IL-21 induces pyroptosis of Treg cells via Akt-mTOR-NLRP3-caspase 1 axis in eosinophilic chronic rhinosinusitis.

BACKGROUND: Regulatory T (Treg) cells, which prevent inflammation-induced eosinophil infiltration, are deficient in nasal polyps (NPs) in patients with eosinophilic chronic rhinosinusitis (ECRS). It is concomitant with loss of Foxp3 after certain inflammatory stimuli. OBJECTIVE: We sought to determine the inflammatory cytokines involved in inducing the loss of Treg cells in NPs. METHODS: The abundance of cytokines in ECRS patients or mice were tested using ELISA, immunochemistry, immunofluorescence, quantitative reverse transcription PCR (qPCR), and/or flow cytometry. Expression of eosinophil cationic protein (ECP), CD4+ T cells, IL-4, and IL-17A and eosinophils in nasal mucosa of mouse model was investigated by immunochemistry, immunofluorescence, and hematoxylin and eosin staining. The percentage and death of induced Treg (iTreg) cells, source of IL-21 in NPs from ECRS and non-ECRS patients, and abundance of different systemic phenotypes of CD4+ T cells in a mouse model were studied by flow cytometry. Western blot analysis, scanning, and transmission electronic microscopy were used to detect pyroptosis of iTreg cells. RESULTS: IL-21 was highly expressed in nasal mucosa of ECRS patients and mice, causing pyroptosis and preventing development of iTreg cells in vitro. The elevated IL-21 in NPs from ECRS patients was mainly produced by CD3+ T cells, including T follicular helper, T peripheral helper, TH2, and TH17 cells and CD3+CD4- T cells. T peripheral helper cells and CD3+CD4- T cells were the predominant source of IL-21 in NPs from non-ECRS patients. Blocking IL-21/IL-21R signaling significantly reduced the number of eosinophils and CD4+ T cells along with ECP, IL-4, and IL-17A expression in the nasal mucosa of ECRS mice. It also increased Treg cell percentage and systemically decreased TH2 and TH17 ratios. Akt-mTOR inhibition prevented IL-21-induced pyroptosis in human and mouse iTreg cells. CONCLUSION: Elevated IL-21 drives pyroptosis and prevents Treg cell development in ECRS patients. IL-21 induced pyroptosis via activating Akt-mTOR-NLRP3-caspase 1 signaling.

Interleukin-19 upregulates fibronectin and collagen I expression via the NF-κB-Smad2/3 pathway in fibroblasts of patients with chronic rhinosinusitis.

BACKGROUND: Tissue remodeling is a prominent characteristic of chronic rhinosinusitis (CRS). Excess deposition of fibronectin (FN) and collagen (Col) I by fibroblasts is crucial for the pathologic tissue remodeling in CRS without nasal polyps (CRSsNP). Increased interleukin (IL)-19 level in patients with CRS had been demonstrated in our previous studies. Here, we aimed to evaluate the role of IL-19 in mediating FN and Col I expression in CRS. METHODS: Nasal mucosal tissue samples were collected from patients with CRS with nasal polyps (CRSwNP), CRSsNP, and controls. The expression of IL-19, vimentin, FN, and Col I were detected using immunohistochemistry and immunofluorescence. Primary human nasal fibroblasts were treated with IL-19, then the activation of Smad2/3, NF-κB and relevant pathways, and the expression of FN and Col I were measured. RESULTS: Expression levels of vimentin, FN, and Col I were significantly increased in nasal tissues from patients with CRSsNP compared with CRSwNP and control subjects. Moreover, IL-19 co-localized with FN and Col Ι in nasal tissues. IL-19-treated fibroblasts had increased production of FN and Col I, which was associated with the activated Smad2/3 and NF-κB pathways. Moreover, Smad2/3 activation was mediated by the NF-κB pathway in IL-19-treated fibroblasts. CONCLUSIONS: IL-19 promotes FN and Col I production via the activated NF-κB-Smad2/3 pathway in fibroblasts, leading to fibrosis and collagen deposition in patients with CRS.

IL-17A mediates pyroptosis via the ERK pathway and contributes to steroid resistance in CRSwNP.

