Production of extracellular traps against Aspergillus fumigatus in vitro and in infected lung tissue is dependent on invading neutrophils and influenced by hydrophobin RodA.
Production of extracellular traps against Aspergillus fumigatus in vitro and in infected lung tissue is dependent on invading neutrophils and influenced by hydrophobin RodA.
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DOI：
10.1371/journal.ppat.1000873
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发表时间：
2010-04-29
期刊：
影响因子：
6.7
通讯作者：
Gunzer M
中科院分区：
文献类型：
--
作者：
Bruns S;Kniemeyer O;Hasenberg M;Aimanianda V;Nietzsche S;Thywissen A;Jeron A;Latgé JP;Brakhage AA;Gunzer M
烟曲霉菌是最重要的空气传播真菌病原体，在免疫功能低下的患者中会引起危及生命的感染。已知巨噬细胞和中性粒细胞可杀死分生孢子，而菌丝主要由中性粒细胞杀死。由于菌丝太大而不能被吞噬，中性粒细胞具有一系列的细胞外杀伤机制，包括形成中性粒细胞胞外陷阱(Net)，该陷阱由带有杀菌蛋白的核DNA组成。然而，到目前为止，对烟曲霉菌的净形成反应只在体外被证明，中性粒细胞在体内产生的重要性还不清楚，这种真菌防御净形成的分子机制也是未知的。在这里，我们展示了人类中性粒细胞在体外遇到烟曲霉菌时产生Net。在延时电影中，网络制作是一个高度动态的过程，然而，只有一小部分细胞表现出这种过程。对菌丝的侵染作用最强，对休眠和膨大的分生孢子的侵染作用减弱。在一个新开发的小鼠模型中，我们可以通过曲霉菌感染的肺部的双光子显微镜来证明存在并测量体内网状结构的形成动力学。我们还观察了中性粒细胞在肺内的巨大动态，以及它们与真菌元素相互作用和吞噬真菌元素的能力。此外，小鼠全身中性粒细胞的耗尽几乎完全抑制了肺部的净形成，从而直接将中性粒细胞的迁移与体内的净形成联系起来。通过使用真菌突变体和纯化的蛋白质，我们证明了疏水蛋白RodA，一种使分生孢子免疫惰性的表面蛋白，导致中性粒细胞遇到曲霉真菌成分时净形成减少。对曲霉菌丝的净依赖杀灭可在以后的时间点表现出来，但仅是温和的。因此，这些数据证实了在寄主对烟曲霉菌的防御过程中，体内发生了网的形成，但表明它在杀死这种真菌方面并不起主要作用。相反，蚊帐可能具有抑菌作用，并可能防止进一步传播。真菌烟曲霉生长在腐烂的有机物上，产生大量的孢子，称为分生孢子，人类不断地吸入这些孢子。这是无害的，因为我们有一个名为中性粒细胞的免疫细胞的正常防御系统，但中性粒细胞太少或没有功能的人可能会死于曲霉感染。中性粒细胞侵入肺部，吞噬/吞噬，从而杀死分生孢子。濒临死亡的中性粒细胞也可以将他们的核DNA扔到菌丝分子上，比如装饰有抗菌蛋白的Net(中性粒细胞胞外陷阱)。因此，更大数量的真菌，包括侵入组织的菌丝，仍然可以控制。然而，直到今天，在曲霉感染的肺中还没有证明Net的形成，中性粒细胞在这一过程中的作用尚不清楚，这种真菌是否有自身的抗网防御策略也不清楚。我们在这里证明了在曲霉菌感染的肺中Net的存在，表明中性粒细胞产生这些结构，并且它们吞噬肺组织中的真菌元素。此外，我们发现曲霉通过表面蛋白疏水蛋白RodA来伪装其孢子，这能够强烈地阻止中性粒细胞的网络形成。这些研究为这一关键的寄主-病原菌相互作用过程的动力学和分子机制提供了新的线索。虽然这些数据证实了在寄主对烟曲霉菌的防御过程中，在体内发生了网状形成，但我们认为网状形成在杀死这种真菌方面并不起主要作用。
Aspergillus fumigatus is the most important airborne fungal pathogen causing life-threatening infections in immunocompromised patients. Macrophages and neutrophils are known to kill conidia, whereas hyphae are killed mainly by neutrophils. Since hyphae are too large to be engulfed, neutrophils possess an array of extracellular killing mechanisms including the formation of neutrophil extracellular traps (NETs) consisting of nuclear DNA decorated with fungicidal proteins. However, until now NET formation in response to A. fumigatus has only been demonstrated in vitro, the importance of neutrophils for their production in vivo is unclear and the molecular mechanisms of the fungus to defend against NET formation are unknown. Here, we show that human neutrophils produce NETs in vitro when encountering A. fumigatus. In time-lapse movies NET production was a highly dynamic process which, however, was only exhibited by a sub-population of cells. NETosis was maximal against hyphae, but reduced against resting and swollen conidia. In a newly developed mouse model we could then demonstrate the existence and measure the kinetics of NET formation in vivo by 2-photon microscopy of Aspergillus-infected lungs. We also observed the enormous dynamics of neutrophils within the lung and their ability to interact with and phagocytose fungal elements in