Staphylococcus aureus induced itch and neuro-immune signaling in skin infections

金黄色葡萄球菌在皮肤感染中引起瘙痒和神经免疫信号传导

• 批准号: 10707178
• 负责人: Isaac Ming-Cheng Chiu
• 金额: $ 73.54万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-19 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10707178
• 关键词: Ablation; Afferent Neurons; Affinity; Atopic Dermatitis; Automobile Driving; Behavior; Binding; Biochemical; Biological; Biological Assay; Calcium; Cells; Cheek structure; Complement; Data; Detection; Disease; Enzyme-Linked Immunosorbent Assay; Esthesia; Family; G-Protein-Coupled Receptors; Genetic; Goals; Human; Image; Immune; Immune response; Immunologics; Impetigo; Infection; Infectious Skin Diseases; Inflammation; Inflammation Mediators; Lesion; Link; Measures; Mediating; Mediator; Microbe; Modeling; Molecular; Molecular Target; Mus; Nerve; Nervous System; Neurobiology; Neuroimmune; Neurons; PAR-1 Receptor; Pain; Pathogenesis; Pathology; Patients; Peptide Hydrolases; Play; Production; Proteinase-Activated Receptors; Proteomics; Pruritus; Recombinants; Role; Serine Protease; Signal Transduction; Site; Skin; Small Interfering RNA; Spinal Ganglia; Staphylococcus aureus; Staphylococcus aureus infection; Symptoms; TRPV1 gene; Therapeutic; Touch sensation; Virulence Factors; Work; antagonist; chronic itch; cytokine; extracellular; genetic approach; glutamyl endopeptidase; host-microbe interactions; immunoregulation; intradermal injection; knock-down; luminescence; microbial; mutant; neural; neurobehavioral; neurotransmission; neutrophil; novel; pain behavior; pathogenic bacteria; pharmacologic; proteinase In; receptor; response; skills; skin barrier; skin damage; skin disorder; skin lesion; subcutaneous

PROJECT SUMMARY Itch is an unpleasant sensation that evokes a desire to scratch. While itch accompanies many skin infections, a causative role for microbes in itch has not been previously investigated. Itch-induced scratching can contribute to significant skin damage and bacterial pathogenesis. Here, we investigate the role of the human bacterial pathogen Staphylococcus aureus and its secreted proteases in driving itch and inflammation. S. aureus colonizes 90% of skin lesions caused by Atopic Dermatitis, a disease characterized by chronic itch. S. aureus also causes impetigo, a contagious skin disease characterized by itchy lesions. We hypothesize that S. aureus secretes proteases that can directly act on host sensory neurons to drive itch, scratch-induced skin damage, and neuroimmune crosstalk. Our preliminary data shows that S. aureus epicutaneous infection of mice induces robust itch behaviors (alloknesis, spontaneous itch) and resulting skin pathology. Using isogenic mutant strains, we find that secreted S. aureus proteases, and in particular the serine protease V8 (SspA) is required for itch production during infection. In Specific Aim 1, we will determine the role of S. aureus V8 protease in driving itch, neuronal activation, and skin inflammation. We will utilize isogenic S. aureus mutant and complemented strains for V8, as well as recombinant V8 to elucidate the necessity and sufficiency of this protease in inducing itch, scratch induced damage, and inflammation caused by S. aureus. Itch is mediated by dorsal root ganglia (DRG) sensory neurons. DRG neuron calcium imaging will be performed to investigate specific neuronal responses to V8 protease. Cheek intradermal injections and neurobehavioral analysis will be performed to distinguish itch vs. pain behaviors in mice. In Specific Aim 2, we will determine whether neurons and host cells detect S. aureus V8 protease through specific protease-activated receptors (PARs). Preliminary data indicates that PAR1 may be an important host receptor that is activated by V8. We will utilize biochemical and luminescence-based approaches to determine the V8 cleavage site on PAR1. We will treat mice with pharmacological antagonists against PAR1 and utilize PAR1-/- mice to determine effects on V8-protease and S. aureus induced itch. In Specific Aim 3, we will utilize genetic approaches to ablate specific skin-innervating neurons (Nav1.8+, Mrgprd+, and Trpv1+) to assay their roles in S. aureus induced inflammation. We hypothesize that neurons will drive both itch/scratch-induced damage, and the release of neural mediators that directly signal to immune cells. We will use targeted approaches and proteomics to assess neuronal release of proinflammatory mediators. Our work could elucidate novel molecular crosstalk between S. aureus, host neurons and immune cells in itch. The three aims of this study leverage the complementary skills of Dr. Chiu and Dr. Horswill, combining neurobiological, immunological, and microbiological approaches to investigate the mechanisms of itch and neuroimmune signaling in S. aureus infection. Given the importance of itch in skin diseases, elucidating a microbial role in inducing this sensation could transform our understanding of host-microbe interactions at the skin barrier.

