2013•Molecular Medicine ReportsOpen access

Suppression of cell proliferation and collagen production in cultured human hypertrophic scar fibroblasts by Sp1 decoy oligodeoxynucleotide

Chen-Liang Deng, Zheng Jianghong, Weidong Wan, SHIXIN ZHANG, Zhi Ding, Guangyu Mao, Songlin Yang

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Abstract

Hypertrophic scars are characterized by the abnormal proliferation of fibroblasts and an overproduction of collagen. The Sp1 transcription factor is involved in the stimulation of collagen synthesis. A decoy oligonucleotide (ODN) targeting Sp1 was designed and transfected into hypertrophic scar fibroblasts (HSFs) cells using cationic liposomes. The transfection efficiency was determined by flow cytometry and was observed to be 85±7% (n=5). Specific binding of the Sp1 decoy ODN was monitored with an electrophoretic mobility shift assay (EMSA). Following transfection with the decoy ODN to Sp1, cell viability and cell proliferation, which were examined by the cell counting kit WST‑8, were decreased by 80% compared with untreated cells. Transforming growth factor‑β (TGF‑β) mRNA and collagen mRNA expression were also reduced by 48% in the transfection decoy ODN group. The cell viability of HSFs after 48 h of transfection with 25, 50, 100 and 150 nM Sp1 decoy ODN was 0.9331±0.0203, 0.7479±0.0868, 0.577±0.0347 and 0.4703±0.0147, respectively. The 100 nM dose of the Sp1 decoy ODN inhibited the expression of types I and III collagen by 32 and 28%, respectively (both P<0.01). TGF‑β mRNA expression was also effectively suppressed by the 100 nM Sp1 decoy ODN (P<0.01). The Sp1 decoy ODN inhibited cell proliferation and the expression of types I and III collagen. Therefore, Sp1 decoy ODNs may be a promising tool for developing and testing novel therapeutic applications for treating hypertrophic scars.

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Hypertrophic scars are characterized by the abnormal proliferation of fibroblasts and an overproduction of collagen. The Sp1 transcription factor is involved in the stimulation of collagen synthesis. A decoy oligonucleotide (ODN) targeting Sp1 was designed and transfected into hypertrophic scar fibroblasts (HSFs) cells using cationic liposomes. The transfection efficiency was determined by flow cytometry and was observed to be 85±7% (n=5). Specific binding of the Sp1 decoy ODN was monitored with an electrophoretic mobility shift assay (EMSA). Following transfection with the decoy ODN to Sp1, cell viability and cell proliferation, which were examined by the cell counting kit WST‑8, were decreased by 80% compared with untreated cells. Transforming growth factor‑β (TGF‑β) mRNA and collagen mRNA expression were also reduced by 48% in the transfection decoy ODN group. The cell viability of HSFs after 48 h of transfection with 25, 50, 100 and 150 nM Sp1 decoy ODN was 0.9331±0.0203, 0.7479±0.0868, 0.577±0.0347 and 0.4703±0.0147, respectively. The 100 nM dose of the Sp1 decoy ODN inhibited the expression of types I and III collagen by 32 and 28%, respectively (both P<0.01). TGF‑β mRNA expression was also effectively suppressed by the 100 nM Sp1 decoy ODN (P<0.01). The Sp1 decoy ODN inhibited cell proliferation and the expression of types I and III collagen. Therefore, Sp1 decoy ODNs may be a promising tool for developing and testing novel therapeutic applications for treating hypertrophic scars.

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Available abstract

Hypertrophic scars are characterized by the abnormal proliferation of fibroblasts and an overproduction of collagen. The Sp1 transcription factor is involved in the stimulation of collagen synthesis. A decoy oligonucleotide (ODN) targeting Sp1 was designed and transfected into hypertrophic scar fibroblasts (HSFs) cells using cationic liposomes. The transfection efficiency was determined by flow cytometry and was observed to be 85±7% (n=5). Specific binding of the Sp1 decoy ODN was monitored with an electrophoretic mobility shift assay (EMSA). Following transfection with the decoy ODN to Sp1, cell viability and cell proliferation, which were examined by the cell counting kit WST‑8, were decreased by 80% compared with untreated cells. Transforming growth factor‑β (TGF‑β) mRNA and collagen mRNA expression were also reduced by 48% in the transfection decoy ODN group. The cell viability of HSFs after 48 h of transfection with 25, 50, 100 and 150 nM Sp1 decoy ODN was 0.9331±0.0203, 0.7479±0.0868, 0.577±0.0347 and 0.4703±0.0147, respectively. The 100 nM dose of the Sp1 decoy ODN inhibited the expression of types I and III collagen by 32 and 28%, respectively (both P<0.01). TGF‑β mRNA expression was also effectively suppressed by the 100 nM Sp1 decoy ODN (P<0.01). The Sp1 decoy ODN inhibited cell proliferation and the expression of types I and III collagen. Therefore, Sp1 decoy ODNs may be a promising tool for developing and testing novel therapeutic applications for treating hypertrophic scars.

Key concepts: Decoy, Transfection, Molecular biology, Hypertrophic scar, Flow cytometry, Cell growth, Viability assay, Cell

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Suppression of cell proliferation and collagen production in cultured human hypertrophic scar fibroblasts by Sp1 decoy oligodeoxynucleotide — Research Paper | ScholarLens