Objective This study aimed to explore the expression trends of innate immune cells and immune-checkpoint molecules validated by data calculation in the process of oral mucosal carcinogenesis,as well as to explore methods of suppressing oral mucosal carcinogenesis based on immunotherapy by predicting their interactions.Me-thods 1)The cancer genome atlas(TCGA)database comprehensively scores immune cells and immune-checkpoint molecules in the process of oral mucosal carcinogenesis and screens out intrinsic immune cells and immune-check-point molecules that interfere with tumor immune escape.2)Clinical patient blood routine data were collected for the statistical analysis of peripheral blood immune cells during the progression of oral mucosal carcinogenesis.Immune cells in peripheral blood that may affect the progression of oral mucosal carcinogenesis were screened.3)Immunohis-tochemical staining was performed on intrinsic immune cells and immune-checkpoint molecules validated based on da-ta calculation in various stages of oral mucosal carcinogenesis.4)Special staining was used to identify innate immune cells in various stages of oral mucosal carcinogenesis based on data-calculation verification.5)Survival analysis was conducted on intrinsic immune cells and immune-checkpoint molecules validated based on data calculation during the process of oral mucosal carcinogenesis.The association of intrinsic immune cells and immune-checkpoint molecules with the prognosis of oral squamous cell carcinoma was verified.Results The expression of monocytes and neutro-phils increased during the process of oral mucosal carcinogenesis.The expression of eosinophils showed a single peak trend of up and down.The expression of mast cells decreased.In the process of oral mucosal carcinogenesis,the ex-pression of the immune-checkpoint molecules cytotoxic T-lymphocyte-associated protein 4(CTLA4)and programmed cell death-ligand(PD-L1)increased.The expression trends of monocytes,neutrophils,and eosinophils were positively correlated with those of CTLA4 and PD-L1 immune-checkpoint molecules.The expression trend of mast cells was negatively correlated with the expression of CTLA4 and PD-L1.Monocytes,neutrophils,and eosinophils may pro-mote tumor immune escape mediated by CTLA4 and/or PD-L1,thereby accelerating the progression of oral mucosal carcinogenesis.Mast cells may inhibit tumor immune escape mediated by CTLA4 and/or PD-L1,delaying the progres-sion of oral mucosal carcinogenesis.Conclusion Therefore,interference with specific immune cells in innate immu-nity can regulate the expression of CTLA4 and/or PD-L1 to a certain extent,inhibit tumor immune escape,and delay the progression of oral mucosal carcinogenesis.

Objective: To study the inhibitory effect of ozone water on bad breath pathogens in vitro. Methods: In vitro cultured bad breath pathogens Porphyromonas gingivalis (Pg), Prevotella intermedia (Pi) and Fusobacterium nucleatum (Fn) were identified by Gram stain and a PCR test. Ozonated water by was prepared by ozone generator and the concentration of ozone in water was measured using iodine titration. Artificial saliva was used to observe its interference on ozone function. Results: Gram stain and PCR results were consisted with strain characteristics. The ozone concentration of ozonated water reached to the maximum of 0. 2 mg/L. Ozone water with the concentration was 0. 05, 0. 1 and 0. 2 mg/L inhibited the proliferation of Pg, Pi and Fn. But the inhibitory effect was weakened when the concentration decreased. The artificial saliva reduced the effect of ozone water. Conclusion: The Pg, Pi and Fn can be inhibited by ozone water. Artificial saliva may reduce the effects of ozone water on the bacteria.

