Effects Of Celecoxib On The Methylation Status,mRNA Expression Of FSTL1, CDH13, TFPI2Promotor And Apoptosis In Squamous Cell Carcinoma Of The Esophagus

Objective: Esophageal cancer is one of the most common malignanttumors that does harm to human health, whose incidence rate rises year byyear. Carcinogenic factors form tumors through different mechanisms andpathways proto oncogene activation and inhibition of tumor suppressor genes.A large number of Clinical studies and experiments show that, in addition togene mutation or deletion, epigenetic is another important reason for thedevelopment of tumor. DNA methylation is one of the important mechanismsof epigenetic, plays an important role in the development of esophageal cancer,and may serve as biomarkers for early diagnosis of esophageal cancer. Thepromoter region of DNA methylation is a chemical modification process, itsintermediate mediating by DNMT (DNA methyl transferase),which belongs toepigenetic aspects. It has been found numerous studies found that, themethylation of FSTL1, CDH13and TFPI2gene promoter results in its lowexpression activities, loss of function. Then the probability of malignant tumorwas significantly increased, which was closely related with the occurrence of,development and metastasis of malignant tumors.In recent years, non steroidal anti-inflammatory drugs (NSAIDS), such asaspirin, ibuprofen, celecoxib, can prevent or reduce the risk of cancer, whichhas become one of the reaearch hot spots in tumor chemical control field. Itcan be show from epidemiological studies that taking the NSAIDS can reducepatients’ mortality of digestive tract cancer, but the specific mechanism is stillin the exploratory stage.This experiment will study the methylation status of FSTL1, CDH13andTFPI2gene promoter regions in esophageal squamous cell carcinoma, and itsinfluence on mRNA expression and cell apoptosis before and after oral administration of celecoxib. I hope this experiment can provide the basictheoretical basis for the antitumor mechanism of NASAIDS and ways to treatesophageal cancer.Methods: Specimens: Twenty-two patients who underwentesophagectomy for squamous cell carcinoma were recruited in the FourthHospital of Hebei Medical University from May2013to January2014. All ofthem were pathologically confirmed esophageal squamous cell carcinoma.There were no history of radiotherapy and chemotherapy, long-term use ofaspirin and other non-steroidal anti-inflammatory drugs. Experimental Group:celecoxib was orally administered in a dose of400mg, bid for10-15daysbefore surgery. Endoscopically biopsied esophageal cancer tissues were takenfrom all recruited patients before celecoxib was administed. Cancer tissueswere taken in the operation for the inveatigation. Control group: Twenty-threepatients who underwent esophagectomy for squamous cell carcinoma wererecruited. There were no history of radiotherapy and chemotherapy, long-termuse of aspirin and other non-steroidal anti-inflammatory drugs. Both thepreoperative and postoperative specimens should be transferred to a-80℃refrigerator storage to spare quickly. The specimens of the control group andthe experimental group must be grinded on ice, and the extracted DNA andRNA were placed in a-80℃refrigerator to prevent degradation. Methylationstatus of DNA were detected by methylation-specific PCR (MSP) technologyand mRNA expression was detected by RT-PCR. Detect the apoptosis rate ofbefore and after celecoxib administration in experimental group. SPSSsoftware package was used for statistical analysis of the experimental data.Rusults:1The methylation status of FSTL1, CDH13, TFPI2gene promoter inexperimental group and control groupFSTL1gene promoter methylation rate was54.55%(12/22) beforecelecoxib administration in experimental group, while that in control groupwas47.83%（11/23）, Pearson χ2=0.203, P=0.652>0.05, the difference was notstatistically significant.The unmethylation rate was50%（6/12）after celecoxib administration, while that in control group was9.1%（1/11）, through Fisher’sexact statistics, P=0.045<0.05, the difference was statistically significant.CDH13gene promoter methylation rate was45.45%(10/22),beforecelecoxib administration in experimental group, while that in control groupwas43.48%（10/23）, Pearson χ2=0.18, P=0.894>0.05, the difference was notstatistically significant. The unmethylation rate was60%（6/10）after celecoxibadministration, while that in control group was0%（0/11）, through Fisher’sexact statistics, P=0.043<0.05, the difference was statistically significant.TFPI2gene promoter methylation rate was50%(11/22) before celecoxibadministration in experimental group, while that in control group was47.83%（11/23）, Pearson χ2=0.021,P=0.884>0.05, the difference was not statisticallysignificant. The unmethylation rate was54.55%（6/11） after celecoxibadministration, while that in control group was9.1%（1/11）, through Fisher’sexact statistics, P=0.032<0.05, the difference was statistically significant.2The comparison of mRNA expression of FSTL1, CDH13, TFPI2before and after celecoxib treatment administrationAfter celecoxib administration, FSTL1gene mRNA relative IOD value(0.675±0.20087) was significantly higher than that (0.292±0.09866) beforecelecoxib administration, and the difference was statistically significant(t=10.427, P=0.000).After celecoxib administration, CDH13gene mRNA relative IOD value(0.303±0.17327) was significantly higher than that (0.098±0.08551) beforecelecoxib administration, and the difference was statistically significant(t=4.921, P=0.001).After celecoxib administration, the TFPI2gene mRNA relative IODvalue (0.476±0.06937) was significantly higher than that (0.219±0.05917)before celecoxib administration and the difference was statistically significant(t=14.709, P=0.000).3The apoptosis status before and after celecoxib administrationThe overall apoptosis rate was15.98±64.298%before celecoxibadministration, while after celecoxib administration it was26.65±3.798%. The difference was statistically significant (t=-11.996, P=0.000<0.05).Conclusion:1Celecoxib can reduce the degree of methylation of FSTL1, CDH13,TFPI2gene promoter methylation.2Celecoxib can improve the expression of FSTL1, CDH13, TFPI2genein esophageal squamous cell carcinoma.3Celecoxib can induce apoptosis in esophageal squamous cellcarcinoma.