Epperly, M. W., Goff, J. P., Zhang, X., Niu, Y., Shields, D. S., Wang, H., Shen, H., Franicola, D., Bahnson, A. B., Nie, S., Greenberger, E. E. and Greenberger, J. S. Increased Radioresistance, G2/M Checkpoint Inhibition, and Impaired Migration of Bone Marrow Stromal Cell Lines Derived from Smad3−/− Mice. Radiat. Res. 165, 671–677 (2006).
Smad3 protein is a prominent member of the Tgfb receptor signaling pathway. Smad3−/− mice display decreased radiation-induced skin fibrosis, suggesting a defect in both Tgfb-mediated fibroblast proliferation and migration. We established bone marrow stromal cell lines from Smad3−/− mice and homozygous littermate / mice. Smad3−/− cells displayed a significant increase in radiation resistance with a D0 = 2.25 ± 0.14 Gy compared to Smad3 / cells with a D0 = 1.75 ± 0.03 (P = 0.023). Radioresistance was abrogated by reinsertion of the human SMAD3 transgene, resulting in a D0 = 1.49 0.10 (P = 0.028) for Smad3−/−(3) cells. More Smad3−/− cells than Smad3 / cells were in the G2/M phase; Smad3−/−(3) cells were similar to Smad3 / cells. Smad3 / cells exhibited increased apoptosis 24 h after 5 Gy (15%) or 8 Gy (43%) compared to less than 1% in Smad3−/− cells exposed to either dose. The movement of Smad3−/− cells, measured in an automated cell tracking system, was slower than that of Smad3 / cells. Smad3−/−(3) cells resembled Smad3 / cells. These studies establish concordance of a defective Tgfb signal transduction pathway, an increased proportion of G2/M cells, and radioresistance. The decreased migratory capacity of Smad3−/− cells in vitro correlates with decreased radiation fibrosis in vivo in mice deficient in Tgfb signaling.