Ionizing Radiation Effect On Proliferation Of Human Umbilical Vein Endothelial Cells

Background and ObjectiveRadiotherapy had become more and more important in saving patient's life, improving the effect of treatment and quality of patient's life as a way of treatment for cancers. But the clinical application of radiotherapy was limited due to the reversible and irreversible damages of bones by ionizing radiation. Osteoradionecrosis of jaw may be resulted frequently and the wound would not heal sustainedly after the radiotherapy for the malignant tumors of the head and neck. However, the mechanism of osteoradionecrosis is not well known yet. So it is significant to explore the mechanism of radiation damages of bone for preventing and treating osteoradionecrosis and improving the quality of patient's life.It was indicated that the functional disorder of vascular is one of the major etiological factors in osteoradionecrosis. It was considered that vascular endothelial cell(VEC) was the target cell in injury of blood vessel. VEC is sensitive to ionizing radiation and its damage by ionizing radiation is the initiation to tissues' radiation damage, which plays a primary role in the occurrence, development and healing of radiation damage. Therefore, it is significant to study the biological changes of VEC after ionizing radiation. In this experiment, the characteristic and mechanism of VEC radiation damage were studied by observing the cell proliferation, the cytoskeleton, cell ultramicrostructure and the secretion of TGF-β1 of HUVE-12 after exposed to various dose of radiation, which would provide theoretical evidence for the mechanism, the treatment and prevention of osteoradionecrosis of jaws.Materials and MethodsHUVE-12 cultivated in vitro were radiated with 60Coγ_ray in different dose, ranging from 0-16Gy. The proliferation of HUVE-12 was detected with MTT Cell Proliferation Assay at 12hs, 24hs and 48hs after radiation. The ultramicrostructure of VEC at 48hs after radiation was observed with transmission electron microscope. The cytoskeleton was observed by Confocal Laser Scanning Microscope at 12hs and 24hs after the radiation. The cytoskeleton protein F-actin of VEC at 6hs,12hs,24hs after radiation and the TGF-β1 at 6hs, 12hs, 24hs and 48hs after radiation were detected with flow cytometry.ResultsHUVE-12, were well cultivated in vitro. The proliferation of VEC was inhibited by ionizing radiation in dose-dependent in the dose of 4-16Gy. Under the electron microscope the swelling or myelin sheath_liked mitochondria and the vacuolus of endocytoplasmic reticulum were found in cells, parts of condensed chromatin were found in cellular nucleus. The destruction of the cytoskeleton was observed with the Confocal Laser Scanning Microscope, and it was more severe as the radiation dose increasing. Detecting by the flow cytometry, as the increase of radiation dose the F-actin decreased and it was in the lowest level at 12hs after the radiation. However, it seemed to have some recovery 24hs later but was not in the normal level. The TGF-β1 increased depending on the dose increasing, evidently at 24hs after the radiation.Conclusion1. Ionizing radiation ofγ-ray may inhibite the proliferation of HUVE-12 with the dose-dependent in the dose of 4-16Gy. This may lower the ability of cells to repair the damage with apoptosis by proliferation.2. Ionizing radiation may make obviously damage to the HUVE-12. With the radiation dose rate increasing, the endocytoplasmic reticulum became vacuolus, the swelling or myelin sheath_liked mitochondria were seen in the cells, the size and appearance of cells changed. The cells lost the homogeneity and became loose. The appearance of some cellular nucleus became irregularity and chromatin in cellular nucleus condensed.3. The cytomembrane skeleton of HUVE-12 were destroyed by ionizing radiation. This effect may be more severe with the dose-time-dependent. The polymerization and depolymerization of F-actin, the protein of the cytoskeleton, may have a balance at 24hs after the radiation. This is probably the other damage pattern to VEC after ionizing radiation, related with the permeability of VEC increasing, ability of proliferation and migration restrained.4. HUVE-12 can secrete TGF-β1 after being ionizing radiated. High concentration of TGF-β1 can restrain HUVE—12 migration and angiogenesis. It indicated that the HUVE-12 growth rate degraded and apoptosis increased may be probably related with the inhibitory action of TGF-β1.