發(fā)布時(shí)間：2018-05-23 11:06

  本文選題：痘苗病毒 + 天壇株��； 參考：《武漢大學(xué)》2005年博士論文

【摘要】：盡管痘苗病毒天壇株曾作為天花病預(yù)防疫苗在中國(guó)大范圍接種使用了幾十年,但其相關(guān)的生物學(xué)特性并沒(méi)有詳細(xì)的研究和報(bào)導(dǎo)。在本文中我們對(duì)痘苗病毒天壇株的細(xì)胞宿主范圍、體外復(fù)制特性和病毒體內(nèi)的毒性等生物學(xué)特性進(jìn)行了系統(tǒng)的鑒定。我們發(fā)現(xiàn)天壇株具有廣泛的宿主范圍,經(jīng)研究的12種哺乳動(dòng)物細(xì)胞中的11種及原代雞胚細(xì)胞CEF均是天壇株的允許細(xì)胞,而中國(guó)倉(cāng)鼠卵巢細(xì)胞CHO-K1是天壇株的非允許細(xì)胞。天壇株在幾乎所有測(cè)試的細(xì)胞中均能產(chǎn)生嚴(yán)重的至細(xì)胞病變效應(yīng)(Cytopathic effects,CPE),包括四種人類來(lái)源的細(xì)胞系。體內(nèi)實(shí)驗(yàn)顯示,盡管天壇株比神經(jīng)毒性的WR株毒性弱很多,但天壇株仍保留相當(dāng)?shù)纳窠?jīng)毒性,而且經(jīng)鼻腔接種可引起小鼠體重嚴(yán)重下降。因而有必要對(duì)天壇株進(jìn)行進(jìn)一步的減毒,使其成為更為安全理想的天花疫苗或作為更安全的病毒載體用于構(gòu)建活病毒載體疫苗。 痘苗病毒的血凝素基因(Hemagglutinin,HA)與痘苗病毒的體內(nèi)毒性密切相關(guān),缺失HA可導(dǎo)致多種痘苗病毒株系在動(dòng)物體內(nèi)不同程度的減毒。在本文中,我們構(gòu)建了HA基因完全失活或部分失活的重組痘苗病毒TT_(zcl)和TT_(pzcl)(HA基因的晚期啟動(dòng)子被插入失活)。我們發(fā)現(xiàn),重組痘苗病毒在12種VTT允許細(xì)胞中的7種中有不同程度的復(fù)制水平的下降,其中TT_(zcl)在RK13和MDCK中完全失去復(fù)制能力。而TT_(pzcl)在MDCK中失去復(fù)制能力。在多數(shù)動(dòng)物細(xì)胞中包括在人源細(xì)胞中重組病毒具有比野生天壇株病毒更低的細(xì)胞到細(xì)胞間的擴(kuò)散水平或更小的對(duì)細(xì)胞的毒性。此外,我們還證實(shí)重組痘苗病毒在連續(xù)傳代中顯示具有外源基因穩(wěn)定插入和穩(wěn)定表達(dá)的特征。在體外實(shí)驗(yàn)中我們進(jìn)一步檢測(cè)了不同感染途徑重組痘苗 病毒在模型動(dòng)物中的毒性。我們發(fā)現(xiàn),重組病毒TT_(zcl)具有經(jīng)多種接種途徑接種在體內(nèi)高度減毒的特征,這可能與TT_(zcl)在動(dòng)物腦中樞神經(jīng)系統(tǒng)中復(fù)制水平顯著下降相關(guān)。而HA部分失活的重組痘苗病毒TT_(pzcl)在體內(nèi)也顯著減毒的特征,但其減毒的程度大大低于重組病毒TT_(zcl)。高度減毒的重組痘苗病毒TT_(zcl)將是更為安全的載體,可用于進(jìn)一步構(gòu)建病毒載體疫苗。
[Abstract]:Although vaccinia virus Temple of Heaven strain has been widely used in China for decades as a smallpox preventive vaccine, its biological characteristics have not been studied and reported in detail. In this paper, we systematically identified the biological characteristics of vaccinia virus Temple of Heaven strain, such as cell host range, in vitro replication characteristics and virulence in vivo. We found that the Temple of Heaven strain has a wide host range, 11 of the 12 mammalian cells studied and the primary chicken embryo cell CEF are all permitted cells of Temple of Heaven strain, while the Chinese hamster ovary cell CHO-K1 is the non-permissible cell of Temple of Heaven strain. Temple of Heaven strain can produce severe cytopathic effects in almost all the tested cells, including four human cell lines. In vivo experiments showed that although the Temple of Heaven strain was much less toxic than the neurotoxic WR strain, the Temple of Heaven strain still retained considerable neurotoxicity, and inoculation through nasal cavity caused severe weight loss in mice. Therefore, it is necessary to further attenuate the Temple of Heaven strain and make it a safer ideal smallpox vaccine or a safer viral vector for construction of live virus vector vaccine. Hemagglutinin gene (Hemagglutinin HA) of vaccinia virus is closely related to virulence of vaccinia virus in vivo. In this paper, we constructed recombinant vaccinia virus (TTSZ) with HA gene inactivation or partial inactivation, and the late promoter of TT_(pzcl)(HA gene was inserted into inactivation. We found that recombinant vaccinia virus has different degrees of reduction in replication level in 7 out of 12 VTT species, and TTS virus completely loses replication ability in RK13 and MDCK. TTT / Pzcls lose the ability to replicate in MDCK. Recombinant viruses in most animal cells, including human cells, have lower cell-to-cell diffusion levels or less cytotoxicity than wild Temple of Heaven strains. In addition, we also confirmed that recombinant vaccinia virus has the characteristics of stable insertion and stable expression of exogenous genes in successive passages. In vitro, we further examined the toxicity of recombinant vaccinia virus with different infection pathways in model animals. We found that the recombinant virus TTS ZL has the characteristics of highly attenuated virus in vivo by multiple inoculation routes, which may be related to the significant decrease of TTS replication in the central nervous system of animals. However, the HA partially inactivated recombinant vaccinia virus TTS / pzcll was also characterized by a marked reduction of virus in vivo, but the degree of the reduction was much lower than that of the recombinant virus. Highly attenuated recombinant vaccinia virus TTSP will be a safer vector for further construction of viral vector vaccine.
【學(xué)位授予單位】：武漢大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2005
【分類號(hào)】：R346

【引證文獻(xiàn)】

相關(guān)期刊論文 前3條

1 朱蓉;黃維金;王佑春;俞永新;;痘苗病毒弱毒株廣9株的免疫原性分析[J];中國(guó)生物制品學(xué)雜志;2011年09期

2 余文博;劉利;陳志偉;;痘苗病毒天壇株的減毒與其作為疫苗載體研究進(jìn)展[J];中國(guó)病毒病雜志;2011年01期

3 朱蓉;黃維金;嚴(yán)子林;溫智恒;王文波;周艷;王佑春;俞永新;;痘苗病毒弱毒株廣9株對(duì)動(dòng)物的致病力研究[J];中國(guó)病毒病雜志;2011年03期

相關(guān)會(huì)議論文 前1條

1 朱蓉;黃維金;王佑春;俞永新;;痘苗病毒弱毒株廣9株的免疫原性分析[A];第五次全國(guó)免疫診斷暨疫苗學(xué)術(shù)研討會(huì)論文匯編[C];2011年

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本文編號(hào)：1924474