AcCystatin在广州管圆线虫感染中的免疫调节作用及其分子机制

The rat lungworm Angiostrongylus cantonensis, a food-borne nematode parasite, is the primary cause of human eosinophilic meningitis, meningoencephalitis and other neurological disorders in many tropical and subtropical regions (Southeast Asia, China and Pacific Basin, etc). Rats serve as the definitive host of the nematode, while human, mice and other non-permissive hosts infected with A. cantonensis will cause eosinophilic meningitis or meningoencephalitis and the parasites will terminate at fifth-stage larvae (L5) in the brain of the hosts. The difference of consequences between infected permissive hosts (rodents) and non-permissive hosts (humans and mice) largely results from the different immune response of host-parasite during A. cantonensis invasion and development. Our previous research data indicated firstly that AcCystatin，an identified immunoregulatory molecule from A. cantonensis, was expressed in a significantly higher level in the fourth- and fifth-stage larvae when compared to that in the third-stage larvae and adults. Interestingly, AcCystatin could augment different Th1/Th2 polarized immune responses to A. cantonesis infection in mice and rats, suggesting a distinct characteristic from cystatins of other parasitic parasites. Further results revealed AcCystatin activated TNF-a production via ERK1/2,PI3K/Akt and NF-κB signaling pathways in mice. Based on our findings, the biotechnics including RNAi, Dual-Luciferase Reporter Array, knock-out animal,microarray chip, point mutation and fixed microinjection were proposed to investigate the molecular mechanism of AcCystatin's augmentation of Th1/Th2 polarized immune responses to A. cantonesis infection in permissive and non-permissive hosts. Our proposed study will contribute greatly to the development of strategies to reduce the neurological damages caused by this parasite.

广州管圆线虫病是严重损伤中枢神经系统的新发食源性寄生虫病。阐明该虫感染所致嗜酸性粒细胞增多性CNS炎症（小鼠、人）或无明显损伤（大鼠）不同病理结局的分子机制仍是广州管圆线虫病防治研究亟需回答的核心科学问题。我们首次发现：免疫调节分子AcCystatin在侵入宿主CNS的虫期显著高表达，并可诱导大、小鼠Th1/Th2免疫应答极化方向及脑部炎性浸润程度迥异，其作用机理与ERK1/2、PI3K/Akt和NF-κB通路相关。本项目拟进一步采用RNAi、基因敲除动物、点突变、芯片检测、显微定位注射等技术系统研究AcCystatin诱导广州管圆线虫宿主不同Th1/Th2免疫应答极化方向的信号通路，全面阐述AcCystatin在广州管圆线虫感染中的免疫调节作用分子机制，为确定AcCystatin为广州管圆线虫病防治的新靶标提供科学依据。

广州管圆线虫病是严重损伤中枢神经系统的新发食源性寄生虫病。阐明该虫感染所致嗜酸性粒细胞增多性CNS炎症（小鼠、人）或无明显损伤（大鼠）不同病理结局的分子机制仍是广州管圆线虫病防治研究亟需回答的核心科学问题。本项目综合利用RNAi、多信号通路报告基因芯片、基因敲除动物、显微定位注射等技术系统研究AcCystatin诱导广州管圆线虫适宜、非适宜宿主不同Th1/Th2免疫应答极化方向的分子机制。研究结果显示：AcCystatin可诱导广州管圆线虫非适宜宿主小鼠巨噬细胞TNF-α显著上调，而对适宜宿主大鼠巨噬细胞TNF-α水平变化无明显影响；亚细胞定位分析首次证实AcCystatin主要定位于巨噬细胞线粒体和高尔基体；确定CARD9是AcCystatin在巨噬细胞RAW264.7胞内的相互作用蛋白。AcCystatin可通过结合CARD9激活下游NF-κB通路，促使转录因子NF-κB入核结合TNF启动子上的相关位点启动TNF表达，进一步阐释了AcCystatin诱导巨噬细胞TNF-α上调的分子机制；第一次发现AcCystatin可显著减轻OVA/氢氧化铝诱导哮喘大鼠肺脏和气道免疫病理损伤。这些研究结果不仅深入AcCystatin诱导广州管圆线虫适宜、非适宜宿主不同Th1/Th2免疫应答极化方向的分子机制，为确定AcCystatin为广州管圆线虫病防治的新靶标提供科学依据，同时首次证实AcCystatin具有免疫治疗哮喘的潜在开发价值。