探討白色念珠菌引起嗜中性白血球產生細胞外網狀結構之機制

Abstract

白色念珠菌是一種雙形真菌，在一般環境下以酵母菌形態存在，當在人類宿主體內，或是處於37度含有血清的環境時會轉變成菌絲形態。對於正常人而言白色念珠菌是一種共生菌，但當宿主免疫不全時他會轉變為病源並導致黏膜或是全身性的念珠菌感染。過去的研究指出慢性肉芽腫以及嗜中性白血球低下症的病人特別容易產生全身性念珠菌感染疾病，顯示嗜中性白血球在抵禦白色念珠菌感染中扮演著重要的角色。Netrophil extracellular traps是由染色質、顆粒蛋白和細胞質蛋白質構成，嗜中性白血球可以借此來清除病源，在過去的研究顯示白色念珠菌可以引起NETs產生，並且可以被NETs清除。 在本篇研究中，我試圖探討opsonized C. albicans和unopsonized C. albicans感染時嗜中性球分別經由什麼機制引起NETs產生。我分別利用opsonized和unopsonized的白色念珠菌刺激野生型以及Ncf-1 KO的嗜中性白血球。實驗結果顯示opsonized C. albicans是經由需要NADPH oxidase活化以及ROS產生的機制引起NETs產生，而unopsonized C. albicans引起的NETs則不需要NADPH oxidase活化。 借由CR3 knockout老鼠實驗，我發現嗜中性球利用CR3辨識opsonized C. albicans並活化PKC、PI3K、Syk的訊息傳遞路徑引起需要ROS的NETs，而unopsonized C. albicans則是借由Dectin-2所辨識並且活化Syk、PKCδ、P38以及ERK的訊息傳遞來引發NETs產生。因此opsonized 和unopsonized 的C. albicans 可以被不同受體辨識並經由不同的路徑引發NETs的產生。

Candida albicans is a dimorphic fungus. It is a commensal in healthy individuals and becomes an opportunistic pathogen in immunocompromised hosts causing mucosal or systemic candidiasis. Previous studies show that, CGD and neutropenia patients are susceptible to systemic candidiasis showing that neutrophil is important to host defense against C. albicans. Neutrophils kill pathogens by neutrophil extracellular traps (NETs), a web like structure composed of chromatins, granular- and cytosolic proteins. Previous studies showed that C. albicans induce NETs via ROS-dependent as well as -independent mechanisms and NETs kill C. albicans. Here, I investigated the mechanism of NETs formation in response to stimulation by opsonized and unopsonized C. albicans. Neutrophils stimulated with both opsonized and unopsonized C. albicans released NETs. Comparing NETs formation of neutrophils from wild type and Ncf-1 knockout mice in response to opsonized and unopsonized C. albicans, we found that opsonized C. albicans induced-NETs formation was via NADPH oxidase-dependent mechanism but unopsonized C. albicans induced NETs did not require NADPH oxidase mediated-ROS production. Comparing neutrophils from CR3 KO and WT mice, I discovered that neutrophils recognized opsonized C. albicans via CR3. CR3 engagement activated PKC, PI3K, and Syk signaling pathway(s) resulting in NADPH oxidase-dependent NETs production. On the other hand, unopsonized C. albicans is recognized by Dectin-2 and activated Syk, PKCδ, P38 and ERK signaling pathway(s). In conclusion opsonized and unopsonized C. albicans are recognized by different receptor and induce NETs production via different pathways.