Microfluidic 3D Cell Culture For Nanodrug Evaluation Against Rabbit VX2 Cells

The traditional two-dimensional(2D)cell culture technology has developed for nearly a century.Although they have important value in biomedical research,they can not support the tissue-specific differentiation of various cell types,and can not accurately predict the tissue function and drug activity in vivo.These limitations make people more and more interested in three-dimensional(3D)cell culture models,because they can better reflect the spatial and physiological complexity of living tissue.Microfluidic technology can provide conditions such as cell interaction,chemical gradient and dynamic mechanical force for 3D cell culture,making microfluidic chip a powerful tool for 3D cell culture.In this work,a microfluidic 3D cell culture platform was designed and constructed for the tumor cell culture of rabbit anaplastic epidermal squamous cell carcinoma(VX2),and the killing effects of two kinds of nanodrugs on VX2 tumor cells were investigated.The specific research work includes:1.Establishment of microfluidic 3D cell culture platformMicrofluidic 3D cell culture platform consists of microfluidic chip and osmotic pump.The microfluidic chip consists of two microchannels,the upper and the lower microchannels molded with PDMS,and a polycarbonate membrane sandwiched with extracellular matrix protein.In order to simulate the microenvironment in which cells are exposed to interstitial flow,an osmotic pump is introduced to generate a stable and slow flow rate,which is used to continuously renew the culture medium and timely remove the metabolic waste.Cell activity test showed that the flow rate produced by osmotic pump with 0.05 M PEG is more suitable for the growth of rabbit VX2 tumor cells;the cell growth curve assay shows that the growth curve of rabbit VX2 tumor cells cultured on the chip is "S" shape,and the growth speed is faster than the traditional 2D cell culture;the cell cycle assay shows that the rabbit VX2 cells cultured on the 3D microfluidic chip are more active than the traditional 2D culture.The above results show that we have successfully built a microfluidic 3D cell culture platform.2.Evaluation of the effect of nanodrugs on rabbit VX2 tumor cells via microfluidic 3D cell culture platformOn the basis of the above research work,two kinds of nanodrugs were prepared and their killing effects on VX2 tumor cells of rabbits on microfluidic 3D cell culture platform were investigated.One of the nanodrugs is a self-assembled nanodrug Dox@HCR based on DNA nanowires(the products obtained by hybridization chain reaction,HCR)embedded with doxorubicin(Dox).The other is a self-assembled drug Dox@β-CDP based on the inclusion of Dox by β-cyclodextrin polymer(β-CDP).The internalization results showed that when incubation time was 3 h,the fluorescence of VX2 cells incubated with Dox@β-CDP was the strongest,followed by Dox,Dox@HCR was the weakest.It indicated that the internalization of Dox@β-CDP in VX2 cells was better than that Dox@HCR.The cytotoxic assay showed that Dox@β-CDP had the strongest toxic effect on VX2 cells in traditional 2D culture,followed by Dox,Dox@HCR was the weakest.Compared with the traditional 2D culture,the toxicity of the three drugs to VX2 cells in microfluidic 3D culture was weakened,which may be due to the flow conditions produced by osmotic pump reducing the accumulation of toxic substances.