McCoy和Hela229细胞沙眼衣原体持续感染模型及omcB与Euo基因的表达
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摘要
目的比较阿奇霉素诱导McCoy细胞与Hela229细胞建立的沙眼衣原体持续性感染模型及omcB和Euo基因表达情况。方法用相同浓度的沙眼衣原体分别感染McCoy和Hela229细胞,在感染22h后加入0.08μg/ml阿奇霉素,构建沙眼衣原体持续性感染模型;通过荧光显微镜观察细胞形态,反转录PCR(RT-PCR)检测omcB和Euo基因表达。结果 McCoy细胞和Hela229细胞内,未加阿奇霉素处理组,沙眼衣原体包涵体较大,加入阿奇霉素处理组,沙眼衣原体包涵体均较小。持续性感染时,Hela229细胞omcB基因的表达较急性感染下调0.138±0.042倍,差异有统计学意义(t=35.120,P<0.001);McCoy和Hela229细胞Euo基因的表达较急性感染分别上调2.445±0.460倍和3.317±0.556倍,差异均有统计学意义(t=5.441,P=0.006;t=7.210,P=0.002);在McCoy和Hela229细胞持续感染时,Euo/omcB比值较急性感染分别上升2.100±0.050倍和24.820±2.630倍,差异均有统计学意义(t=32.201,11.240;均P<0.001)。结论在阿奇霉素诱导下,Hela229比McCoy细胞更容易建立沙眼衣原体持续性感染模型。
OBJECTIVE To compare the McCoy and Hela229 cell models of Chlamydia trachomatis persistent infection induced by azithromycin and observe the expression of omcB and Euo genes.METHODS McCoy and Hela229 cells were infected with the same concentration of Ct.rachomatis,azithromycin(0.08μg/ml)was added 22 hours after chlamydial infection to construct the model of Ct.rachomatis persistent infection.The morphology of the cells was observed by the fluorescence microscopy,and the expressions of omcBand Euo genes were detected by reverse transcription PCR(RT-PCR).RESULTS The size of Ct.rachomatis inclusion bodies in McCoy cells and Hela229 cells became smaller with the treatment of azithromycin but became larger without treatment of azithromycin.The expression of omcBgene of Hela229 cells of the persistent infection was downregulated 0.138±0.042 times as the acute infection,and there was significant difference(P<0.01);the expressions of Euo gene of McCoy and Hela229 cells of the persistent infection were respectively upregulated 2.445±0.460 times and 3.317±0.556 times as the acute infection,and there were significant differences(t=5.441,P=0.006;t=7.210,P=0.002).The ratios of Euo to omcBin MyCoy cell and Hela229 cells of the persistent infection were respectively upregulated 2.10±0.05 times and 24.82±2.63 times as the acute infection,and there were significant differences(t=32.201,11.240;all P<0.001).CONCLUSION Under the induction of azithromycin,Hela229 cell is more convenient than Mycoy cell to establish the model of Ct.rachomatis persistent infection.
OBJECTIVE To compare the McCoy and Hela229 cell models of Chlamydia trachomatis persistent infection induced by azithromycin and observe the expression of omcB and Euo genes.METHODS McCoy and Hela229 cells were infected with the same concentration of Ct.rachomatis,azithromycin(0.08μg/ml)was added 22 hours after chlamydial infection to construct the model of Ct.rachomatis persistent infection.The morphology of the cells was observed by the fluorescence microscopy,and the expressions of omcBand Euo genes were detected by reverse transcription PCR(RT-PCR).RESULTS The size of Ct.rachomatis inclusion bodies in McCoy cells and Hela229 cells became smaller with the treatment of azithromycin but became larger without treatment of azithromycin.The expression of omcBgene of Hela229 cells of the persistent infection was downregulated 0.138±0.042 times as the acute infection,and there was significant difference(P<0.01);the expressions of Euo gene of McCoy and Hela229 cells of the persistent infection were respectively upregulated 2.445±0.460 times and 3.317±0.556 times as the acute infection,and there were significant differences(t=5.441,P=0.006;t=7.210,P=0.002).The ratios of Euo to omcBin MyCoy cell and Hela229 cells of the persistent infection were respectively upregulated 2.10±0.05 times and 24.82±2.63 times as the acute infection,and there were significant differences(t=32.201,11.240;all P<0.001).CONCLUSION Under the induction of azithromycin,Hela229 cell is more convenient than Mycoy cell to establish the model of Ct.rachomatis persistent infection.
引文
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[18]Skilton RJ,Cutcliffen LT,Barlow D,et al.Penicillin induced persistence in Chlamydia trachomatis:high quality time lapse video analysis of the developmental cycle[J].PLoSOne,2009,4(11):e7723.
[19]黄凯凯,刘隽华,韩建德,等.沙眼衣原体生殖道持续性感染的实验室诊断和治疗进展[J].国际皮肤性病学杂志,2017,43(1):54-57.
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[21]朱慧玲,李红梅,陈木开,等.体外沙眼衣原体持续感染快速鉴定方法探索[J].中国皮肤性病学杂志,2013,27(4):411-421.
[22]Meyer T.Diagnosis and treatment of Chlamydia trachomatis infections[J].Hautarzt,2012,63(1):16-23.
[23]Schoborg RV.Chlamydia persistence-a tool to dissect chlamydiahost interactions[J].Microbes Infect,2011,13(7):649-662.
[24]Hammerschlag MR,Kohlhoff SA.Treatment of chlamydial infections[J].Expert Opin Pharmacother,2012,13(4):545-552.
[25]Ong VA,Marsh JW,Lawrence A,et al.The protease inhibitor JO146 demonstrates a critical role for CtHtrA for Chlamydia trachomatis reversion from penicillin persistence[J].Front Cell Infect Microbiol,2013,3:100.
