Evaluating the Role of CCL20 in Prostate Cancer Progression

CCK-8, EDU, colony formation, scratch assay, transwell migration, and invasion assays were performed as previously reported [20 (link)]. Specifically, for cell proliferation assays, stably transfected PCa cells were treated with the conditioned medium (CM) from TAMs (co-cultured with PCa cells), human recombinant CCL20 protein (rh-CCL20, 20 ng/ml, #0511102, Peprotech) and neutralizing antibody to CCL20 (anti-CCL20, 5 μg/ml; ab9829, Abcam). The cell proliferative rate was assessed using the Cell Counting Kit-8 (CK-04, Dojindo). For tumor cell migration assays, the stable cells lines treated with or without 5 μg/ml CCL20 neutralizing antibody were seeded in the upper chambers; the lower chambers were filled with medium containing 10% FBS with or without 20 ng/ml CCL20 recombinant protein. In addition, the CM of TAMs was placed in the lower chamber and used as an attractor for PCa cell migration and invasion experiments.