| Microfluidic chip technology is a technology that uses an internal micron-sized chip as an operating platform and can integrate sample preparation,separation,and reaction in the fields of biology,physics,medicine,and chemistry on this small chip.Due to their special size,the chips have many advantages such as controllable liquid flow,lightweight size,low sample usage and less consumption of drugs,etc.It has become an important research direction in many subject areas.At present,the cost of oncology drug research is very high,and most of the reasons are from the in vitro detection stage.A large number of repeated animal experiments are required before entering the clinical test.In addition to the above-mentioned cost problems,there are still ethical problems and animal uncertainty in animal experiments,which is the difficulty of the entire drug development chain.Therefore,this topic was based on microfluidic chip as a platform for cell culture,combined with three-dimensional cell culture technology and hydrogel technology to establish a tumor evaluation model,in order to gradually replace animal experiments for drug in vitro detection.The research contents are as follows:A semi-closed chip was designed with a microarray column as the cell culture chamber,which was to reduce the damage of the fluid shearing force to the cells and to ensure that the cells are not washed away by the liquid.The upper layer was polydimethylsiloxane(PDMS)with good air permeability and biocompatibility,and the lower layer was covered with a glass slide to provide an operating platform for three-dimensional culture of cells.In order to provide a good three-dimensional scaffold to the cells,thermosensitive PNA hydrogels were prepared by copolymerizing N-isopropylacrylamide(NIPAM)and acrylic acid(AA),where N,N’-methylenebis(acrylamide)(BIS)as the crosslinker.Since the thermosensitive hydrogel has a phase change characteristic with the temperature,it was more convenient to operate.Furthermore,the RGD was grafted onto the PNA hydrogels to explore the effects of RGD peptides on the behaviors of cells.SK-OV-3 and HepG2 cells were separately mixed with PNA hydrogel in equal volumes and added to the chip culture chamber for culture.According to the results,it was determined that SK-OV-3 cells were in the logarithmic growth phase on the 4th day and HepG2 cells were in the logarithmic growth phase on the 6th day,and the best time to administrate drug was the logarithmic growth phase.SK-OV-3 cells and HepG2 cells were mixed with PNA-RGD hydrogels in a chip,and the absorbance of the cells was compared with the data of PNA.Both results showed that the growth status of cells in PNA-RGD was better than that in PNA,indicating the preliminary establishment of the two multi-cell sphere evaluation models and the promotion of cell growth and adhesion by RGD.SK-OV-3 cells were cultured in PNA for 4 days and then administrated different concentrations of doxorubicin.The cell growth was slowed after administration according to the results,and the drug resistance was observed after 72 h in the experimental group of 5.00 μg/mL.SK-OV-3 was tested for drug susceptibility in PNA-RGD,and the cells were more obviously affected by the drug,and the drug resistance appeared in the 2.00 μg/mL group after 72 h.HepG2 cells were cultured in PNA and PNA-RGD for 6 days and then doxorubicin was administered for drug sensitivity test.According to the results,the inhibitory effect of the drug on the cell mass was enhanced with the increase of the concentration both in the PNA or PNARGD,and the drug resistance was exhibited at the concentration of 5.00 μg/mL.The results analysis revealed that the two simple tumor SK-OV-3 and HepG2 evaluation models were successfully established and could be used for preliminary detection of drugs.In addition,the RGD peptide in this experiment could not only promote cell adhesion and growth,but also promote the inhibition of the drug doxorubicin to HepG2 cell mass. |
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