Expression Of Tight Junction Proteins In Mucosal Margin Of Oral Squamous Cell Carcinoma Determined By Iodine Staining And Their Effects On Proliferation And Migration Of Oral Squamous Carcinoma Cells

Objective: The aim of this study was to compare and analyze the expression of tight junction proteins and infiltration of iodine in mucosal margin of oral squamous cell carcinoma(OSCC),then to determine the targeted tight junction proteins related to epithelial permeability and investigate their effects on proliferation and migration of oral squamous cell carcinoma cells in vivo.Methods:Immunohistochemical stain method was carried out to detect the expression of claudin-1,2,3,4,5,6,7,8,10,11,14,17,23,ZO-1,ZO-2,occludin and then compare iodine and glycogen distribution measured by iodine stain and Periodic Acid-Schiff stain(PAS)respectively in mucosal margin of 20 oral squamous cell carcinoma samples and preliminarily confirmed the possibly involved cell tight junction proteins.Western blot was performed for the expressions of proteins mentioned above in HSC-3,HSC-4,SAS,Ca9-22,Tca-8133,CAL-27 cells.When the key protein and the optimal cell line were finally determined,the lentivirus vector harbouring target gene si RNA was transfected into cultured cells.The down-regulated expression level was examined by real time-polymerase chain reaction(RT-PCR),and the cell proliferation and migration were assessed by the MTT assay and wound healing assay.Results:1.Claudin-1,4,6,7,8,10,occludin and ZO-2 instead of claudin-2,3,5,11,17,23,ZO-1 were expressed at different degrees.And claudin-1 was expressed strongly in all cases.Claduin-1,4,7,8,10 were mainly distributed in basal and parabasal layer of normal epithelium and below the surface of abnormal epithelium.Iodine distribution in the epithelium of oral mucosal margin of OSCC was not completely consistent with that of glycogen but generally complementary to the distribution of claudin-1,4,7,8,10,especially claudin-1.2.Western blot showed that Claudin-1 was obviously expressed in the HSC-3,HSC-4,SAS and CAL-27 cell lines and mildly expressed in Ca9-22,Tca-8113 cell lines.The cell infection efficiency of SAS was higher than that of HSC-4 cells.3.MTT assay and wound healing assay showed that the proliferation and migration rate of SAS cell transfected by claudin-1 siRNA were obviously decreased compared to control group(P<0.05).Conclusions:1.Claudin-1 is a reliable molecular marker for distinguishing mucosal margin of OSCC.2.Claudin-1 influences the infiltration of vital stain in the oral mucosa and distinguish effect of mucosal margin of OSCC measured by vital stain further.3.Down-regulation of claudin-1 prohibits the proliferation and migration of OSCC cells,which indicated that claudin-1 has a close relationship with biological behaviour of neoplasm and maybe a potential target for the treatment of oral squamous cell carcinoma.