The Role Of PTCH1Gene Methylation In The Development Of Esophagus Cancer

ObjectiveTo study the relation between PTCH1gene methylation and esophagealcancer occurrence and development and it’s mechanism of action, meanwhilediscussit’s treatment and prognosis which methylateon transferase inhibitor5-Aza-dC to esophageal cancer.METHODS(1)Bisulfite sequencing PCR （BSP）was employed to test the situation ofPTCHl gene methylate in people’s Eca109esophagus cancer cell lines,esophageal gland cancer of esophagus cancer tissue and normal tissue adjacentcarcinoma from patientswho did not after radiation and chemotherapy treatment,methylated CpG sites of the total number of detection of the CpG site whichhappend is used to be the quantitative indicator, use matching t test for PTCHlexpression and its gene methylation degree of correlation analysis and therelation between PTCH1gene methylation and esophageal cancer occurrenceand development.(2) Using different concentration of methylation transferaseinhibitor5-Aza-dC on esophageal cancer cell line Eca109to do the drugintervention, observed the cell type under inverted microscope, AnnexinV/PIapoptosis detection kit cell apoptosis, real-time quantitative PCR detection ofdifferent concentrations of5-Aza-dC drug intervention Eca109cells afterPTCHlmRNA relative to express the quantity change, the data pf each treatmentgroup and control group were statistically analyzed by t test.RESULTS(1) Use the NBCI Find PCTH1transcription body found7transcripts andtranscription protein Methyl Primer Express software v1.0PTCH11b mRNAtranscripts-2580~5341bp CpG island analysis found six CpG island-820~ 426bp amplified BSP primers designed according to the needs, containing19CpG islands.(2)30patients with cancer organization next the PTCH1gene methylationseven cases, not fully methylated cancer adjacent tissues are not fullymethylated the the esophageal tissue PTCH1gene methylation in29cases,which completely methylated in19cases,11cases of partially methylated bythe statistical comparison, esophageal tissue with paraneoplastic organizationmethylation rate exist significant differences (P <0.05).(3) esophageal cancer cell line Eca109DNA is almost complete methylation,the degree of methylation (99.6±0.72)%, the majority of the esophageal tissueDNA methylation, the degree of methylation of18%to99%the average degreeof methylation (64.6±5.72)%, the cancer tissue methylation level of0%to37%,the average degree of methylation (16.6±2.72)%next after comparing cancerorganizations and esophagus the degree of methylation in cancer tissue therewere significant differences (P <0.01).(4) different concentrations of5-Aza-dC drugs interfere with the Eca109cellline measured490nm OD at concrete results are shown in Table1,1um,2um,5um group compared with the control group OD value, there are significantdifference (P <0.05),10um OD value of the control group, no significantdifference (P>0.05).(5) different concentrations after the intervention, the apoptosis rate of5-Aza-dC group compared with the control,1um,2um,5um group comparedwith the control group, the apoptosis rate there is a statistically significantdifference (P <0.05),10um comparison with the control group, the apoptosisrate was no significant difference (P>0.05).CONCLUSION(1) Esophageal cancer PTCHl gene promoter region CpG island happenmethylation is the molecular events which esophagus cancer organizationdifferent from normal esophageal, it can to participate in the occurrence ofesopha--gus cancer development by inhibiting the expression of PTCHl gene.PTCHl gene methylation level can be used as a new molecular markers todiagnosis and differential diagnosis of esophagus cancer early.(2) PTCHl gene methylation in esophageal cancer cell lines Eca109can usemethylation transferase inhibitor5-Aza-dC drugs for methylation processing,but 5-Aza-dC drugs maye functionin in a certain concentration rang,so it could beused for esophageal cancer treatment and prognosis.