| Objectives:Microfluidic chip system is the main means recognized by the scientists. Lung cancer is the most frequently occurring malignancies the leading cause of death in the world. Chemotherapy is one of the common used methods. But because of the resistance, patients are always dead. Based on the Microfluidic chip system, this study does the research on the function of a stress protein-Glucose Regulated Protein 78 (GRP78) on the resistance of the lung cancer cells. Microchip-based systems have many desirable characteristics and can be used in much cellular biochemical analysis. Glucose-regulated protein 78 (GRP78), an endoplasmic reticulum chaperone, has a critical role in chemotherapy resistance of some cancers. This work aimed at analyzing the correlation between the expression of GRP78 and anticancer drug VP-16 in human non-small lung cancer cell line SK-MES-1 using this microchip-based system. Meanwhile, this work may provide an ideal and high-throughput platform for cell culturing and cell biological study.Methods:An integrated microfluidic device which contains an upstream network called concentration gradient generator (CGG) module and a downstream cell culture chamber module was used. The cell suspension was injected into the microfluidic device at 5×106 cells/ml via the downstream cell inlet by syringe pump. A continuous concentration gradient of stimulator (A23187) was generated in the upstream network through a binary input chamber and used to cultured downstream cells that have been cultured for 24h. The activity of cells before induced was detected by 0.2% trepan blue staining. The expression of GRP78 at protein levels is detected by using immunofluorescence. Then the cells were treated by medium that contained VP-16 for 6h, and then change the medium and cultured the cells for 48h. Apoptosis of cells was assessed morphologically by DAPI—a nuclear staining in order to survey cytoplasm concentration and karyopyknosis, and observe the state of apoptosis cells under the different concentration of the inducer. The resultant fluorescence of cellular population was monitored with fluorescence microscope.Results:The cells cultured on the integrated microfluidic device were in good condition. The result with less than 1% of dead cells suggested that the cell culture chambers provide a suitable environment for cell maintenance. The generated concentration gradient was good coherence between the experimental and theoretical data. The expressions of GRP78 at protein levels for the A23187-induced cells that detected by the immunofluorescence on the microchip were increased. After the treatment by VP-16, the percentage of apoptotic cells decreased with the concentration increasing of the inducer. GRP78 plays an important role in the resistance to VP-16 in human lung cancer SK-MES-1 cell line.Conclusion:This study integrated cell culture, generation of multi-drug concentration, immunofluorescence and detection of apoptosis on a microfluidic chip. The results demonstrated that overexpression of GRP78 induced by calcium ionophore A23187 directly conferred resistance to VP-16 induced apoptosis in human non-small cell lung carcinoma SK-MES-1 cell line. This microfluidic device offers a unique platform to characterize cellular responses in a high-throughput fashion, whereas impossible with conventional methods. This system was valuable in the cell culture and the detection of cell components and deserved to be studied further. |
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