中性粒细胞在牛分枝杆菌感染宿主免疫反应中作用机制的研究
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摘要
牛结核病是由牛分枝杆菌感染引起的疾病,对世界养牛业乃至全世界的食品安全和人类健康均产生很大的影响。长久以来的研究都将巨噬细胞作为结核菌感染的主要宿主细胞,但是随着中性粒细胞生理功能的逐步研究明了,其在结核菌感染中的作用也越来越受到人们的重视。中性粒细胞是机体受到感染后最先游走到感染部位的细胞成分,属于机体天然免疫反应的第一道防线,其在对抗胞内菌感染中也起着不可忽视的作用。本研究着眼于中性粒细胞在牛分枝杆菌感染的不同阶段中所发挥作用的不同,研究了在急性感染和慢性感染阶段,即中性粒细胞在机体的抗结核免疫反应中包括固有免疫和获得性免疫阶段所发挥的抗菌防御和致病机理中可能发挥的作用。
1.本研究利用体外感染试验来模拟在牛结核病急性感染的早期阶段,检测了牛中性粒细胞的功能改变如表型变化、凋亡率以及炎性因子表达等。实验数据表明中性粒细胞受到牛分枝杆菌刺激后,发挥吞噬功能并启动抗菌程序,但是并不能完全清除感染,反而在牛分枝杆菌的刺激下能够促使中性粒细胞以坏死的形式走向死亡；自噬可能参与中性粒细胞杀菌的过程。
2.本研究以从小鼠分离的原代的中性粒细胞细胞为模型,检测了经过牛分枝杆菌感染的小鼠中性粒细胞其上清液作为刺激剂对巨噬细胞、树突状细胞、淋巴细胞、Ⅱ型肺泡上皮细胞等已经被证实参与在肺脏微环境下抗结核感染中的细胞成分的激活效应。数据表明,中性粒细胞感染结核菌后的旁分泌效应能够有效的激活巨噬细胞并增强其杀菌能力；在一定程度上促进树突状细胞的激活和成熟；能够增强Ⅱ型肺泡上皮细胞抗炎因子等的分泌活性。
3.本研究对正常牛外周血中性粒细胞体外经牛分枝杆菌感染刺激后以及牛结核病感染牛的外周血中性粒细胞其表达干扰素诱导转录基因以及中性粒细胞的PTX3基因和MPO基因的变化进行了检测。实验数据显示在牛结核病慢性感染过程中,外周血中性粒细胞的干扰素诱导转录基因的表达具有一定的特异性。
Bovine tuberculosis (BTB), a zoonotic disease caused by Mycobacterium bovis(M. bovis) infection, continues to be a major problem in cattle industry and pose a potential threat to food safety and human health worldwide. Interactions between M. bovis and bovine macrophages have been extensively characterized in various studies, while similar analyses in neutrophils, which are one of the other types of white blood cells in mammals, were often overlooked. Neutrophils form the front line of immune defense and are the first inflammatory cells to arrive at sites of infection to eliminate pathogenic bacteria by presenting a diverse collection of antimicrobial molecules. Latest research results imply the participation of neutrophils also in the elimination of intracellular bacterial pathogens. Our reaserch concentrated on investigating the role neutrophils might play in the acute and chronic infection with M. bovis, providing a further understanding of their exact participation in host anti-BTB immunity and tuberculosis progression.
1. The study was aimed to investigate in vitro modulation of neutrophils functions including phenotypic changes, apoptosis rate, and inflammatory cytokines expression after infection with M. bovis, micking the acute early infection phase. It was demonstrated that phagocytosis of M. bovis activated and enhanced bovine neutrophils functions as well as initialed their defense mechanism, but failed to eliminate the mycobacteria. Moreover, autophagy might get involved in the defense infection process functioning as a protective mechanism.
2. The supernatant of isolated primary mouse neutrophils infected with M. bovis was used as stimuli to challenge macrophages cells, dendritic cells, lymphocytes, type II alveolar epithelial cells, which have been shown to participate in the effect of anti-tuberculosis infection in the lung micro-environment. Data indicated that neutrophil paracrine effect after M. bovis infection could effectively activate macrophages and enhance the bactericidal activity of macrophages, promote activation and dendritic cell maturation to a certain degree. Meanwile, Type II alveolar epithelial cells display enhanced activity of secreting cytokines..
