新型促排剂WSC-DTPA纳米粒对内污染铀促排和防护作用的研究

发布时间：2018-05-05 21:55

本文选题：铀 + WSC-DTPA纳米粒　；参考：《苏州大学》2013年硕士论文

【摘要】：目的：通过细胞和动物实验研究水溶性低分子量壳聚糖-二乙烯三氨五乙酸（WSC-DTPA）纳米粒对硝酸铀酰染毒的HK-2细胞和SD大鼠促排效果和防护作用。方法：根据实验室已建立的方法制备WSC-DTPA聚合物及其纳米粒，再利用电镜和粒径仪检测纳米粒大小。细胞实验：采用MTT法检测WSC-DTPA纳米粒对硝酸铀酰染毒的正常肾近曲小管上皮细胞(HK-2)存活率的影响；流式细胞仪PI单染法检测WSC-DTPA纳米粒作用硝酸铀酰染毒的HK-2细胞的细胞周期分布份额变化；流式细胞仪荧光法检测WSC-DTPA纳米粒作用硝酸铀酰染毒的HK-2细胞内的ROS含量；ICP-AES检测经过WSC-DTPA纳米粒作用硝酸铀酰染毒后的HK-2细胞内铀含量。动物实验：采用腹腔注射硝酸铀酰（0.5mgU/kg）方式对SD大鼠进行染毒，即刻尾静脉注射92.7%WSC-DTPA纳米粒、1.59%WSC-DTPA纳米粒、WSC纳米粒及促排灵（DTPA-Na_3Ca），给药剂量均为60mg/kg。ICP-AES测量SD大鼠第1天、第2天、第3天的尿和粪中铀的排出量，以及给药3天后肝、脾、肾和骨中铀蓄积量；采用腹腔注射硝酸铀酰（2.0mgU/kg）方式对SD大鼠进行染毒，相同方式处理，5天后，取肝、肾、脾做病理切片，骨髓作骨髓细胞涂片，查看病理损伤程度。评价WSC-DTPA纳米粒的对内污染铀的促排和辐射防护效果。结果：制备出的WSC-DTPA纳米粒经检测，符合实验要求。细胞实验，MTT法结果表明，铀浓度在12.5μg/mL左右HK-2细胞的存活率接近50%。MTT实验发现DTPA-CaNa_3和1.59%WSC-DTPA纳米粒不能提高12.5μg/mL铀染毒HK-2细胞的存活率，而92.7%WSC-DTPA纳米粒和WSC纳米粒能明显提高细胞的存活率（P㩳0.05），并且存活率随着给药浓度的增加而提高。流式细胞PI单染法结果表明，12.5μg/mL铀染毒细胞出现G0/G1期阻滞，该期的DNA含量显著增高（P㩳0.05），而S期含量明显减少，G2/M期的含量大致不变，维持在15%左右；DTPA-CaNa_3和1.59%WSC-DTPA纳米粒不能改善铀染毒造成的G0/G1期阻滞现象，而92.7%WSC-DTPA纳米粒和WSC纳米粒能显著地缓减G0/G1期阻滞。细胞促排实验结果表明，与单纯染毒（3.13μg/mL）组相比， DTPA-CaNa_3、1.59%WSC-DTPA纳米粒、92.7%WSC-DTPA纳米粒和WSC纳米粒均有促排效果（P㩳0.05），，其中1.59%WSC-DTPA纳米粒的促排效果最佳，不同时间给药实验发现立刻给药的促排效果相对较好。细胞内ROS含量实验结果表明，铀染毒可造成细胞内的ROS含量明显增加（P㩳0.05），92.7%WSC-DTPA纳米粒和WSC纳米粒能有效地减少细胞内的ROS含量（P㩳0.05），而1.59%WSC-DTPA纳米粒和DTPA-CaNa_3不能减少细胞内的ROS含量。可见92.7%WSC-DTPA纳米粒和WSC纳米粒具有抗辐射效果。动物实验结果显示，在0.5μg/ml铀染毒实验中，92.7%WSC-DTPA纳米粒、1.59%WSC-DTPA和DTPA-CaNa_3均有促排效果，且1.59%WSC-DTPA纳米粒促排效果明显优于DTPA-CaNa_3组，统计学上有显著性差异（P㩳0.05）；给药组的主要组织铀蓄积量明显低于单纯染毒组，有显著性统计学差异（P㩳0.05）。在2.0μg/ml铀染毒实验中，92.7%WSC-DTPA纳米粒和WSC纳米粒的主要组织铀蓄积量明显低于DTPA-CaNa_3和1.59%WSC-DTPA纳米粒，统计学上有显著性差异（P㩳0.05）；病理组织切片显示92.7%WSC-DTPA纳米粒和WSC纳米粒对肝、肾有保护作用，损伤明显要比其他药物组轻。可见在较高剂量铀染毒的情况下对组织的防护作用相对比较重要的。骨髓图片的细胞分类未见有明显异常。结论：WSC-DTPA纳米粒对内污染铀有较好促排效果，其中1.59%WSC-DTPA纳米粒的促排效果最好；92.7%WSC-DTPA纳米粒不仅有促排作用，而且对铀染毒的大鼠的肝、肾组织有辐射防护作用。
[Abstract]:Objective: To study the effect and protective effect of water-soluble low molecular weight chitosan, two ethylene three ammonia five acetic acid (WSC-DTPA) nanoparticles, on HK-2 cells and SD rats infected with uranyl nitrate.
Methods: the WSC-DTPA polymer and its nanoparticles were prepared according to the method established in the laboratory. The size of the nanoparticles was detected by electron microscopy and particle size analyzer. The effect of WSC-DTPA nanoparticles on the survival rate of the normal renal proximal tubular epithelial cells (HK-2) was detected by the MTT method. The flow cytometry PI single staining method was used to detect WSC. The changes in the cell cycle distribution of the HK-2 cells infected with -DTPA nanoparticles; the flow cytometer fluorescence assay was used to detect the ROS content in HK-2 cells infected with WSC-DTPA nanoparticles; ICP-AES was used to detect the content of uranium in HK-2 cells infected by WSC-DTPA nanoparticles in the presence of uranyl nitrate. Animal experiments: SD rats were poisoned by intraperitoneal injection of uranyl nitrate (0.5mgU/kg). 92.7%WSC-DTPA nanoparticles, 1.59%WSC-DTPA nanoparticles, WSC nanoparticles and DTPA-Na_3Ca (DTPA-Na_3Ca) were injected into the tail vein. The dosage of the drug was measured by 60mg/kg.ICP-AES in SD rats for first days, second days, third days of urine and excretion of uranium, and 3 days after the administration of the liver and spleen. The amount of uranium accumulated in the kidney and bone; the SD rats were treated by intraperitoneal injection of uranyl nitrate (2.0mgU/kg) and treated in the same way. After 5 days, the liver, kidney and spleen were taken to make pathological sections. The bone marrow smear was used as the bone marrow cell smear to examine the degree of pathological damage. The effect of WSC-DTPA nanoparticles on internal pollution of uranium and radiation protection were evaluated.
