纳米材料对小鼠肺毒性的研究和甲型H1N1流感病毒在人肺上皮细胞及雪貂中的致病性研究

发布时间：2018-04-03 22:16

本文选题：急性肺损伤　切入点：地塞米松　出处：《北京协和医学院》2012年博士论文

【摘要】：纳米技术已经是当今最重要及新兴的科技发展之一,但因为它的发展而导致的安全问题也引起了越来越多的关注。在它们的发展过程中对纳米材料在人体暴露和引起的环境问题方面的危害评估对于纳米材料将来的发展至关重要。本文所研究的纳米材料是具有可溶性的修饰化单壁碳纳米管(functionalized single-walled carbon nanotubes,f-SWCNTs)。碳纳米管(Carbon NanoTubes,CNT)是一种特殊形式的碳分子,是碳原子之间形成化学键组成的管状结构。单壁碳管(single-walled carbon nanotubes, SWCNT),由一层碳原子网组成,直径范围0.6～2.4nm；由于其特有的理化性质(重量较轻,较好的延伸性能,热力学/化学稳定性以及导电能力),碳纳米管被大量用于制造组织支架。同时,由于具有较大的表面积,表面可以被各种功能基团修饰而具有不同的功能,碳纳米管被广泛用于递送肽段、蛋白质、基因、药物或疫苗等[4]。本研究针对表面功能基团为AMIDE, COOH, PABS, PEG的单壁碳管在小鼠中的肺毒性做了评估并阐述了相关的分子机制,进一步检测了修饰化单壁碳纳米管处理后小鼠的自我修复能力和小鼠肺纤维化程度。在气管注射修饰化单壁碳纳米管18h或14d后,检测了小鼠肺组织病理、肺泡支气管灌洗液(Bronchial Alveolar Lavage Fluid, BALF)中细胞因子、肺血管通透性以及血氧分压,另外也做了免疫组化检测。我们发现部分修饰化单壁碳纳米管能在小鼠中通过MyD88-NF-κB信号通路能引起促炎性细胞因子风暴而造成急性肺损伤(Acute Lung Injury, ALI)。本研究中,我们也证实了皮质类固醇类药物能减轻修饰化单壁碳纳米管在小鼠中诱导的急性肺损伤。另外,单次给药14d后,小鼠的急性肺损伤几乎完全恢复正常,而同时轻度至中度的肺纤维化、肉芽肿、DNA损失在14d时仍然存在。综上所述,我们的研究为将来处理随着应用而不断增长的纳米材料安全性问题提供了可能的参考。背景：2009年在北美爆发了以一株新的猪源性甲型H1N1流感病毒造成的的流行性感冒[1,2,3],该病毒随后在人类中流行并短时间内传播至全世界大部分地区。然而此病毒在人呼吸道上皮细胞和哺乳动物中的致病机制仍然不清晰。实验方法及实验结果：在本研究中,我们发现一株从病人身上分离的2009年甲型H1N1流感病毒,A/Beijing/501/2009,能在雪貂中引起典型的流行性感冒症状,包括体重减轻、体温波动、肺组织病理变化。我们的研究也证实了人肺腺癌上皮细胞A549细胞对甲型H1N1流感病毒易感,同时被感染的细胞在早期即发生了凋亡。与季节性H1N1流感病毒相比,甲型H1N1流感病毒株A/Beijing/501/2009能在A549细胞中引起更多的细胞死亡,且细胞死亡方式是依赖caspase-3信号通路的凋亡。另外,感染了甲型流感A/Beijing/501/2009H1N1病毒株的雪貂表现出体温上升、体重下降更厉害、肺组织中病毒滴度更高等症状。因此,A/Beijing/501/2009型甲型H1N1流感病毒株能成功感染雪貂肺组织,并能造成比季节性流感病毒更严重的病理损伤。结论：本研究发现2009年甲型H1N1流感病毒与季节性H1N1流感病毒在体内和体外表现出不同的毒力,可能是通过不同的机制导致的。细胞凋亡参与甲型H1N1流感病毒造成的体外细胞病变、甲型H1N1流感病毒感染的雪貂肺组织中病理变化比季节性流感病毒更严重的这些发现拓宽了人类对甲型H1N1流感病毒在感染人类和动物的致病机制方面的知识。
[Abstract]:Nanotechnology is one of the most important and emerging scientific and technological developments in the present day , but the security problems caused by its development have attracted more and more attention . In the course of their development , the damage assessment of nanomaterials in human exposure and environmental problems is essential to the future development of nanomaterials .

Nanomaterials studied in this paper are soluble modified single - walled carbon nanotubes ( f - SWCNTs ) . Carbon NanoTubes ( CNT ) is a special form of carbon molecule , which is a tubular structure composed of chemical bonds between carbon atoms . Single - walled carbon nanotubes ( SWCNT ) are composed of a layer of carbon network with a diameter ranging from 0.6 to 2.4 nm .
The carbon nanotubes are widely used to deliver peptide segments , proteins , genes , drugs or vaccines because of their specific physical and chemical properties ( lighter weight , better elongation properties , thermodynamic / chemical stability , and conductivity ) .

In this study , the lung toxicity of single - walled carbon nanotubes in mice was assessed and the relative molecular mechanism was assessed and the self - healing ability and the degree of pulmonary fibrosis in mice were also described . After tracheal injection - modified single - walled carbon nanotubes for 18 h or 14 d , the expression of cytokines , pulmonary vascular permeability and oxygen partial pressure in lung tissues of mice were examined .

In this study , we also demonstrated that corticosteroids can alleviate acute lung injury induced by modified single - walled carbon nanotubes in mice . In addition , after a single dose of 14 days , acute lung injury in mice is almost completely normal , while mild to moderate pulmonary fibrosis , granuloma , DNA loss still exist at 14d .

In conclusion , our research provides a potential reference for future handling of the safety of nanomaterials with growing applications .

Background : In 2009 , a new swine - derived influenza A ( H1N1 ) virus , caused by a new swine - derived influenza A ( H1N1 ) virus , was outbreak in North America , which was then prevalent in humans and disseminated in a short period of time to most parts of the world . However , the pathogenic mechanism of the virus in human respiratory epithelial cells and mammals remains unclear .

In this study , we found that influenza A / A / Beijing / 501 / 2009 , isolated from a patient , can cause typical influenza symptoms , including weight loss , body temperature fluctuation and pathological changes in lung tissues . In addition , the influenza A / Beijing / 501 / 2009 H1N1 virus strain can cause more cell death in A549 cells . In addition , A / Beijing / 501 / 2009 H1N1 virus strain can successfully infect ferret lung tissue , and can cause more serious pathological damage than seasonal influenza virus .

Conclusion : This study shows that influenza A ( H1N1 ) virus and seasonal H1N1 influenza virus exhibit different virulence in vivo and in vitro , which may be caused by different mechanisms . Apoptosis is involved in the in vitro cellular pathological changes caused by influenza A H1N1 virus , and the pathological changes in mink lung tissues infected by influenza A H1N1 influenza virus are more serious than those of seasonal influenza virus . These findings broaden the knowledge of influenza A ( H1N1 ) virus in infection of human and animal pathogenic mechanisms .

【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R114

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