Embryonic stem cell-based fibroblast model of innate immunity development

描述（由申请人提供）：胚胎干细胞（ESC）被认为是再生医学的一种有前途的细胞来源。在过去的十年中，深入的研究已经导致了ESC分化细胞（ESC-DCs）可用于治疗人类疾病的可能性。然而，越来越多的证据表明，目前的分化方法产生的ESC-DCs是不完全功能。最近的研究表明，ESC-DCs缺乏对广泛的感染因子的先天免疫。当用于患者时，ESC-DCs将被放置在暴露于各种病原体和炎性细胞因子的伤口部位;因此，如果细胞不具有有效的免疫力，它们的活力和功能可能会受到损害。我们最近证明，ESCs是内在缺陷的表达I型干扰素和炎性细胞因子。这一发现解释了ESC-DCs中先天免疫的缺乏，并导致了我们的中心假设，即先天免疫在ESC中不发育，并且不能通过目前的分化方法有效诱导，但如果在分化期间提供适当的“免疫刺激”，则可以诱导先天免疫。该研究旨在了解控制先天免疫的分子机制，并开发产生具有免疫能力的ESC-DCs的分化策略。成纤维细胞（FB）是在组织形成和调节组织免疫/炎症反应中起关键作用的主要细胞。使用ESC分化的FB（ESC-FB）作为模型系统，我们将确定ESC先天免疫缺陷的分子基础，并确定刺激先天免疫发育的因素（目的1）。然后，我们将开发利用IFN？的新策略，病毒RNA类似物和脂多糖作为“免疫刺激剂”，在分化过程中促进先天免疫（目的2）。ESC-FB的先天免疫将通过体外模型与天然分化的FB进行比较分析，并将使用同基因小鼠体内模型进行评价，其中在宿主受到细菌和病毒感染攻击后，将评估植入ESC-FB中的免疫和炎症反应（目的3）。我们预计，这些结果将导致分化策略的发展，从根本上改变目前的方法，并更好地了解胚胎发育过程中控制先天免疫发育的机制。

DESCRIPTION (provided by applicant): Embryonic stem cells (ESCs) are considered a promising cell source for regenerative medicine. Intensive research over the past decade has led to the possibility that ESC-differentiated cells (ESC-DCs) could be used for the treatment of human diseases. However, increasing evidence indicates that ESC-DCs generated by the current differentiation methods are not fully functional. Recent studies indicate that ESC-DCs lack innate immunity to a wide range of infectious agents. When used in the patient, ESC-DCs would be placed in a wound site that is exposed to various pathogens and inflammatory cytokines; therefore, their viability and functionality could be compromised if the cells do not have competent immunity. We recently demonstrated that ESCs are intrinsically deficient in expressing type-I IFN and inflammatory cytokines. This finding explains the lack of innate immunity in ESC-DCs and has led to our central hypothesis that innate immunity is not developed in ESCs and cannot be effectively induced by current methods of differentiation, but it could be induced if proper "immunostimulation" is provided during differentiation. The proposed study aims to understand the molecular mechanisms that control innate immunity and to develop differentiation strategies that generate ESC-DCs with competent immunity. Fibroblasts (FBs) are major cells that play key roles in tissue formation and in modulating tissue immune/inflammatory responses. Using ESC-differentiated FBs (ESC-FBs) as a model system, we will determine the molecular basis for innate immunity deficiency in ESCs and identify the factors that stimulate innate immunity development (aim 1). We will then develop novel strategies that utilize IFN?, viral RNA analogs, and lipopolysaccharide as "immunostimulants" to promote innate immunity during differentiation (aim 2). The innate immunity of ESC-FBs will be comparatively analyzed with naturally differentiated FBs by in vitro models and will be evaluated with a syngeneic mouse in vivo model, where immune and inflammatory responses in implanted ESC-FBs will be assessed after the host is challenged with bacterial and viral infections (aim 3). We expect that the results will lead to the development of differentiation strategies that could fundamentally transform the current methods and achieve a better understanding of mechanisms that control innate immunity development during embryogenesis.