MOLECULAR PATHOGENESIS OF BACTERIAL CYSTITIS

细菌性膀胱炎的分子发病机制

• 批准号: 6105693
• 负责人: JOHN W WARREN
• 金额: $ 18.7万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2000-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6105693
• 关键词: Escherichia coli; adhesin; gene mutation; hemolysin; host organism interaction; interstitial cystitis; laboratory mouse; molecular cloning; molecular pathology; urinary bladder epithelium; virulence

Bacterial cystitis is overwhelmingly a syndrome of women and the kidney or urologic disease prompting the most visits to physician offices in the US. That the pathogenesis of this disease is poorly understood is ironic. It is the intention of this new project to direct the tools of molecular biology at pertinent E. coli strains in order to examine mechanisms of pathogenesis using human bladder epthelial cells (HBEC) in culture and an in vivo animal model of UTI modified to minimize kidney involvement. The majority of cystitis strains do not have P fimbriae, S fimbriae, hemolysin, and other E. coli characteristics identified as virulence factors for pyelonephritis. Therefore, we have developed an experimental plan that slows the study not only of those genes modestly epidemiologically associated with cystitis but also the identification, by functional roles, of heretofore unknown genes. Preliminary studies demonstrate the E. coli hemolysin, a pore-forming cytotoxin is active against HBEC, that HBEC internalize E. coli, and that mouse UTI virulence is not explained solely by known E. coli virulence factors. We will test the following hypotheses: 1) That strains causing cystitis in humans adhere to HBEC via specific known or as yet unrecognized adhesins. This will be demonstrated by mutagenesis of pertinent strains: isogenic for known adhesins and transposon to identify unknown adhesins. 2) That F54, a potent E. coli cystitis strain with no known virulence factors, causes cystitis in part through internalization into bladder epithelial cells. This will be approached via transposon mutagenesis to seek mutants with diminished internalization and virulence. 3) That the activities of E. coli hemolysin and cytotoxic necrotizing factor (CNF) in the living bacterium can be dissociated by genetic manipulation. Hemolysin is epidemiologically linked to cystitis and CNF is an interesting protein identified in 2/3 of hemolytic strains which enhances bacterial internalization and causes multinucleation of host epithelial cells. Our intent will be to dissociate hemolysin and CNF via isogenic mutagenesis and to study the four permutations of these two putative virulence factors in adherence, internalization, and cytotoxicity assays in order to generate hypotheses of the action and interaction of hemolysin and CNF.

细菌性膀胱炎绝大多数是女性和肾脏或肾脏疾病的综合征。 在美国，泌尿系统疾病导致就诊人数最多。 具有讽刺意味的是，人们对这种疾病的发病机制知之甚少。 它 这个新项目的目的是指导分子工具 对相关大肠杆菌菌株进行生物学研究，以检查其机制 使用培养物中的人膀胱上皮细胞 (HBEC) 的发病机制和 修改尿路感染体内动物模型以尽量减少肾脏受累。 这 大多数膀胱炎菌株没有 P 菌毛、S 菌毛、溶血素、 和其他被确定为毒力因子的大肠杆菌特征 肾盂肾炎。 因此，我们制定了一个实验计划 不仅在流行病学上适度减缓了对这些基因的研究 与膀胱炎相关，但也通过功能作用进行识别， 迄今为止未知的基因。 初步研究表明大肠杆菌溶血素是一种成孔物质 细胞毒素对 HBEC 有活性，HBEC 内化大肠杆菌，并且 小鼠尿路感染的毒力不能仅用已知的大肠杆菌毒力来解释 因素。 我们将测试以下假设： 1) 导致人类膀胱炎的菌株通过特定的途径粘附在 HBEC 上 已知或尚未认识的粘附素。 这将通过以下方式证明 相关菌株的诱变：已知粘附素和 转座子识别未知的粘附素。 2) F54，一种强效大肠杆菌膀胱炎菌株，无已知毒力 因素，部分通过内化到膀胱引起膀胱炎 上皮细胞。 这将通过转座子诱变来实现 寻找内化和毒力减弱的突变体。 3) 表明大肠杆菌溶血素和细胞毒性坏死活性 活细菌中的CNF因子可以通过遗传解离 操纵。 溶血素在流行病学上与膀胱炎有关，而 CNF 是 在 2/3 的溶血菌株中发现了一种有趣的蛋白质 增强细菌内化并导致宿主多核 上皮细胞。 我们的目的是通过溶血素和 CNF 分离 等基因突变并研究这两个的四种排列 粘附、内化和细胞毒性的假定毒力因子 分析以产生作用和相互作用的假设 溶血素和CNF。