Tumor-Stroma-Inflammation Networks Promote Pro-metastatic Chemokines and Aggressiveness Characteristics in Triple-Negative Breast Cancer

In this study, MSCs of four different healthy donors were used. Patient-derived CAFs from a primary breast tumor (used in ELISA and their accompanying signaling experiments) and from a lung metastasis (used in tumor cell invasion assays) were kindly provided by Dr. Bar, Sheba Medical Center, Ramat Gan, Israel). The cells were grown, identified and immortalized as described in Katanov et al. (67). TNFα and IL-1β Concentrations Used in Different Analyses Titration studies were initiated by determining the ability of rhTNFα (#300-01A, PeproTech, Rocky Hill, NJ), and rhIL-1β (#200-01B, PeproTech) to elevate in MDA-MB-231 cells and/or MSCs/CAFs the expression of CXCL8, CCL2 and/or CCL5 to levels that enabled us to perform the required comparisons between different cell combinations in ELISA studies (concentrations studied - TNFα: 100 pg/ml, 1 ng/ml, 10 ng/ml; IL-1β: 20, 100, 250, 350, 500, 750 pg/ml). The selected concentrations of 10 ng/ml TNFα and 350 pg/ml IL-1β were appropriate also for MSC and CAF experiments. Therefore, in all MDA-MB-231 studies, alone or with MSC/CAF, these selected concentrations were used in in vitro and in vivo experiments. In parallel, titration studies indicated that the above selected concentrations were not optimal for ELISA responses of BT-549 and MDA-MB-468 cells; thus, based on additional analyses, the concentrations of cytokines were raised in these two cell types:...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research