BACKGROUND: Pyroptosis is closely related to inflammation. However, the molecular mechanisms and pathologic contributions of pyroptotic epithelial cell are not yet fully understood. OBJECTIVE: This study aimed to explore the function and molecular mechanisms of IL-17A on human nasal epithelial cell (hNEC) pyroptosis. METHODS: The expression of pyroptosis-related biomarkers and IL-17A was assessed in sinonasal mucosa from control individuals, patients with chronic rhinosinusitis without nasal polyps, and patients with chronic rhinosinusitis with nasal polyps (CRSwNP) by using quantitative RT-PCR. Their localization was analyzed via immunohistochemistry and immunofluorescence. The ultrastructural characteristics of IL-17A-induced pyroptosis in hNECs were visualized by using electron microscopy. IL-17A functional assays were performed on hNECs and airway epithelial cell lines. Cytokine levels were quantified via ELISA. The signaling pathways involved in IL-17A-induced pyroptosis were studied via unbiased RNA sequencing and Western blotting. RESULTS: The expression of IL-17A and the pyroptotic biomarkers NOD-like receptor family, pyrin domain containing 3 (NLRP3), caspase-1, gasdermin D, and IL-1ß was increased in nasal mucosa from patients with CRSwNP compared with in those with chronic rhinosinusitis without nasal polyps and the control subjects. IL-17A was positively correlated and colocalized with the pyroptotic biomarkers. IL-17A treatment induced pyroptosis in the hNECs and cell lines analyzed, primarily through the extracellular signal-regulated kinase (ERK)-NLRP3/caspase-1 signaling pathway, and increased IL-1ß and IL-18 secretion in hNECs. Moreover, IL-17A-induced pyroptosis contributed to steroid resistance by affecting glucocorticoid receptor-α and glucocorticoid receptor-ß expression, and the inhibition of pyroptotic proteins partially abolished IL-17A-induced steroid resistance in hNECs. CONCLUSION: Elevated IL-17A level promotes pyroptosis in hNECs through the ERK-NLRP3/caspase-1 signaling pathway and contributes to glucocorticoid resistance by affecting glucocorticoid receptor homeostasis in patients with CRSwNP.

Tetrandrine sensitizes nasopharyngeal carcinoma cells to irradiation by inducing autophagy and inhibiting MEK/ERK pathway.

Radioresistance was the main reason for local recurrence and metastasis of nasopharyngeal carcinoma. Tetrandrine is reported as an antitumor drug via inducing cell cycle arrest and apoptosis. In this study, the radiosensitization effects of maximum noncytotoxic doses of tetrandrine in nasopharyngeal carcinoma were analyzed both in vitro and in vivo, using MTT assay, western blot, TUNEL, and HE staining. It was found that the maximum dose of tetrandrine inhibited the phosphorylation of ERK and MEK induced by irradiation, and significantly enhanced irradiation-induced cell growth inhibition in nasopharyngeal carcinoma cells CNE1, CNE2, and C666-1. The ERK activator and overexpression of ERK reversed the radiosensitization effect of tetrandrine. About 50 mg/kg of tetrandrine which was used as the maximum noncytotoxic dose of tetrandrine in vivo, enhanced the radiosensitivity of the xenograft tumor and increased the apoptosis rate of the xenograft tumor cells caused by irradiation, while did not raise the side effect of the treatment. Moreover, tetrandrine increased autophagy in nasopharyngeal carcinoma cells. These results suggested that the maximum noncytotoxic dose of tetrandrine sensitized nasopharyngeal carcinoma to irradiation by inhibiting MEK/ERK pathway and inducing autophagy.

A low dose of AZD8055 enhances radiosensitivity of nasopharyngeal carcinoma cells by activating autophagy and apoptosis.

Activation of the PI3K/mTOR pathways is significantly correlated with a poor prognosis in nasopharyngeal carcinoma (NPC). Inhibition of these pathways was reported to be effective in restoring radiosensitivity. In this study, the activity of the novel ATP-competitive, orally bioavailable mTOR inhibitor AZD8055 was found to inhibit the phosphorylated mTOR and NPC cells proliferation. The IC50 doses in CNE1 and CNE2 cell lines were 60 and 100 nanomolar, respectively. AZD8055 significantly enhanced the inhibitions of cell growth and colony formation induced by irradiation (P < 0.05 for all). AZD8055 at the IC50 doses prolonged G2/M arrest (P < 0.05) and promoted the apoptosis (P < 0.01) induced by irradiation and autophagy in NPC cells. Blocking autophagy weaken the cell growth inhibition and decreased apoptosis induced by AZD8055 combined with irradiation. Treatment with AZD8055 at 5, 10 and 20 mg/kg/d significantly enhanced NPC cell radiosensitivity in vivo and significantly induced apoptosis and autophagy in tumor tissues, Neither 5 nor 20 mg/kg/d AZD8055 induced significantly pro-apoptosis bax expressions in mouse livers and kidneys. 5 mg/kg/d produced good radiosensitivity but had little impact on body weight. We concluded that AZD8055 was a promising candidate radiosensitizer for NPC.