二甲双胍对造血免疫系统辐射损伤防护作用及分子机制研究
发布时间：2019-03-22 11:01
【摘要】：核事故受照人员以及临床上接受放射治疗的肿瘤患者,会出现各种辐射损伤,其中造血系统对辐射高度敏感,尤其是造血干细胞(HSCs)损伤会导致持久性骨髓抑制甚至个体死亡,目前尚无有效治疗手段,因此研发造血系统辐射损伤预防和治疗药物具有重要的社会意义和临床价值。我们前期研究发现氧化应激水平升高是介导亚致死剂量(6-6.5 Gy)受照小鼠造血干细胞衰老、骨髓长期抑制的主要机制。应用抗氧化制剂具有一定的治疗作用。二甲双胍是广泛应用的Ⅱ型糖尿病治疗药物,近年来研究发现有抗氧化和抗衰老的作用,具有防止糖尿病心血管并发症及肿瘤预防作用。本项研究通过体外、体内照射模型,探讨二甲双胍对辐射引起的造血免疫损伤的防护作用,尤其是在接受4 Gy全身照射(TBI)小鼠HSCs功能损伤的保护作用,并探讨其作用机制。为观察二甲双胍对造血免疫细胞辐射损伤的保护作用,分离C57BL/6小鼠骨髓、胸腺及脾脏细胞,加入不同浓度二甲双胍孵育30 min后接受不同剂量γ射线照射,孵育过夜后进行细胞活力测定,结果显示二甲双胍对照射引起的造血免疫细胞损伤具有一定的保护作用。为了研究二甲双胍对TBI小鼠造血免疫系统辐射损伤的保护作用,我们将C57BL/6小鼠分为对照组、照射组和照射给药组,在30d生存率实验中,二甲双胍能够提高接受致死剂量(7.2 Gy)TBI小鼠的生存率。利用4 Gy全身照射模型,探讨了二甲双胍不同剂量、不同给药时间对骨髓细胞辐射损伤的保护作用,确定小鼠受照前24 h灌胃给药250mg/kg剂量,受照后持续给药7 d,能够提高受照小鼠外周血细胞计数、骨髓有核细胞计数和脾结节计数；CFU-GM和CAFC实验表明4 Gy照射可以引起造血祖细胞和造血干细胞功能损伤,而二甲双胍能够提高受照小鼠造血祖细胞和造血干细胞的增殖能力,骨髓竞争移植实验表明,二甲双胍可以提高受照小鼠造血干细胞长期再植能力。二甲双胍辐射防护机制研究发现,二甲双胍能够降低受照小鼠造血细胞中活性氧(ROS)水平,qRT-PCR实验表明二甲双胍可以降低受照小鼠造血细胞中NOX4mRNA表达水平,这与二甲双胍能够降低受照小鼠造血细胞中ROS水平有关。对造血细胞的p16mRNA表达和凋亡水平的研究表明,二甲双胍可以降低受照小鼠造血细胞中p16mRNA表达水平、不影响细胞的凋亡率,提示二甲双胍对造血系统辐射损伤的防护作用与改善辐射诱导造血细胞的衰老有关,而与凋亡途径无关。全基因组表达谱芯片分析结果显示,接受4 Gy照射小鼠HSCs中mTOR、S6K、S6基因的表达均上调,说明照射可能激活了HSCs中mTOR信号通路,而给药后能够抑制这些基因表达的上调,提示二甲双胍可能能够抑制受照小鼠HSCs中mTOR通路的活性。总之,我们首次系统地研究了二甲双胍对造血免疫系统辐射损伤的防护作用,初步阐明了二甲双胍对4 Gy受照小鼠造血干细胞的损伤有保护作用,阐明了这一保护作用与二甲双胍能够降低造血干细胞氧化应激水平和衰老有关,降低造血干细胞氧化应激水平与二甲双胍调节NOX4mRNA表达水平有关。也有可能与二甲双胍对mTOR通路调节作用有关。我们的研究提示二甲双胍有可能成为临床上造血系统辐射损伤防护药,对改善肿瘤放疗患者预后和提高意外辐射事故受害人员生存率都具有潜在的应有价值。
[Abstract]:In patients with a nuclear accident and a tumor with a clinically acceptable radiation treatment, various radiation damage may occur, in which the hematopoietic system is highly sensitive to radiation, in particular hematopoietic stem cells (HSCs), which may lead to persistent bone marrow suppression, or even death of the individual, and there is no effective means of treatment at this time, Therefore, the research and development of the radiation damage prevention and treatment medicine of the hematopoietic system has important social significance and clinical value. We found that the level of oxidative stress in the early stage of our study was the main mechanism of mediating sublethal dose (6-6.5 Gy) on the aging of hematopoietic stem cells and long-term inhibition of bone marrow. The antioxidant preparation has certain therapeutic effect. Dimethyldicyandiamide is widely used in the treatment of type II diabetes. In recent years, the effects of anti-oxidation and anti-aging have been found, and it has the function of preventing the cardiovascular complications of diabetes and the prevention of tumor. In this study, by in vitro and in vivo irradiation model, the protective effect of the dimethoxy-2 on the hematopoietic immune injury induced by radiation, in particular the protective effect of the function injury of the HSCs in the mice receiving the whole body irradiation (TBI) of 4 Gy, was discussed, and its mechanism of action was discussed. In order to observe the protective effect of the dimethoxy dicyandiamide on the radiation injury of the hematopoietic immune cells, the bone marrow, thymus and spleen cells of the C57BL/6 mice were isolated, and the cells of the bone marrow, the thymus and the spleen of the C57BL/6 mice were isolated and incubated for 30 min with different concentrations of dimethyldiisopropyl alcohol and then irradiated with different doses of X-ray, and the cell viability was measured after the incubation overnight. The results showed that the dimethoxy-2-2 was a protective effect on the damage of the hematopoietic immune cells caused by the irradiation. In order to study the protective effect of the dimethoxy dicyandiamide on the radiation damage of the hematopoietic immune system in the TBI mice, the C57BL/6 mice were divided into the control group, the irradiation group and the irradiation group. In the 30-day survival experiment, the survival rate of the TBI mice receiving the lethal dose (7.2Gy) was increased. The protective effects of different doses and different administration time on the radiation injury of bone marrow cells were discussed by using a 4 Gy whole-body irradiation model. The mice were given a dose of 250 mg/ kg for 24 h before the irradiation, and 7 days after