Sahar Esmailzadeh1, Behzad Mansoori1,2, Ali Mohammadi1, Dariush Shanehbandi1, Behzad Baradaran3. 1. Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. 2. Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran. 3. Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. behzad_im@yahoo.com.
Abstract
PURPOSE: Overexpression of HMGA2, known as small non-histone chromosomal protein, is associated with progression of various tumors, including colorectal cancers. The aim of this study was to investigate the effect of a specific HMGA2 siRNA on apoptosis and cell cycle of HCT-116 (colorectal carcinoma) cells. METHODS: The cells were transfected with siRNAs using a transfection reagent. The cytotoxic effects of HMGA2 siRNA on colorectal carcinoma cells were determined using MTT assay. Relative HMGA2 mRNA and protein levels were measured by QRT-PCR and western blotting, respectively. Apoptosis was measured by a TUNEL test based on labeling of DNA strand breaks. We also evaluated caspase-3, caspase-8, and caspase-9 expression by QRTPCR to determine which pathway is involved in apoptosis. Cell cycle was assessed by FACS and cell cycle analysis using PI DNA staining. RESULTS: HMGA2 siRNA significantly reduced both mRNA and protein expression levels 48 h after transfection and dose-dependent manner in colorectal carcinoma cells. We also showed that the silencing of HMGA2 led to the induction of apoptosis through intrinsic pathway and cell cycle arrest in G2/M phases of interphase in HCT-116 cells in vitro. CONCLUSIONS: These results propose that HMGA2 might play an important role in the progression of colorectal carcinoma and might be a potential therapeutic target for trigger apoptosis and cell cycle arrest in colorectal carcinoma.
PURPOSE: Overexpression of HMGA2, known as small non-histone chromosomal protein, is associated with progression of various tumors, including colorectal cancers. The aim of this study was to investigate the effect of a specific HMGA2 siRNA on apoptosis and cell cycle of HCT-116 (colorectal carcinoma) cells. METHODS: The cells were transfected with siRNAs using a transfection reagent. The cytotoxic effects of HMGA2 siRNA on colorectal carcinoma cells were determined using MTT assay. Relative HMGA2 mRNA and protein levels were measured by QRT-PCR and western blotting, respectively. Apoptosis was measured by a TUNEL test based on labeling of DNA strand breaks. We also evaluated caspase-3, caspase-8, and caspase-9 expression by QRTPCR to determine which pathway is involved in apoptosis. Cell cycle was assessed by FACS and cell cycle analysis using PI DNA staining. RESULTS:HMGA2 siRNA significantly reduced both mRNA and protein expression levels 48 h after transfection and dose-dependent manner in colorectal carcinoma cells. We also showed that the silencing of HMGA2 led to the induction of apoptosis through intrinsic pathway and cell cycle arrest in G2/M phases of interphase in HCT-116 cells in vitro. CONCLUSIONS: These results propose that HMGA2 might play an important role in the progression of colorectal carcinoma and might be a potential therapeutic target for trigger apoptosis and cell cycle arrest in colorectal carcinoma.
Entities:
Keywords:
Apoptosis; Cell cycle; Colorectal carcinoma; HMGA2 (high mobility group A2); Small interference RNA (siRNA)
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