Affiliations: [a] Laboratory of Oral Biomedical Science and Translational Medicine, School of Stomatology, Tongji University, Shanghai 200072, China | [b] State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041, Sichuan, China | [c] The State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, Sichuan, China
Correspondence:
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Corresponding author: Xiaohui Hou, Lecturer for Oral Biomedical Science and Translational Medicine, School of Stomatology, Tongji University. 399 Yanchang Road, Shanghai 200072, China. Tel.: +86 21 66315500; E-mail: xiaohui_hou@yahoo.com.
Abstract: Oral submucous fibrosis (OSF) is a chronic insidious disease which predisposes to oral cancer. Understanding the molecular markers for OSF is critical for diagnosis and treatment of oral cancer. In this study, the proteins expression profile of OSF tissues was compared to normal mucous tissues by 2 dimensional electrophoresis (2-DE). The 2-DE images were analyzed through cut, spot detection and match analysis using mass spectrometry (MS). Differentially expressed genes were identified as candidates. RT-PCR, Western Blot and immunohistochemistry were performed to validate the difference in expression of the candidates between OSF and normal mucous tissues. The shRNA targeted to the candidates were then transfected by Lipofectamine2000 to the 3T3 cells to study gene function. Cell proliferation and apoptosis were measured by MTT, clonogenic formation, PI and TUNEL staining. From the proteomic analysis, 94 of the 182 selected spots with differential expression were identified by MS analysis and Cyclophilin A (CYPA) was determined to be the OSF-associated protein candidate. The significant differences in expression between OSF and normal tissues were verified and confirmed by RT-PCR, Western blot and Immunohistochemical analysis. Inhibition of CYPA expression by RNA interference suggested its potential activities involved in cell proliferation and apoptosis process. In conclusion, these results indicated a novel molecular mechanism of OSF pathogenesis and demonstrated CYPA as a potential biomarker and gene intervention targets of OSF. These data may help the development for therapeutics of oral cancer.
Keywords: Oral submucous fibrosis, proteomics, 2-D electrophoresis, mass spectrum, Cyclophilin A, small hairpin RNA
