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背景:当各级生殖细胞DNA受到损伤而发生突变时,均可能通过受精卵遗传给后代,苯妥英钠作为传统抗癫痫药物,已被许多研究证实其对体细胞具有诱变效应,而是否对生殖细胞也具有诱变作用,并通过受精卵遗传给后代,尚需进一步研究。目的:检测苯妥英钠对人精子染色体的诱变效应。设计:以离体人精子染色体为观察对象的随机对照观察。单位:重庆医科大学附属第一医院心理卫生中心。材料:苯妥英钠购自Sigma公司,人精液标本取自近半年内未接触过理化诱变因子的健康成年男性。鼠卵取自6～8周龄的雌金黄地鼠,用含0.3%人血清白蛋白的BWW液培养基洗涤精卵和受精,在含3.5%血清白蛋白的BWW液培养基中使精子获能,受精后培养基用含10%地鼠血清的卵培养基。方法:将精子洗涤、离心、获能后,制作成5mL精子悬液,平均分置于5支离心管中,一支加入终浓度为40mg/L的平阳霉素作为阳性对照,一支不加任何试剂作为空白对照,其余3支分别加入终浓度为10mg/L、20mg/L、40mg/L的苯妥英钠。按常规方法制作无透明带的地鼠卵后,平均分为5等份,分别与上述5组精液混合,使地鼠卵受精,并用受精的地鼠卵制备人精子染色体。采用染色体结构畸变精子率(简称畸精率)和断裂数2个指标,应用人精子染色体离体测试系统检测不同浓度苯妥英钠对人精子染色体的诱变作用,并与平阳霉素40mg/L组和空白对照组比较。主要观察指标:各染色体精子染色体结构畸变情况,结构畸变精子率和断裂数。结果:①染色体结构畸变:不同浓度苯妥英钠组、阳性对照组和空白对照组均显示染色体断裂和单体断裂、断片、相互易位、双着丝粒体、环状染色体等多种精子染色体结构畸变,尤以苯妥英钠40mg/L组和平阳霉素40mg/L组更为显著。②染色体结构畸变精子率和染色体断裂数:不同浓度苯妥英钠组和平阳霉素40mg/L组的染色体结构畸变精子率和断裂数均较空白对照组高,其中苯妥英钠40mg/L组和平阳霉素40mg/L组差异有显著和高度显著性(P<0.005,P<0.05～0.01)。结论:苯妥英钠对人精子染色体有诱变作用。
BACKGROUND: When all levels of germ cell DNA are damaged and mutated, they may be passed on to offspring through fertilized eggs. Phenytoin, as a traditional antiepileptic drug, has been proved by many studies to have a mutagenic effect on somatic cells, Cells also have a mutagenic effect, and passed on to the offspring of fertilized eggs, need further study. Objective: To detect the mutagenic effect of phenytoin on human sperm chromosome. Design: A Randomized Controlled Observation of In Vitro Human Sperm Chromosomes. Unit: First Affiliated Hospital of Chongqing Medical University Mental Health Center. MATERIALS: Phenytoin sodium was purchased from Sigma, and human semen samples were taken from healthy adult men who had not been exposed to physico-chemical mutagens for nearly six months. The mouse eggs were taken from 6-8 week-old female golden hamsters and the sperm eggs were washed with BWW broth containing 0.3% human serum albumin and fertilized, and sperm were harvested in BWW broth containing 3.5% serum albumin Can, after fertilization medium with 10% hamster serum egg medium. Methods: The sperm were washed, centrifuged, and harvested to make a 5 mL sperm suspension. The cells were evenly divided into 5 centrifuge tubes and one was added with Pingyangmycin at a final concentration of 40 mg / L as a positive control. Any reagent as a blank control, the remaining three were added to a final concentration of 10mg / L, 20mg / L, 40mg / L of phenytoin sodium. The ophthalmophilic zygotic eggs were prepared according to a conventional method, and were equally divided into 5 equal portions, each of which was mixed with the above-mentioned 5 groups of semen to fertilize the hamster eggs and prepare human sperm chromosomes with fertilized hamster eggs. Chromosome structural aberration sperm rate (referred to as the rate of abnormality) and the number of fracture 2 indicators, the application of human sperm chromosome in vitro test system to detect different concentrations of phenytoin sodium on human sperm chromosome mutagenesis, and Pingyangmycin 40mg / L group Compared with the blank control group. MAIN OUTCOME MEASURES: Chromosome structural aberration of each chromosome, structural aberration sperm ratio and number of fractures. Results: (1) Chromosomal structural aberrations: The phenytoin sodium group, the positive control group and the blank control group all showed multiple chromosome structures including chromosome rupture and monomer rupture, fragment, reciprocal translocation, dicenosome and circular chromosome Distortion, especially in 40mg / L sodium phenytoin and Pingyangmycin 40mg / L group was more significant. ② Chromosome structural aberration Sperm ratio and number of chromosome rupture: The chromosome aberration rate of sperm motility and number of rupture in the phenytoin sodium group and the pingyangmycin 40 mg / L group were higher than those in the blank control group, in which 40 mg / L phenytoin group and The difference was statistically significant (P <0.005, P <0.05 ~ 0.01). Conclusion: Phenytoin has a mutagenic effect on human sperm chromosome.