RORα Suppresses Cancer-associated Inflammation And Metastasis Of Breast Cancer

Breast cancer development is associated with macrophage infiltration and differentiation in the tumor microenvironment.Our previous study highlights the crucial function of reactive oxygen species(ROS)in enhancing macrophage infiltration during the disruption of mammary tissue polarity.However,the regulation of ROS and ROS-associated macrophage infiltration in breast cancer has not been fully determined.Objective:To study the effects of RORα in breast cancer and its mechanism.Methods:We examined the effect of RORα in inhibiting activation of inflammation gene signature and expression of cytokines in breast cancer cells by 3D culture,microarray assay,q PCR and ELISA assay.We examined the effect of RORα in ROS and superoxide production by Facs and q PCR.We used Seahorse assay to analysis the effect of RORα in oxygen consumption rate(OCR).We used Ch IP assay to identified complex I subunits NDUFS6 and NDUFA11 as RORα targets.We used co-culture assay,4T1 xenografts experiment and facs analysis of tumor-associated macrophages to determine how RORα inhibits breast cancer metastasis and macrophage infiltration.Results:1.The expression of RORα in breast cancer cell lines significantly suppressed invasive tumor growth in 3D culture.Microarray assay and q PCR results showed that the inflammatory pathway and expression of cytokine genes were inhibited by RORα in 3D culture.ELISA assay showed that RORα decreased the IL-6 protein levels in breast cancer cells.2.RORα inhibited ROS production by repressing the expression of complex I genes.Seahorse assay showed that the inactivation of RORα in breast cancer cells enhances electron leakage in mitochondria by increasing NDUFS6 and NDUFA11 expression.Using gene co-expression and chromatin immunoprecipitation(Ch IP)analyses,we identified complex I subunits NDUFS6 and NDUFA11 as RORα targets that mediated its function in suppressing superoxide generation in mitochondria.3.RORα expression in mammary epithelial cells inhibited macrophage infiltration by repressing ROS generation in the co-culture assay.4.The expression of RORα in 4T1 cells significantly inhibited cancer metastasis,reduced macrophage accumulation,and enhanced M1-like macrophage differentiation in tumor tissue.Conclusion:Our data inditated that RORα is a tumor suppressor that can significantly reduce the level of ROS in breast cancer cells and inhibit ROS-induced cytokine expression.RORα inhibits ROS production by repressing the expression of Complex I genes.RORα expression inhibits mammary tumor metastasis and macrophage infiltration in tumor tissue.40 figures,14 tables,118 references.