Study Of Circadian Rhythm Genes HPer2 And HCry1 On Chemosensitivity Of Esophageal Cancer

Objectives: To investigate the expression of PER2、CRY1 protein inesophageal cancer tissues and adjacent normal tissues to show its clinicalrelevance．To detect of the expression level of hPer2 and hCry1 in Eca-109esophageal cancer cell lines at different time points,and then to observe thechemosensitivity of Eca-109 cell lines to docetaxel and cisplatin at the peakand nadir time point respectively.Methods: The expressions of PER2 and CRY1 protein from 40 surgicalspecimens of esophageal cancer tissue and adjacent normal tissue mucosawere determined by imunohistochemistry.Circadian gene expression ofEca-109 cell lines were induced by dark-culture,total RNA were extractedevery two hours,and then expression level of the two circadian weredetermined by RT-PCR. Cells were divided into three groups according to thedifferent culture conditions and dosing time points.Docetaxel plus cisplatinsolutions which were diluted to the following concentration gradients10^-8,10^-7,10^-6,10^-5,10^-4,10^-3μM,were given at the peak time point,nadir timepoint and conventional time （8:00⁹:00） respectively.The inhibition ofEca-109 cell proliferation was measured using a CCK-8 kit.Results: The weak-positive/positive（which score≧2） expression rate ofCRY1,PER2 protein from esophageal carcinoma tissue is 17.5%,40%respectively,while the positive expression rate of CRY1, PER2 protein fromadjacent normal tissue is 75%,92.5% respectively.The peak expression ofhPer2 mRNA was at 24:00 and the nadir at 10:00;The peak expression of hCry1 mRNA was at about 4:00 and the nadir value was at about 14:00. Aftertreating with the docetaxel plus cisplatin all the three groups known as thegroup1（dark-culture,24:00²:00）,group2（dark-culture,12:00¹4:00）,group3（r-outine culture,8:00⁹:00）.All the three groups known as group1,2,3,showedinhibition of cell proliferation under the concentration of 10^-6,10^-5,10^-4,10^-3μM.The maximum inhibitory concentration was found to be 10-5μM. There aresignificent differences in inhibitory rate among the three groups under eacheffective inhibitory concentration. The maximum inhibitory rate is in group2（at the nadir time point）.Conculsion: There is significent differences in expression of CRY1 andPER2 between esophageal cancer tissues and adjacent normal tissues.Wefound a phase fluctuations in the expression of hPer2 and hCry1 gene.Whenchemotherapy drugs were given at nadir time point of the two genes’expression,there is a higher inhibition rate of cell proliferation compared withthat given at peak and conventional time.