3D Printed Hydroxyapatite Microfluidic Chip For Bone Related Drug Screening And Bone Vascularization Research

Bone is an important part of human body structure and plays a vital role in human health.The development of a biological microfluidic chip that simulates bone structure and function will provide a good platform for in vitro drug screening and bone-related biological research.However,the most commonly used polydimethylsiloxane(PDMS)microfluidic chip is not suitable for simulating bone microenvironment.Hydroxyapatite(HAp),as a bioactive ceramic,has a structure and composition similar to bone mineralization products.In this study,HAp was first proposed as materials for the preparation of microfluidic chips.This study explored the photocuring printing parameters of the ceramic substrate and the bonding method of the ceramic microfluidic chip,hoping to find a new method for building a biological microfluidic chip with a highly bionic bone environment.First,in order to obtain a microfluidic chip substrate with a fine micro-grooved structure,the 3D printing parameters of the hydroxyapatite material were continuously optimized.The debinding curve of the green sample was determined by thermogravimetry(TG)and differential scanning calorimetry(DSC),and the highest sintering temperature of the sample was determined to be 1280 ? by density and compressive strength tests.After printing and sintering,the ceramic microfluidic chip had a regular shape and controllable dimensional accuracy,and the density of the substrate reached 96.8%,which can meet the requirements of the microfluidic chip.Cell experiment results showed that the ceramic chip substrate had no cytotoxicity,which was more conducive to cell adhesion,proliferation and osteogenic differentiation than PDMS,and was closer to the bone microenvironment in the human body.Subsequently,the preparation technology of hydroxyapatite microfluidic chip substrate was used in the study of bone-related gradient drug screening model and bone vascularization cell co-culture model.On the one hand,in order to prepare a gradient drug screening microfluidic chip,a PDMS prepolymer was used as glue,and the ceramic microfluidic chip substrate with a Christmas tree structure and the blank PDMS cover were heat-sealed and bonded.The concentration gradient formed by the hydroxyapatite microfluidic chip was verified by fluid simulation,fluorescent ink perfusion and the concentration measurement experiment of the anti-tumor drug doxorubicin hydrochloride(DOX).The results showed that the gradient concentration was stable and controllable,increasing equally from left to right.Using the hydroxyapatite microfluidic chip with Christmas tree structure,real-time monitoring of the apoptosis process of DOX drug-induced osteosarcoma cells(UMR-106)was achieved.The measured drug half-inhibition concentration value was accurate and the method was more efficient.On the other hand,so as to study bone vascularization,using the advantages of hydroxyapatite with good biocompatibility and printability,a bioceramic chip with a complex hollow pipe structure was prepared.The hollow ceramic chip imitated the structure of blood vessels in the bone,and the internal tube diameter ranged from 400 ?m to 1 mm.The results of cell experiments showed that human umbilical vein endothelial cells(HUVEC)adhered well to the hollow tube of the ceramic chip.After 7 days of cultivation,the cells grew into a ring on the inner wall of the tube.When HUVEC was co-cultured with human osteoblast cell line(h FOB),the expression level of vascular endothelial growth factor(VEGF)was significantly up-regulated,which was beneficial to the proliferation of endothelial cells and angiogenesis.To sum up,this study aimed to build a microfluidic chip with a bionic bone microenvironment,used hydroxyapatite for the first time to prepare microfluidic chips,and carried out bone-related drug screening research based on Christmas tree structure and bone vascularization studies based on cell co-culture.These studies are expected to provide a new strategy for the preparation of bone-related microfluidic chips,and promote the further application of microfluidic chips in the fields of bone-related drug screening and bone biology.