3. The expression levels of ten IFN-inducible transcriptional genes in neutrophils from healthy cattle stimulated by M. bovis and neutrophils isolated from three groups of cattle of different infection status was assessed. Additionnally, the changes in the expression of myeloperoxidase and pentraxin-related protein pentraxin-inducible protein genes were also tested during bovine tuberculosis infection. Our results demonstrated a specific expression pattern of IFN-inducible transcriptional genes and MPO and PTX3genes in neutrophils during bovine tuberculosis infection.
1.本研究利用体外感染试验来模拟在牛结核病急性感染的早期阶段,检测了牛中性粒细胞的功能改变如表型变化、凋亡率以及炎性因子表达等。实验数据表明中性粒细胞受到牛分枝杆菌刺激后,发挥吞噬功能并启动抗菌程序,但是并不能完全清除感染,反而在牛分枝杆菌的刺激下能够促使中性粒细胞以坏死的形式走向死亡；自噬可能参与中性粒细胞杀菌的过程。
2.本研究以从小鼠分离的原代的中性粒细胞细胞为模型,检测了经过牛分枝杆菌感染的小鼠中性粒细胞其上清液作为刺激剂对巨噬细胞、树突状细胞、淋巴细胞、Ⅱ型肺泡上皮细胞等已经被证实参与在肺脏微环境下抗结核感染中的细胞成分的激活效应。数据表明,中性粒细胞感染结核菌后的旁分泌效应能够有效的激活巨噬细胞并增强其杀菌能力；在一定程度上促进树突状细胞的激活和成熟；能够增强Ⅱ型肺泡上皮细胞抗炎因子等的分泌活性。
3.本研究对正常牛外周血中性粒细胞体外经牛分枝杆菌感染刺激后以及牛结核病感染牛的外周血中性粒细胞其表达干扰素诱导转录基因以及中性粒细胞的PTX3基因和MPO基因的变化进行了检测。实验数据显示在牛结核病慢性感染过程中,外周血中性粒细胞的干扰素诱导转录基因的表达具有一定的特异性。
Bovine tuberculosis (BTB), a zoonotic disease caused by Mycobacterium bovis(M. bovis) infection, continues to be a major problem in cattle industry and pose a potential threat to food safety and human health worldwide. Interactions between M. bovis and bovine macrophages have been extensively characterized in various studies, while similar analyses in neutrophils, which are one of the other types of white blood cells in mammals, were often overlooked. Neutrophils form the front line of immune defense and are the first inflammatory cells to arrive at sites of infection to eliminate pathogenic bacteria by presenting a diverse collection of antimicrobial molecules. Latest research results imply the participation of neutrophils also in the elimination of intracellular bacterial pathogens. Our reaserch concentrated on investigating the role neutrophils might play in the acute and chronic infection with M. bovis, providing a further understanding of their exact participation in host anti-BTB immunity and tuberculosis progression.
1. The study was aimed to investigate in vitro modulation of neutrophils functions including phenotypic changes, apoptosis rate, and inflammatory cytokines expression after infection with M. bovis, micking the acute early infection phase. It was demonstrated that phagocytosis of M. bovis activated and enhanced bovine neutrophils functions as well as initialed their defense mechanism, but failed to eliminate the mycobacteria. Moreover, autophagy might get involved in the defense infection process functioning as a protective mechanism.
2. The supernatant of isolated primary mouse neutrophils infected with M. bovis was used as stimuli to challenge macrophages cells, dendritic cells, lymphocytes, type II alveolar epithelial cells, which have been shown to participate in the effect of anti-tuberculosis infection in the lung micro-environment. Data indicated that neutrophil paracrine effect after M. bovis infection could effectively activate macrophages and enhance the bactericidal activity of macrophages, promote activation and dendritic cell maturation to a certain degree. Meanwile, Type II alveolar epithelial cells display enhanced activity of secreting cytokines..
3. The expression levels of ten IFN-inducible transcriptional genes in neutrophils from healthy cattle stimulated by M. bovis and neutrophils isolated from three groups of cattle of different infection status was assessed. Additionnally, the changes in the expression of myeloperoxidase and pentraxin-related protein pentraxin-inducible protein genes were also tested during bovine tuberculosis infection. Our results demonstrated a specific expression pattern of IFN-inducible transcriptional genes and MPO and PTX3genes in neutrophils during bovine tuberculosis infection.
引文
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