Interaction of actinobacillus pleuropneumoniae with host epithelial cells

胸膜肺炎放线杆菌与宿主上皮细胞的相互作用

• 批准号: 3428-2006
• 负责人: Jacques, Mario
• 金额: $ 4.1万
• 依托单位: Université de Montréal
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2008
• 资助国家: 加拿大
• 起止时间: 2008-01-01 至 2009-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=388758
• 关键词: Interaction; actinobacillus; pleuropneumoniae; host; epithelial

The long-term objective of this research program is to gain a better understanding of the virulence factors involved in the Gram-negative bacterial pathogens colonization of the host's respiratory tract. Actinobacillus pleuropneumoniae (App), the agent of porcine pleuropneumonia, is our model. Our hypothesis is that App is able to express multiple adhesins and  that these adhesins are involved, at one time or another, during the colonization and infection process. We have shown that lipopolysaccharide (LPS) is one of the adhesins of App involved in adherence to porcine respiratory tract cells. In order to identify which part of the LPS molecule (core oligosaccharide or O-antigen) is involved in binding to these cells, we have generated LPS mutants of App serotype 1 using mini-Tn10 transposon mutagenesis. Our data suggest that deletion of specific residues in the outer LPS core affects colonization and results in the attenuation of the virulence of App serotype 1 and that an intact core-lipid A region is required for optimal protection of App against host cationic antimicrobial peptides. The short-term objectives of this research program are: (1) to further characterize the biological properties of the LPS mutants we have generated in our laboratory. We will compare the parental strain and LPS mutants' capacity to stimulate cytokines production by lung epithelial cells; (2) to develop an in vitro adherence assay using porcine lung (SJPL) and trachea (NPTr) epithelial cell lines; (3) to determine the transcriptome profiling of App in contact with host cells using DNA microarrays which will enable us to get for the first time a global view on App genes that are specifically expressed after interaction with host cells; (4) to generate novel mutants in putative adhesins by allelic exchange (on selected genes) or by random transposon mutagenesis. A better understanding of the adhesin-receptor systems of App and of the genes expressed upon contact with host epithelial cells will lead to the development of more efficient strategies to control infection.

这项研究计划的长期目标是更好地了解革兰氏阴性细菌在宿主呼吸道的定植所涉及的毒力因素。猪胸膜肺炎的病原--胸膜肺炎放线杆菌(App)就是我们的模型。我们的假设是，App能够表达多个粘附素，并证实这些粘附素在定植和感染过程中或多或少参与其中。我们已经证明，脂多糖(LPS)是App的粘附素之一，参与了与猪呼吸道细胞的黏附。为了确定与这些细胞结合的是内毒素分子的哪一部分(核心低聚糖或O-抗原)，我们利用微型Tn10转座子突变产生了App血清1型的内毒素突变体。我们的数据表明，内毒素外区特定残基的缺失会影响App的定植，并导致App 1型毒力的减弱，而完整的核心-脂类A区是App对宿主阳离子抗菌肽的最佳保护所必需的。这项研究计划的短期目标是：(1)进一步鉴定我们实验室产生的内毒素突变体的生物学特性。我们将比较亲本株和脂多糖突变体刺激肺上皮细胞产生细胞因子的能力；(2)利用猪肺(SJPL)和气管(NPTR)上皮细胞系建立体外黏附试验；(3)利用DNA微阵列确定App与宿主细胞接触后的转录组图谱，这将使我们首次能够对与宿主细胞相互作用后特异表达的App基因进行全局剖析；(4)通过等位基因交换(在选定的基因上)或通过随机转座子突变来产生可能粘附素的新突变体。更好地了解App的粘附素受体系统以及与宿主上皮细胞接触时表达的基因将导致开发更有效的感染控制策略。