Overexpression Of TLR-9 In Oral Squamous Cell Carcinoma And Its Effect On Tumor Infiltrating PDC

[Objective] Chronic inflammation is closely related to the occurrence and development of malignant tumors.This study aims to investigate the Overexpression of TLR-9 in oral squamous cell carcinoma and its effect on the function of tumor infiltrating pDC[Methods] We first used Q-PCR,western blotting and immunohistochemistry to detect the expression and trend of TLR-9 mRNA and protein in normal oral epithelial tissues,oral squamous cell carcinoma and oral epithelial cancer cell lines.Subsequently,the bioinformatics analysis and promoter luciferase reporter gene assay was used to map the TLR-9 gene,and the candidate promoter region of the TLR-9 gene was preliminarily identified.The ABI3730 sequencing and subsequently promoter luciferase reporter gene assay were continued to Screen significant mutation sites of TLR-9 gene.Finally,immunomagnetic beads sorting combined with flow cytometry and real-time PCR techniques were used to study the overexpression of TLR-9 in oral squamous cell carcinoma and its effect on tumor infiltrating pDC.[Results] The expression of TLR-9 receptor is up-regulated during oral carcinogenesis,and the sequence from-1432 to-725 may be the core region of the TLR-9 gene promoter;subsequent ABI3730 sequencing revealed high frequency mutations of DNA occurred in the range of 250-310 bp upstream of the TLR-9 promoter core region.The TLR-9 promoter-luciferase reporter gene expression vector experiments indicated that MUT4 may play an important role in TLR-9 overexpression;The number of pDC cells in normal oral epithelial tissues is very small and however,the number in cancerous tissues is obviously increased,but the function is significantly reduced.The overexpression of TLR-9 in cancerous cells can mediate the inhibition of TLR-9 receptor expression in pDC cells;the culture supernatant of oral cancer cells(OCDS)can significantly reduce the expression of TLR-9 in pDC cells and the secretion of pDC IFNa and IL-6 before and after CpG-ODN stimulation;OCDS-mediated pair of the inhibition of IFN-? production was largely reversed when IL-10 and TGF-? are simultaneously blocked,suggesting that the effect of the oral cancer microenvironment on the specific inhibition of pDC immune function is mainly mediated through IL-10 and TGF-?.[Conclusion] The expression of TLR-9 is up-regulated during oral carcinogenesis.The abnormal transcriptional regulation of TLR-9 gene caused by mutation of DNA in promoter region may be an important reason of TLR-9 overexpression.Over-expressed TLR-9 can inhibit pDC intracellular TLR-9 expression and inhibits its secretion of IFN-a;and the secretion of IL-10 and TGF-? in cancer cell supernatant inhibits the immunoregulatory and secretory function of oral squamous cell carcinoma infiltrating pDC,which can be used for early monitoring and prevention of oral squamous cell carcinoma.