Effect Of MUC1 On The Biological Behaviour Of Salivary Gland AdCC And MEC And Related Molecular Mechanisms

Salivary gland cancers are important parts of head and neck cancers.The complicated pathological patterns and varied prognosis of salivary gland cancers pose a great challenge to the clinical diagnosis and treatment.Clinical experiences and researches show that advanced salivary gland cancers(e.g.mucoepidermoid carcinoma)characterized with invasiveness and poorly outcome due to poor sensitive to radiotherapy and chemotherapy.Hence,seeking for therapeutic target for the treatment of salivary gland cancers is extremely urgent.MUC1 is a highly-glycosylated transmembrane glycoprotein which aberrantly glycosylated and over-expressed in many carcinomas and associated with poor outcomes.Researches shows that MUC1 may be a potential biomarker and therapeutic target for different carcinomas.The biological effect and related molecular mechanisms in salivary cancer cells and the relationship of MUC1 and mucoepidermoid carcinoma clinical features are still unclear.In this study,we aimed at investigating the molecular mechanism underlying the biological effects and relationship between MUC1 and mucoepidermoid carcinoma clinical features.This work will uncover the roles of MUC1 in salivary gland cancers and the important therapeutic significance in MUC1-positive salivary cancers.PURPOSES:1.To determine the expression of MUC1 in salivary AdCC and MEC cancer cells and investigate the effect of MUC1 knockdown or overexpression on the ability of cell proliferation,migration and invasion.2.To evaluate the effect of MUC1 expression on the growth of salivary AdCC and MEC cancer cells xenograft tumors in different groups.3.To investigate the effect of MUC1 in regulating the activity of EGFR signaling pathway in salivary AdCC and MEC cancer cells.4.To investigate the effect of MUC1 on the expression of HIF-1?and the effect of Co Cl2 on the ability of cell proliferation and migration.To evaluate the effect of treatment of Digoxin on ability of cell proliferation and migration in MUC1 overexpression salivary cancer cells.To comprehensive understand the underlying mechanisms of MUC1 in salivary AdCC and MEC cancer cells based on the RNA-seq techniques.5.To determine the expression of MUC1 in normal salivary gland and mucoepidermoid carcinoma.To assess the correlation between MUC1expression and clinical parameters in mucoepidermoid carcinoma.METHODS:1.qRT-PCR and WB to detect the expression of MUC1 in salivary AdCC and MEC cancer cells;MUC1 knockdown and overexpression were conducted to investigate the effect on the ability of proliferation,migration and invasion in salivary cancer cells;2.Salivary xenograft models were established to investigate the effect of MUC1 expression on the growth of tumors in different groups.3.WB was carried out to detect the effect of MUC1 expression on the activity of EGFR signaling pathway;q RT-PCR,WB and IF were carried out to detect the effect of MUC1 on the expression levels of HIF-1?;the expression of HIF-1?was detected by WB after treatment of Co Cl₂ and colony formation assay and transwell assay were carried out to detect the ability of proliferation and migration in A-253 cells;WB was carried out to detect the expression of HIF-1?after treatment of Digoxin and colony formation and transwell assay were carried out to evaluate the ability of proliferation and migration after treatment of Digoxin in A-253-OE cells.4.RNA-seq techniques were carried out to discover the significantly different expressed genes and GO analysis and KEGG analysis were employed to summarized the different expressed genes.q RT-PCR was carried out to verify the degree of accuracy of RNA-seq.5.Collecting cases of MEC and using WB and IHC to determine the relationship of MUC1 expression and clinical parameters.RESULTS:1.MUC1 is expressed in salivary AdCC and MEC cancer cells;knockdown of MUC1 expression suppress cell proliferation,migration and invasion and elevated MUC1 expression promote cell proliferation,migration and invasion.2.MUC1 knockdown suppress tumor growth and MUC1 overexpression promote tumor growth in xenograft models of salivary AdCC and MEC cancer cells.3.MUC1 knockdown suppress the activity of EGFR signaling pathway and MUC1 overexpression promote the activity of EGFR signaling pathway.4.MUC1 could stability the expression of HIF-1?;the treamment of Co Cl₂could improve the expression of HIF-1?and promote the ability of proliferation and migration in A-253 cells;the treatment of Digoxin could decrease the expression of HIF-1?and inhibited the ability of proliferation and migration in A-253-OE cells.The results of RNA-seq revealed that there were 349 and 1355 significance different expressed genes in MUC1knock-down cells and MUC1 overexpression cells,respectively.The different expressed genes involved in many signaling pathway,such as MAPK signaling pathway.The q RT-PCR analysis of genes were consistent with the results of RNA-seq.5.The expression of MUC1 was pronounced elevated in MEC compared with normal salivary gland and elevated expression of MUC1 correlated with higher lymphatic metastasis and advanced tumor in MEC.CONCLUSIONS:1.MUC1 acts as proto-oncogene role in salivary AdCC and MEC cancer cells and play important roles in EGFR signaling pathway;MUC1 can stability the expression of HIF-1?and Digoxin could effective weaken the effect of MUC1 overexpression.2.High-expressed MUC1 correlated with higher lymphatic metastasis and advanced tumor in MEC.