| Objective:Lung cancer is a highly malignant tumor with high incidence and poorprognosis, which is a disease of serious threat to human health and life. In recentyears, many countries have reported the incidence and mortality of lung cancerwere significantly increased, and the treatment of lung cancer with surgery,radiotherapy, chemotherapy and immunotherapy. Surgical treatment has beenrecognized as the preferred method for prophase lung cancer, chemotherapy arethe main treatments for unresectable lung cance（rmore than90%of lung cancerrequire chemotherapy treatment）, while, in recent years, immunotherapy is alsoprogressing rapidly.Studies have revealed that the levels of vascular endothelial growth factor(VEGF), transforming growth factor-β1(TGF-β1), interleukin-10(IL-10) andother immunosuppressive molecules in lung cancer cells of culture supernatantsare increased. Tumor cells secrete immunosuppressive molecules inhibitingimmune function,is an important mechanism for tumor cells to escape immunesurveillance and tumor to occur, develop and metastasize. Can we inhibit thesecretion of immune molecules by some substances, to achieve the purpose oftreating cancer?Ganoderma is a traditional Chinese medicine, and it has a history of2000years. Ganoderma lucidum polysaccharides (Gl-PS) is one of the main activecomponents of Ganoderma lucidum, and Gl-PS has actions includingimprovement of immune function and anti-tumor; a number of studies showthat Gl-PS has the effect to inhibit tumor cell directly; and have the effect toinhibit the secretion of immune molecules has not been fully elucidated. Thereis a experiment show that Ganoderma lucidum polysaccharides can antagonizeimmunosuppression of B16F10melanoma cells, but whether Ganoderma lucidum polysaccharides can antagonize immunosuppressive effect of othertumor cells remains to be demonstrated. This study is to investigate the effectof Ganoderma lucidum polysaccharides on immunosuppressive moleculessecreted by LA795cells, to consummate the mechanisms of anti-tumor, and toprovide new ideas for the immunotherapy of lung cancer.Methods:1. Cells: LA795cells is a derived from mouse lung adenocarcinoma celllines.2. Experimental group and the intervention: five groups have been set upin the experiment according to the concentrations of GL-PS in RPMI-1640complete medium of LA795cells, namely0μg/ml group,0.2μg/ml group,0.8μg/ml group,3.2μg/ml group, and12.8μg/ml group.3. ELISA: the expression of VEGF, TGF-β1, IL-10in each group weredetected by ELISA essay.4. RT-qPCR: the expression of VEGF, TGF-β1, IL-10in each group weredetected by qRT-PCR.5. The statistical analysis was performed with SPSS17.0using one-wayanalysis of variance (ANOVA). P<0.05was considered statistically significant.Result:1. The effect of Gl-PS on VEGF secretion in LA795cells was detected byELISA assay and RT-qPCR test: under the action of different concentrations ofGl-PS (0μg/ml,0.2μg/ml,0.8μg/ml,3.2μg/ml,12.8μg/ml), each group ofLA795cells cultured for48h, then detected by ELISA assay. The relativeexpression of VEGF in each group of LA795cells were147.98±0.28、120.25±14.24、103.11±10.57、91.27±13.62、84.66±10.41. The control groupstill express VEGF highly, and with the addition of Gl-PS, the expression ofVEGF decreased (F=15.912, P<0.005). Mutiple comparisons, the differenceswere statistically significant (P<0.05) between0μg/ml group vs0.2μg/mlgroup、0.8μg/ml group、3.2μg/ml group、12.8μg/ml group. These resultssuggest that the level of VEGF in LA795cells can be inhibited byGl-PS.RT-qPCR test showed that the VEGF mRNA expression in each group of LA795cells were1.00±0.00、0.67±0.24、0.68±0.24、0.48±0.21、0.50±0.23.And the control group also express VEGF highly. With the addition of Gl-PS,the VEGF mRNA expression decreased. The difference was significant byanalysis of variance (F=3.034, P<0.1). Mutiple comparisons, the differenceswere statistically significant (P<0.05) between0μg/ml group vs3.2μg/mlgroup、12.8μg/ml group. These results suggest that VEGF gene transcription inLA795cells can be inhibited by Gl-PS.2. The effect of Gl-PS on TGF-β1secretion in LA795cells was detectedby ELISA assay and RT-qPCR test: under the action of different concentrationsof Gl-PS (0μg/ml,0.2μg/ml,0.8μg/ml,3.2μg/ml,12.8μg/ml), each group ofLA795cells cultured for48h, then detected by ELISA essay. The relativeexpression of TGF-β1in each group of LA795cells were98.78±12.02、87.01±5.97、71.98±8.54、58.96±13.24、54.08±14.84. The control group andgroup still high express TGF-β1, and with the addition of Gl-PS, theexpression of TGF-β1decreased. The analysis of variance indicated thedifference was significant (F=8.152, P<0.005). Mutiple comparisons, thedifferences were statistically significant (P<0.05) between0μg/ml group vs3.2μg/ml group、12.8μg/ml group. These results suggest that the level of TGF-β1in LA795cells can be inhibited by Gl-PS.RT-qPCR test showed that theTGF-β1mRNA expression in each group of LA795cells were1.00±0.00、1.24±0.25、0.84±0.22、0.56±0.15、0.46±0.06. The control group and the0.2μg/ml group high express TGF-β1. With the addition of Gl-PS, the TGF-β1mRNA expression decreased. The difference was significant by analysis ofvariance (F=11.145, P<0.005). Mutiple comparisons, the differences werestatistically significant (P<0.05) between0μg/ml group vs3.2μg/ml group、12.8μg/ml group.These results suggest that TGF-β1gene transcription inLA795cells can be inhibited by Gl-PS.3. The effect of Gl-PS on IL-10secretion in LA795cells was detected byELISA assay and RT-qPCR test: under the action of different concentrations ofGl-PS (0μg/ml,0.2μg/ml,0.8μg/ml,3.2μg/ml,12.8μg/ml) group, each group ofLA795cells cultured for48h, then detected by ELISA essay. The relative expression of IL-10in each group of LA795cells were57.04±3.45、46.53±4.23、41.54±1.59、41.54±4.21、36.49±0.80. The control group still highexpressed IL-10, and with the addition of Gl-PS, the expression of IL-10decreased. The analysis of variance indicated the difference was significant(F=17.975, P<0.001). Mutiple comparisons, the differences were statisticallysignificant (P<0.05) between0μg/ml group vs0.2μg/ml、0.8μg/ml、3.2μg/mlgroup、12.8μg/ml group. These results suggest that the level of IL-10in LA795cells can be inhibited by Gl-PS. RT-qPCR test showed that the IL-10mRNAexpression in each group of LA795cells were1.00±0.00、0.98±0.09、0.64±0.17、0.36±0.10、0.39±0.10,. And the control group also high expressIL-10. With the addition of Gl-PS, the IL-10mRNA expression decreased. Thedifference was significant by analysis of variance (F=24.319, P<0.001).Mutiple comparisons, the differences were statistically significant (P<0.05)between0μg/ml group vs0.8μg/ml、3.2μg/ml group、12.8μg/ml group. Theseresults suggest that IL-10gene transcription in LA795cells can be inhibited byGl-PS.Conclusion:1. LA795cells can secrete immunosuppressive molecules VEGF, TGF-β1,IL-10.2. The ability that LA795cells secrete immunosuppressive moleculesVEGF,TGF-β1, IL-10can be reduced by Ganoderma lucidum polysaccharides. |
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