Thalidomide attenuates oral epithelial cell apoptosis and pro-inflammatory cytokines secretion induced by radiotherapy via the miR-9-3p/NFATC2/NF-κB axis

Oral mucositis is the most common oral complication of cancer patients receiving radiotherapy or chemotherapy, leading to poor quality of life. Increasing clinical studies demonstrated that thalidomide (THD) can effectively ameliorate radiation-induced oral mucositis (RIOM). Here we established an experimental mouse model, radiation-induced human oral epithelial cells (HOECs), and further investigate the underlying mechanism the THD protective effect against RIOM. Combined with RNA sequencing result, we selected the gene nuclear factor of activated T cells c2 (NFATC2) as the most interesting candidate. THD downregulated NFATC2 expression, attenuated human oral epithelial cells (HOECs) apoptosis and promoted pro-inflammatory factors secretion. Further studies show that overexpression of NFATC2 in HOECs promotes cells apoptosis and pro-inflammatory cytokines level, while inhibition of NFATC2 present an opposite effect. Additionally, the regulatory miRNA of NFATC2 was predicted using StarBase, and the targeting relationship between miR-9-3p and NFATC2 was confirmed using a dual-luciferase reporter gene assay. miR-9-3p mimic reversed the elevated cell apoptosis and pro-inflammatory cytokines level by radiation or NFATC2-overexpression. Furthermore, NFATC2 upregulated the phosphorylation of p65, thus activating the NF-kappa B pathway in RIOM; while miR-9-3p reduced this effect. In conclusion, THD attenuates oral epithelial cell apoptosis and pro-inflammatory cytokines secretion induced by radiotherapy via the miR-9-3p/NFATC2/NF- NF-kappa B axis. (C) 2022 The Authors. Published by Elsevier Inc.

Automated summary

Oral mucositis, the most common complication in cancer patients undergoing radiotherapy or chemotherapy, can significantly reduce the patients' quality of life. Thalidomide has been shown to effectively alleviate radiotherapy-induced oral mucositis based on increasing clinical studies. In this study, a mouse model of radiation-induced human oral epithelial cells (HOECs) was established to investigate the mechanism underlying the protective effect of thalidomide against oral mucositis. Through RNA sequencing, the gene NFATC2 was identified as a potential target. Thalidomide downregulated NFATC2 expression, reduced HOECs apoptosis, and promoted the secretion of pro-inflammatory factors. Further experiments demonstrated that NFATC2 overexpression promoted apoptosis and pro-inflammatory cytokine production in HOECs, while NFATC2 inhibition had the opposite effect. The miRNA miR-9-3p was predicted as a regulator of NFATC2 and its targeting relationship with NFATC2 was confirmed. miR-9-3p mimic reversed the increased apoptosis and pro-inflammatory cytokine levels induced by radiation or NFATC2 overexpression. Moreover, NFATC2 upregulated the phosphorylation of p65, activating the NF-kappa B pathway in radiation-induced oral mucositis, while miR-9-3p reduced this effect. In conclusion, thalidomide attenuates oral epithelial cell apoptosis and pro-inflammatory cytokine secretion induced by radiotherapy through the miR-9-3p/NFATC2/NF-kappa B axis.