situ. Furthermore, systemic neutrophil depletion in mice almost completely inhibited NET formation in lungs, thus directly linking the immigration of neutrophils with NET formation in vivo. By using fungal mutants and purified proteins we demonstrate that hydrophobin RodA, a surface protein making conidia immunologically inert, led to reduced NET formation of neutrophils encountering Aspergillus fungal elements. NET-dependent killing of Aspergillus-hyphae could be demonstrated at later time-points, but was only moderate. Thus, these data establish that NET formation occurs in vivo during host defence against A. fumigatus, but suggest that it does not play a major role in killing this fungus. Instead, NETs may have a fungistatic effect and may prevent further spreading. The fungus Aspergillus fumigatus grows on decaying organic matter and produces large numbers of spores, called conidia, which are constantly inhaled by humans. This is harmless, because we have a functioning defence system of immune cells called neutrophil granulocytes, but people with too few or non-functioning neutrophils can die of Aspergillus infections. Neutrophils invade the lung, engulf/phagocytose and thereby kill conidia. Dying neutrophils can also throw their nuclear DNA on hyphal elements as NETs (Neutrophil Extracellular Traps) that are decorated with antimicrobial proteins. Thus, larger fungal amounts, including tissue-invading hyphae, can still be controlled. However, until today the formation of NETs has not been demonstrated in Aspergillus-infected lungs, the role of neutrophils for this process was unknown and whether the fungus has anti-NET defence strategies on its own was not clear. We demonstrate here the existence of NETs in Aspergillus-infected lungs, show that neutrophils produce these structures and that they phagocytose fungal elements within the lung tissue. Furthermore, we show that Aspergillus camouflages its spores by means of the surface protein hydrophobin RodA, which is able to strongly prevent NET formation by neutrophils. These studies shed new light on the dynamics and molecular mechanisms of this key process of host-pathogen interaction. Although these data establish that NET formation occurs in vivo during host defence against A. fumigatus, we suggest that NET formation does not play a major role in killing this fungus.
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DOI：
10.1086/600380
发表时间：
2009-08-15
期刊：
The Journal of infectious diseases
影响因子：
--
作者：
Mircescu MM;Lipuma L;van Rooijen N;Pamer EG;Hohl TM
通讯作者：
Hohl TM
感染期间淋巴结中中性粒细胞迁移的动态。
影响因子：
32.4
作者：
Chtanova, Tatyana;Schaeffer, Marie;Han, Seong-Ji;van Dooren, Giel G.;Nollmann, Marcelo;Herzmark, Paul;Chan, Shiao Wei;Satija, Harshita;Camfield, Kristin;Aaron, Holly;Striepen, Boris;Robey, Ellen A.
通讯作者：
Robey, Ellen A.
脊髓细胞细胞募集会导致急性病毒性脑膜炎期间致命的中枢神经系统损伤。
影响因子：
64.8
作者：
Kim, Jiyun V.;Kang, Silvia S.;Dustin, Michael L.;McGavern, Dorian B.
通讯作者：
McGavern, Dorian B.
影响因子：
30.5
作者：
Buerckstuemmer, Tilmann;Baumann, Christoph;Superti-Furga, Giulio
通讯作者：
Superti-Furga, Giulio
影响因子：
3.4
作者：
Dague, Etienne;Alsteens, David;Dufrene, Yves F.
通讯作者：
Dufrene, Yves F.