ABSTRACT
It has been demonstrated that circ_0001874 and circ_0001971 are potential biomarkers for the diagnosis of oral squamous carcinoma (OSCC). MiR-186 was reported to serve as a tumor suppressor in OSCC, and the down-regulation of miR-186 was reported to lead to higher expression of oncogenic factor SHP2 and the activation of growth promoting signaling. In this study, we aimed to explore the possible molecular role of circ_0001874 and circ_0001971 signaling in the pathogenesis of OSCC. RT-qPCR, Western blot, online bioinformatics tools and luciferase assay were utilized to study the molecular signaling pathways of circ_0001874 and circ_0001971. MTT assay and FCM assay were performed to investigate the synergistic effect of circ_0001971 and circ_0001874 on cell proliferation and apoptosis. By observing the effect of different miRNAs on the levels of circ_0001847 and circ_0001971, it was identified that circ_0001847 and circ_0001971 respectively sponged the expression of miR-296 and miR-186 via binding to these miRNAs. Also, SHP2 mRNA and PLK1 mRNA were respectively targeted by miR-186 and miR-296-5p. We also established two signaling pathways, i.e., circ_0001971/miR-186/SHP2 and circ_0001874/miR-296-5p/PLK1, and validated the synergistic effect of circ_0001971 and circ_0001874 via observing their positive effect on cell proliferation and negative effect on cell apoptosis. The expression of miR-186 and miR-296-5p was generally lower in saliva of OSCC patients compared with that in OLK patients, while the expression of miR-186 and miR-296-5p was specifically up-regulated in saliva of OSCC patients. In conclusion, the finding of this study demonstrated that the relative level of hsa_circ_0001971 and hsa_circ_0001874 were different in the saliva of OSCC patients and could be used as predictive biomarkers for the development of OSCC. Furthermore, oncogenic effects of hsa_circ_0001971 and hsa_circ_0001874 in the development of OSCC might be, at least partially, mediated by its downstream signaling pathways including hsa_circ_0001971/microRNA-186/SHP2 and hsa_circ_0001874/microRNA-297/PLK1.
Subject(s)
MicroRNAs/genetics , RNA, Circular/genetics , Squamous Cell Carcinoma of Head and Neck/genetics , Apoptosis/genetics , Carcinoma, Squamous Cell/pathology , Cell Cycle Proteins/metabolism , Cell Line, Tumor , Cell Proliferation/genetics , Gene Expression/genetics , Gene Expression Regulation, Neoplastic/genetics , Humans , Mouth Neoplasms/pathology , Protein Serine-Threonine Kinases/metabolism , Protein Tyrosine Phosphatase, Non-Receptor Type 11/metabolism , Proto-Oncogene Proteins/metabolism , Signal Transduction/genetics , Squamous Cell Carcinoma of Head and Neck/metabolism , Squamous Cell Carcinoma of Head and Neck/pathology , Polo-Like Kinase 1ABSTRACT
Objective:To explore the cooperation and clinical significance of HIF-1α,ALDH1 and Hedgehog signaling pathway in the activation of cancer stem cell(CSC) in triple negative breast cancer(TNBC).Methods: ALDH1+(Aldehyde dehydrogenase1)breast cancer stem cells and ALDH1-breast cancer cells were selected from MDA-MB-231 cells by magnetic activated cell sorting system(MACS),qRT-PCR method was employed to analyze the expression differences of HIF-1α and Hedgehog signaling molecules Sonic hedgehog(SHH),patched1(PTCH1),Smoothened(SMO) and Glioma-associated oncogene homoglog1(GLI1) in ALDH1+ breast cancer stem cells and ALDH1-breast cancer cells.Immunohistochemical method was applied to study the expressions of HIF-1α and ALDH1 and the relationships among HIF-1α,ALDH1 and Hedgehog signaling molecules in TNBC.Results: The expressions of HIF-1α mRNA,SMO mRNA and GLI1 mRNA in ALDH1+ breast cancer stem cell were higher than those in ALDH1-breast cancer cell(P all<0.05).The positive expression rates of HIF-1α were 90.0% and 70.0%,and the positive rates of ALDH1 were 93.3 % and 66.7 % in TNBC and non-TNBC,respectively(P all<0.05).Spearman rank correlation analysis showed that the expression of HIF-1α was positively related with that of ALDH1 in TNBC(r=0.53,P<0.01).HIF-1α expression was correlated with lymph node metastasis and TNM stage(P all<0.05),ALDH1 expression was correlated with histological grade and TNM stage(P all<0.05).In addition,the expression of HIF-1α was positively related with that of Hedgehog signaling molecules SHH(r=0.584,P<0.01),SMO(r=0.467,P<0.01) and GLI1(r=0.439,P<0.05),the expression of ALDH1 was positively related with that of SHH(r=0.426,P<0.05) and GLI1(r=0.394,P<0.05).Conclusion: HIF-1α and Hedgehog signaling pathway were activated in ALDH1+ breast cancer stem cell.HIF-1α,ALDH1 and Hedgehog molecules may cooperate with each other to activate breast CSC to promote the malignant progression of TNBC.