【作者】 孙建玲；

【导师】 张斌；

【作者基本信息】 山东大学 ， 分析化学， 2013， 硕士

【摘要】 化学和材料科学的飞速发展使得大批新材料涌现,纳米材料作为新型材料的代表,目前已渗透到工业和人类日常生活的众多领域。随着纳米材料在生态环境中有意或无意暴露的增多,人们对纳米材料是否会影响人类的健康和生态环境这一问题的关注越来越多。2003年,Science和Nature相继发表文章,探讨了纳米材料的生物效应和对环境的影响。碳纳米管凭借独特的电、磁、热和机械性能,成为纳米材料中的研究热点之一。碳纳米管目前已用于复合材料、电子和能源等领域,以生物医学应用为目标的探索研究也正迅速发展成为一个新的研究领域。碳纳米管在医药领域深入广泛应用的前提是其良好的生物相容性和无毒副作用等。碳纳米管可经血液循环在肝、脾、肺、肾和脑中积累,并产生不同程度的损伤,但是碳纳米管是否能转移至雌性生殖器官并造成脏器功能障碍进而损伤生殖力的研究目前尚无报道。由于受到工业污染等方面的影响,人类女性生殖健康状况呈不断下降趋势。了解纳米材料对雌性生殖健康的影响,作为纳米材料安全性评价的重要部分势在必行。已有报道指出金纳米和磷酸钙纳米颗粒可进入卵巢颗粒细胞,诱发细胞凋亡,抑制雌激素的分泌。孕期暴露单壁碳纳米管可造成胎儿畸形等形态学上的损伤。然而,还未有报道对碳纳米管的非孕期雌性生殖系统的体内毒性进行全面的评价。碳纳米管作为一种工业及医药应用前景非常广阔的纳米材料,且在实际生产、生活中主动或者被动进入人体的几率极高,研究其对雌性生殖健康的影响具有现实意义。本研究采用水溶性较好的羧基化多壁碳纳米管为研究对象,以雌性ICR小鼠为模型,首次评价了多壁碳纳米管对非孕期雌性生殖健康的影响。依据雌性生殖系统的内分泌调控轴(下丘脑-垂体-卵巢轴),系统研究了羧基化多壁碳纳米管在雌性生殖系统的分布和毒性,并初步进行了机制研究。实验结果表明,尾静脉单针注射64Cu标记的羧基化碳纳米管可在卵巢、子宫和脑中分布,并在24h内呈积累下降趋势。碳纳米管的暴露造成了小鼠动情周期的在给药期间的延长和血清性激素水平紊乱,说明在受试条件下,雌鼠的生殖内分泌功能受到了碳纳米管的干扰。然而,对第13天和第60天卵巢和子宫组织进行光镜观察发现,多次暴露碳纳米管并未造成明显的病理学改变,各级卵泡和黄体数目、子宫内膜和肌层厚度均无显著性变化。对雌性生殖功能进行的评价结果显示,羧基化多壁碳纳米管对怀孕期母鼠和子代发育中的各指标(交配率、怀孕率、生殖率、孕鼠体重和行为；仔鼠出生数目和性别比、第1天和第10天生存率、体重变化等)均未产生负面影响。毒性机制研究表明,实验组小鼠子宫和卵巢在第13天表现有免疫因子TNFα水平的明显升高,氧化应激指标MDA和GSH与对照组相比有显著改变。卵巢切除实验证实,多壁碳纳米管不能维持小鼠正常的动情周期和子宫变化,因此不具备类雌激素效应。综上所述,多壁碳纳米管可进入雌性生殖系统并影响生殖相关激素的分泌,但组织结构和生殖力均未受到影响。多壁碳纳米管在体内并不具备类雌激素效应,因此对内分泌的影响与碳管与雌激素的拮抗作用无关,而可能是多壁碳纳米管在卵巢内产生过量的氧化应激的结果。碳纳米管可进入小鼠脑部,但是碳纳米管是否会对与生殖内分泌相关的下丘脑和垂体造成影响尚不明确。因此,我们推测过量的活性氧影响了激素产生相关酶的活性或者碳纳米管对生殖内分泌相关的其他靶器官的影响干扰了激素的产生和分泌。本研究拓宽了纳米毒理学的广度,为认识纳米材料的生殖安全提供评价,并对碳纳米管的广泛生物医学应用提供了重要的实验参考。更多还原

