de Almeida Lucia, Dorfleutner Andrea, Stehlik Christian
Division of Rheumatology, Department of Medicine, Feinberg School of Medicine, Northwestern University, Chicago, USA.
Bio Protoc. 2016 Oct 5;6(19). doi: 10.21769/BioProtoc.1944.
In response to pathogen infection and tissue damage, inflammasome sensors such as NLRP3 and AIM2 are activated, which triggers PYRIN domain (PYD)-mediated ASC nucleation, followed by self-perpetuating ASC polymerization, which ultimately culminates in caspase-1 activation, interleukin (IL)-1β and IL-18 processing and release and pyroptosis (Ratsimandresy ., 2013; Cai ., 2014). Inflammasomes release not only cytokines, but also the polymeric ASC danger particles (pASC) by pyroptosis, which perpetuate and propagate inflammasome responses to bystander cells to engage cell intrinsic ASC and caspase-1 (Baroja-Mazo ., 2014; Franklin ., 2014). In this protocol we describe intraperitoneal injection of polymeric ASC particles as a danger signal and measure neutrophil infiltration and levels of the pro-inflammatory cytokine IL-1β by ELISA in the peritoneal lavage (de Almeida ., 2015).
作为对病原体感染和组织损伤的反应，诸如NLRP3和AIM2等炎性小体传感器被激活，这会触发含PYRIN结构域（PYD）介导的凋亡相关斑点样蛋白（ASC）成核，随后ASC进行自我延续的聚合，最终导致半胱天冬酶-1激活、白细胞介素（IL）-1β和IL-18的加工与释放以及细胞焦亡（拉齐曼德雷西，2013年；蔡，2014年）。炎性小体不仅通过细胞焦亡释放细胞因子，还会释放聚合的ASC危险颗粒（pASC），这些颗粒使炎性小体对旁观者细胞的反应持续并扩散，从而激活细胞内源性ASC和半胱天冬酶-1（巴罗亚-马佐，2014年；富兰克林，2014年）。在本实验方案中，我们描述了将聚合的ASC颗粒腹腔注射作为一种危险信号，并通过酶联免疫吸附测定法（ELISA）测量腹腔灌洗中嗜中性粒细胞浸润和促炎细胞因子IL-1β的水平（德阿尔梅达，2015年）。
