Clusterin silencing inhibits proliferation and reduces invasion in human laryngeal squamous carcinoma cells

• Published in: World journal of surgical oncology 2014-04, Vol.12 (1), p.124-124, Article 124
• Main Authors: Wang, Qianjin, Cao, Weiyan, Su, Quancai, Liu, Zimin, Zhang, Lin
• Format: Article
• Language: English
• Subjects: Analysis; Animals; Apoptosis; Blotting, Western; Breast cancer; Carcinoma, Squamous Cell - metabolism; Carcinoma, Squamous Cell - pathology; Carcinoma, Squamous Cell - prevention & control; Cell Adhesion; Cell culture; Cell Movement; Cell Proliferation; Clusterin - antagonists & inhibitors; Clusterin - genetics; Clusterin - metabolism; Data analysis; Female; Gene expression; Gene Silencing; Humans; Laryngeal cancer; Laryngeal Neoplasms - metabolism; Laryngeal Neoplasms - pathology; Laryngeal Neoplasms - prevention & control; Lung Neoplasms - metabolism; Lung Neoplasms - prevention & control; Lung Neoplasms - secondary; Membranes; Metastasis; Mice; Mice, SCID; Neoplasm Invasiveness; Physiological aspects; Plasmids; Proteins; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; RNA, Small Interfering - genetics; Squamous cell carcinoma; Tumor Cells, Cultured; Tumors; Wound Healing
• ISSN: 1477-7819; 1477-7819
• DOI: 10.1186/1477-7819-12-124

Clusterin is, in its major form, a secreted heterodimeric disulfide-linked glycoprotein (sCLU), which plays important roles in cell survival and death. In laryngeal squamous cell carcinomas (LSCC), sCLU is up-regulated and its expression is related to the invasiveness of these tumors. The purpose of this study was to explore the inhibiting role of sCLU gene silence in the invasive ability and growth of Hep-2 human laryngeal squamous carcinoma cells (Hep-2) by transfection of short hairpin RNA expression plasmids against sCLU (sCLU-shRNA) (in vivo) or small interference RNA (sCLU-siRNA) (in vitro). sCLU-siRNA and the control siRNA were transfected into Hep-2 cells using Lipofectamine 2000. RT-PCR and Western blot were used to detect the effect of siRNA transfection on sCLU mRNA and sCLU protein expression. The invasive activity of sCLU-siRNA-transfected Hep-2 cells was measured with the modified Boyden chamber assay and wound healing assay. The effects of sCLU-siRNA on cell proliferation were evaluated by MTT assay. Apoptosis was measured by Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) double-staining methods. We next evaluated the effects of sCLU silencing by sCLU-shRNA transfection in vivo on tumor growth and metastatic properties to the lung. Terminal deoxytransferase-mediated dUTP nick end labeling (TUNEL) staining was used to observe the apoptosis in the xenografts. It showed that siRNA-mediated down-regulation of sCLU expression in Hep-2 cells significantly inhibited cell proliferation and promoted apoptosis in vitro. Furthermore, siRNA-mediated down-regulation of sCLU expression decreases in vitro cell migration and invasion ability. In vivo, the average volume of tumors in the sCLU-shRNA transfected group was significantly lower than in the control group (P