Molecular Mechanism Of OPN Inhibiting NK Cell Function In Head And Neck Squamous Cell Carcinoma

Objective: Recurrence and metastasis are the leading causes of death in patients with head and neck squamous cell carcinoma(HNSCC).Natural killer(NK)cells are innate immune lymphocytes that respond to viral infection and tumor cells,and play a key role in tumor therapy.However,the specific regulatory mechanism of the inhibition of infiltrating NK cell function in head and neck tumors is unclear.The purpose of this study was to systematically analyze the effects of immune-related genes screened on the TCGA database on NK cells in head and neck squamous cell carcinoma,construct a combined prognostic diagnostic model,and reveal the underlying molecular mechanism.Methods: First,this study analyzed key prognostic markers in head and neck tumors by bioinformatics.Then,10 pairs of tumor tissue samples and peripheral blood from HNSCC patients,10 peripheral blood from healthy people,and 10 tonsil samples from snoring patients were collected.The expression of osteopontin(OPN)and the level of NK cell infiltration in each sample were analyzed by flow cytometry and multiplex immunohistochemistry(m IHC).Antibody array experiments were used to identify and quantify downstream key cytokine expression levels.Further in vitro experiments,including tumor-lymphocyte co-culture,immunoblotting,co-immunoprecipitation,ELISA,LDH release assay,etc.,further studied the potential molecular mechanism of TGF-β1 inhibiting NK cell function.Finally,the above findings were verified by establishing a xenograft mouse model and performing m IHC experiments on the xenograft specimens.Results: The expression of OPN was significantly increased in the HNSCC microenvironment,and the high expression of OPN was associated with low infiltration of NK cells and poor prognosis of patients.The results of clinical sample validation showed that compared with healthy controls,infiltrating NK cells in tumor patient tissues were in a suppressed state and the number was lower.In vitro experiments showed that OPN promoted the growth,migration and invasion of cancer cells.In addition,OPN promotes the secretion of TGF-β1 by tumor cells to inhibit the activation and killing activities of NK cells.Importantly,OPN activates the downstream NF-k B pathway to promote the secretion of TGF-β1 from HNSCC cells by binding to the tumor cell surface receptor integrin αvβ3.Finally,the xenograft mouse model demonstrated that the tumor volume and tumor were reduced after knockout of OPN,and the expression of intercellular TGF-β1 was reduced with high infiltration of NK cells.The OPN combined with TGF-β1 prediction model has better diagnostic performance.Conclusion: OPN secreted by tumor cells can impair NK cell activation and killing activities through integrin avβ3/NF-k B/TGF-β1signaling pathway,and lead to the malignant progression of HNSCC.