Mouse-on-a-chip systems to evaluate pancreas-adipose tissue dynamics in vitro

 描述(申请人提供)：脂肪组织(FAT)不仅仅是甘油三酯的储存场所，现在人们认识到，脂肪组织是一个复杂的、多细胞的内分泌器官，具有深刻的全身影响，几乎改变了几乎所有其他器官系统的功能。然而，尽管它很重要，但缺乏关于脂肪组织中脂肪因子分泌和营养吸收的动态性质的信息，突出了方法学上的几个未得到满足的需求。很少有技术可以询问少量的脂肪组织，也缺乏探索器官动态功能的方法。具体地说，我们对葡萄糖、胰岛素和脂肪功能之间的动态关系的看法有限，这突显了迫切需要更好的体外技术来研究胰腺-脂肪组织的串扰。正如我们之前的资助期所表明的那样，我们建议我们的微流控系统是满足这些持续需求的理想选择。我们的研究小组开发了微流控方法，用于培养C57BL/6J小鼠的内分泌组织，即胰岛和脂肪组织，以及采集激素分泌。这些系统允许以标准技术不可能实现的方式对组织进行动态询问。这项研究的长期目标是开发内分泌系统的体外模型，用于营养、新陈代谢和药物开发。在短期内，我们的目标是开发一种芯片上的小鼠微流控系统，允许对初级组织的激素分泌和营养吸收进行动态和定量测量。微流控设备将与小体积方法同时开发，以分析胰岛和脂肪组织的分泌或营养吸收，并将使用3D打印来改进我们新的设备接口和组织培养方法。该提案的目标1旨在开发一种自动微流控输入/输出多路复用器(µMUX)，用于对进出内分泌组织的激素和营养进行通用的动态控制。目标2将导致有针对性的小容量相容的激素和脂肪组织中游离脂肪酸摄取的分析。AIMS 3和AIMS 4本质上是生物学的，使用µMUX系统来确定具有不同血糖动态的内分泌组织中激素分泌、脂肪酸摄取和脂肪分解的动态(AIMS 3)，并使用共培养的µMUX系统来确定组织之间的动态反馈的作用(AIM 4)。这项研究的基本原理是提供一个灵活可编程的体外胰腺-脂肪动力学微观模型，以测试与肠道-胰腺信号动力学、胰岛素/脂肪分解/脂肪酸摄取动力学以及低胰岛素和低葡萄糖(空腹或生酮代谢)下脂肪分解调节相关的几个重要生物学假说。这项拟议的工作具有重大意义，这是此类研究在体外的首次。 模拟胰腺-脂肪生理学，我们预计这将导致关于人类饮食干预的更好信息。因此，该提案在技术和生物方法上都是创新的。初步证据有力地支持了这些建议的可行性，研究小组已经证明了成功的记录。

 DESCRIPTION (provided by applicant): Rather than a mere storage site for triglycerides, it is now understood that adipose tissue (fat) is a complex, multicellular endocrine organ that has profound systemic effects, altering the function of nearly all other organ systems. Despite its importance, however, there is a lack of information on the dynamic nature of adipokine secretion and nutrient uptake in adipose tissue, highlighting several unmet needs in methodology. Few techniques exist to interrogate small amounts of adipose tissue, and there is a shortage of methods to explore dynamic function of the organ. Specifically, we have a limited view of the dynamic relationship between glucose, insulin, and adipose function, highlighting an immediate need for better in vitro techniques to study pancreas-adipose tissue crosstalk. As demonstrated in our previous funding period, we propose that our microfluidic systems are ideal to meet these ongoing needs. Our research team developed microfluidic approaches for culture of endocrine tissue, namely pancreatic islets and adipose tissue from C57BL/6J mice, as well as for sampling of hormone secretion. These systems permit dynamic interrogation of the tissues in ways not possible with standard techniques. The long-term goal of this research is to develop in vitro models of the endocrine system for applications in nutrition, metabolism, and drug discovery. In the short term, our objective is to develop a mouse-on-a-chip microfluidic system that permits dynamic and quantitative measurements of both hormone secretion and nutrient uptake from primary tissue. Microfluidic devices will be developed concurrently with small-volume methodology to assay secretion or nutrient uptake from pancreatic islets and adipose tissue, and 3D printing will be used to improve our novel device interfacing and tissue culture methods. Aim 1 of the proposal seeks to develop an automated microfluidic input/output multiplexer (µMUX) for generalizable dynamic control over hormones and nutrients to/from endocrine tissue. Aim 2 will result in targeted small-volume compatible assays for hormones and free fatty acid uptake in adipose tissue. Aims 3 and 4 are biological in nature, using the µMUX system to determine the dynamics of hormone secretion, fatty acid uptake, and lipolysis in endocrine tissues with varied glycemic dynamics (Aim 3), and determining the role of dynamic feedback between the tissues using a co-culture µMUX system (Aim 4). The rationale for this research to provide a flexibly programmable, in vitro micro-model of pancreas-adipose dynamics to test several important biological hypotheses related to gut-pancreas signaling dynamics, insulin/lipolysis/fatty acid uptake dynamics, and regulation of lipolysis at low insulin and glucos (fasting or ketogenic metabolism). The proposed work is significant as a first-of-its-kind in vitro mimic of pancreas-adipose physiology, which we expect will lead to better information on human dietary interventions. The proposal is thus innovative in its technological and its biological approaches. Preliminary evidence strongly supports the feasibility of these proposals, and the research team has a proven track-record of success.