Construction And Application Of Microfluidic Bio-Mimetic Hepatic-Sinusoid-on-a-Chip

In vitro liver models play an important role in liver biology,liver-related disease research,drug screening and evaluation.However,the existing in vitro liver models were too simple in structure which far from that in vivo,so they cannot maintain the activities and functions of the liver cells.Organ-on-a-chip is a new technique for in vitro animal cells culture that has been proposed in recent years.It can simulate the complex microstructures,microenvironments and physiological functions of specific organs.This study focused on the construction of hepatic-sinusoid-on-a-chip,and carried out a series of drug toxicology evaluations,liver disease and toxicity mechanism studies,as follows:By applying microfluidic technology,micromachining technology,and the principle of bionics,according to the spatial arrangement and proportion in vivo,a hepatic sinusoid chip which could bionic both blood and bile flows was constructed.In addition,four major functional hepatic cell lines were co-cultured in the chip.All cells in the chip maintained good activity throughout,and had a certain selective permeability to foreign substances.HepG2 cells showed obvious polarization and formation of bile ducts in the chip.The hepatic sinusoid chip maintained good anabolic and metabolic functions,and presented high phase I and phase II metabolic capacity and sensitivity to drugs.The chip was used to investigate the effects of shear force on liver functions and drug metabolism enzyme activities in vitro.Compared with the traditional in vitro liver models,hepatic-sinusoid-on-chip are more similar in structure and function to that in vivo.Next,acetaminophen,rifampicin and amiodarone were used as model drugs to evaluate the hepatotoxicity and study drug-drug interactions with the hepatic-sinusoid-on-a-chip.The evaluation of the hepatotoxicity of drugs by the hepatic-sinusoid-on-a-chip showed a better in vivo-in vitro correlation.The liver-chip can reflect the faint changes of acetaminophen-induced hepatotoxicity after combination,and the results were comparable with that from rat primary cells.The hepatic-sinusoid-on-a-chip also presents great potential in the liver disease mechanism studies.In this work we studied alcoholic liver disease as an example.The chip can reproduce the does-dependented and time-dependented of alcohol-induced liver injury.How do the two types of non-parenchymal cells(hepatic sinusoidal endothelial cells and hepatic stellate cells)participate in the process of alcoholic liver injury in the early stage was studied.We found that,for hepatic sinusoidal endothelial cells,low concentration of alcohol can destroy the tight connection of liver sinusoidal microvessels and reduce the synthesis of nitric oxide(NO),thus weakening the inhibition of hepatic stellate cells.On the other hand,low concentration of alcohol can directly activate hepatic stellate cells,resulting in a large proliferation of cells,increasing expression of vascular endothelial growth factor(VEGF)and ?-smooth muscle actin(?-SMA).Moreover,alcohol can also induce apoptosis of hepatic stellate cells when the concentration reaches 150 mM.Furthermore,T lymphocytic leukemia cells(HuT-78 cells)and myelogenous leukemia cells(HL-60 cells)were used to construct immune liver chip with immune stress response based on the hepatic-sinusoid-on-a-chip.The immune liver chip could reproduce the migration and infiltration process of immune cells in the liver under the action of chemokines.A series of defective liver chips were designed.With acetaminophen,sulfamethoxazole,ethinyl estradiol and abacavir as model drugs,the key role of each knockout factor in drug induced hepatotoxicity was discussed and then the mechanism of drug hepatotoxicity was studied.In summary,comparing with the existing in vitro liver platforms,the hepatic sinusoid chip has higher simulation in structure and function as in vivo.The hepatic-sinusoid-on-a-chip is expected to improve the pre-clinical hepatotoxicity and the ability to predict metabolism-based drug interactions,and is used for liver-related disease research and drug screening,and is expected to be used for the discovery of new drugs and improve the preclinical research of drugs.