Influence Of Hypoxic Preconditioning On Cell Apoptosis Induced By Radiation And Its Possible Mechanisms

With the development of medical care and the extensive use of nuclear technology, people are more likely to be irradiated. At the same time, the possibility of radiation damage caused by medical irradiation or nonmedical irradiation is also increasing. Radiation damage usually results in gene mutation, cell apoptosis, tissue degeneration and necrosis, organ dysfunction and immunological malfunction. Some severe damage even leads to multiple organ failure and radiation damage syndrome. Hence, study of radiation damage and its prevention and care has arrested much attention from many scholars. Recently, how to induce endogenous protective reaction against radiation damage has been a new subject in the field of prevention and care of radiation damage. As a ubiquitous endogenous protective phenomenon, preconditioning has common internal mechanism as well as cross protection among different environment factor. Hypoxia preconditioning can easily be performed and does have authentic protection. In order to prove that hypoxia preconditioning could induce cross protection against radiation damage and reveal its possible mechanism, especially to discuss the protective function of hypoxia-inducible factor-1, an important nuclear transcription factor with tight relation to hypoxia, against radiation damage preconditioned by hypoxia, and provide new clues to prevention and care of radiation damage, we designed this subject and investigated the expression of HIF-1, Bcl-2/Bax and their protective function against cell apoptosis induced by radiation. In this study, we treated the radiosensitive IEC-6 cell line, which derived from rat jejunum epithelium cells, with the condition of 37℃, 5%CO2 and 95%N2 mixture for 2h and reoxygen for 24h as hypoxia preconditioning model. After irradiated by 35Gy 60Coγray, we detected the cell apoptosis rate and proliferation, and explored the protective mechanism of hypoxia preconditioning with Western Blot and RT-PCR technique. The main results of the study are as follows: Compared with the simple 35Gy 60Coγray irradiation group, the apoptosis rate and micronucleus frequency of cells with the 35Gy 60Coγray plus hypoxic predonditioning separately decreased from 20.16±0.96%, 85±5.16‰ to 13.65±1.17%,61±4.32‰,while survival rate increased from 52.36±3.51% to 67.58±2.72%,and the morphological damage of cells was apparently relieved .Compared with control group, the oxygen concentration and free radical production had no notable change 24h after hypoxic preconditioning, which exclude the direct influence of oxygen effect and free radical changes induced by hypoxia on radiation. 24h after HPC, SOD activity increased significantly, which relieved free radicals' injury of radiation. Hypoxia preconditioning remarkably up-regulated the expression of HIF-lα protein. The protective function of HPC was attenuated when cells were treated with PI-3K selective inhibitor LY294002 to inhibit HIF-lα protein expression. The same result was observed when HIF-lα selective antagonist was used.Hypoxia preconditioning could up-regulate the expression of Bcl-2 mRNA and protein during 24h to 48h after HPC, which was consistent with the occurrenc of the second window of protection. LY294002 and HIF-lαantagonist markedly attenuated the expression of Bcl-2 protein, while hypoxia preconditioning had no notable influence on Bax expression.Based on all the results ,we concluded that:Cell apoptosis decreased and its survival rate increased under the condition of 35Gyγradiation 24h after HPC for 2h, showing that HPC had the cross-protective function against 35Gyγradiation, and this effect excluded the direct influence of oxygen effect and free radical changes induced by hypoxia on radiation. SOD activity accumulation induced by HPC might be included in delayed protection. HIF-1 might play an important role in delayed protection of HPC, and this effect was realized partially by indirect up-regulation of Bcl-2.