Microfluidic Cell Chip Construction For Tumor Microenvironment Research

Tumor and its metastasis involve a series of complicated pathological processes and are closely related to its microenvironment.A very complex and comprehensive system of tumor microenvironment is composed of tumor cells,non-tumor cells,and intercellular substances.Conventional in vitro cell experiments and animal experiments often deviate from the actual in vivo environment,and there are ethical issues that are difficult to solve.And it cannot meet the needs of tumor-related research.Microfluidic chip technology can design two-dimensional or three-dimensional channel network structure,the diameter of the microchannel is generally 10 ~ 100?m,and the size of the cell is equivalent,with less sample consumption,fast analysis speed,high degree of automation,easy integration,can be miniaturized,etc.advantage.The microfluidic cell chip allows cell culture,observation and analysis to be completed on the chip platform.It provides a new research platform for cell research,such as cell-cell interactions,tissue engineering,drug screening,biosensors,and organ microchips.In this dissertation,microfluidic technology is used as a platform to construct lung cancer microenvironmental microarrays using human lung adenocarcinoma cells(A549),small cell lung cancer(H446),human embryonic lung fibroblasts(HLF-1)and human endothelial cells(HUVECs).Using the "soft lithography" process,two kinds of microfluidic cell chips that manipulate the cells inside the chip through different physical structures were designed and manufactured,and multicellular co-cultivation was realized.Observe the survival state and migration of endothelial cells under the influence of different cell and growth factors.And we also explore the biological effects of related cytokines on endothelial cells in the tumor micro-environment.The paper is mainly divided into three parts:1.To sum up the challenges faced by the study of tumor microenvironment and the current research status of microfluidic chip and its application advantages in the field of cell biology,the research ideas for the construction of microfluidic cell chip are proposed and used for simulation of tumor microenvironment.2.Design and preparation of three kinds of cell co-cultivation microfluidic chip: A microfluidic chip with a “micro dam” and a “micro gap” structure was designed to physically isolate different cells while small molecules of nutrients in the medium is free to circulate.A silicon-based mold was fabricated by a soft lithography process using SU-8 negative photoresist,and the process parameters such as spin speed control,exposure time,and development time of the spin-coated photoresist were studied,and a cell microfluidic chip fabrication process was established.Multi-cell co-cultivation studies were performed on the chip.The results showed that the cells were in good growth condition and could initially simulate endothelial vascular wall function.3.A novel microfluidic cell co-cultured microarray was designed to simulate the tumor microenvironment and the biocompatibility of the microarray was evaluated.The directional migration of HUVECs co-cultured with A549 and HLF-1 and different lung cancers(A549,H446)were studied.The results showed that compared with fibroblasts,HUVECs tend to migrate to cancer cells;compared with A549 with weaker invasion ability,HUVECs tend to migrate with stronger invasion of H446.And we explore the possible mechanisms for this process...