探討細菌與宿主交互作用在金黃色葡萄球菌引起的感染性心內膜炎致病機轉中扮演的角

Role of bacteria-host interactions in the pathogenesis of Staphylococcus aureus-induced infective endocarditis

許智傑(Chih-Chieh Hsu)

指導教授 ： 賈景山

共同指導教授 ： 鍾筱菁(Chiau-Jing Jung)

國立臺灣大學/醫學院/口腔生物科學研究所/博士(2021年)

https://doi.org/10.6342/NTU202100937

摘要

急性感染性心內膜炎，主要是由金黃色葡萄球菌(Staphylococcus aureus)及表皮 葡萄球菌(Staphylococcus epidermidis)黏附於心臟瓣膜上所引起的感染性疾病，具有 高致死率及再發性。此疾病的病徵是在心臟瓣膜上會有細菌與敗血性血栓所組成 的贅疣(vegetation)，面對這種贅疣的治療必須持續性的給予抗生素，但效果有限。 當細菌進入血液循環後，能經由感應環境變化並快速的重組蛋白質，並且與宿主因 子作用例如:血小板，以利細菌躲避宿主免疫監控系統、形成生物膜及引發心內膜 炎。利用大鼠心內膜炎模式，我們證明了金黃色葡萄球菌誘導的贅生物中，嗜中性 球胞外結構(neutrophils extracellular traps, NETs)會包覆著細菌及血小板的團塊。給 予去氧核醣核酸酶(DNase I)能顯著的減少甲氧西林具有抗性(MRSA)及不具有抗性 的金黃色葡萄球菌(MSSA)引起贅疣形成，這個結果顯示 NETs 在金黃色葡萄球菌 引起的心內膜炎中扮演關鍵角色。有趣的是，在體外及活體實驗證明，在有血小板 的條件下 MRSA 和 MSSA 會表現不同的生物膜型態。為了進一步鑑定出會與這些 宿主因子交互作用的細菌調控系統，我們分析了十五套雙分子調控系統。結果顯示 arlSR 和 vraSR 的缺失，降低了細菌抵抗吞噬作用、誘導 NET 釋放及引發贅疣形 成的能力。我們利用核糖核酸定序鑑定出 VraSR 所調控的下游蛋白 PrsA，此蛋白 質在細菌抵抗免疫球蛋白介導的吞噬作用扮演重要角色，並且透過調節葡萄球菌 蛋白質 A (Spa)的表現誘導 NETs。另外，arlSR 缺失的菌株降低分選酶 (sortase A) 的表現，這也影響了錨定在細胞壁上 Spa。此外，ArlSR 在細菌引起的血小板凝集、 血小板介導的生物膜形成和誘導 NETs 中扮演重要角色。綜合以上，金黃色葡萄球 菌透過與血小板的交互作用誘導 NET 的形成促進感染性心內膜炎的發生，而 ArlSR 及 VraSR 這兩套調控系統參與了細菌與宿主因子交互作用。針對細菌與宿主因子 交互作用(例如:DNase I)，可為臨床上控制急性感染性心內膜炎提供新的策略。

關鍵字

急性感染性心內膜炎 ； 金黃色葡萄球菌 ； 噬中性球胞外結構 ； 血小板 ； 雙分子調控系統

並列摘要

Acute infective endocarditis (IE) is primarily caused by Staphylococci (S. aureus or S. epidermidis) on the heart valves, with high mortality and recurrent rates. IE is characterized by the formation of vegetation, septic thrombus with the embedded bacteria forming biofilm, which is refractory to routine antibiotic treatment. When entering into the circulations, bacteria can sense the environmental changes and rapidly remodel protein expression to interact with host factors, such as platelets, for escaping immune surveillance or forming biofilm on the heart valve in IE. Using an experimental endocarditis rat model, we demonstrated that S. aureus-induced vegetation was composed of bacterial floes encased in aggregated platelets and surrounded by neutrophil extracellular traps (NETs). Interestingly, we also found S. aureus methicillin resistant (MRSA) and sensitive (MSSA) strains express different biofilm phenotypes in the presence of platelets in vitro and in vivo. Administration of DNase I significantly reduced the size of vegetation induced by MRSA or MSSA isolates, suggesting the crucial role of NETs in S. aureus-induced vegetation formation. In order to identify the bacterial regulatory systems involved in S. aureus-host interactions, fifteen isogenic mutant strains in S. aureus two component systems (TCSs) were investigated. The arISR and vraSR- deficient mutant strains decreased abilities to resist phagocytosis, to induce NET release, and to cause bacterial vegetation formation. By RNA-sequencing analysis, we identified a VraSR-downstream protein, PrsA, which plays roles in bacterial resistance to immunoglobulin-dependent phagocytosis and to induce NETs by modulating the expression of staphylococcal protein A (Spa). In addition, deletion of arlSR reduced the expression of sortase A, which affects the Spa anchoring on bacterial cell wall. Meanwhile, ArlSR also plays roles in bacterial platelet aggregation, platelet-dependent biofilm formation, and NET induction. Taken together, S. aureus interact with platelets to induce NET formation, contributing to vegetation formation in IE, which are at least partly modulated by the bacterial TCSs, ArlSR and VraSR. Targeting to the interaction of S. aureus and host factors, such as DNase I, may provide new strategies for the clinical control of S. aureus-induced acute IE.
Keywords: Acute infective endocarditis, Staphylococcus aureus, neutrophil extracellular traps, platelets, two-component regulatory system

並列關鍵字

Acute infective endocarditis ； Staphylococcus aureus ； neutrophil extracellular traps ； platelets ； two-component regulatory system

參考文獻

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