IMMUNE REGULATION AND VACCINE DEVELOPMENT IN LEISHMANIASIS

利什曼病的免疫调节和疫苗开发

• 批准号: 10692024
• 负责人: David Sacks
• 金额: $ 77.51万
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10692024
• 关键词: Ablation; Address; Apoptosis; Bone Marrow; C57BL/6 Mouse; CASP3 gene; CCL24 gene; Cell Count; Cell Death; Cells; Chronic; Clinical Research; Complex; Confocal Microscopy; Cutaneous; Cutaneous Leishmaniasis; Dermal; Dermis; Detection; Development; Disease; Disease Progression; Eotaxin; Flow Cytometry; Genetic; Goals; Granuloma; Growth; Hamsters; Hepatic Granuloma; Homeostasis; Human; ITGAX gene; Immunologics; Impairment; India; Infection; Infectious Skin Diseases; Inflammation; Inflammatory; Interleukin-4; Interleukin-5; Kupffer Cells; Lead; Leishmania; Leishmania donovani; Leishmaniasis; Lesion; Lipid Peroxidation; Liver; Lymphoid Cell; Maintenance; Measures; Mediator of activation protein; Modeling; Molecular; Mus; Organ; Parasites; Persons; Population; Preventive vaccine; Property; Resistance; Resistance to infection; Signal Transduction; Skin; Source; Spleen; Stains; TSLP gene; Tissues; Tropical Disease; Vaccines; Visceral Leishmaniasis; blastomere structure; chronic infection; congenic; cytokine; eosinophil; experimental study; healing; immunoregulation; inflammatory milieu; macrophage; mannose receptor; microbiota; monocyte; rational design; recruit; repaired; response; single-cell RNA sequencing; therapeutic vaccine; therapy development; vaccine development

Tissue-resident macrophages (TRMs) are critical for tissue homeostasis/repair, and their development is governed by tissue niche-specific signals. Although the importance of these nurturing signals for TRMs to perform tissue-specific functions has been extensively studied, it remain poorly addressed how TRMs maintain their homeostatic properties during inflammation, in particular infection-driven inflammation in the skin. We previously showed the importance of eosinophil-TRM cooperative interactions for the maintenance of the M2-like properties of dermal TRMs within the strong pro-inflammatory environment of the L. major infected dermis; eosinophils provide IL-4 to dermal TRMs, and IL-4-stimulated dermal TRMs produce CCL24 (eotaxin) which in turn functions to recruit more eosinophils. In our current studies, we have identified two independent subsets of dermal TRMs, MHCII+ mannose receptor (MR) low and MHCII- MR hi populations. By single cell RNA seq, the MHCII- MR hi subset of dermal TRMs was shown to be the sole producer of CCL24 in the infected skin, and was also found to be the sole source of the alarmin thymic stromal lymphopoietin (TSLP) required to activate innate lymphoid cells 2 (ILC2), which produce IL-5 to amplify eosinophil-TRM interactions. Both selective depletion of IL-5+ ILC2 or genetic ablation of TSLP impaired the localized TH2 circuitries required to maintain the numbers and functionality of M2-like dermal TRMs, and disease progression was ameliorated. Thus, in the setting of the strong pro-inflammatory environment of the L. major infected dermis, dermal TRMs actively maintain themselves by producing CCL24 and TSLP, and any disruption of the localized TH2 circuitries will alter their number and activation states, and promote stronger resistance to infection. Visceral leishmaniasis (VL) is a tropical disease caused by Leishmania species within the L. donovani complex (L. donovani/L. infantum). Macrophages in the liver, spleen, and bone-marrow are the principal target cells for infection. Kupffer cells (KCs) are the embryonic-derived, liver resident macrophages, and are characterized by Clec4f and Tim4 expression. In homeostasis, KCs maintain their numbers via self-proliferation, but in some inflammatory settings they can die and be replaced by monocyte-derived cells (moKCs). In the murine VL model, KCs are important for both the initial growth of the parasite in the liver and for granuloma formation, which is associated with the eventual protective response in this tissue. KC death, their replacement by moKCs, and the functionality of KCs vs moKCs has never been investigated in VL. In C57BL/6 mice infected with L. infantum, at 42 days post-infection we found evidence of KC apoptosis, measured by cleaved caspase 3 staining by confocal microscopy, and ferroptosis, by lipid peroxidation detection by flow cytometry. Consistently, Clec4f and Tim4 expression in KCs was reduced, while monocyte markers such as Ly6C and CD11c were enhanced. As further evidence of their monocytic origin during chronic infection, KC numbers were lower in CCR2-/- mice when compared to WT mice, and in experiments involving congenic parabiotic mice, the KCs present within the granulomas expressed both congenic markers. Selective KC depletion and replacement by moKCs prior to infection using Clec4f-Cre-DTR mice, demonstrated that the predominance of moKCs resulted in lower parasite loads, indicating that moKCs are more effective in controlling infection. Thus, KCs death and their replacement by monocyte-derived cells appears to be central to the protective response in this organ.

