Co-culture Of Probiotics And Colorectal Cancer Cells Based On Microfluidic Chip Technology

Intestinal flora plays an important role in human’s immune system,helping to digest and stop their disease-causing counterparts from invading.A large number of bacteria adheres the intestinal wall to prevent the epidermal cells from contacting with the harmful substances directly.The change of host health is accompanied by the diversification of Intestinal microorganism.The complexity of the intestinal environment led to a lot of factors which were researched.Therefore,it is very important to explore the specific mechanism of intestinal bacteria and its host.It is necessary to develop an appropriate 3D model to simulate intestinal microenvironment aiming at exploring the relationship between microorganism in the intestine and human health.Based on the microfluidic chip technology,we constructed a model of co-culture of intestinal cells and microbial to simulate intestinal microenvironment.In the experiment,Clostridium butyricum MIYAIRI 588 and colon cancer cells HCT116 and Caco-2 were selected as the representatives.The main research content and results are as follow.(1)To investigate its tolerance to gastrointestinal tract environment,Clostridium butyricum MIYAIRI 588 was activated and analyzed.The metabolic status of the strain showed good tolerance to gastrointestinal tract.Therefore,it had good value.Detection of metabolites of culture medium DMEM indicated Clostridium butyricum MIYAIRI 588 could ferment glucose into a variety of short chain fatty acids and regulated the generation of amino acids as well as generated various B vitamins at the same time.(2)A microfluidic chip model was constructed to simulate intestinal structure.The reliability of the device was detected.By testing,105 cells/mL was selected as the optimum initial culture concentration of the injection cells and the perfusion velocity of microorganism medium was 5 μL/min.Furthermore,co-culture experiment was carried out by injecting cells and bacteria into the chip in succession.The results showed that cells grew well with 4 hours,demonstrating that the chip can be used for the simulation of intestinal microenvironment.(3)Cell morphology,activity,cell cycle and apoptosis,the co-culture of Clostridium butyricum MIYAIRI 588 and colon cancer cells showed a different immune effect.The apoptosis of HCT116 cells was accelerated by the effect of co-culture,but Caco-2 cells were not exposed to serious damage.(4)To further understood the effect of Clostridium butyricum MIYAIRI 588,this study explored the effect of butyrate on cells,butyrate was the main product of Clostridium butyricum MIYAIRI 588.The result showed life activities of two kinds of cells which were disturbed in varying degrees.Butyric produced by Clostridium butyricum MIYAIRI 588 played a role in regulating the apoptosis of colon cancer cells.At the same time,there might be other mechanisms that played a role in the growth of colon cancer cells.