the irradiation, the peripheral blood cell count of the irradiated mice can be improved. Bone marrow nucleated cell count and spleen nodule count; CFU-GM and CAFC experiment showed that 4 Gy irradiation could cause functional damage of hematopoietic progenitor cells and hematopoietic stem cells, and the proliferation ability of hematopoietic progenitor cells and hematopoietic stem cells in irradiated mice can be improved, and the bone marrow competition transplantation experiment shows that, The ability of the long-term replantation of the hematopoietic stem cells of the irradiated mice can be improved. The results of the study on the radiation protection mechanism of dimethoxy-2-(2-(2-)-2-(2-(2-)-2-(2-(2-)-2-(2-(2-)-2-(2-)-(2-(2-)-(2-)-(2-)-(2-)-(2-)-(2-)-(2-(2-)- This is related to the reduction of the level of ROS in the hematopoietic cells of the irradiated mice. The study of the expression of p16mRNA and the level of apoptosis in the hematopoietic cells showed that the expression of p16mRNA in the hematopoietic cells of the irradiated mice can be reduced, and the apoptosis rate of the cells is not affected, and the protective effect of the dimethoxy dicyandiamide on the radiation damage of the hematopoietic system is related to the improvement of the aging of the radiation-induced hematopoietic cells. Regardless of the pathway of apoptosis. The results of the whole genome analysis showed that the expression of mTOR, S6K and S6 in the HSCs irradiated by 4 Gy was up-regulated, indicating that the irradiation could activate the mTOR signaling pathway in the HSCs, and the upregulation of the expression of these genes can be inhibited after administration. It is suggested that the dicyandiamide may be able to inhibit the activity of the mTOR pathway in the HSCs of the irradiated mice. In conclusion, we first systematically studied the protective effect of the dimethoxy dicyandiamide on the radiation damage of the hematopoietic immune system. This protective effect is related to the reduction of the oxidative stress level and the aging of the hematopoietic stem cells, and the level of the oxidative stress of the hematopoietic stem cells is related to the level of the NOx4mRNA expression regulated by the dimethoxy-2-1. It is also possible to be related to the regulation of the mTOR pathway with the di-dimethoxy. Our study suggests that the disidiolone has the potential to be a protective agent for radiation damage in the clinical hemopoietic system, and it is of potential value to improve the prognosis of patients with tumor radiotherapy and to improve the survival rate of patients with accidental radiation accident.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2013
【分类号】：R818
本文编号：2445532
[Abstract]:In patients with a nuclear accident and a tumor with a clinically acceptable radiation treatment, various radiation damage may occur, in which the hematopoietic system is highly sensitive to radiation, in particular hematopoietic stem cells (HSCs), which may lead to persistent bone marrow suppression, or even death of the individual, and there is no effective means of treatment at this time, Therefore, the research and development of the radiation damage prevention and treatment medicine of the hematopoietic system has important social significance and clinical value. We found that the level of oxidative stress in the early stage of our study was the main mechanism of mediating sublethal dose (6-6.5 Gy) on the aging of hematopoietic stem cells and long-term inhibition of bone marrow. The antioxidant preparation has certain therapeutic effect. Dimethyldicyandiamide is widely used in the treatment of type II diabetes. In recent years, the effects of anti-oxidation and anti-aging have been found, and it has the function of preventing the cardiovascular complications of diabetes and the prevention of tumor. In this study, by in vitro and in vivo irradiation model, the protective effect of the dimethoxy-2 on the hematopoietic immune injury induced by radiation, in particular the protective effect of the function injury of the HSCs in the mice receiving the whole body irradiation (TBI) of 4 Gy, was discussed, and