Expressions And Effects Of Granolocyte-macrophage-colony Stimulating Factor Combined With Its Receptor In Esophageal Cancer

Objective:Granolocyte-macrophage-colony stimulating factor(GM-CSF)is one of the most common growth factors in hematological system,which functions to promote maturation and differentiation of blood cells and facilitate body immune effect on binding with its specialized receptor(GM-CSFR).Studies demonstrated that GM-CSF expression level was up-regulated in different malignancies including head and neck cancer,breast cancer,lung cancer,colorectal cancer and prostate cancer.Moreover,GM-CSF could regulate tumor growth and progression through immune-dependent and independent manners.But investigation of GM-CSF in esophageal cancer(EC)is rarely seen.Thus,we performed this study to explore expression levels of GM-CSF/-R in EC on aspects of protein and biopsy,and to illustrate its biological effect by laboratory experiments and clinical retrospective studies.Methods:1.Detection of GM-CSF/-R expression in EC1.1 Western blot was applied for detecting expression levels of GM-CSF/-R in Eca109 and 9706 cells.1.2 Immunochemistry was performed on paraffin-embedded sections for detecting GM-CSF/-R expression in EC patients,scores were evaluated according to staining intensity and proportion of positive tumor cells,and the outcomes were classified into negative,low-expression and high expression.2.Research of biological effect of GM-CSF in EC2.1 Influences of exogenous GM-CSF on Eca109 cell biological behaviors2.1.1 Cell Counting Kit-8(CCK-8)essay was used to compare differences of proliferation ability between Eca109 cells with and without exogenous GM-CSF application at 24h、48h、72h and 96h;2.1.2 Flow cytometric essay was utilized to analyze differences of cell apoptosis between Eca109 cells with and without exogenous GM-CSF application;2.1.3 Cell migration and invasion capacity differences between Eca109 cells with and without exogenous GM-CSF application were observed by counting cell numbers traverse through the Transwell chamber membrane;2.2 Predictive value of serum GM-CSF change during radiotherapy on treatment efficacy in EC patients2.2.1 Enzyme-linked immunosorbent assay(Elisa)was applied to detect serum GM-CSF change during radiotherapy,and Log-rank test was used to analyze its prognostic effect;2.2.2 We used Cox proportional hazard model to determine whether serum GM-CSF change during radiotherapy could be an independent prognostic factor;Results1.Detection of GM-CSF/-R expression in EC1.1 Western blot revealed significant GM-CSF expression in Eca109 cells and 9706 cells;GM-CSFR was high in Eca109 cells,and the expression level was higher than the result observed in 9706 cells(p<0.05);1.2 Pathological sections from 93 EC patients were analyzed in this study.All patients showed positive GM-CSF expression,41 were high-expressed and 52 were low-expressed.For GM-CSFR expression,55.9%patients suggested positive outcomes,45 were high-expressed,7 were low-expressed and 41 were negative.2.Research of biological effect of GM-CSF in EC2.1 Influences of exogenous GM-CSF on Eca109 cell biological behaviors2.1.1 Proliferation ability of Eca109 cells with GM-CSF stimulation were significantly better than the group without GM-CSF at 72(p=0.001)and 96 hours(p=0.003),but no difference was detected at 24 and 48 hours;2.1.2 After being cultured in complete medium for 48 hours,no significant difference of Eca109 cell apoptosis was observed between GM-CSF stimulated group and the contrast group(p>0.05),indicating that GM-CSF may exert no effect on cell apoptosis;2.1.3 In GM-CSF stimulated group,Eca109 cell counts traversing the Transwell chamber membrane were significantly reduced compared with the control group(all p<0.05),suggesting that GM-CSF could significantly repress cell migration and invasion.2.2 Predictive value of serum GM-CSF change during radiotherapy on treatment efficacy in EC patients2.2.1 A total of 54 EC patients were enrolled in this study,23 indicated elevated serum GM-CSF level during radiotherapy,while 31 showed the opposite results.Patients with elevated serum GM-CSF level suggested significant longer overall survival(p=0.002)and progression-free survival(p=0.001)than down-regulated patients.2.2.2 GM-CSF change during radiotherapy was an independent prognostic factor for EC patients(p=0.001,HR:0.244,95%CI:0.106-0.564),which indicated stronger prognostic effect than disease stage(p=0.001,HR:0.275,95%CI:0.127-0.596).ConclusionGM-CSF and its receptor both showed certain expression levels in EC.Exogenous use of GM-CSF could significantly repress cell proliferation,migration and invasion,but showed no effect on cell apoptosis.Moreover,elevated serum GM-CSF level during radiotherapy predicted favorable outcomes in EC patients.Therefore,GM-CSF demonstrated anti-tumor effect in this study,further exploration of potential mechanisms under GM-CSF may play a critical role in EC treatment.