Cytotoxic Molecules in Invertebrate Immunity

昆虫和其他节肢动物是许多具有医学和兽医重要性的寄生虫病的媒介，包括疟疾、锥虫病（昏睡病、恰加斯病）、丝虫病（蠕虫、象皮病）和犬心丝虫病。 了解昆虫媒介中调节感染过程的因素对于制定保护人类和牲畜免受昆虫媒介寄生虫不利影响的对策至关重要。昆虫具有有效的先天细胞和体液免疫机制，这些机制与哺乳动物的先天免疫反应有着惊人的相似之处，这表明这些至关重要的过程有着共同的祖先。 除了记录和表征几种昆虫物种的各种免疫效应器应答的研究外，对初始非自我识别应答的潜在机制或免疫激活细胞使用的杀伤分子的身份和作用模式知之甚少或一无所知。 拟议的研究旨在阐明昆虫产生的细胞毒性成分的性质，这些成分专门针对并摧毁其内部寄生虫。研究人员计划研究果蝇Drosophila melanogaster对后生动物寄生虫Leptopilina boulardi的细胞防御反应，以及reduviid bug Rhodnius prolixus对原生动物寄生虫Trypanosoma的细胞毒性反应。 使用这两种昆虫-寄生虫模型系统，将监测各种活性氧中间体（超氧阴离子、过氧化氢和羟基自由基）的产生，以及活性氮中间体（例如，一氧化氮和过氧亚硝酸盐），以及两种重要酶NADPH氧化酶和一氧化氮合酶的活性水平。 为了研究果蝇和Rhodnius对各自的后生动物和原生动物寄生虫引起的免疫反应期间发生的生物化学和酶活性的变化，研究人员将采用高效液相色谱法，电化学检测，分光光度法，荧光测定法和分子技术。 这种全面的方法，积极确定特定的细胞毒性分子和酶的机制，它们是在昆虫免疫反应过程中产生的，将是非常有用的，提供重要的见解细胞信号和免疫识别昆虫。

Insects and other arthropods are vectors of numerous parasitic diseases of medical and veterinary importance, including malaria, trypanosomiasis (sleeping sickness, Chagas' disease) filariasis (eye worms, elephantiasis), and dog heartworm. Knowledge of the factors that regulate the infection process in the insect vector is essential for the development of counter strategies to protect humans and livestock from the adverse effects of insect-vectored parasites. Insects possess effective innate cellular and humoral immune mechanisms that exhibit striking parallels between the innate immune responses of mammals, suggesting a common ancestry for these critically important processes. With the exception of studies documenting and characterizing various immune effector responses for several insect species, little or nothing is known of the mechanisms underlying the initial non-self recognition response, or the identity and mode of action of killing molecules used by immune activated cells. The proposed research is designed to elucidate the nature of the cytotoxic components generated by insects that specifically target and destroy their internal parasites. The researchers plan to study the cellular defence responses made by the fruit fly Drosophila melanogaster against the metazoan parasite Leptopilina boulardi, and the cytotoxic responses of the reduviid bug Rhodnius prolixus against the protozoan parasite Trypanosoma. Using these two insect-parasite model systems, the production of various reactive oxygen intermediates (superoxide anion, hydrogen peroxide, and the hydroxyl radical) will be monitored, as will the production of reactive nitrogen intermediates (e.g., nitric oxide and peroxynitrite), and the activity levels of two important enzymes, NADPH oxidase and nitric oxide synthase. To investigate the changes in biochemistry and enzyme activity that occur during the immune responses elicited by Drosophila and Rhodnius against their respective metazoan and protozoan parasites, the investigators will employ high performance liquid chromatography with electrochemical detection, spectrophotometry, fluorometry and molecular techniques. This comprehensive approach for positively identifying specific cytotoxic molecules and the enzymatic mechanisms by which they are produced during insect immune responses will be extremely useful by providing important insights concerning cell-cell signaling and immune recognition in insects.