Objective To investigate the therapeutic effect of modified powder and vitamin B12 on the treatment of recurrent oral ulcer in rats and its effect on serum interleukin-6 (IL-6) and tumor necrosis factor alpha(TNF-α).Methods Forty-eight male Sprague-Dawley (SD) rats were randomly divided into normal group (equal distilled water),model group (equal distilled water),positive control group (levamisole 20 mg/kg gavage) and experimental group (containing dcsgd crude drugs 1 g/mL,1.278 mL/200 g+ B12 20 g/kg gavage),each group with 12 rats.Model group,positive control group and experimental group were treated with immunological method to establish the recurrent oral ulcer mode.Each group treated with the corresponding treatment methods for 20 days.Results The oral ulcer number,duration time in positive control group and the experimental group were significantly lower than that in model group rats(P＜0.05),ulcer interval time was significantly longer than the rats in the model group (P＜0.05);the serum IL-6,TNF-α detection level in positive control group and experimental group were significantly lower than in the model group(P＜0.05);serum IL-6,TNF-α detection level in model group were significantly higher than those in normal group(P＜0.05);the peripheral blood SOD,GSHPx levels in positive control group and experimental group rats were significantly higher than that in model group(P＜0.05),the level of MDA was significantly lower than that in model group (P＜0.05).Conclusion Modified powder and vitamin B12 have therapeutic effects on recurrent oral ulcer in rats.The mechanism may be related to reducing the inflammatory reaction and improving the antioxidant capacity of the tissue.

Objective To evaluate the expression of epithelial cadherin (E-cad) and proliferating cell nuclear antigen (PC-NA) in various stages of tongue carcinogenesis and explore its relevance.Methods SP immunohistochemical method was employed to detect the expression of E-cad and PCNA protein in 82 rat tongue carcinogenesis specimens which induced by 4-nitroquinoline 1-oxide(4NQO).Chi square test for trend and the spearman correlation were used to analysis the correlation between E-cad and PC-NA.Results In normal mucosa,epithelial hyperplasia,mild dysplasia,severe dysplasia and squamous cell carcinoma,the positive rate of E-cad were 100％,95.24％,92.86％,80％,68.75％,the differences were statistically significant(P＜0.05);The positive rate of PCNA were 9.52%,14.29%,35.71%,50%,56.25%,the difference were statistically significant(x2 =16.676,P＜0.05).The expression of E-cad and PCNA has negative correlation(r=-0.614,P＜0.01).Conclusion E-cad and PCNA may be one of the biomarkers of carcinogenesis of tongue mucosa.

Objective To investigate whether the effects of rapamycin on promoting oral squamous cancer cell apoptosis is related with inhibiting TLR4 expression and inflammatory factor IL-6 and PGE2 expression.Methods Human oral squamous carcinoma SCC-15 cell line was cultured and interfered by rapamycin.Then the activity of SCC-15 cells was detected by CCK 8,the SCC-15 cells apoptosis was detected by the Hochest33342/PI double staining,the invasion ability was determined by the transwell method.The TLR4 protein expression level was detected by Western blot and IL-6 and PGE2 expressions were detected by ELISA.Results Rapamycin could promote cell apoptosis,the invasion ability of SCC-15 cells was significantly decreased.After rapamycin intervention,the TLR4 protein expression was decreased and expression levels of IL-6 and PGE2 were reduced,showing statistical difference as compared with the negative group (P＜0.05).Conclusion Rapamycin promotes oral squamous cancer SCC-15 cell apoptosis,which may be related with inhibiting TLR4,IL-6 and PGE2 expression.

