Migration and differentiation of epicardial cells stimulated by TGFβ1, TGFβ2, or BMP‐2 do not require the Type III Transforming Growth Factor β receptor

Abstract

Transforming Growth Factor β (TGFβ) Receptor III (TGFβR3) binds all three TGFβ ligands, inhibin, and Bone Morphogenic Protein‐2 (BMP‐2). Since TGFβ causes epicardial cell differentiation into smooth muscle in vitro we asked if Tgfbr3 null epicardial cells differentiate in response to TGFβ or BMP‐2. Wiltype and Tgfbr3−/− immortalized mouse epicardial cells (ED 11.5) were cultured in the presence or absence of ligands for 72 hours and scored by immunohistochemistry, QT‐PCR, or a reportor assay for Smooth Muscle 22 alpha (SM22α). Wildtype and Tgfbr3−/− cells lose epithelial character and undergo smooth muscle differentiation in response to 250 pM TGFβ1 or TGFβ2. Induction of SM22α in wildtype cells occurred with an EC50 of 12.5 pM. BMP‐2 (5 nM) induced the loss of Zonula Occludens‐1 (ZO‐1) but failed to induce the smooth muscle markers SM22α, smooth muscle α‐actin (SMαA) or calponin. Inhibition of activin receptor‐like kinase (ALK) 5 with 2.5 μM SB431542 blocked the effects of TGFβ1 and TGFβ2, but not BMP‐2. Pre‐ and co‐incubation of cells with 5 nM BMP‐2 did not block the ability of 250 pM TGFβ1 or TGFβ2 to induce smooth muscle markers. These data demonstrate that BMP2 induces epicardial cell motility without smooth muscle differentiation and that TGFβR3 is not required for epicardial cell responsiveness to TGFβ or BMP‐2.Funding: HL085708, NIGMS 5K12GM068543Grant Funding Source: NIH HL