MiR-944/CISH Mediated Inflammation <i>via</i> STAT3 Is Involved in Oral Cancer Malignance by Cigarette Smoking

Abstract

Background: The cytokine-inducible Src homology 2-containing protein (CISH) is an endogenous suppressors of signal transduction and activator of transcription (STAT) and acts as a key negative regulator of inflammatory cytokine responses. Downregulation of CISH has been reported to associate with increased activation of STAT and enhanced inflammatory pathways. However, whether microRNAs (miRNAs) play a crucial role in CISH/STAT regulation and its underlying mechanism in oral squamous cell carcinoma (OSCC) remains unknown. Methods: The expression of CISH on OSCC patients was determine by quantitative real-time PCR (qRT-PCR) and immunohistochemistry. Specific targeting by miRNAs was determined by software prediction and luciferase reporter assay. The functions of miR-944 and CISH were accessed by transwell migration and invasion analyses using gain- and loss-of-function approaches. Enzyme-linked immunosorbent assay and qRT-PCR were used to evaluate the pro-inflammation cytokines expression under the miR-944, CISH, NNK or combinations treatment. Findings: CISH protein was significantly down-regulated in OSCC patients and cell lines and its level was inversely correlated with miR-944 expression. MiR-944-induced STAT3 phosphorylation, pro-inflammation cytokines secretion, migration and invasion were abolished by CISH restoration. Furthermore, tobacco extract (NNK) may contribute to the miR-944 induction and the activation of STAT3 in OSCC. MiR-944 inhibitor prevented the NNK-induced STAT3 phosphorylation and pro-inflammation cytokines secretion, indicating that miR-944 is required for NNK-induced cellular inflammatory pathways. Interpretation: Altogether, these data demonstrate that NNK-induced miR-944 expression plays an important role in CISH/STAT3-mediated inflammatory response and activation of tumor malignancy. Funding: This work was supported by National Health Research Institutes (NHRI) grants from Taiwan (NHRI-CA107-PP03, NHRI-CA108-PP03), and the Ministry of Health and Welfare (MOHW) grants from Taiwan (MOHW 108-TDU-B-212-124026, MOHW 109-TDU-B-212-134013). Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: The study protocol was approved by the Research Ethics Committee of National Health Research Institutes (EC1040409-E) and Institutional Human Experiment and Ethic Committee of National Cheng Kung University Hospital (HR-97-100) for the use of clinical materials for research purpose. Informed consent was obtained from each patient.