Chemotherapy Vector-mediated Expression Of Immunogenicity Of Renal Carcinoma Cells In Vitro

Objective To investigate the immunogenicity associated protein of calreticulin(CRT) exposure in renal carcinoma cell line 786-0 by the chemotherapeutants Gemcitabine(GMC) and 5-fluorouracil(5-FU). To explore the optional concentrations with which the chemotherapeutants can induce CRT translocation to the cell surface effectively. To collaborate therapy of biological immune, provides experimental data, strengthens theoretical support for the treatment of kidney cancer, and provides a new approach to the method. Methods The inhibition of cell line 786-0 by GMC and 5-FU was tested with the CCK-8 assay. The CRT exposure of the cell cultured in different chemotherapeutants were measured by flow cytometry (FCM).Results1. The results of CCK-8 assay: The inhibiting rates of 786-0 cell line were increasing as the concentrations of each of two chemotherapeutants got higher. The median inhibition concentration (IC50) of GMC for 24h was 1.2ug/ml, 5-FU was 40ug/ml.2. CRT expression on 786-0 cell surface treatmented with two chemotherapeutants of IC50: 786-0 cells were treated for 24h with 0.01mol/L PBS,1.2ug/ml GMC,40ug/ml5-FU, then the surface expression of CRT were determined by FCM, the Average fluorescence intensity of the group of PBS was: 59.22±3.27, the group of GMC was : 179.21±13.22, the group of 5-FU was : 145.6±8.23. GMC and 5-FU can increase CRT expression effectively . there is a significant difference compared with control group (P <0.01). CRT exposure were induced effectively by two chemotherapeutants, the expression of CRT was stronger compared with 5-FU.3. CRT expression on 786-0 cell surface treated with different concentrations of GMC and 5-FU :①the Average fluorescence intensity of CRT expression was increasing with increased GMC concentrations , But when the concentration reach 8 x IC50, its higher amplitude no longer increased significantly.②the Average fluorescence intensity of CRT expression was increasing with increased 5-FU concentrations too, But when the concentration reach 8 x IC50, its higher amplitude no longer increased significantly. Through the statistical analysis, each of the groups of GMC and 5-FU was significantly higher than the control group (P<0.01). the group of GMC was significantly higher than the 5-FU (P<0.01). it was no statistical significance between the groups of the GMC of 4×IC50 and 8×IC50 (t=0.676,P=0.876>0.05). it was no statistical significance between the groups of the 5-FU of 4×IC50 and 8×IC50 (t=-1.308,P=0.187>0.05). Except that, there is a significant difference among other concentrations (P<0.01).ConclusionsIn our study, we processed renal clear lines of 786-0 cells With different concentrations of the GMC and 5-Fu, to test the expression of CRT on the cell membrane surface by FCM. To explore the optional concentrations with which the chemotherapeutants can induce the pression of CRT on the cell surface effectively. Explore the way and dose of the combination therapy of the chemotherapy and biological immune therapy , summarizes these.1. GMC and 5-FU can induce CRT exposure to stimulate the immunogenicity of tumor cell death, and can improve CRT expression on 786-0 cell surface effectively.2. treated with different concentrations of GMC and 5-FU ,the CRT expression upon renal clear lines of 786-0 cells surface is different. The expression of CRT was promoted obviously when Two drugs in lower concentrations processed. So, it is appropriate with lower doses chemotherapy drugs to induce tumor cells immunogenicity death.