Construction And Application Of Microfluidic Lung Cell System

Human industrial and agricultural production activities inevitably cause environmental pollution.Among various environmental pollutions,air pollution has received much attention.It has not only caused various ecological problems,but also seriously affected human health.Various pollutants induce various respiratory diseases such as chronic obstructive pulmonary disease and asthma,which not only bring physical pain to the living individual,but also impose a heavy burden on the family and society.The mechanism and development of pollutant-induced respiratory diseases have always been the focus of biological and medical research.At present,there are two main research methods in this field at home and abroad:one is based on an in vitro two-dimensional cell culture model,and the other is an in vivo experimental animal model.Although the in vitro cell culture model can simplify the experimental analysis factors,it can not realize the spatio-temporal control and reconstruction of the extracellular microenvironment of micro-scale organ tissues.In vivo animal experiments can provide complete in vivo drug metabolism pathways,but it is difficult to achieve lesions in human body.Both of these research methods have the disadvantages of cumbersome operation,time-consuming and labor-intensive and high consumption of reagents,and the latter has to face the ethical problems faced by animal experiments.It can be seen that the effective in vitro reconstruction and methodological improvement of various pollutant-induced respiratory pathological events(such as bronchial and alveolar inflammation)is still an important research direction.Microfluidic has the characteristics of miniaturization,integration,high throughput,low consumption,high sensitivity and rapid analysis,and has shown great application potential in the fields of drug screening,clinical diagnosis and bionics.In view of this,this research project considers the effective in vitro reconstruction mechanism of two pollutant-induced pathological events based on microfluidic biomimetic chip technology,in order to evaluate the application potential of microfluidic technology in the mechanism and development of pollutant-induced respiratory diseases.In this study,the microfluidic cell chip and the two-channel layered microfluidic chip microfluidic chip,which are assisted by chemical concentration gradient generation,are designed and constructed by photolithography and soft etching.Then two kinds of biomimetic chip systems are used.Microscale manipulation and real-time dynamic analysis of benzopyrene and nicotine-induced lung injury and inflammatory response were studied.1.Construction and application of microfluidic cell chip assisted by chemical concentration gradient.(1)A microfluidic cell chip assisted by chemical concentration gradient was designed and constructed by photolithography and soft etching.The chip consists of a flow layer and a glass substrate layer,wherein the flow layer comprises a chemical gradient generator and a microcavity array.Chemical concentration gradient generators are used for the generation of multi-concentration gradients in the chip,while microcavity arrays are used for perfusion input of cells and reagents,cell retention,cell culture,cell detection,and liquid output.The chip is capable of achieving high stability with concentration gradients in the flow rate range of no more than 1 ?L min-1.(2)Based on the established chemical concentration gradient assisted microfluidic cell chip,the real-time and flux analysis of bromine epithelial damage induced by different concentrations of benzopyrene in a single chip was performed.Compared with conventional in vitro research and analysis methods,the chip not only achieves a variety of positive and significant inflammatory responses or even apoptotic responses induced by benzopyrene in the lung epithelium,and the high dose of benzopyrene in the chip stimulates the epithelium to lead to more significant apoptosis.2.Construction and application research of two-channel layered microfluidic chip.(1)A two-channel layered microfluidic chip was designed and constructed by photolithography and multi-layer soft etching.The chip can meet the regional inoculation and adhesion growth of human alveolar epithelial cells and human lung microvascular endothelial cells,and achieve sub-regional and high-activity co-culture of the two cells;further form different spatial regions of epithelial cell layer and vascular endothelium.Cell layer formation and tight connection between monolayer cells;thus,a microfluidic chip-based alveolar tissue barrier miniaturization model system was constructed.(2)Based on the constructed two-channel layered microfluidic chip,the multi-indicator parameters related to the induction of inflammatory damage and apoptosis of alveolar epithelium and microvascular endothelial cells induced by benzopyrene and nicotine were studied.A comparative analysis of on chip coculture and monoculture and off chip cell culture was carried out.The on chip coculture microenvironment can reduce the inflammatory damage of alveolar tissue during the stimulation of environmental pollutants benzopyrene and nicotine,and realize the dynamic analysis of the cell and molecular level of the alveolar tissue barrier chip induced by different environmental pollutants in the alveolar tissue.This research project designed and constructed two kinds of microfluidic chips,and used the built chips to study the effective in vitro reconstruction mechanism and methodological improvement of pollutant-induced pathological events,and demonstrated the microfluidic technology in pollutant-induced respiratory diseases.The application mechanism in the research mechanism of development mechanism and development process has broadened the application field of microfluidic technology.