The Role Of NOX4 And SPOCK1 Involving In The Development And Progression Of Colorectal Cancer

Aims:NADPH oxidase 4(NOX4),as a major source of reactive oxygen species(ROS)production,has been increasingly reported to be involved in tumorigenesis and/or tumor progression,but limited data is available regarding the role of NOX4 in colorectal carcinoma(CRC).We firstly retrieved six independent investigations from Oncomine database and found that NOX4 is highly expressed in CRC tissues compared to corresponding normal controls.The relationship of NOX4 expression with clinicopathologic parameters including prognostic significance,and the biologic function of NOX4 in CRC patients,then caught our attention.Methods: 1)Using clinical samples,we profiled the expression level of NOX4 at both the m RNA and protein level,determining by RT-PCR and Western blotting,respectively;2)A tissue microarrays containing 82 paired CRC specimens was used to evaluate the correlation between NOX4 expression and clinicopathologic characteristics including prognostic markers.We performed NOX4 expression profiling in large CRC cohorts from Gene Expression Omnibus(GEO)and The Cancer Genome Atlas(TCGA)dataset;3)To gain further explore the biological pathways involved in CRC tumorigenesis and/or tumor progression through NOX4 pathway,Gene Set Enrichment Analysis(GSEA)was performed;4)Applied si RNA to knock down NOX4 of the HCT116 and LOVO cells,we studies the possible functions of NOX4 involved in CRC;5)Western blotting analysis was performed to validate the expression level of the gene involved in GSEA analysis.Results: 1)Based on Oncomine database,similar results were also found in clinical specimens;2)We found that NOX4 overexpression was significantly correlated with T classification,N classification,distant metastasis.Interestingly,we also observed that T stage significantly associates with NOX4 expression in TCGA database.Kaplan-Meier analysis indicate that overexpression of NOX4 is associated with poor prognosis of CRC patients,which was also confirmed by GSE14333 and GSE17536 datasets from the Gene Expression Omnibus;3)GSEA analysis revealed that cell proliferation,metastasis,apoptosis,EMT and angiogenesis–related genes were enriched in NOX4 higher expression patients;4)We demonstrated that when NOX4 expression was knocked down by si RNAs,cell proliferation,cell-cycle and apoptosis,migration and invasion in CRC cell lines HCT116 and LOVO were obviously altered;5)NOX4 promoted cancer cell proliferation and apoptosis,migration and invasion by regulating the expression of relevant genes in the light of GSEA analysis.Conclusions: By these approaches we aim to elucidate NOX4 may be a reliable prognostic factor or therapeutic target in CRC.