[26]薛耀华,麦志达,覃晓琳,等.四种细胞系培养沙眼衣原体易感性的比较[J].皮肤性病诊疗学杂志,2017,24(1):8-13.
[27]Fadel S,Eley A.Chlamydia trachomatis omcB protein is a surface exposed glycosaminoglycan-depent adhesin[J].J Med Microbiol,2007,56(pt1):15-22.
[28]Hou S,Lei L,Yang Z,et al.Chlamydia trachomatis outer membrane complex protein B(OmcB)is processed by the protease CPAF[J].J Bacteriol,2013,195(5):951-957.
[2]邱晓丽,龚向东,滕菲,等.2008-2015年中国性病监测点生殖道沙眼衣原体感染流行特征分析[J].中华皮肤科杂志,2016,49(5):308-313.
[3]World Health Organization.Global health sector strategy on sexually transmitted infections,2016-2021[M].Geneva:World Health Organization,2016:7-11.
[4]唐卫民,薜耀华,郑和平,等.生殖道沙眼衣原体感染的挑战与应对[J].中华皮肤科杂志2017,50(5):313-315.
[5]Malhotra M,Sood S,Mukherjee A,et al.Genital Chlamydia trachomatis:an update[J].Indian J Med Res,2013,138(3):303-316.
[6]Hammerschlag MR.Treatment of Chlamydia trachomatis infections[M].Basel:Karger,2013,7:89-96.
[7]叶顺章.性传播疾病的实验室诊断[M].2版.北京:科学出版社,2009:109.
[8]Bavoil PM.What′s in a word:the use,misuse,and abuse of the word"persistence"in Chlamydia biology[J].Front Cell Infect Microbiol,2014,4:27.
[9]黄敏,李建红,段行武,等.生殖道沙眼衣原体持续性感染研究进展[J].中华皮肤科杂志,2013,46(8):607-609.
[10]Shima K,Klinger M,Solbach W,et al.Activities of first-choice antimicrobials against gamma interferon-treated Chlamydia trachomatis differ in hypoxia[J].Antimicrob Agents Chemother,2013,57(6):2828-2830.
[11]Menon S,Timms P,Allan JA,et al.Human and pathogen factors associated with Chlamydia trachomatis-related infertility in women[J].Clin Microbiol Rev,2015,28(4):969-985.
[12]Rosario CJ,Hanson BR,Tan M.The transcriptional repressor EUO regulates both subsets of Chlamydia late genes[J].Mol Microbiol,2014,94:888-897.
[13]Rosario CJ,Tan M.The early gene product EUO is a transcriptional repressor that selectively regulates promoters of Chlamydia late genes[J].Mol Microbiol,2012,84:1097-1107.
[14]Rosario CJ,Tan M.Regulation of Chlamydia gene expression by tandem promoters with different tempora patterns[J].JBacteriol,2016,198(2):363-369.
[15]Lewis ME,Belland RJ,AbdelRahman YM,et al.Morphologic and molecular evaluation of Chlamydia trachomatis growth in human endocervix reveals distinct growth patterns[J].Front Cell Infect Microbiol,2014,4:71.
[16]Fechtner T,Stallmann S,Moelleken K,et al.Characterization of the interaction between the chlamydial adhesin OmcB and the human host cell[J].J Bacteriol,2013,195:5323-5333.
[17]薜耀华,白顺,郑磊,等.IFN-γ体外诱导沙眼衣原体持续性感染细胞模型的建立[J].皮肤性病诊疗学杂志,2015,22(1):10-13,17.
[18]Skilton RJ,Cutcliffen LT,Barlow D,et al.Penicillin induced persistence in Chlamydia trachomatis:high quality time lapse video analysis of the developmental cycle[J].PLoSOne,2009,4(11):e7723.
[19]黄凯凯,刘隽华,韩建德,等.沙眼衣原体生殖道持续性感染的实验室诊断和治疗进展[J].国际皮肤性病学杂志,2017,43(1):54-57.
[20]Kintner J,Lajoie D,Hall J,et al.Commonly prescribedβ-lactam antibiotics induce C.trachomatis persistence/stress in culture at physiologically relevant concentrations[J].Front Cell Infect Microbiol,2014,4:44.
[21]朱慧玲,李红梅,陈木开,等.体外沙眼衣原体持续感染快速鉴定方法探索[J].中国皮肤性病学杂志,2013,27(4):411-421.
[22]Meyer T.Diagnosis and treatment of Chlamydia trachomatis infections[J].Hautarzt,2012,63(1):16-23.
[23]Schoborg RV.Chlamydia persistence-a tool to dissect chlamydiahost interactions[J].Microbes Infect,2011,13(7):649-662.
[24]Hammerschlag MR,Kohlhoff SA.Treatment of chlamydial infections[J].Expert Opin Pharmacother,2012,13(4):545-552.
[25]Ong VA,Marsh JW,Lawrence A,et al.The protease inhibitor JO146 demonstrates a critical role for CtHtrA for Chlamydia trachomatis reversion from penicillin persistence[J].Front Cell Infect Microbiol,2013,3:100.
[26]薛耀华,麦志达,覃晓琳,等.四种细胞系培养沙眼衣原体易感性的比较[J].皮肤性病诊疗学杂志,2017,24(1):8-13.
[27]Fadel S,Eley A.Chlamydia trachomatis omcB protein is a surface exposed glycosaminoglycan-depent adhesin[J].J Med Microbiol,2007,56(pt1):15-22.
[28]Hou S,Lei L,Yang Z,et al.Chlamydia trachomatis outer membrane complex protein B(OmcB)is processed by the protease CPAF[J].J Bacteriol,2013,195(5):951-957.
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