组织居民巨噬细胞（TRM）对于组织稳态/修复至关重要，它们的发育受组织小众特异性信号的控制。尽管已经对这些培育信号对执行组织特异性功能的重要性进行了广泛的研究，但它仍然很差解决TRM在炎症期间如何保持其稳态特性，特别是皮肤中感染驱动的炎症。我们先前表明了嗜酸性粒细胞-TRM合作相互作用对维持在强烈感染真皮的强烈促炎环境中维持真皮TRM的M2样性质的重要性；嗜酸性粒细胞向真皮TRM提供IL-4，而IL-4刺激的皮肤TRM产生CCL24（eotaxin），进而起作用以募集更多的嗜酸性粒细胞。在我们目前的研究中，我们已经确定了两个独立的皮肤TRM的独立子集，即MHCII+ Mannose受体（MR）低和MHCII-MR HI人群。 通过单细胞RNA SEQ，真皮TRM的MHCI- MR HI子集被证明是感染皮肤中CCL24的唯一生产者，并且也被发现是危及胸膜基质淋巴细胞增生蛋白（TSLP）的唯一来源，以激活生动淋巴样细胞2（iLc2），从而激活iMply-illc-ilm imply imply im iL iL iL iL iL iL iL， IL-5+ ILC2的选择性耗竭或TSLP的遗传消融都损害了维持M2样皮肤TRM的数量和功能所需的局部TH2电路，并改善了疾病进展。 因此，在主要感染真皮的强烈促炎环境的环境中，真皮TRM通过产生CCL24和TSLP积极维持自己，任何局部TH2电路的破坏都会改变其数量和激活状态，并促进对感染的强烈抵抗力。 内脏利什曼病（VL）是一种由L. donovani综合体（L. donovani/l. intantum）内的利什曼原虫物种引起的热带疾病。 肝脏，脾脏和骨髓中的巨噬细胞是感染的主要靶细胞。 Kupffer细胞（KCS）是胚胎衍生的肝遗传巨噬细胞，其特征是Clec4f和Tim4表达。在体内平衡中，KCS通过自我增殖保持数量，但是在某些炎症环境中，它们可能死亡并被单核细胞衍生的细胞（MOKC）取代。在鼠VL模型中，KCS对于肝脏中寄生虫的初始生长和肉芽肿形成都很重要，这与该组织中最终的保护反应有关。 KC死亡，MOKC的替代以及KCS与MOKC的功能从未在VL中进行研究。在感染婴儿乳杆菌的C57BL/6小鼠中，在感染后42天，我们发现了Kc凋亡的证据，通过裂解的caspase 3通过共焦显微镜染色，通过共聚焦显微镜染色，通过脂质过氧化作用，通过脂质过氧化检测，通过流式细胞仪检测。一致地，降低了KC中的CleC4F和Tim4的表达，而单核细胞标记（如LY6C和CD11C）得到了增强。作为其在慢性感染期间单核细胞起源的进一步证据，与WT小鼠相比，CCR2 - / - 小鼠的KC数量较低，并且在涉及先天性抛害小鼠的实验中，颗粒中存在的KCS表示，表达了这两种先天性标记。选择性的KC耗竭和使用Clec4f-Cre-DTR小鼠感染之前MOKC替换，表明MOKC的主要率导致寄生虫负荷较低，这表明MOKC在控制感染方面更有效。因此，KCS死亡及其被单核细胞衍生细胞的替代似乎是该器官保护性反应的核心。