【Abstract】 The rapid development of chemistry and materials science produced a large number of new materials, in which nanomaterials represent the most striking example have entered into many aspects of people’s life. Nowadays, more and more people begin to concern the effects of nanomaterials on human health and ecological environment after intentional or unintentional exposure of nanoparticles. In2003, Science and Nature published articles one after another to explore the biological effects, environmental and healthy impacts cause by nanomaterials.Carbon nanotubes (CNTs) are collections of induvidual or several concentric graphite cylinders, and have attracted the attention of many scientists worldwide. CNTs represent the ideal, perfect and ordered carbon fiber. The size, structure, and topology make them possess unique electrical, magnetic, thermal and mechanical properties, and fascinating in plastics and composite materials, electronics, energy. The exploration in biomedicine is also rapidly developed into a new research field. However, the promise of the wide use of CNTs in biomedicine is their good compatibility and no toxic or side effects. The research in biological toxicity have found that CNTs can translocate in liver, spleen, kidney, and brain through blood circulation, and produce different degrees of damage. However, the study on female reproductive toxicity caused by CNTs is rare and need further study.The health of female reproduction shows a declining trend, due to various effects like industrial pollution. As an important part in safety evaluation of nanomaterials, it is imperative to understand the impact of nanomaterials on female reproductive system. Studies have been reported that gold nanopaticles and calcium phosphate nanoparticles can translocated into ovarian granulosa cell, and induced cell apoptosis and inhibited the secretion of estradiol. Prenatal exposure of single-walled carbon nanotubes caused fetal abnormalities such as morphology damage. Nevertheless, the effects of CNTs on female reproductiv system in non-pregnant period have not been investigated systematacially.CNTs have extensive prospect in manufacture and medicine, and have high risk of entering the human body actively or passively. Study the effects of CNTs on female reproductive health has a realistic significance. In view of the above considerations, we use ICR mouse as the mammalian model to investigate the effects of carboxylic multi-walled carbon nanotubes (MWCNT-COOH) on female reproductive system. According to the regulatory mechanism of the female reproductive system (hypothalamus-pituitary-ovarian axis), we investigated the distribution and effects of MWCNT-COOH on female reproductive organs, and further discussed the toxic mechanisms. The results showed that64Cu-labelled MWCNT-COOH can translocate in ovary, uterus and brain through tail vein injection and the accumulation showed a decreased trend within24h. Repeated administration of MWCNT-COOH prolonged the estrous cycle during the exposure time and inhibited the secretion of hormones (FSH, LH, estradiol, progesterone), which indicated that MWCNT-COOH disturbed the endocrine system of female mice. However, histologic studies of ovary and uterus showed little alteration after exposure. The number of corpus luteum and each stage follicles, thickness of the endometrium and myometrium from treated mice were characterized no difference compared with control mice. Fertility studies at day13and day60showed that MWCNT-COOH do not have detrimental effect on successful conception and healthy growth of offspring (litter size, survival rate, gain of body weight). To explore the mechanism of toxicity, we found that MWCNT-COOH increased the level of TNFa and changed the level of MDA and GSH in ovary and uterus. Further more, MWCNT-COOH do not possess estrogen-like activity because they are unable to maintain normal estrous cycle and cyclical change of uterus in ovariectomized mice.In conclusion, CNTs can translocate into the female reproductive organs and inhibite hormonal secretion, but leave the structure of organs and fertility not affected. CNTs do not possess estrogen-like effect in vivo, so their influence on endocrine system was not the antagonism effect with estrogen but may be the results of excessive reactive oxygen species caused on steroidogenic enzymes. Due to the complexity and importance of the female reproductive system, understanding of the impact of nanomaterials on reproductive health has a realistic significance. Our study has broadened the knowledge of nanotoxicology on female reproducive system and provided important experimental reference on safe application of carbon nanotubes in biological medicine.更多还原