毒理学杂志

目的 运用体外哺乳动物细胞微核试验和Hprt基因突变试验，以及斑马鱼胚胎毒性试验，评价了一种新型3D打印义齿样品的生物相容性。方法 用RPMI 1640培养液和超纯水分别对义齿进行浸提，浸提原液(100%)分别用RPMI 1640培养液和超纯水稀释至75%、50%和25%。体外哺乳动物细胞微核试验用CHL细胞，分为添加S₉培养6 h、不添加S₉培养6 h和不添加S₉培养24 h 3组试验，每组分别加入100%、75%、50%和25%的样品浸提液，培养至预定时间后观察细胞内微核的形成并统计微核率。体外哺乳动物细胞Hprt基因突变试验用V79细胞，分为添加S₉培养6 h和不添加S₉培养6 h 2组试验，分别加入100%、75%、50%和25%的样品浸提液，培养至突变型细胞表达后，观察并统计集落形成率和突变率。斑马鱼胚胎毒性试验分为4组，分别加入100%、75%、50%和25%的样品浸提液，培养后观察胎体发育情况，并测量96 hpf的斑马鱼体长。结果 体外哺乳动物细胞微核试验中，细胞阻滞率均低于10%,微核率均低于0.5%,所有的处理组与阴性对照组相比差异无统计学意义(P>0.05)。体外哺乳动物细胞Hprt基因突变试验中，相对集落率均高于92%,且突变率低于1.63×10^-6,所有的处理组与阴性对照组相比差异无统计学意义(P>0.05),但与各自的阳性对照相比差异有统计学意义(S₉组：F=92.5,P<0.001;-S₉组：F=199.1,P<0.001)。斑马鱼胚胎毒性试验中，各处理的胚胎均能正常发育，未观察到死亡终点和亚致死发育终点，96 hpf的幼鱼体长与阴性对照相比差异无统计学意义(P>0.05)。结论 在本试验条件下，该新型义齿浸提液未显示有致诱变性和致突变性，也未观察到浸提液有斑马鱼胚胎毒性。

Objective In vitro micronucleus test of mammalian cells in vitro Hprt gene mutation test of mammalian cells, and the zebra fish embryo toxicity test were conducted to evaluate the biocompatibility of a novel 3D printed denture. Methods The denture was extracted by RPMI 1640 culture solution and ultrapure water. And then the extraction solutions were diluted to 75%, 50%, and 25% with RPMI 1640 medium and ultrapure water as extraction reagents, respectively. For In vitro micronucleus of mammalian cells test, CHL cells were divided into three groups. In group 1, cells were cultured with S₉ for 6 h, while in group 2 and group 3, cells were cultured without S₉ for 6 h and for 24 hours, respectively. Each group was added with 100%, 75%, 50%, and 25% diluted sample extract. After culture, the formation of micronucleus in cells was observed and the micronucleus rate was calculated. For In vitro Hprt gene mutation of mammalian cells test, V79 cells were divided into group cultured with S₉ for 6 hours and group cultured without S₉ for 6 h, respectively. Each group was also added with 100%, 75%, 50%, and 25% diluted sample extract, respectively. After culture to the status that mutant gene fully expressed in cells, the colony formation was observed and the mutation rate was calculated. For the Zebrafish embryo toxicity test, Zebrafish embryos were divided into four groups, adding 100%, 75%, 50%, and 25% diluted sample extract, respectively. After culture, Zebrafish embryos development were carefully checked, and the body length of all Zebrafishes at 96 hpf were measured. Results In vitro micronucleus test of mammalian cells showed that the block rates of all cells were less than 10%, and the micronucleus rates were less than 0.5%. There was no significant difference between all treatment groups and the negative control group(P > 0.05). In vitro Hprt gene mutation test of mammalian cells showed that the relative colony formation rates were higher than 92%, and the mutation rates of Hprt gene were lower than 1.63×10^-6. No significant difference in mutation rate between all treatment groups and the negative control group was observed(P > 0.05). However, there was a highly significant difference compared with their respective positive controls(S₉ group: F=92.5,P<0.001;-S₉ group: F=199.1,P<0.001). In the zebra fish embryo toxicity test, the embryos in all treatment groups developed normally, and no death and sublethal endpoints were observed. There was no significant difference in body lengths of the Zebra fish between all treatment groups and the negative control group(P > 0.05). Conclusion Under the experimental conditions, the denture manufactured using 3D technology has no genotoxicity and no zebra fish embryo toxicity.