Modeling of human HSV infection: development of immune-competent 3D skin-on-chip with vascular perfusion

摘要 传统的机理研究和药物开发严重依赖体内动物模型和体外 细胞培养系统。尽管传统的体外培养 2D 或 3D 细胞能够分析信号 与感染或各种条件反应相关的途径和特征基因的鉴定 和治疗，它们无法重现组织中发生的复杂的细胞-细胞和细胞-基质相互作用 微环境。动物模型提供了对体内综合多器官反应的理解，但 人们对其预测价值和对人类的生物学相关性存在严重担忧。事实上，越来越多 候选药物未能从一期临床试验进入三期临床试验并进入市场。关键 减少临床试验中代价高昂的失败所面临的挑战凸显了为临床试验生成更好的模型系统的紧迫性 人类药物功效和安全性的临床前测试，以及了解分子机制 数以千计的人类疾病的根源。三维 (3D) 人体组织模型有望引人注目 预测复杂生理功能、对传染病的免疫反应的优势，以及 对治疗剂的药理学反应。这种合成组织模型具有巨大的潜力 可靠的药效测试和作为动物模型的优越替代品，特别是对于那些具有传染性的动物模型 没有足够的动物模型的疾病。皮肤是人体最大的器官，形成 保护身体免受病原体和潜在有害物质渗透的屏障。为了 模仿人类器官水平的皮肤病理生理学，我们建议利用生物工程方法 开发一种体外 3D“芯片皮肤”，将循环免疫细胞纳入正常的结构中 皮肤表皮和真皮用于模拟人类单纯疱疹病毒 (HSV) 的病毒-宿主相互作用 感染。我们的具体目标是： 1) 使用以下技术建立并验证血管化 3D“芯片上皮肤”平台 供体来源的原代细胞用于 HSV 感染建模并识别关键的免疫反应以进行保护。 2) 概括 3D 血管化“芯片上皮肤”中的组织驻留记忆 T 细胞区室，用于建模 组织驻留记忆 T 细胞介导的局部免疫中的抗原特异性、细胞密度和 TCR 多样性 保护。

SUMMARY Conventional mechanistic research and drug development relies heavily on in vivo animal models and in vitro cell culture systems. Although conventional in vitro cultured 2D or 3D cells enable the analysis of signaling pathways and the identification of signature genes associated with responses of infection or various conditions and treatments, they can't reproduce complex cell-cell and cell-matrix interactions occurring in the tissue microenvironment. Animal models provide understanding on in vivo integrated multi-organ responses but serious concerns exist over their predictive value and biological relevance to humans. In fact, more and more drug candidates have failed to advance from Phase I to Phase III clinical trials and to reach the market. Critical challenges to reduce costly failures in clinical trials highlight the urgency to generate better model systems for preclinical testing of drug efficacy and safety in humans, and for understanding molecular mechanisms underlying thousands of human diseases. A three-dimensional (3D) human tissue model promises compelling advantages to predict complex physiological functions, immune responses to infectious diseases, and pharmacological responses to therapeutic agents. Such a synthetic tissue model has outstanding potential for reliable drug efficacy testing and as a superior replacement for an animal model, especially for those infectious diseases that do not have adequate animal models. Skin is the largest organ of the human body and forms a barrier to protect the body against pathogens and penetration of potential harmful substances. In order to mimic the organ-level skin pathophysiology in humans, we propose to harness bioengineering approaches to develop an in vitro 3D `skin-on-chip' that incorporates circulating immune cells into the normal architecture of skin epidermis and dermis for modeling of viral-host interactions in human herpes simplex virus (HSV) infection. Our Specific Aims are: 1) Establish and validate a vascularized 3D `skin-on-chip' platform using donor-derived primary cells for modeling of HSV infection and identifying key immune responses for protection. 2) Recapitulate a tissue resident memory T-cell compartment in 3D vascularized `skin-on-chip' for modeling of antigenic specificity, cell density and TCR diversity in tissue resident memory T-cell mediated local immune protection.