UNDERSTANDING MOLECULAR AND PHENOTYPIC EFFECTS OF CYCLO-OXYGENASE IN ZEBRAFISH

本研究旨在阐明考克斯抑制剂对斑马鱼免疫和胃肠系统的影响，并探讨两者之间的相互作用。使用不同的非甾体抗炎药，在他们的能力，以抑制这两种考克斯酶，学生将寻求分子和表型的签名，定义在斑马鱼幼虫的行动。将特别注意免疫细胞的激活和迁移的动力学及其与组织损伤表现的关系。将使用健康鱼以及发生炎症反应的鱼进行COX抑制剂暴露。后者将通过用脂多糖（LPS）（革兰氏阴性菌外膜的主要组分）预处理来触发。在健康与“发炎”鱼的药物诱导的反应的比较将有助于阐明生物体的生理状态在确定效果类型和幅度所发挥的作用。对免疫和胃肠道系统的影响将使用不同的成像，流式细胞术和分子技术进行量化。斑马鱼是炎症生物学的一个很好的模型，因为它的生物学的许多方面与人类系统非常相似。由于COX介导的途径的调节与人类、动物和环境健康的高度相关性，斑马鱼的使用也将最大限度地提高该项目在各个领域产生的数据的转化价值。这种高转化潜力完全符合迅速出现的“一个健康”愿景，该愿景认识到人类与环境健康之间不可分割的联系。非甾体抗炎药（NSAID）每天被全世界数百万人用于治疗各种各样的疾病，包括疼痛、发热和炎症。这些化合物通过抑制催化前列腺素类合成的酶环加氧酶（COX 1、COX 2）的一种或两种同工型起作用。在健康的人类中，低水平的前列腺素类发挥重要的稳态功能;然而，它们的生物合成在炎症期间急剧增加，导致疼痛和体温调节不良等。因此，通过考克斯抑制剂抑制前列腺素类生物合成是疼痛和发热管理中的高效治疗策略。然而，NSAID的长期治疗性使用可对胃肠道（例如消化不良、胃溃疡、出血）以及肾、肝和心血管系统产生不利影响。已分配了大量工作来了解NSAID的毒性机制并减轻这些风险。例如，许多NSAID调节COX 1和COX 2。已经假设，尽管胃肠道中COX 2的抑制是有益的，但是COX 1的抑制显著地有助于药物诱导的毒性。这种机制的观点代表了选择性COX 2抑制剂的发展，最大限度地减少胃肠道毒性的风险的基本原理。尽管在理解NSAID药理学和毒理学方面取得了很大进展，但考克斯抑制剂的作用机制及其生物学意义的许多方面仍有待阐明。了解这些机制的重要性不仅限于人类健康，而且对环境健康和保护动物健康也至关重要。

The aim of this project is to elucidate the effects of COX inhibitors on immune and gastro-intestinal system of zebrafish and to explore the interplay between the two responses. Using NSAIDs that differ in their ability to inhibit the two COX enzymes, the student will seek molecular and phenotypic signatures that define action in zebrafish larvae. Particular attention will be given to the dynamics of activation and migration of immune cells and their relationship with the manifestation of tissue damage.Exposure to COX-inhibitors will be carried out using healthy fish as well as fish undergoing inflammatory responses. The latter will be triggered by pre-treatment with lipopolysaccharide (LPS), a major component of the outer membrane of Gram-negative bacteria. The comparison of the drug-induced responses in healthy versus "inflamed" fish will help to elucidate the role played by the physiological state of the organism in determining effect type and magnitude. The effects on immune and gastro-intestinal systems will be quantified using a diverse set of imaging, flow cytometry, and molecular techniques. Zebrafish is an excellent model for inflammation biology as many aspects of its biology closely resemble the human system. Due to the high relevance of the modulation of COX-mediated pathways for both human, animal and environmental health, the use of zebrafish will also maximise the translational values of the data generated in this project across fields. This high translational potential is fully in line with the rapidly emerging "One Health" vision, which recognizes the inextricable link between human and environmental health. Project background Non-steroidal anti-inflammatory drugs (NSAIDs) are used by millions of people worldwide every day to treat a wide variety of conditions that involve pain, fever and inflammation. These compounds act by inhibiting one or both isoforms of the enzyme cyclo-oxygenase (COX1, COX2), which catalyse the synthesis of prostanoids. In healthy humans, low levels of prostanoids play important homeostatic functions; however, their biosynthesis increases dramatically during inflammation, leading, among other things, to pain and poor thermoregulation. The inhibition of prostanoids biosynthesis via COX inhibitors is therefore a highly effective therapeutic strategy in pain and fever management. However, long-term therapeutic use of NSAIDs can adversely affect the gastro-intestinal tract (e.g. dyspepsia, gastric ulcer, bleeding), but also kidney, liver and cardiovascular system. Significant efforts have been allocated to understand the mechanisms of toxicity of NSAIDs and to mitigate these risks. For example, many NSAIDs modulate both COX1 and COX2. It has been hypothesised that whereas the inhibition of COX2 in the gastrointestinal tract is beneficial, the inhibition of COX1 contributes significantly to the drug-induced toxicity. This mechanistic perspective represented the rationale for the development of selective-COX2 inhibitors that minimise the risk of gastro-intestinal toxicity. Despite the great advancements achieved in the understanding of NSAIDs pharmacology and toxicology, many aspects of the mechanisms of action of COX inhibitors and of their biological significance still remain to be elucidated. The importance of understanding these mechanisms is not only limited to human health, but it has also critical importance for environmental health and for the protection of animal health.