Function Of PDC Affected By Overexpression Of TRIM21 In Oral Lichen Planus

Objectives: Oral lichen planus(OLP)is a chronic inflammatory disorder of oral mucosa of unknown etiology.Plasmacytoid dendritic cells with TLR9 expressed dominantly on endosomal membrane were significantly higher in OLP lesions and the downstream IFN-?signaling pathway was activated.The present research is to screen the interacting proteins of TLR9 and to be verified on the basis of quantity and distribution as well as functional changes of IFN-? from pDC cultured in vitro and T cell related cytokine profiles form pDCT co-culture.Methods: Research mediums including whole unstimulated saliva,buccal mucosa samples and PBMCs were collected from both OLP and health control groups for the following studies:(1)IFN-? levels of tissue homogenates were assessed by ELISA as indicator of pDCs activation;T cell related cytokine profiles in whole unstimulated saliva were detected by CBA as reference standards of T cells activation;(2)Co IP followed by mass spectrometry to screen interacting proteins of TLR9 in OLP lesions;(3)Reverse Co IP,immunofluorescence co-localization analysis and pDC simulating experiments were used to verify the protein interaction;q PCR used to quantify its gene expression;immunohistochemistry used to delineate its spatial distribution;(4)Candidate for TLR9 interacting protein,elute of TLR9 Co IP samples,homogenate form mucous epithelium/whole mucosa of the lesions of OLP patients and agonist,antagonist of TLR9 were added into the system of pDC-T co-culture to detect proliferation of T cells by CFSE proliferation assay and CCK-8 method,to analyze differentiation of T cells by change of T cell related cytokine profiles.Results:(1)Compared with health control group,elevated IFN-? levels in oral lesions and increased levels of IL-6,IL-10 and IFN-?,IFN-?/ IL-4 in whole unstimulated saliva from the patients with OLP were found;salivary IL-10 and especially IL-6 correlated positively with severity of OLP;(2)Including TRIM21,there were 7 potential TLR9 interacting proteins predominantly presented in OLP samples;(3)The possible interaction between TRIM21 and TLR9 was verified by reverse Co IP and immunofluorescence co-localization analysis;significantly increase of IFN-? secretion from pDC stimulated by TRIM21 was not found;TRIM21 was revealed significantly higher transcription through q PCR;it was indicated that a larger amount of TRIM21 positive cells mainly infiltrated in the lamina propria of OLP lesions than those of health control group by immunohistochemistry;TRIM21 expression increased in T cells among PBMCs,lymph nodes and OLP lesions respectively;(4)Quantity and secretion of IFN-? from pDC induced by ODN did not differ statistically between groups;no difference existed in proliferation and migration of T cells;stimulated by TRIM21 and various extracts of mucosa origin,no significant cytokine profiles differences ensued.Conclusions:(1)There's evidence of the IFN-? signaling pathway activation;cytokine profiles changes indicated activation of T cells with interpopulationally mixed and intrapopulationally inhomogeneous trends;(2)As to pDC,TRIM21 is possible interacting protein of TLR9,which maybe acts as an immunogen or its component;(3)As to T cell,TRIM21 may be a secondary and inducible but important regulatory factor;(4)Compared to health controls,peripheral pDCs and T cells of patients with OLP may be relatively normal.