Objective To investigate ff the γ-H12AX foci is a precise index for the DSB formation and repair in mature neurons of brain in vivo after clinically relevant doses irradiation. Methods For the DSB formation experiment, the mature neurons in the neocortex of brain tissue of C57BL/6 mice were analyzed at 10 rain after whole-body irradiation with 0.1, 0.5 and 1.0 Gy. For the DSB repair kinetics experiment, the mature neurons in the neocortex of brain tissue of repair-proficient (C57BL/6 mice) and repair-deficient mouse strains (BALB/c, A-T and SCID mice) were analyzed at 0.5, 2.5, 5, 24 and 48 h after whole-body irradiation with 2 Gy. The mature neurons in the neocortex of brain tissue of sham-irradiated mice of each strain served as controls. γ-H2AX immunohistochemistry and γ-H2AX and NeuN double immunofluorescence analysis was used to measure DSBs formation and repair in the mature neurons in the neocortex of brain tissue of the different mouse strains. Results For the DSB formation experiment, γ-H2AX foci levels with a clear linear dose correlation and very low backgrounds in the nuclei in the neocortex of brain tissue were observed. Scoring the loss of γ-H12AX foci allowed us to verify the different, genetically determined DSB repair deficiencies, including the minor impairment of BALB/c mice. Repair-proficient C57BL/6 mice exhibited the fastest decrease in foei number with time, and displayed low levels of residual damage at 24 h and 48 h post-irradiation. In contrast, SCID mice showed highly increased γ-H2AX foci levels at all repair times (0.5 h to 48 h) while A-T mice exhibited a lesser defect which was most significant at later repair times (≥ 5 h). Radiosensitive BALB/c mice exhibited slighdy elevated foei numbers compared with C57BI./6 mice at 5 h and 24 h but not at 48 h post-irradiation. Conclusion Quantifying the γ-H2AX loci in normal tissue represents a sensitive tool for the detection of induction and repair of radiation-induced DSBs at clinically relevant doses in vivo.

To investigate the effects of TGF-beta1 on the two gelatinases (MMP-2 and MMP-9), and their roles in lung remodeling after irradiation-induced lung injury. Expressions of TGF-beta1 were measured with western blot, and expressions of MMP-2 and MMP-9 were analyzed with zymography in a -TGF-beta1 transgenic mouse model after thoracic irradiation with 12 Gy. We found expressions of TGF-beta1 in the lung from the transgenic mice were three folds as compared to those from control mice. With densitometrical analysis, we found a significant decrease in MMP-9 activity in lung homogenates from the transgenic mice as compared with those from non-transgenic control mice 8 weeks after sham-irradiation (relative MMP-9 activity: C: 1.000 0.1091; TG: 0.4772 +/- 0.470 (n = 8, P < 0.05). But MMP-2 was constitutively expressed in the lung homogenates from the transgenic mice as compared to those from control mice 8 weeks after sham-irradiation (relative MMP-2 activity 8 weeks after sham-irradiation: C: 1.000 +/- 0.1556, TG: 1.0075 +/- 0.1472). Eight weeks after thoracic irradiation with 12 Gy, we observed a significant increase of MMP-2 and MMP-9 activity in lung homogenates from both transgenic and normal mice. In TGF-beta1 transgenic mice relative MMP-9 activity was increased to 1.5321 +/- 0.2217 folds 8 weeks after thoracic irradiation with 12 Gy as compared to those after sham-irradiation (1.000 +/- 0.2153), and relative MMP-2 activity was increased to 1.7142 +/- 0.4231 folds. Our results show that TGF-beta1 itself down-regulates activity of MMP-9, thereby decreases ECM degradation in lungs of TGF-beta1 transgenic mice. Also we find that ionizing irradiation upregulates both MMP-2 and MMP-9 activity. Over-expressions of MMP-9 and MMP-2 after lung irradiation are involved in the inflammatory response associated with radiation-induced lung injury, and maybe further in radiation-induced lung fibrosis.

To investigate the effects of TGF-β1 on the two gelatinases (MMP-2 and MMP-9), and their roles in lung remodeling after irradiation-induced lung injury. Expressions of TGF-β1 were measured with western blot, and expressions of MMP-2 and MMP-9 were analyzed with zymography in a TGF-β1 transgenic mouse model after thoracic irradiation with 12 Gy. We found expressions of TGF-β1 in the lung from the transgenic mice were three folds as compared to those from control mice. With densitometrical analysis, we found a significant decrease in MMP-9 activity in lung homogenates from the transgenic mice as compared with those from non-transgenic control mice 8 weeks after sham-irradiation (relative MMP-9 activity: C: 1.000±0.1091; TG: 0.4772± 0.470 (n=8, P＜0.05). But MMP-2 was constitutively expressed in the lung homogenates from the transgenic mice as compared to those from control mice 8 weeks aftersham-irradiation (relative MMP-2 activity 8 weeks after sham-irradiation: C: 1.000±0.1556, TG: 1.0075±0.1472). Eight weeks after thoracic irradiation with 12 Gy, we observed a significant increase of MMP-2 and MMP-9 activity in lung homogenates from both transgenic and normal mice. In TGF-β1 transgenic mice relative MMP-9 activity was increased to 1.5321±0. 2217 folds 8 weeks after thoracic irradiation with 12 Gy as compared to those after sham-irradiation (1.000±0.2153), and relative MMP-2 activity was increased to 1. 7142±0. 4231 folds. Our results show that TGF-β1 itself down-regulates activity of MMP-9, thereby decreases ECM degradation in lungs of TGF-β1 transgenic mice.Also we find that ionizing irradiation upregulates both MMP-2 and MMP-9 activity. Over-expressions of MMP-9 and MMP-2 after lung irradiation are involved in the inflammatory response associated with radiation-induced lung injury, and maybe further in radiation-induced lung fibrosis.