| Background and objective:Esophageal cancer(EC)is one of the most common gastrointestinal malignancies in the world.According to the latest survey statistics,the number of new cases and deaths of EC in the world in 2018 were 572,034 and 508,585,ranking ninth and sixth in malignant tumors.China is a country with a high incidence of EC,and the number of cases of EC each year ranks third and fourth among malignant tumors,respectively.At present,the treatment of EC mainly includes traditional surgery,radiotherapy,chemotherapy,etc.The emergence of immunotherapy provides new treatment options for patients with EC.In recent years,with the improvement of medical technology,the disease control rate and quality of life of patients with EC have been significantly improved,but the five-year survival rate is still very low,only 15%to 20%.According to the classification of pathology,EC can be divided into two types:esophageal adenocarcinoma and esophageal squamous cell carcinoma(ESCC).In my country,ESCC is the most common,accounting for about 90%of the total number of patients with esophageal cancer.Therefore,it is very important to clarify the pathogenesis and progression mechanism of ESCC at the molecular level,find effective influencing factors and predictors of the prognosis of patients with ESCC,and thus make early and targeted therapeutic intervention.According to the recent studies,arachidonic acid(AA)and its metabolism play an important role in tumor progression.The purpose of this study was to investigate the role of AA’s metabolic enzyme-12-lipoxygenase(12-LOX)and its metabolite 12-HETE in the progress of ESCC.Methods:1.ESCC cells were treated with 12-LOX inhibitor-Baicalein to reduce its 12-LOX expression level,and group with 12-HETE pretreatment was set as a compensation group.Twenty-four hours later,the Transwell invasion and migration test was used to detect the invasion and migration ability of cells in different treatment groups,and Western blot was used to detect the expression of TGF-β1 and epithelial-to-mesenchymal transition(EMT)markers(vimentin,N-cadherin,E-cadherin)2.12-LOX-specific siRNA(Si 12-LOX)was designed to knock down the expression of 12-LOX in ESCC cells,and the group pretreated with 12-HETE/TGF-β1was also set as the compensation group,and after 24 h,the expressions of TGF-β1 and EMT markers in cells of different treatment groups were detect by Western blot.3.Immunohistochemical staining was used to detect the expression levels of 12-LOX,TGF-β1,and EMT markers in tumor tissues of ESCC patients,and the correlation between 12-LOX and clinicopathological characteristics,12-LOX and prognosis,as well as the correlation of 12-LOX and TGF-β1,EMT markers in ESCC tissues and was statistically analyzedResults:1.Western blot results showed that compared with the control group,20 μM and 40 μM Baicalein can inhibit the expression of 12-LOX in ESCC cells,and the inhibitory effect was more significant at the concentration of 40 μM.Transwell invasion and migration experiments showed that 40 μM Baicalein can inhibit the invasion and migration ability of ESCC cells,while 12-HETE pretreatment can partially alleviate the above inhibition.2.Western blot results showed that after Baicalein/Si12-LOX treatment,the expression level of 12-LOX in ESCC cells decreased,and the expressions of TGF-β1 and EMT were suppressed(vimentin and N-cadherin expression were down-regulated,and E-cadherin expression was up-regulated),12-HETE pretreatment can partially alleviate the above inhibition.TGF-β1 pretreatment can also partially relieve EMT levels,while TGF-β1 expression did not change significantly.This indicates that 12-LOX/12-HETE may improve the invasion and migration ability of ESCC cells by promoting TGF-β1-mediated EMT process.3.Immunohistochemical staining of tumor tissues of patients with ESCC showed that the expression of 12-LOX was positively correlated with TGF-β1,vimentin,N-cadherin,but negatively correlated with the expression of E-cadherin.This result further confirms that there is a close correlation between 12-LOX,TGF-β1 and EMT.At the same time,clinical analysis showed that the expression level of 12-LOX was positively correlated with N stage(P=0.001)and pathological TNM stage(P=0.022),but was not related to the patient’s gender,age,smoking,drinking,T stage(P>0.05).4.Kaplan-Meier and log-rank survival analysis showed that compared with the 12-LOX low expression group,the overall survival of patients in the 12-LOX overexpression group was significantly reduced(P=0.002).Multivariate analysis showed that 12-LOX overexpression was an independent prognostic risk factor for patients with ESCC[HR(95%CI):2.371(1.250,4.496),P=0.008].Conclusion:1.12-LOX can enhance the migration and invasion of ESCC tumor cells by promoting the TGF-β1-mediated EMT process;2.The expression of 12-LOX in ESCC patients is positively correlated with clinical pathological characteristics such as tumor stage and lymph node metastasis;3.The expression level of 12-LOX was negatively correlated with the overall survival of ESCC patients;4.12-LOX expression is expected to become a new target and prognostic biomarker for ESCC patients. |
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