IL-21 isoform transgenic mice spontaneously develop mammary tumors: possible involvement of IL-21-induced osteopontin expression in tumorigenesis

Introduction Interleukin-21 (IL-21) is a pleiotropic cytokine produced by CD4 + T cells that critically regulates the differentiation and functions of various adaptive and innate immune cells. Emerging evidence suggests that IL-21 production increases in CD4 + T cells during aging. However, the long-term impact of sustained IL-21 exposure on peripheral tissues remains unclear. Methods To address this question, we generated transgenic mice expressing a membrane-bound IL-21 isoform specifically in T cells (IL-21isoTg mice) and analyzed age-associated tissue alterations. Results Female IL-21isoTg mice developed progressive mammary gland dysplasia beginning at approximately 14 weeks of age, and more than 45% developed mammary tumors after 12 months. Comprehensive gene expression analysis of mammary epithelial cells (MECs) isolated prior to tumor onset (10 weeks of age) revealed robust upregulation of the Spp1 gene encoding osteopontin (OPN) in IL-21isoTg mice. Quantitative RT-PCR confirmed a significant increase in Spp1 expression in both MECs and mammary stromal cells of IL-21isoTg mice. Given the established role of OPN in shaping the tumor microenvironment (TME) and promoting immunosuppression, we investigated the mechanism underlying Spp1 induction. Using splenocytes as a surrogate model for mammary stromal immune cells, we found that IL-21 stimulation selectively enhanced OPN expression in macrophages and B cells. In contrast, cytokines previously reported to induce Spp1 , including IL-1β, IL-6, IL-17A, and TNF-α, failed to significantly upregulate Spp1 in this context, indicating an IL-21-specific effect. Although IL-21 did not directly affect mammary epithelial tumor cell lines, it induced splenocyte-derived IL-6 and TNF-α, which subsequently promoted Spp1 expression in tumor cell lines. These findings suggest that in IL-21isoTg mice, T cells accumulating in the mammary gland produce IL-21iso, which directly induces OPN expression in macrophages and B cells, and indirectly enhances OPN expression in MECs through IL-6 and TNF-α. Age-associated increases in IL-21 production may, therefore, promote mammary tumor development and progression by enhancing OPN expression, leading to local inflammation and immunosuppression within the mammary tissue.