Perturbations of host cell signaling by a complex hepatotropic pathogen

复杂的嗜肝病原体对宿主细胞信号传导的干扰

• 批准号: 10677933
• 负责人: Alexis Kaushansky
• 金额: $ 63万
• 依托单位: SEATTLE CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-01 至 2028-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10677933
• 关键词: Animals; Antigen-Presenting Cells; Antigens; Apoptosis; Apoptotic; Biological; Blood; CD8-Positive T-Lymphocytes; CRISPR screen; Cell Death; Cell Survival; Cells; Cessation of life; Characteristics; Collection; Complex; Culicidae; Data; Development; Disease; Dissection; Environment; Equilibrium; Erythrocytes; Genetic; Genetic Transcription; Growth and Development function; Hepatocyte; Host Defense; Human; Immune; Immune response; Immunity; Immunologic Stimulation; Immunologics; In Situ; Individual; Infection; Inflammatory; Integration Host Factors; Internet; Intervention; Invaded; Investigation; Knock-out; Lipid Peroxidation; Lipid Peroxides; Liver; LoxP-flanked allele; Macrophage; Malaria; Mediating; Mediator; Medical; Mitochondria; Molecular; Mus; Nature; Outcome; Parasites; Parasitic infection; Pathway interactions; Persons; Plasmodium; Process; Proteomics; Research; Resources; Robot; Role; Signal Transduction; Site; Sterility; Stimulus; Stream; System; T cell response; T-Cell Development; T-Lymphocyte; Technology; Testing; Tissues; Vaccination; Vaccines; cell killing; cell type; comparative; cytokine; design,build,test; experimental study; genome-wide; in vivo; insight; liver infection; malaria infection; migration; pathogen; prevent; recruit; response; small molecule; tool; transcriptomics; transmission process; whole genome

ABSTRACT During Plasmodium parasite liver stage infection, parasites protect their host hepatocyte by preventing its death and exploiting host resources for growth and development. This process is imperfect, as some parasites succumb to host defenses and are eliminated by host cell death; this, in turn, elicits an immune response. This proposal is focused on the investigation of molecular mechanisms in the hepatocyte that curtail, or promote, malaria parasite infection and the immunological consequences of those outcomes. After transmission by a mosquito, Plasmodium parasites are carried by the blood stream to the liver and invade a single hepatocyte to form a liver stage. Throughout the liver stage we have shown that both apoptosis and lipid peroxide mediated death pathways curtail infection. Whether the infected cell survives or succumbs to host cell death has consequences that expand beyond the individual infection; parasites that are eliminated via hepatocyte cell death can provide potent stimuli for the subsequent immune response and the nature of the immune stimulation is partially dependent on the type of regulated cell death engaged. Indeed, we show that immune cell subsets, in particular immune cells with markers consistent with tolerogenic macrophages, sequester around the parasite during liver infection. Additionally, eliminating apoptosis, a sterile form of cell death, reduces immunity in response to whole parasite vaccination. This proposal aims to test the hypothesis that multiple forms of cell death can control Plasmodium infection and that altering the balance between sterile and inflammatory forms of cell death impacts subsequent immunity to parasite infection. To test this hypothesis, we will make extensive use of animal and technological tools. First, we will use mice with cell death pathways eliminated exclusively in hepatocytes to examine the specific role of each type of cell death in regulating wild type parasites and parasites with multiple biological deficiencies. We will use genome wide CRISPR/Cas9 screens to identify specific genetic regulators of cell death in infected cells and we will make use of a mosquito dissection robot that we have recently designed, built, and tested to perform larger, more unbiased analyses than previously possible. We will also take advantage of state-of-the-art spatial profiling approaches that facilitate the collection of proteomic and transcriptomic information in situ for Plasmodium-infected livers to examine infected cells, and the surrounding immune environment. In addition to identifying mediators of initial infection, will use these tools to ask how altered hepatocyte death primes CD8 T cell responses and informs immunity to subsequent infection. Accomplishing our aims enables altering key host factors with small- molecules that could prevent a wild-type parasite from progressing to symptomatic erythrocytic infection or eliminate infection in a way that elicits a potent immune response against subsequent challenge.

摘要 在疟原虫肝期感染过程中，疟原虫通过阻止肝细胞的增殖来保护宿主肝细胞。 死亡和开发宿主资源以促进生长和发展。这个过程是不完美的，因为一些寄生虫 屈服于宿主防御并被宿主细胞死亡消除;这反过来又激发免疫应答。这 该提案集中于研究肝细胞中减少，或促进， 疟疾寄生虫感染和这些结果的免疫后果。经过A 在蚊子身上，疟原虫寄生虫通过血流进入肝脏，侵入单个肝细胞， 形成肝脏阶段。在整个肝脏阶段，我们已经表明，细胞凋亡和脂质过氧化物介导的 死亡途径减少了感染。感染的细胞是存活还是死于宿主细胞的死亡， 超出个体感染范围的后果;通过肝细胞消除的寄生虫 死亡可以为随后的免疫反应和免疫反应的性质提供强有力的刺激。 刺激部分地依赖于参与的调节性细胞死亡的类型。事实上，我们证明免疫 细胞亚群，特别是具有与致耐受性巨噬细胞一致的标志物的免疫细胞， 在肝脏感染时围绕寄生虫。此外，消除细胞凋亡，细胞死亡的不育形式， 降低对整个寄生虫疫苗接种的免疫力。这项提议旨在检验以下假设： 多种形式的细胞死亡可以控制疟原虫感染，改变不育和 炎性形式的细胞死亡影响随后对寄生虫感染的免疫力。为了验证这个假设，我们 将广泛使用动物和技术工具。首先，我们将使用具有细胞死亡途径的小鼠， 专门在肝细胞中消除，以检查每种类型的细胞死亡在调节野生型 型寄生虫和具有多种生物缺陷的寄生虫。我们将使用全基因组CRISPR/Cas9 筛选来识别受感染细胞中细胞死亡的特定遗传调节因子， 解剖机器人，我们最近设计，建造和测试，以执行更大，更公正的分析， 比以前可能的。我们还将利用最先进的空间分析方法， 促进疟原虫感染肝脏原位蛋白质组和转录组信息的收集， 检查受感染的细胞和周围的免疫环境。除了确定调解人的初步意见外， 感染，将使用这些工具来询问如何改变肝细胞死亡引发CD 8 T细胞反应，并告知 对后续感染的免疫力。实现我们的目标能够改变关键的东道国因素与小- 可以防止野生型寄生虫发展为有症状的红细胞感染的分子，或 消除感染的方式，激发了一个强大的免疫反应，对随后的挑战。