Transcriptomics of Tuberculosis Latency and Reactivation in Primates

描述(申请人提供)：结核病(TB)，由结核分枝杆菌(Mtb)引起，是一种全球性紧急传染病。抗击结核病的一个主要障碍是结核杆菌能够在宿主组织中以静止状态持续很长一段时间。当免疫系统受损时，这些细菌能够重新激活并导致肺结核病。因此，为了有效地控制结核病，需要对结核病的潜伏期和重新激活有一个全面的了解。进行这些研究所需的研究模型和工具现已可用。非人灵长类动物是研究结核病的优秀模型，特别是在研究实验性感染到潜伏期的进展，以及研究肉芽肿性病变的病理学和生物学--结核病感染的特征。我们通过将NHP暴露在真实的结核分枝杆菌气雾剂中，建立了人类结核病的模型。虽然许多研究小组专注于潜伏期和重新激活的细菌因素，但我们希望利用我们高度易处理的模型来确定这一过程的宿主签名和介体。我们表明，最初在NHP结核病变中诱导的促炎免疫信号通路在接下来的几周内被压倒性地沉默。这种转录重编程可能是宿主对细菌复制和生理变化的反应。此外，这些反应可能反映了病原体在肺部感染期间防止过度免疫病理的努力。我们建议的中心假设是，宿主肉芽肿反应可以用来预测潜伏和再激活的结核病。我们建议对NHP肺部病变的“转录组”和“miRNAome”进行系统研究。将从感染了低剂量结核分枝杆菌气雾剂的NHP获得时间分布，准确地模拟长期潜伏感染。还将从潜伏结核病被猿猴艾滋病重新激活的国家卫生政策中获得概况。这些全系统范围的概况，与从受感染的NHP获得的临床、微生物学和免疫学数据相结合，将产生潜伏和重新激活结核病的统计学习算法和混合效应计算模型。从收集的数据中获得的一些最具信息量的遗传预测因子的相关性将在NHP模型和人类患者中进行测试。CCL24、CCL25和CCL27与其他趋化因子配体和受体在灵长类结核肉芽肿中的表达呈负相关。这三种趋化因子的表达在晚期明显增加，而不是在早期。我们假设这些趋化因子对于长期维持含有潜伏的结核分枝杆菌的灵长类病变是重要的。LAG3在灵长类动物早期病变中的表达是晚期病变的40多倍。LAG3是一种新的Treg细胞标志物。我们假设LAG3负责负调控灵长类结核病变中效应器T细胞产生的保护性免疫反应。利用一种新的基于脂化siRNA纳米颗粒的方法，“潜伏期”特定基因CCL24/25/27和“活性结核：特定基因LAG3”的表达将在NHP中被沉默。然后将在这些动物身上研究潜伏性疾病的进展及其免疫学和分子相关性。最后，将确定对这些和其他“潜伏期”和“重新激活”特定特征的免疫反应在潜伏期和活动性肺结核患者以及结核/艾滋病混合感染患者中的表达。这些系统生物学研究可能会成倍地提高我们对结核病潜伏期和在一个尽可能接近人类的模仿结核病和艾滋病的宿主中重新激活的理解。最终，这些进展可能使临床医生能够更好地发现和治疗潜伏性结核病。

DESCRIPTION (provided by applicant): Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is a global infectious disease emergency. A major hurdle in combating TB is the fact Mtb is able to persist for long periods of time in host tissues, in a quiescent state. These bacilli are able to reactivate and cause pulmonary TB, when the immune system is compromised. Hence, a complete understanding of TB latency and reactivation is required for the effective control of TB. The research models and tools necessary to perform these studies are now available. Nonhuman Primates (NHPs) are excellent models of TB, especially to study the progression of experimental infection to latency, and to study the pathology and biology of granulomatous lesions - the hallmarks of TB infections. We have established a model of human TB, by exposing NHPs to true Mtb aerosols. While many research groups focus on the bacterial factors of latency and reactivation, we would like to leverage our highly tractable model to identify host signatures and mediators of this process. We show that pro-inflammatory immune signaling pathways, initially induced in NHP TB lesions, are overwhelmingly silenced over the course of next several weeks. This transcriptional reprogramming could be a host response to changes in bacterial replication and physiology. Further, these responses could reflect the efforts of the pathogen to prevent excessive immunopathology during the infection of lungs. The central hypothesis of our proposal is that host granuloma responses can be used to predict latent and reactivation TB. We propose to perform a systematic study of the "transcriptome" and the "miRNAome" of NHP lung lesions. Temporal profiles will be obtained from NHPs infected with a low-dose of Mtb aerosols, accurately modeling long-term latent infection. Profiles will also be obtained from NHPs in which latent TB is reactivated by simian AIDS. These system-wide profiles, in conjugation with the clinical, microbiological and immunological data obtained from infected NHPs will generate statistical learning algorithms and mixed effects computational models of latent and reactivation TB. The relevance of some of the most informative set of genetic predictors available from the data collected will be tested back in both the NHP model, as well as in human patients. The expression profiles of CCL24, CCL25 and CCL27 show negative correlation with all other chemokine ligands and receptors in primate TB granulomas. The expression of these three chemokines is significantly increased in late, rather than early lesions. We hypothesize that these chemokines are important for the long-term maintenance of primate lesions harboring latent Mtb bacilli. The expression of LAG3 was induced more than 40-fold in early primate lesions relative to late ones. LAG3 is a novel marker of Treg cells. We hypothesize that LAG3 is responsible for negatively regulating protective immune responses generated by effector T cells in primate TB lesions. The expression of "latency" specific genes CCL24/25/27 and the "active-TB: specific gene LAG3 will be silenced in NHPs using a novel lipidated-siRNA nanoparticle based approach. The progression of latent disease and its immunological and molecular correlates will then be studied in these animals. Finally, the expression of an immune response to these and other "latency" and "reactivation"- specific profiles will be determined in human patients of latent and active TB, as well as TB/AIDS co- infected patients. These systems-biology studies will likely exponentially enhance our understanding of TB latency and reactivation in a host that mimics both TB and AIDS in the closest possible manner to humans. Eventually, these advances may empower clinicians better to detect and treat latent TB.