Morteza Akbari; Dariush Shanehbandi; Milad Asadi; Navid Shomali; Afsaneh Faraji; Vahid Khaze; Abbas Pakdel; Ahad Mokhtarzadeh; Ali Asghar Ebrahimi; Aliakbar Shabani; Behzad Baradaran
Abstract
Background: Colorectal cancer (CRC) is attributed as one of the most common malignancies worldwide. CD133 molecule, as a pentaspan transmembrane glycoprotein, confers stem cell-related characteristics, including self-renewal and multi-directional differentiation capability. CD133 plays important roles ...
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Background: Colorectal cancer (CRC) is attributed as one of the most common malignancies worldwide. CD133 molecule, as a pentaspan transmembrane glycoprotein, confers stem cell-related characteristics, including self-renewal and multi-directional differentiation capability. CD133 plays important roles in the progression of CRC by conferring apoptotic resistance and migration ability. Objective: To investigate the anti-apoptotic and anti-angiogenic effect of CD-133 targeted siRNA in a colorectal cancer cell line. Methods: In this study, CD133-targeted siRNA transfection was conducted into HT-29 cells. MTT assay was employed to evaluate the cytotoxic effects of transfection on the cells. Flow cytometry was used to evaluate the apoptosis rate. The mRNA expression of apoptosis and metastasis related genes were assessed by quantitative Real-Time PCR (qRT-PCR). Wound healing assay was used to assess the migration potency of the infected cells. Results: Expression of CD133 was significantly downregulated after transfection of CD133-specific siRNA. Moreover, the rate of apoptosis was significantly increased after transfection. The migration potential of cells was diminished after transfection. siRNA delivery resulted in the modulation of expression of apoptosis and metastasis-related genes. Conclusion: siRNA mediated targeting of CD133 could be considered as a promising approach to treat CRC through suppressing the cancerous behavior of tumor cells.
Padideh Ebadi; Mohammad Hossein Karimi; Ali Akbar Pourfathollah; Saheb Ghadam Lotfi; Zahra Soheila Soheili; Shahram Samiee; Smerdis Hajati; Fatemeh Nadali; Bita Geramizadeh; Seyyed Mohammad Moazzeni
Volume 6, Issue 1 , March 2009, , Pages 1-11
Abstract
Background: Dendritic cells (DCs) are ideal accessory cells in the field of gene therapy. Delivery of DNA and siRNA into mammalian cells is a useful technique in treating various diseases caused by single gene defects. Selective gene silencing by small interfering RNAs (siRNAs) and antisense oligodeoxynucleotides ...
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Background: Dendritic cells (DCs) are ideal accessory cells in the field of gene therapy. Delivery of DNA and siRNA into mammalian cells is a useful technique in treating various diseases caused by single gene defects. Selective gene silencing by small interfering RNAs (siRNAs) and antisense oligodeoxynucleotides (ODN)s is an efficient method for the manipulation of cellular functions. An efficient, functional delivery system with no toxicity problems would be attractive. Objective: We compared two commercially available cationic lipids, Lipofectamine and FuGENE6, in the delivery of both siRNA and antisense ODNs into mice spleen-derived DCs. Methods: Cellular uptake was measured by the means of fluorescein-labelled non-silencing siRNA and antisense ODNs as a model system using flow cytometry. Cytotoxicity of the two delivery systems was compared with propidium iodide and annexin-V staining, and quantified with flow cytometry. The efficiency of our oligonucleotide delivery systems was compared by measuring CD40 expression by flow cytometry. Results: CD40 expression in DCs was 38%. After siRNA transfection by Lipofectamine, CD40 expression decreased to 13%, and after transfection by FuGENE6, it decreased to 18%. The difference was statistically significant. CD40 down regulation in DCs transfected with the two different antisense sequences by Lipofectamine was 21% and 23%, and down regulation after transfection by FuGENE6 was 19% and 18%, respectively. The differences were not statistically significant. The effects of siRNA and antisense ODNs were specific. Conclusion: Lipofectamine was a more potent delivery system in siRNA effect, followed by FuGENE6. There was no significant difference between Lipofectamine and FuGENE6 as a delivery system of antisense ODNs.