Mechanism Of Inflammation Mediated Epigenetic Regulation Of SOCS3 Gene In Pancreatic Cancer And Its Significance

Background: Inflammatory microenvironment plays an important role in pancreatic cancer which is characterized by pronounced fibro-inflammatory reaction.STAT3 signaling pathway is regarded as the "bridge" between inflammation and cancer,and involved in the development of pancreatic cancer.SOCS3 is a key negative feedback regulator of JAK/STAT signaling pathway.Recent years,more and more studies focused on the abnormal expression of SOCS3 in various tumors.However,studies about SOCS3 gene in pancreatic cancer are rarely reported.Our study aimed to explore the regulatory mechanism of SOCS3 expression in pancreatic cancer and its role in pancreatic cancer development,which may offer new theoretical basis and potential therapeutic targets for the treatment of pancreatic cancer.Methods: 1.Differential expression of IL-6/pSTAT3/SOCS3 signaling and DNMT1 /DNMT3 a among pancreatic cancer tissues,matched pericancerous tissues and cell lines were detected by Western blot and immunohistochemistry tests.2.Effects on expression of SOCS3 and DNMT1 /DNMT3 a in pancreatic cancer cell lines after treatment with IL-6 and S3I-201 were studied by Western blot and qRT-PCR.3.Methylation status of SOCS3 gene promoter region was detected by MSP after treatment with IL-6 and S3I-201.We also evaluated SOCS3 protein expression induced by IL-6 in the presence of 5-Aza-cdR or DNMT1 knocked-down.4.We evaluated the roles of IL-6/STAT3 signaling pathway and DNMT1 on transcription of SOCS3 gene promoter by immune co-precipitation and dual luciferase reporter gene experiments.5.We observed the effects of SOCS3 gene on biological behavior of pancreatic cancer cells through apoptosis,proliferation,migration,invasion and tumor formation in nude mice.6.We observed SOCS3 expression in pancreatic cancer tissue microarray and correlated with the clinical pathological characteristics and prognosis of pancreatic cancers.Results: 1.We observed that SOCS3 expression was down-regulated in pancreatic cancer tissues,while IL-6,pSTAT3 and DNMT1 expression were up-regulated by immunohistochemistry and Western blot.2.Inflammatory cytokines IL-6 can promote the expression of DNMT1 through activation of STAT3 signaling pathways and also increase the global DNA methylation level of pancreatic cancer cells;STAT3 activation negatively regulated SOCS3 expression in pancreatic cancer cells.3.STAT3 activation promoted hypermethylation of SOCS3 gene promoters.Silence DNMT1 expression or 5-Aza-cdR treatment can reverse IL-6/STAT3 signaling pathway mediated down-regulation of SOCS3 in pancreatic cancer cells.4.We observed that phosphorylated STAT3 recruited DNMT1 to SOCS3 promoter regions and suppressed its transcription activity by immune co-precipitation and dual luciferase report gene tests.5.Overexpression of SOCS3 inhibited proliferation,migration,invasion and tumorigenetic ability of pancreatic cancer cells and the possible mechanisms included induction of G1/S cell cycle arrest and cell apoptosis,inhibition of STAT3 activation and expression of relevant downstream target genes.6.Low expression of SOCS3 in pancreatic cancer tissue microarray by immunohistochemical analysis correlated with lymph node metastasis and advanced clinical stage.Patients with high expression of SOCS3 in pancreatic cancers often indicated a relatively good prognosis.Conclusions: We demonstrated that activated IL-6/STAT3 signaling could induce SOCS3 methylation via DNMT1,which led to imbalance and sustained activation of STAT3 signaling pathway.The reduced expression of SOCS3 promoted the growth and metastasis of pancreatic cancer.Thus,targeting IL-6/STAT3 signaling pathway or DNMT1 may become an important treatment strategy of pancreatic cancer.