its mechanism of action was discussed. In order to observe the protective effect of the dimethoxy dicyandiamide on the radiation injury of the hematopoietic immune cells, the bone marrow, thymus and spleen cells of the C57BL/6 mice were isolated, and the cells of the bone marrow, the thymus and the spleen of the C57BL/6 mice were isolated and incubated for 30 min with different concentrations of dimethyldiisopropyl alcohol and then irradiated with different doses of X-ray, and the cell viability was measured after the incubation overnight. The results showed that the dimethoxy-2-2 was a protective effect on the damage of the hematopoietic immune cells caused by the irradiation. In order to study the protective effect of the dimethoxy dicyandiamide on the radiation damage of the hematopoietic immune system in the TBI mice, the C57BL/6 mice were divided into the control group, the irradiation group and the irradiation group. In the 30-day survival experiment, the survival rate of the TBI mice receiving the lethal dose (7.2Gy) was increased. The protective effects of different doses and different administration time on the radiation injury of bone marrow cells were discussed by using a 4 Gy whole-body irradiation model. The mice were given a dose of 250 mg/ kg for 24 h before the irradiation, and 7 days after the irradiation, the peripheral blood cell count of the irradiated mice can be improved. Bone marrow nucleated cell count and spleen nodule count; CFU-GM and CAFC experiment showed that 4 Gy irradiation could cause functional damage of hematopoietic progenitor cells and hematopoietic stem cells, and the proliferation ability of hematopoietic progenitor cells and hematopoietic stem cells in irradiated mice can be improved, and the bone marrow competition transplantation experiment shows that, The ability of the long-term replantation of the hematopoietic stem cells of the irradiated mice can be improved. The results of the study on the radiation protection mechanism of dimethoxy-2-(2-(2-)-2-(2-(2-)-2-(2-(2-)-2-(2-(2-)-2-(2-)-(2-(2-)-(2-)-(2-)-(2-)-(2-)-(2-)-(2-(2-)- This is related to the reduction of the level of ROS in the hematopoietic cells of the irradiated mice. The study of the expression of p16mRNA and the level of apoptosis in the hematopoietic cells showed that the expression of p16mRNA in the hematopoietic cells of the irradiated mice can be reduced, and the apoptosis rate of the cells is not affected, and the protective effect of the dimethoxy dicyandiamide on the radiation damage of the hematopoietic system is related to the improvement of the aging of the radiation-induced hematopoietic cells. Regardless of the pathway of apoptosis. The results of the whole genome analysis showed that the expression of mTOR, S6K and S6 in the HSCs irradiated by 4 Gy was up-regulated, indicating that the irradiation could activate the mTOR signaling pathway in the HSCs, and the upregulation of the expression of these genes can be inhibited after administration. It is suggested that the dicyandiamide may be able to inhibit the activity of the mTOR pathway in the HSCs of the irradiated mice. In conclusion, we first systematically studied the protective effect of the dimethoxy dicyandiamide on the radiation damage of the hematopoietic immune system. This protective effect is related to the reduction of the oxidative stress level and the aging of the hematopoietic stem cells, and the level of the oxidative stress of the hematopoietic stem cells is related to the level of the NOx4mRNA expression regulated by the dimethoxy-2-1. It is also possible to be related to the regulation of the mTOR pathway with the di-dimethoxy. Our study suggests that the disidiolone has the potential to be a protective agent for radiation damage in the clinical hemopoietic system, and it is of potential value to improve the prognosis of patients with tumor radiotherapy and to improve the survival rate of patients with accidental radiation accident.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2013
【分类号】：R818
【参考文献】
相关期刊论文 前1条
1 董小英;张如意;;二甲双胍对糖尿病大鼠心肌病变的抗氧化保护作用[J];宁夏医学院学报;2008年02期
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