Irradiated homozygous TGF- β 1 knockout fibroblasts show enhanced clonogenic survival as compared with TGF- β 1 wild-type fibroblasts

Abstract

Purpose : To study the role of transforming growth factor β 1 (TGF- β 1) on cellular radiation sensitivity by analysing mouse lung fibroblasts of different TGF- β 1 genotypes. Materials and methods : Heterozygous TGF- β 1 knock-out mice were mated to produce offspring of different TGF- β 1 genotypes as confirmed by PCR-genotyping. Primary lung fibroblast populations were established from new-born animals of specific genotypes (TGF- β 1 +/+, TGF- β 1 +/-, TGF- β 1 -/-) . Production of TGF- β 1 was tested by ELISA. TGF- β 1 receptor-II mRNA expression was analysed by RT-PCR. Colony formation of untreated, TGF- β 1-treated and/or irradiated primary lung fibroblasts was determined under different medium conditions. Results : Plating efficiencies under different medium conditions were independent of TGF- β 1 genotype. Production of TGF- β 1 correlated with the genotype: heterozygous TGF- β 1 knock-out fibroblasts (TGF- β 1 +/-) produced 60-65% of wild-type (TGF- β 1 +/+ cells). As expected, homozygous TGF- β 1 knock-out fibroblasts (TGF- β 1 -/-) did not produce TGF- β 1. Radiation exposure significantly enhanced TGF- β 1 production in TGF- β 1 +/+ cells by a factor of 2. No such stimulation was observed in TGF- β 1 +/- cells. TGF- β 1 +/- and especially TGF- β 1 -/- cells were significantly more radioresistant than TGF- β 1 +/+ cells. TGF- β 1 treatment significantly reduced clonogenic survival for both TGF- β 1 +/+ and TGF- β 1 -/- cells. TGF- β 1 treatment of TGF- β 1 -/- cells resulted in an enhancement of radiation sensitivity. Conclusion : The data are the first direct evidence that TGF- β 1 is a major autocrine regulator of intrinsic radiation sensitivity of mouse lung fibroblasts.