Effect of radiation and repeated sub-culturing on the transforming growth factor-ß1 signaling pathway in FRTL-5 cells.

BACKGROUND/AIM: Fisher rat thyroid cells (FRTL-5) display increased proliferation, reduced follicularization and decreased thyroxin release with repeated sub-culturing. These changes occur earlier and more rapidly following exposure to ionizing radiation. We hypothesized that altered transforming growth factor-ß1 (TGF-ß1) signaling contributes to these differences. MATERIALS AND METHODS: Assessments included FRTL-5 cell growth rate and quantification of TGF-ß1 ligand and receptors. The levels and activity of Smads2, 3 and 4 were measured by western blotting and the ability of TGF-ß1 to regulate cyclin A and plasminogen activator inhibitor type 1 (PAI-1) activity was assessed using transfection assays. RESULTS: TGF-ß1 production increased after radiation but returned to control levels after repeated sub-culturing. There was no difference in TGF-ß1 levels between un-irradiated cells at low versus high-passage number. TGF-ß1 receptors and basal levels of Smads2, 3 and 4 remained unchanged. However, there were significant changes in cell proliferation, TGF-ß1-mediated Smads2 and 3 activation and in TGF-ß1's ability to regulate cyclin A and PAI-1 transcription in irradiated and repeatedly sub-cultured cells (p<0.05). CONCLUSION: Collectively, these results support the conclusion that alterations in the TGF-ß1 pathway contribute to phenotypic changes in FRTL-5 cells as a function of passage number and radiation.

Alterations in glutamate uptake in NT2-derived neurons and astrocytes after exposure to gamma radiation.

Currently, the cellular and molecular mechanisms that underlie radiation-induced damage in the CNS are unclear. The present study began investigations of the underlying mechanism(s) for radiation-induced neurotoxicity by characterizing glutamate transport expression and function in neurons and astrocytes after exposure to gamma rays. NTera2-derived neurons and astrocytes, isolated as pure cultures, were exposed to doses of 10 cGy, 50 cGy and 2 Gy gamma rays, and transporter expression and function were assessed 3 h, 2 days and 7 days after exposure. In neurons, at 7 days after exposure, a significant increase was detected in EAAT3 after 50 cGy (P < 0.05) and a dose-dependent increase in GLT-1 expression was seen between doses of 10 and 50 cGy (P < 0.05). Functional assays of glutamate uptake revealed that neurons and astrocytes respond in a reciprocal manner after irradiation. Neurons responded to radiation exposure by increased glutamate uptake, an effect still evident at our last time (7 days) after exposure (P < 0.05). The astrocyte response to gamma radiation was an initial decrease in uptake followed by recovery to baseline levels at 2 days after exposure (P < 0.05). The observations made in this study demonstrate that neurons and astrocytes, while part of the same multifunctional unit, have distinct functional and reciprocal responses. The response in neurons appears to indicate a protracted response with potential long-term effects after irradiation.