HO-1 Effect And Its Mechanism Of Tumor Cell Biology Function Of PPARα And YY1 Regulation

Part one: PPARα activation sensitizes cancer cells toepigallocatechin-3-gallate (EGCG) and ionizingradiationObjective: To investigate the effect of PPARα activation on EGCG and ionizingradiation efficacy and its underlying mechanisms. Methods: CCK-8kit was used todetect cell viability. Western Blotting and real-time PCR was used to measure theprotein and mRNA level, respectively. The PPARα agonist clofibrate and inhibitorGW6471were used to alter PPARα expression. Luciferase reporter system andchromatin immunoprecipitation (ChIP) were used to investigate the effect of PPARα onHO-1expression. Clogenic assay was used to investigate the radiosensitivity of cancercells. mRNA microarray was used to explore the involved genes. Results: When cancercells were exposed to EGCG, the expression of PPARα was increased at the proteinlevel in a dose-dependent manner. The PPARα agonist clofibrate blocked hemeoxygenase (HO-1) induction and sensitized cancer cells to EGCG-induced cell death.However, the PPARα inhibitor GW6471increased HO-1expression. In vivo ChIPconfirmed that PPARα interacts with the peroxisome proliferator-responsive element(PPRE) of the HO-1promoter. PPARα activation can increase the radiosensitivity ofcancer cells via complex mechanisms. In addition, clofibrate could significantlyincrease the radiosensitivity of pancreatic cells. Microarray revealed hundreds of genesin multiple pathways were involved especially receptor signaling pathwys, indicatingcomplex mechanisms. Conclusions: PPARα is a direct negative regulator of HO-1activation by EGCG and confers cell susceptibility to EGCG. PPARα activation canincrease the radiosensitivity of cancer cells via complex mechanisms. Part two: Upregulation of Ying Yang1(YY1)suppresses esophageal squamous cell carcinomadevelopment through heme oxygenase-1Objective: To investigate the expression of YY1in normal and cancer-ousesophageal tissues and its function in ESCC development. Methods: The mRNA levelsof YY1were determined by reverse transcription-PCR and real-time PCR; The proteinlevels of YY1gene in102esophageal squamous cell carcinomas (ESCC) weredetermined by immunohistochemistry. YY1overexpression vector and YY1silencingvectors were constructed. Protein levels of YY1HO-1and Nrf2were determined byWestern Blot. Protein interaction was determined by co-immunoprecipitation.Regulation of HO-1by YY1was determined by luciferase assay HO-1-overexpressingvirus Ad-HO-1or control virus Ad-EGFP. to explore the impacts of HO-1onesophageal squamous cell carcinomas. Results: The expression of YY1mRNA wassignificantly increased in the tumor tissues, compared with the para-tissues or normalesophageal tissues. The increased expression of YY1in tumor samples was furtherconfirmed by immunohistochemistry. Furthermore, the overexpression of YY1conferred radioresistance to the ESCC TE-1cells and resulted in markedly reduced cellproliferation. Accordingly, the small interfering RNA-mediated silencing of YY1expression in TE-1cells resulted in increased pro-liferation by enhancing the binding ofP21to Cyclin D1and CDK4, a protein complex known to mediate cell cycleprogres-sion. Moreover, besides P21, heme oxygenase-1(HO-1) was identified as aYY1downstream effector, as YY1stimulated HO-1expression in esophageal cancercells. YY1mediated biological function through transcription of HO-1. Forcedexpression of HO-1could moderately suppress proliferation of TE-1cells. Theexpression of YY1significantly correlated with that of HO-1in ESCC tissues.Conclusions: Overexpression of YY1in esophageal carcinoma suppressed ESCCproliferation via HO-1.