltration, IL-17, and enrichment of IL-17 pro-tumorigenic pathways. Conclusion: Eosinophils likely protect against ESCC through ROS release during degranulation and suppression of IL-17.

A method for scoring 4-nitroquinoline 1-oxide-induced murine esophageal squamous neoplasia.

A mouse model for esophageal squamous cell carcinoma (ESCC) is induced by oral administration of the carcinogen 4-nitroquinoline 1-oxide (4-NQO). There is not an objective method for determining histopathologic severity of disease in this model. We aim to create a clearly defined and easily applied scoring system that can quantify the severity of 4-NQO-induced murine ESCC. Fifteen wild-type C57BL/6J mice were treated with 4-NQO for 8 (n = 8) or 16 (n = 7) weeks, while the rest (n = 9) were treated with vehicle, as 8 weeks of 4-NQO typically results in dysplasia and 16 weeks in carcinoma. We identified histologic abnormalities of the esophagus in this model and developed metrics to grade severity of dysplasia, papillomas, and invasion. Scores were then calculated using quantitative digitized image analysis for measuring depth and extent of each feature within the entire sample. Each feature was also assigned a weight based on its relation to cancer severity. Histology scores were significantly different in the three groups, suggesting that this method can discriminate dysplasia from carcinoma. This model can be applied to any mouse treated with 4-NQO.