肠型放射病小鼠模型中小肠细胞死亡类型的鉴定和修饰性p53蛋白的表达研究

发布时间：2019-02-25 13:50

【摘要】：目的：放射治疗是肿瘤治疗的三大主要手段之一，其中70%的恶性肿瘤可以进行放射治疗。在许多腹腔恶性肿瘤放疗过程中病人会出现肠道症状，如腹泻、消化不良等，这是放疗对肠道正常细胞的损伤引起的副作用。这种副作用的存在极大地影响了放疗的进行和病人的康复。再者，目前没有有效的药物救治核泄漏事故中的肠型放射病。通过对照射后肠道细胞死亡形式及其机制的研究，寻找相应的救治措施，提高对正常组织的保护，以期提高放射治疗的疗效和救治肠型放射病。 方法：小鼠接受15Gy全身照射后，观察记录小鼠的生存，，绘制生存曲线；在照后指定的时间点处死小鼠，取出回肠，观察结肠粪便形成、HE染色以及透射电镜等形态学指标；免疫荧光法分析活性氧（ROS）的水平；ELISA法测定谷胱甘肽（GSH）、一氧化氮（NO）、过氧化氢（H2O2）以及超氧化物歧化酶（SOD）的水平；免疫印迹法分析Caspase-3，LC3B以及p53的磷酸化、乙酰化水平；免疫荧光法分析HMGB1；酶联免疫法测定血清中的IL-6水平。 结果：照后小鼠回肠中活性氧明显增加，而氧化指标（GSH, NO, H2O2, SOD）却没有明显改变。通过形态学以及生化指标反映：在15Gy全身照射后的小鼠回肠中均显示出凋亡、坏死以及自噬的特征。剪切的Caspase-3（凋亡标志物）的出现，LC3-Ⅰ向LC3-Ⅱ（自噬标志物）的转换，血清IL-6的分泌以及HMGB1（坏死标志物）的表达。此外，在15Gy照后出现p53基因379位赖氨酸的乙酰化以及第6、20、392位的丝氨酸的磷酸化，这种现象在照后2小时后消失。 结论：在照射后的肠道细胞的死亡不应被视为单一的死亡形式，而应该是一种“混合型死亡”。放疗中的同步治疗药物如能促进p53基因379位赖氨酸的乙酰化以及第6、20、392位的丝氨酸的磷酸化，则可能降低放疗后肠道细胞的死亡。
[Abstract]:Objective: radiotherapy is one of the three main methods of tumor therapy, 70% of which can be treated by radiotherapy. Intestinal symptoms such as diarrhea, dyspepsia and so on will occur in many patients with abdominal cancer during radiotherapy, which is a side effect of radiation therapy on normal intestinal cells. The presence of this side effect has greatly affected radiotherapy and patient recovery. Furthermore, there are no effective drugs available for the treatment of intestinal radiation sickness in the event of nuclear leakage. Through the study of the death form and mechanism of intestinal cells after irradiation, the corresponding treatment measures were found to improve the protection of normal tissues, so as to improve the curative effect of radiation therapy and the treatment of intestinal radiation sickness. Methods: after the mice were irradiated with 15Gy, the survival of the mice was observed, the survival curve was drawn, the mice were killed at the designated time point after irradiation, the ileum was taken out, and the colonic feces formation, HE staining and transmission electron microscopy were observed. The levels of reactive oxygen species (ROS), glutathione (GSH), (GSH), nitric oxide (NO), hydrogen peroxide (H2O2) and superoxide dismutase (SOD) were determined by immunofluorescence assay. The levels of phosphorylation and acetylation of Caspase-3,LC3B and p53 were detected by immunoblotting assay, and the levels of IL-6 in serum were determined by immunofluorescence assay for HMGB1; enzyme linked immunosorbent assay. Results: after irradiation, the active oxygen species (Ros) in the ileum increased significantly, but the oxidation index (GSH, NO, H2O2 (SOD) did not change significantly. Morphological and biochemical results showed that apoptosis, necrosis and autophagy were observed in the ileum of mice irradiated with 15Gy. The appearance of shearing Caspase-3 (apoptotic marker), the conversion of LC3- 鈪

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