基于γ-H2AX免疫荧光显微镜观察CT检查后外周血淋巴细胞内DNA双链断裂的研究

发布时间：2018-05-30 22:29

  本文选题：CT + DNA双链断裂　； 参考：《南京大学》2017年硕士论文

【摘要】：第一部分CT检查对外周血淋巴细胞DNA双链断裂影响的研究目的:评价腹部CT增强检查对外周血淋巴细胞内DNA双链断裂(double strand breaks,DSBs)水平的影响。材料与方法:选取12例健康志愿者,各抽取8 ml静脉血,将每例志愿者的静脉血分为A组、B组、C组和D组,每组2 ml血液,进行体外实验;其中A组外周血作为对照组,不接受CT扫描;B-D组外周血接受相当于1、2、3次腹部CT辐射剂量的扫描。选取30例腹部增强CT检查的患者进行体内实验。在CT检查前(基线组)及检查后5 min(辐射组)分别抽取2ml静脉血,并记录剂量长度乘积(dose-length product,DLP)和 CT 容积剂量指数(volume CT dose index,CTDIvol)。所有血样通过淋巴细胞分离及γ-H2AX蛋白的免疫荧光染色,标记出DSBs。通过重复测量方差分析比较A、B、C、D组间DSBs水平的关系。通过配对样本t检验比较基线组和辐射组间DSBs水平的关系。采用Pearson相关分析分析体内实验组辐射剂量与γ-H2AX焦点增加的关系。结果:在体外试验中,接受CT辐射暴露的B、C、D组血样中DSBs水平均较A组明显升高,升高幅度分别为49.4%、96.6%和149.4%(所有P0.001),且辐射剂量分组与DSBs水平存在线性趋势(P0.001)。在体内实验中,CT检查前γ-H2AX焦点的平均值及标准差为0.62±0.35焦点/细胞,检查后为0.89±0.42焦点/细胞,辐射组血样中淋巴细胞内DSBs水平较基线组DSBs水平升高约43.5%(P0.001);但CT检查后DSBs的增加量与DLP或CTDIvol未发现明显相关性。结论:外周血淋巴细胞内DSBs水平在进行腹部CT增强检查后明显升高,体外研究表明CT检查辐射剂量与细胞内DSBs水平存在线性趋势。第二部分口服维生素C预防CT检查后DNA双链断裂的研究目的:研究口服维生素C对腹部对比增强CT检查后患者外周血中DNA双链断裂(double strand breaks,DSBs)损伤的预防作用。材料与方法:本研究共入组60例腹部对比增强CT检查患者,分为对照组(30例)及预防组(30例)。预防组在CT检查前30～120 min 口服1 g维生素C,对照组不使用任何药物。预防组根据口服维生素C时间点的不同分为3组(30 min,n=16;60min,n=10;120min,n=4)。所有患者在CT检查前及检查后5 min抽取2 ml外周血。将所有血样分离出淋巴细胞,进行免疫荧光染色,标记出细胞核内的代表DSBs的y-H2AX荧光焦点。两组间的计数资料(包括性别和合并症)的组间比较使用卡方检验。两组间计量资料,包括剂量长度乘积(dose-length product,DLP)、CT容积剂量指数(volume CT dose index,CTDIvol)、y-H2AX焦点基线水平间的比较使用t检验及u检验。不同时间点口服维生素C的预防组间γ-H2AX焦点增加的差异性使用ANOVA检验。辐射剂量与γ-H2AX焦点增加的关系使用Pearson相关及Spearman相关分析。结果:对照组和预防组两组间的CTDIvol及DLP没有统计学差异(P值均0.05)。对照组检查后γ-H2AX焦点较检查前平均增加0.49个/细胞,预防组检查后平均增加0.19个/细胞,检查后预防组较对照组y-H2AX焦点增加量显著减少(p0.001),减少量占对照组增加量的61%。在预防组中,γ-H2AX焦点增加量分别与DLP及CTDIvol呈显著相关(r值分别为0.449和0.403,P值均0.05)。不同时间点口服维生素C的预防组间γ-H2AX焦点增加没有表现出差异性(所有P0.05)。结论:检查前口服维生素C能够显著减少腹部对比增强CT检查后的DSBs水平,对CT所致的DNA损伤起到一定保护作用。
[Abstract]:The first part of CT examined the effect of DNA double strand breaks on peripheral blood lymphocytes. Objective: To evaluate the effect of abdominal CT enhancement on the level of DNA double strand breaks (double strand breaks, DSBs) in peripheral blood lymphocytes. Materials and methods: 12 healthy volunteers were selected to take 8 ml venous blood each, and the venous blood of each volunteer was divided into A group. In group B, group C and group D, each group of 2 ml blood was tested in vitro, and the peripheral blood of group A was taken as the control group and no CT scan was accepted; the peripheral blood of the group B-D received the equivalent of the CT radiation dose in the abdomen of 1,2,3 times. The patients of the abdominal enhanced CT examination were selected in the body experiment. Before CT examination (baseline group) and after examination 5 min (radiation groups) were extracted 2, respectively. Ml venous blood was recorded and the dose length product (dose-length product, DLP) and CT volume dose index (volume CT dose index, CTDIvol) were recorded. All blood samples were stained by lymphocyte separation and immunofluorescence staining of gamma -H2AX protein. The relationship between the baseline group and the DSBs level between the radiation groups was compared. The relationship between the radiation dose of the experimental group and the increase of the focus of gamma -H2AX was analyzed by Pearson correlation analysis. Results: in the in vitro test, the levels of DSBs in the B, C and D groups receiving CT radiation exposure were significantly higher than those in the A group, and the elevation was 49.4%, 96.6% and 149.4% respectively (all P0.001), respectively. And there was a linear trend (P0.001) in the radiation dose group and the DSBs level. In the experiment in vivo, the average value and standard deviation of the focus of gamma -H2AX before CT examination were 0.62 + 0.35 focus / cell, and 0.89 + 0.42 focus / cell after examination. The level of DSBs in the lymphocyte of radiation group increased by 43.5% (P0.001) than that in the baseline group (P0.001); but DSBs after CT examination, DSBs There was no significant correlation between the increase of DLP or CTDIvol. Conclusion: the level of DSBs in peripheral blood lymphocytes increased significantly after abdominal CT enhanced examination. In vitro studies showed that there was a linear trend in the radiation dose of CT and the level of intracellular DSBs. The second part of the study was to study the aim of the study of DNA double strand breaks after the oral vitamin C pre control of CT. Study the preventive effect of oral vitamin C on DNA double strand breaks (double strand breaks, DSBs) injury in peripheral blood of patients after abdominal contrast enhanced CT examination. Materials and methods: 60 cases of abdominal contrast enhanced CT examination were included in this study. The control group was divided into control group (30 cases) and prevention group (30 cases). The prevention group had 1 g 30~120 min before CT examination. The prevention group was divided into 3 groups (30 min, n=16; 60min, n=10; 120min, n=4) according to the time points of oral vitamin C. All the patients were extracted with 2 ml peripheral blood before and after the examination of CT. All the blood samples were separated from the lymphocytes, and the immunofluorescence staining was carried out to mark out the DSBs in the nucleus. Fluorescent focus. The count data between groups of two groups (including sex and complication) were compared with the chi square test. Two groups of measurement data, including the dose length product (dose-length product, DLP), the CT volume dose index (volume CT dose index, CTDIvol), and the comparison between the y-H2AX focus baseline level using t test and U test. The differential use of ANOVA test for the increase of gamma -H2AX focus between the prevention groups of vitamin C was used. The relationship between the radiation dose and the increase of gamma -H2AX focus was associated with Pearson and Spearman related analysis. Results: there was no statistical difference between the two groups of the control group and the prevention group, CTDIvol and DLP (P is 0.05). The focus of gamma -H2AX after the control group was compared to the check before the check. The increase of 0.49 / cell was increased by 0.49 / 0.19 cells, and the increase in the focus of the prevention group was significantly reduced (p0.001), and the decrease in the amount of 61%. in the control group was significantly correlated with DLP and CTDIvol (R values were 0.449 and 0.403, P values were 0.05 respectively). The increase of gamma -H2AX focus between the prophylactic group of oral vitamin C at the same time did not show a difference (all P0.05). Conclusion: oral vitamin C before examination can significantly reduce the level of DSBs after abdominal contrast-enhanced CT examination, and can play a protective role in DNA damage caused by CT.
【学位授予单位】：南京大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R816
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本文编号：1957101