Objective To study the expression of vimentin in the tongue mucosa carcinogenesis and to explore its significance in the invasion and metastasis of tongue squamous cell carcinoma .Methods The occurrence of tongue squamous cell carcinoma in rat was induced by means of 4NQO water solution ,and 56 cases in total were collected in the cancerous process ,including normal tongue mucosa ,epithelial hyperplasia ,mild dysplasia ,moderate dysplasia ,severe dysplasia and the tongue tissue specimen of squa‐mous cell carcinoma .The immunohistochemistry was used to evaluate protein expression and real‐time fluorescent quantitative PCR method was used to obtain the expression quantity of mRNA .Results In immunohistochemistry ,with the increase of degree of rat tongue mucosa dysplasia ,the positive rate of vimentin expression increases obviously .The difference between groups was statistical‐ly significant (χ2 =10 .685 ,P<0 .05) .Lesion groups compared with normal group ,their mRNA expression differences all hold sta‐tistical significance(P<0 .05);Mild dysplasia ,moderate dysplasia ,severe dysplasia and squamous cell carcinoma groups were com‐pared with epithelial hyperplasia group .The difference between squamous cell carcinoma group and epithelial hyperplasia group was statistically significant (P<0 .05) .mRNA expression of epithelial hyperplasia ,mild dysplasia ,moderate dysplasia and severe dys‐plasia were respectively 1 .22 times ,1 .28 times ,1 .29 times and 1 .42 times of that of the normal group .Conclusion During rat tongue carcinogenesis induced by 4NQO ,the expression of vimentin was increased with the increase of the degree of pathological change ,which is closely related to the invasion of tumor and could be regarded as a predictor of tongue squamous cell carcinoma .

Objective:To study the effects of a potassium channel blocker 4-Amino pyridine(4-AP)on the proliferation of human tongue squamous cell carcinoma Tca8113 cells.Methods:CCK-8 assay was used to detect the proliferation of Tca8113 cells cultured with 4-AP at the concentration of 1,5,10,20,50 and 100 mmol/L for 12,24 and 48 h respectively,the cell proliferation inhibition rate was calculated,the cell cycle distribution was examined by flow cytometry.Data were analyzed by SPSS19.0 software.Results:With the increase of 4-AP concentration and culture time,cells showed some morphologic changes.4-AP at 5 -100 mmol/L dose and time dependently inhibited the proliferation,with 24 h exposure dose dependenty decreased S-phase population and increased G0 /G1 phase population of Tca8113 cells.Conclusion:4-AP may inhibit Tca8113 cell proliferation by regulation of the cell cycle distribution.

Patients with burning mouth syndrom(BMS,n =12)and patients with wisdom tooth removal(controls,n =9)were included. Immunohistochemistry and image analysis were used to quantify P2X3 receptor expression in lingual nerve fibres.A pain history and score were recorded on a visual analogue scale(VAS)prior to obtaining a lingual biopsy.The value of P2X3 receptor positive fibres in BMS and control subjects was 0.56 ±0.29 and 0.15 ±0.06 respectively(P <0.001).There was no significant correlation between P2X3 and VAS scores(R2 =0.012).Increased P2X3 may play a role in BMS but not correlated with the VAS score.

Objective:To investigate the expression of HERG1 and Kv3.4 in normal oral mucosa(NOM)and oral lichen planus (OLP).Methods:20 OLP and 1 6 NOM specimens were collected and immunohistochemically stained(IHC)by SP method for the detection of HERG1 and Kv3.4 protein expression.Results:HERG1 and Kv3.4 were negative or weak-positive in the sapmles of bas-al layer of NOMand non-erosive OLP,but positive in basal layer,spinous layer and granular layer of erosive OLP.The expression of HERG1 and Kv3.4 was higher in OLP than in NOM tissues (P <0.05);and higher in erosive OLP than in non-erosive OLP(P <0.05).In OLP HERG1 expression was positively related to Kv3.4(P <0.05).Conclusion:HERG1 and Kv3.4 may be related to the development of OLP.

Objective To investigate the expression of the Kv3 .4 protein and Kv3 .4 mRNA in various stages of oral carcino‐genesis .Methods The expression of Kv3 .4 protein and Kv3 .4 mRNA were detected by immunohistochemistry (SP method) and RT‐PCR technique respectively in the oral carcinogenesis of SD rat which were induced by 4NQO .Results With the aggravation of the epithelial dysplasia ,Kv3 .4 protein and Kv3 .4 mRNA expression increased gradually .They were strongly positive in oral squa‐mous cell carcinoma .Conclusion The expression of Kv3 .4 protein and Kv3 .4 mRNA levels increased consistently with the aggra‐vation of the epithelial dysplasia .