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.