Results: the prepared WSC-DTPA nanoparticles were tested and met the experimental requirements. Cell experiments and MTT results showed that the survival rate of uranium concentration at about 12.5 u g/mL near 50%.MTT showed that DTPA-CaNa_3 and 1.59%WSC-DTPA nanoparticles could not increase the survival rate of HK-2 fine cells infected by 12.5 u g/mL uranium, while 92.7%WSC-DTPA nanoparticles and WSC Na The rice grain survival rate (P? 0.05) was significantly increased, and the survival rate increased with the increase of drug concentration. The results of PI single staining in flow cytometry showed that the G0/G1 phase block appeared in the 12.5 u g/mL infected cells, and the DNA content in this period increased significantly (P? 0.05), while the content of S phase decreased significantly, and the content of G2/M phase remained approximately 15%; DT; DT PA-CaNa_3 and 1.59%WSC-DTPA nanoparticles did not improve the G0/G1 phase blocking phenomenon caused by uranium exposure, while 92.7%WSC-DTPA nanoparticles and WSC nanoparticles could significantly reduce the G0/G1 phase block. The results of cell promotion experiments showed that DTPA-CaNa_3,1.59% WSC-DTPA nanoparticles, 92.7%WSC-DTPA nanoparticles and WSC nanoparticles were compared with those of the group of pure (3.13 mu g/mL). The effect of P? 0.05 was the best, and the effect of 1.59%WSC-DTPA nanoparticles was the best. The effect of immediate delivery was better. The results of intracellular ROS content showed that the content of ROS in the cells increased significantly (P? 0.05), and the 92.7%WSC-DTPA nanoparticles and WSC nanoparticles could be effectively reduced. The intracellular ROS content (P? 0.05) and 1.59%WSC-DTPA nanoparticles and DTPA-CaNa_3 did not reduce the ROS content in the cells. The 92.7%WSC-DTPA nanoparticles and WSC nanoparticles had radiation resistance. The experimental results showed that the 92.7%WSC-DTPA nanoparticles, 1.59%WSC-DTPA and DTPA-CaNa_3 had the effect of stimulating, and 1.5 in the 0.5 u g/ml uranium exposure test. The effect of 9%WSC-DTPA nanoparticles was significantly better than that in the DTPA-CaNa_3 group (P? 0.05); the main tissue uranium accumulation of the drug group was significantly lower than that of the pure exposure group (P? 0.05). In the 2 UU g/ml uranium exposure experiment, the main tissue uranium accumulation of 92.7% WSC-DTPA nanoparticles and WSC nanoparticles Significantly lower than DTPA-CaNa_3 and 1.59%WSC-DTPA nanoparticles, statistically significant differences (P? 0.05); pathological tissue sections show that 92.7%WSC-DTPA nanoparticles and WSC nanoparticles have protective effects on the liver and kidney, and the damage is significantly lighter than that of other drug groups. There were no obvious abnormalities in the cell classification of bone marrow images.
Conclusion: WSC-DTPA nanoparticles have better effect on promoting and discharging of internal contaminated uranium, and the effect of 1.59%WSC-DTPA nanoparticles is the best, and 92.7%WSC-DTPA nanoparticles not only have the effect of stimulating, but also have radiation protection against the liver and kidney of rats infected with uranium.

【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R818.05

【参考文献】