项目摘要 痒是一种令人不快的感觉，它会引起抓挠的欲望。虽然瘙痒伴随着许多皮肤感染， 微生物在瘙痒中的致病作用以前没有研究过。瘙痒引起的抓挠 严重的皮肤损伤和细菌致病。在这里，我们研究了人类细菌的作用， 病原体金黄色葡萄球菌及其分泌的蛋白酶在驱动瘙痒和炎症。S.金黄色 寄生于90%由特应性皮炎引起的皮肤病变，特应性皮炎是一种以慢性瘙痒为特征的疾病。S.金黄色 也会导致脓疱病，一种以瘙痒为特征的传染性皮肤病。我们假设S.金黄色 分泌蛋白酶，可以直接作用于宿主感觉神经元，以驱动瘙痒，抓挠诱导的皮肤损伤， 神经免疫串扰初步数据表明，S.金黄色葡萄球菌表皮感染小鼠诱导 强烈的瘙痒行为（异感、自发性瘙痒）和导致的皮肤病理学。使用同基因突变株， 我们发现分泌型S.金黄色葡萄球菌蛋白酶，特别是丝氨酸蛋白酶V8（SspA）是瘙痒所需的 在感染期间生产。在具体目标1中，我们将确定S的作用。金黄色葡萄球菌V8蛋白酶在驱痒中， 神经元激活和皮肤炎症。我们将利用等基因S。金黄色葡萄球菌突变株和互补株 为阐明该蛋白酶诱导瘙痒的必要性和充分性， 划痕引起的损伤和S.金黄色。瘙痒由背根神经节（DRG）介导 感觉神经元将进行DRG神经元钙成像，以研究特定的神经元反应， V8蛋白酶。将进行面颊皮内注射和神经行为分析以区分瘙痒与 小鼠的疼痛行为。在具体目标2中，我们将确定神经元和宿主细胞是否检测到S。金黄色 V8蛋白酶通过特异性蛋白酶激活受体（PAR）。初步数据表明，PAR 1可能是 一种由V8激活的重要宿主受体。我们将利用生物化学和发光技术 确定PAR 1上V8切割位点的方法。我们将用药理学拮抗剂治疗小鼠 并利用PAR 1-/-小鼠测定对V8-蛋白酶和S.金黄色葡萄球菌引起瘙痒。在 具体目标3，我们将利用遗传方法消融特定的皮肤神经支配神经元（Nav1.8+，Mrgprd+， 和Trpv 1+），分析它们在S.金黄色葡萄球菌诱导的炎症。我们假设神经元会驱动 瘙痒/抓挠诱导的损伤，以及直接向免疫细胞发出信号的神经介质的释放。我们将 使用靶向方法和蛋白质组学来评估促炎介质的神经元释放。我们的工作 可以阐明S.金黄色葡萄球菌、宿主神经元和免疫细胞参与瘙痒。三 这项研究的目的是利用Chiu博士和Horswill博士的互补技能，结合神经生物学， 免疫学和微生物学方法来研究瘙痒和神经免疫的机制 在S.金黄色葡萄球菌感染。鉴于瘙痒在皮肤病中的重要性，阐明微生物在皮肤病中的作用， 诱导这种感觉可能会改变我们对皮肤屏障处宿主-微生物相互作用的理解。