Microfabricated Biosensor Platform for Monitoring of Cell Metabolism

描述（由申请人提供）：肝脏是重要的生物系统，在调节机体能量供应中起关键作用。该器官维持葡萄糖释放和储存的程序，以响应激素刺激，建立葡萄糖稳态。毒物对能量代谢的破坏具有严重的病理生理后果，并与糖尿病和肥胖症有关。用于测量细胞代谢的传统方法需要大量的细胞，对代谢动力学的揭示很少，不允许将细胞平行暴露于多种刺激，并且不允许研究一起培养的多种细胞类型之间的相互作用。因此，传统的细胞培养/代谢物检测方法不适合于监测稀缺细胞（例如原代人肝细胞）和筛选候选药物和毒物对肝细胞的影响。本研究的目的是开发一种新型的芯片实验室生物传感器平台，用于体外监测肝细胞代谢。所提出的平台的新奇是基于我们将用于多种代谢物的微生物传感器与功能性微制造肝细胞培养物相结合的能力。细胞和传感元件的并置将通过制造聚（乙二醇）（PEG）水凝胶微结构来实现，该微结构作为细胞培养室和酶包封基质具有双重功能。含酶的PEG水凝胶微室将建立在细胞培养基质的顶部，创建可单独寻址的细胞培养单元，用于葡萄糖的电化学检测和基于荧光的过氧化氢（H2 O2）检测。所提出的生物传感器平台将提供几个优点，包括同时监测两种分析物，观察两种不同细胞表型的动态和相互代谢相互作用，以及利用少量细胞进行分析。微流体递送系统将与感测模块结合以控制可溶性环境的组成。将功能性肝细胞与微型生物传感器相结合的新型微加工装置将成为研究肝毒物对肝功能影响的重要技术。公共卫生相关性肝脏是人体的代谢中心，主要负责解毒和能量调节功能。能量调节是指糖如葡萄糖的消耗和生产的程序。能量调节失衡与糖尿病和肥胖症密切相关。我们建议建立一个微芯片来监测肝细胞中的糖消耗。这种微芯片将用于确定有害物质如何影响肝脏及其调节体内糖水平的能力。

DESCRIPTION (provided by applicant): Liver is an important biological system that plays a key role in regulating energy supply of the body. This organ maintains a program of glucose release and storage in response to hormonal stimulation establishing glucose homeostasis. The disruption of energy metabolism by toxicants has serious pathophysiological consequences and is connected to diabetes and obesity. Traditional approaches for measuring cellular metabolism require large cell numbers, reveal little about metabolism dynamics, do not allow exposure of cells to multiple stimuli in parallel and do not permit to study the interplay between multiple cell-types cultured together. Thus, the traditional cell culture / metabolite detection approaches are not suited for monitoring scarce cells (e.g. primary human hepatocytes) and screening the effects of drug candidates and toxicants on the liver cells. The goal of this proposal is to develop a novel lab-on-chip biosensor platform for in vitro monitoring of liver cell metabolism. The novelty of the proposed platform is based on our ability to integrate micro-biosensors for multiple metabolites with functional microfabricated liver cell cultures. The juxtaposing of cells and sensing elements will be achieved by fabricating poly(ethylene glycol) (PEG) hydrogel microstructures serving dual function as cell culture chambers and matrices for enzyme encapsulation. Enzyme-containing PEG hydrogel microchambers will be built on top of cell culture substrates, creating individually addressable cell culture units for electrochemical detection of glucose and fluorescence-based detection of hydrogen peroxide (H2O2). The proposed biosensor platform will offer several advantages including monitoring two analytes simultaneously, observing dynamic and reciprocal metabolic interactions of two distinct cell phenotypes, and utilizing small numbers of cells for analyses. A microfluidic delivery system will be incorporated with the sensing module to control composition of the soluble environment. A novel microfabricated device integrating functional liver cells with miniature biosensors will be an important enabling technology in the study of the effects of hepatotoxicants on liver function. PUBLIC HEALTH RELEVANCE Liver is the metabolic center of human body and is largely responsible for detoxification and energy regulation functions. Energy regulation refers to the program of consumption and production of sugars such as glucose. Imbalance in energy regulation is closely connected diabetes and obesity. We propose to build a microchip for monitoring sugar consumption in liver cells. This microchip will be used to determine how harmful substances affect the liver and its ability to regulate sugar levels in the body.