RAW 264.7 macrophages induce tumor-initiating behavior in metastatic but not nonmetastatic mammary carcinoma clones.

Abstract

Macrophages promote primary tumor growth and metastatic dissemination through a variety of mechanisms. This thesis examined the ability of RAW 264.7 murine macrophages to elicit malignant behaviors from a series of murine mammary carcinoma cell lines (4T1, 66cl4, 4TO7, 67NR) that differ extensively in their metastatic potential when transplanted in vivo. It was hypothesized that the RAW 264.7 macrophages would evoke malignant behaviors from the carcinoma cells in vitro, but in a way that correlated with the metastatic nature of each line. Conditioned media experiments revealed that paracrine cues from the RAW 264.7s induced the expression of Sca-1, a murine surface marker known to represent tumor-initiating cells, in the most metastatic 4T1 and 66cl4 carcinoma cell lines. This effect was attainable in carcinoma cell co-culture with the RAW 264.7s, where the metastatic 4T1 and 66cl4 cell lines also formed spheroid structures amid direct cell contact from the macrophages. A subsequent gene expression analysis performed on 4T1/RAW 264.7 co-cultures suggested that the RAW 264.7s induced Sca-1 expression in the metastatic cell lines through anti-tumoral actions. Finally, a functional study of Sca-1(+) 4T1 cells indicated that they proliferated faster but tended to be less motile than Sca-1(-) cells in response to RAW 264.7-conditioned media, further supporting their tumorinitiating capability. By integrating these findings into a metastatic narrative, this thesis proposes that macrophages enhance metastasis through the expansion of a tumor-initiating subpopulation in metastatic mammary carcinoma cell lines.