Preliminary Study On The Role And Mechanism Of USP25 Inhibiting TSLP Expression In Nasal Mucosal Epithelial Cells In Allergic Rhinitis

Objective: Allergic rhinitis(AR)is a common allergic disease.Ubiquitination is the main way for cells to mediate selective protein degradation.Ubiquitin-specific protease(USP)is the largest family of human deubiquitinating enzymes(DUBs).It has been found in the past that USP25 is involved in immune-related pathological and physiological processes such as virus immunity and tumorigenesis.However,the role and mechanism of USP25 in allergic diseases such as AR are still unclear.Recent literature shows that USP25 can maintain the high expression of TRAF3 through deubiquitination,and TRAF3,as a negative regulator of the NF-?B pathway,plays an important role in immune regulation.This study aims to explore the role of USP25 in AR immune imbalance and its regulatory mechanism in nasal mucosal epithelial cells.Methods: Nasal mucosal tissues from AR patients and healthy controls were collected,and the expression of USP25 in nasal mucosal tissues was detected by immunohistochemistry and Western blotting,and the expression of Thymic Stromal Lymphopoietin(TSLP)in nasal mucosal tissues was also detected by Western blotting,respectively.The AR mouse models of wild-type mice and USP25 knockout mice sensitized by ovalbumin(OVA)were constructed.The expression levels of IL-4,IL-5,IL-10,IL-13 and IFN-? in nasal irrigation solution and serum of mice were measured by ELISA,and the histological changes of nasal mucosa were observed by HE staining.Western blotting and immunohistochemistry were applied to detect the expression of USP25 and TSLP in mice nasal mucosa tissues.HNEp C cells were stimulated with OVA to mimic the allergic inflammatory environment,and the expression of USP25 or TRAF3 was inhibited by si RNA,and TRAF3 or USP25 was overexpressed by pc DNA,respectively,and finally the expression of USP25,TRAF3 and TSLP in the cells was detected by WB.Results: The expression of USP25 in the nasal mucosa of AR patients was significantly lower than that of the control group,while the expression of TSLP was significantly higher than that of the control group.Compared with wild-type mice stimulated by PBS,the expression levels of IL-4,IL-5,IL-10,and IL-13 in the nasal cavity lavage fluid and serum of AR model mice sensitized by OVA were significantly increased,while the expression level of IFN-? decreased significantly.The number of eosinophils was significantly increased in the nasal mucosal tissue of AR mice,and the expression of UPS25 was decreased,while the expression of TSLP was increased.Compared with wild-type AR mice,the expression levels of IL-4,IL-5,IL-10,and IL-13 in the nasal cavity lavage fluid and serum of USP25 knockout mice sensitized by OVA were significantly increased,while the expression of IFN-? was significantly lower;UPS25 expression was absent in the nasal mucosa,and TSLP expression further increased.Relative to the blank control group,OVA stimulation resulted in a decrease in USP25 and TRAF3 expression and an increase in TSLP expression in HNEp C cells.Inhibition of USP25 expression with si RNA also resulted in a simultaneous decrease in TRAF3 expression,while while TSLP expression was further increased.On this basis,the addition of pc DNA caused TRAF3 overexpression,and relative to the NECs+OVA+USP25 si RNA+ TRAF3 pc NC group,the expression of USP25 did not change significantly,but the expression of TSLP was significantly reduced.In contrast,after OVA stimulation,the addition of pc DNA to overexpress USP25 was followed by a simultaneous increase in TRAF3 expression and a decrease in TSLP expression.On this basis,the addition of si RNA inhibited the expression of TRAF3.Compared with the NECs+OVA+ USP25 pc DNA+ TRAF3 si RNA group,the expression of USP25 was not significantly changed,but TSLP expression increased significantly.Conclusion: Our results demonstrated that USP25 was significantly low in AR and TSLP was significantly high expression.The loss of USP25 further increased the expression of TSLP in the nasal mucosa epithelium,and aggravated the allergic inflammatory response in AR mice.In human nasal mucosal epithelial cells cultured in vitro,USP25 inhibited TSLP by maintaining the expression of TRAF3,thereby playing a key regulatory role in allergic inflammation.