Macrophage-Derived Endotrophin Supports Tumor Migration Potentials in Thyroid Caner

Abstract

Abstract Endotrophin (ETP), a cleaved fragment of the C5 domain of the Type VI collagen α3 (Col6α3), has been shown to play pro-tumorigenic roles in breast and liver cancers. However, the ETP actions in tumor microenvironment (TME) is still undetermined. This study aimed to investigate the role and the mechanism of ETP in macrophage-enriched thyroid cancer TMEs. First, the expression of ETP on various human thyroid tissues was studied. Immunohistochemical staining showed that the ETP was expressed in papillary (PTC) or anaplastic (ATC) thyroid cancer tissues but not in benign adenoma or autoimmune thyroiditis. Among 146 PTCs, tumors were divided into two groups according to their ETP expression, ETPlow and ETPHigh. Tumor size was bigger and LN metastasis rates were higher in ETPHigh than ETPlow group. Interestingly, the ETP expression showed positive correlations with the CD163-positive macrophages in thyroid tumors. To clarify the origin of ETP in TME of thyroid cancer, xenograft tumor from FRO (ATC) cells was co-stained with anti-ETP and anti-F4/80 antibodies. Immunofluorescent staining showed that the expression of ETP and F4/80-postive macrophages were co-localized in the peritumoral area. Additionally, human monocytic THP-1 cells were treated with conditioned media (CM) from FRO or BCPAP cells (designated FRO-CM or BCPAP-CM) and demonstrated that ETP expression was more up-regulated in the FRO-CM or BCPAP-CM treated group than in the control group. Collectively, ETP produced from macrophages in thyroid cancer microenvironments. Next, the pro-tumorigenic functions of ETP has been studied. CMs from co-cultures of FRO or BCPAP cells with THP-1 cells were harvested and treated to FRO or BCPAP cells. Treatment of CMs from co-cultures increased cell migration potentials than that of the single cell alone, and these effects were reduced by anti-ETP neutralizing antibody, indicating that ETPs play essential role in macrophage-induced tumor cell migration potentials. Moreover, MMP-9 and MMP-14, the candidates of Col6α3 protease to produce ETP were increased in THP-1 cells by treatment of CM. Additionally, treatment of CMs from MMP-9 inhibitor-treated co-cultures of FRO and THP-1 cells decreased cancer cells migration potentials than the control CMs. Finally, the RNA-seq dataset of human thyroid cancer showed that higher expressions of ETP positively correlated with macrophage-related genes such as CD163 and MMPs including MMP-9 and MMP-14. In conclusion, macrophage contributes ETP productions by modulating MMP expressions and ETP supports pro-metastatic potentials of human thyroid cancer cells in TMEs. Thus ETP can be a good therapeutic target for macrophage-enriched advanced thyroid cancers.