Abstract
BackgroundTo investigate the effect of DACH1 over-expression on proliferation and invasion of laryngeal squamous cell carcinoma (LSCC).MethodsThe 120 cases of LSCC tumors and 114 adjacent non-neoplastic tissues were collected to detect the expression of DACH1 by immunohistochemistry. The changes of DACH1 expression from each group were assessed and correlated to the clinical parameters of the patients. Plasmid-DACH1 was transfected into Hep-2 cells to up-regulate the expression of DACH1C. Real-time PCR, Western blot, CCK8 and transwell assay were used to verify the cell proliferation and invasion after plasmid-DACH1 transfection.ResultsThe results indicated that DACH1 was downregulated in LSCC tissues as compared to corresponding adjacent non-neoplastic tissues. Decreased expression of DACH1 was found in the tumors upraglottic tumor, lymph node metastases, T3–4 stage and advanced clinical stage. In Hep-2 cells, transfection with plasmid-DACH1 could suppress cell proliferation, invasion and induce G1 phase extension in cell cycle.ConclusionsDACH1 may act as a tumor suppressor gene and could be a potential target for therapeutic intervention of LSCC.
Highlights
To investigate the effect of DACH1 over-expression on proliferation and invasion of laryngeal squamous cell carcinoma (LSCC)
DACH1 was mainly found in the nucleus (Fig. 1), and its expression was associated with several clinical parameters, including smoking, drinking, T classification, lymph node metastases, primary location and clinical stage in LSCC paraffin specimens (Table 1)
DACH1 expression was up-regulated after DACH1 plasmid transfection in Hep-2 cells Plasmid-DACH1 was transfected in Hep-2 cells to up-regulate the expression of DACH1 in order to further explore the functional roles of DACH1 in LSCC (Fig. 2)
Summary
DACH1 plasmid with GFP as a reporter gene was transfected into Hep-2 cells with Lipofectamine 2000 (Invitrogen) according to the manufacturer’s instructions. Real-time PCR and Western blotting Total RNA was isolated using Trizol reagent (Invitrogen) according to the manufacturer’s instructions. Untreated Hep-2 cells and GFP-plasmid groups were collected and analyzed by western blot to assess the expression of DACH1. Cell cycle analysis After transfecting with plasmid-DACH1 for 72 h, the cells were washed twice with cold PBS and fixed in 75% ethanol for 24 h. The positive cancer cells were evaluated on a 5-point scale according to the fraction of stained cells (0, < 1%, 1, 1–10%, 2, 10–50%, 3, 50–80%, 4, 80–100%). DACH1 low-expression based on the percentage of positive cells was lower than the mean value.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
