Type III Transforming Growth Factor‐ β Receptor regulates proliferation and apoptosis in epicardial cells

Epicardial cells transform into the myocardium and form coronary vessels. The Type III Transforming Growth Factor‐ β Receptor (Tgfbr3) is required for coronary vessel development. Tgfbr3‐/‐ embryos have an irregular epicardium, few coronary vessels, and die at E14.5. To explore TGF βR3 signaling we made immortalized epicardial cell lines from E11.5 embryos. Tgfbr3+/+, +/‐, and ‐/‐ cells form a tight epithelium. TGF β1 or TGF β2 (250 pM) caused transformation and induced the smooth muscle marker, SM22α. Circular wounds were made in cell monolayers and the percent wound closure calculated. Tgfbr3 +/+ and +/‐ cells closed the wound by 48h. Tgfbr3‐/‐ cells required 72h (p<0.05). Tgfbr3 +/+ and +/‐ cells gave similar proliferation rates when measured by BrdU, peaking at 48h (33.1% and 33.93%) and returning to basal levels (19.9% and 20.85%) by 72h. Tgfbr3‐/‐ cells sustained a basal proliferation rate 13% lower at 48h (p=0.001). As a second measure of proliferation, we used the in vivo reduction of MTS tetrazolium. Tgfbr3‐/‐ cells had a rate of proliferation 2‐fold lower at 48h and 2.6 fold lower at 72h (p<0.05). Apoptosis was determined by Apo‐One Caspase 3/7 Homogenous assay. The apoptosis rate in Tgfbr3‐/‐ cells was 1.8, 3.3 and 5.9‐ fold higher than Tgfbr3+/+ cells at 24, 48, 72h, respectively (p<0.05). In summary, TGFβR3 is not required for transformation in epicardial cells, but does regulate proliferation and apoptosis.Source: HL085708, HL076133 (AFA), AHA0655129 (JVB), GM007628 (NSS), & GM07347 (LAC).