Regulation of early growth response 2 expression by secreted frizzled related protein 1

Kelly J. Gregory, Stephanie M. Morin, Sallie S. Schneider

Secreted frizzled-related protein 1 (SFRP1) expression is down-regulated in a multitude of cancers, including breast cancer. Loss of Sfrp1 also exacerbates weight gain as well as inflammation. Additionally, loss of SFRP1 enhances TGF-β signaling and the downstream MAPK pathway. TGF-β has been shown to increase the expression of Early Growth Response 2 (EGR2), a transcription factor implicated in immune function in a wide variety of cell types. The work described here was initiated to determine whether SFRP1 modulation affects TGF-β mediated EGR2 expression in mammary tissues as well as macrophage polarization. Real-time PCR analysis was performed to examine EGR2 expression in human and murine mammary epithelial cells and tissues in response to SFRP1 modulation. Chemical inhibition was employed to investigate the roles TGF-β and MAPK signaling play in the control of EGR2 expression in response to SFRP1 loss. Primary murine macrophages were isolated from Sfrp1(-/-) mice and stimulated to become either M1 or M2 macrophages, treated with recombinant SFRP1, and real-time PCR was used to measure the expression of murine specific M1/M2 markers [Egr2 (M2) and Gpr18 (M1)]. Immunohistochemical analysis was used to measure the expression of human specific M1/M2 markers [CD163 (M2) and HLA-DRA (M2)] in response to rSFRP1 treatment in human mammary explant tissue. Knockdown of SFRP1 expression increases the expression of EGR2 mRNA in human mammary epithelial cells and addition of rSFRP1 decreases the expression of EGR2 when added to explant mammary gland tissues. Chemical inhibition of both TGF-β and MAPK signaling in Sfrp1(-/-) or knockdown mammary epithelial cells results in decreased expression of EGR2. Stimulated murine macrophages obtained from Sfrp1(-/-) mice and treated with rSFRP1 exhibit a reduction in Egr2 expression and an increase in Gpr18 mRNA expression. Human mammary explant tissue treated with rSFRP1 decreases CD163 protein expression whereas there was no effect on the expression of HLA-DRA. Loss of SFRP1 likely contributes to tumor progression by altering the expression of a critical transcription factor in both the epithelium and the immune system.