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Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA
BACKGROUND: Proliferation and migration of vascular smooth muscle cells (VSMCs) play a key role in neointimal formation which leads to restenosis of vein graft in venous bypass. STAT-3 is a transcription factor associated with cell proliferation. We hypothesized that silencing of STAT-3 by siRNA wil...
Autores principales: | , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3286375/ https://www.ncbi.nlm.nih.gov/pubmed/22216901 http://dx.doi.org/10.1186/1479-5876-10-2 |
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author | Sun, Jiangbin Zheng, Jinhua Ling, Kaitelynne H Zhao, Keyan Xie, Zhongshang Li, Bo Wang, Tiance Zhu, Zhicheng Patel, Amit N Min, Weiping Liu, Kexiang Zheng, Xiufen |
author_facet | Sun, Jiangbin Zheng, Jinhua Ling, Kaitelynne H Zhao, Keyan Xie, Zhongshang Li, Bo Wang, Tiance Zhu, Zhicheng Patel, Amit N Min, Weiping Liu, Kexiang Zheng, Xiufen |
author_sort | Sun, Jiangbin |
collection | PubMed |
description | BACKGROUND: Proliferation and migration of vascular smooth muscle cells (VSMCs) play a key role in neointimal formation which leads to restenosis of vein graft in venous bypass. STAT-3 is a transcription factor associated with cell proliferation. We hypothesized that silencing of STAT-3 by siRNA will inhibit proliferation of VSMCs and attenuate intimal thickening. METHODS: Rat VSMCs were isolated and cultured in vitro by applying tissue piece inoculation methods. VSMCs were transfected with STAT 3 siRNA using lipofectamine 2000. In vitro proliferation of VSMC was quantified by the MTT assay, while in vivo assessment was performed in a venous transplantation model. In vivo delivery of STAT-3 siRNA plasmid or scramble plasmid was performed by admixing with liposomes 2000 and transfected into the vein graft by bioprotein gel applied onto the adventitia. Rat jugular vein-carotid artery bypass was performed. On day 3 and7 after grafting, the vein grafts were extracted, and analyzed morphologically by haematoxylin eosin (H&E), and assessed by immunohistochemistry for expression of Ki-67 and proliferating cell nuclear antigen (PCNA). Western-blot and reverse transcriptase polymerase chain reaction (RT-PCR) were used to detect the protein and mRNA expression in vivo and in vitro. Cell apoptosis in vein grafts was detected by TUNEL assay. RESULTS: MTT assay shows that the proliferation of VSMCs in the STAT-3 siRNA treated group was inhibited. On day 7 after operation, a reduced number of Ki-67 and PCNA positive cells were observed in the neointima of the vein graft in the STAT-3 siRNA treated group as compared to the scramble control. The PCNA index in the control group (31.3 ± 4.7) was higher than that in the STAT-3 siRNA treated group (23.3 ± 2.8) (P < 0.05) on 7d. The neointima in the experimental group(0.45 ± 0.04 μm) was thinner than that in the control group(0.86 ± 0.05 μm) (P < 0.05).Compared with the control group, the protein and mRNA levels in the experimental group in vivo and in vitro decreased significantly. Down regulation of STAT-3 with siRNA resulted in a reduced expression of Bcl-2 and cyclin D1. However, apoptotic cells were not obviously found in all grafts on day 3 and 7 post surgery. CONCLUSIONS: The STAT-3 siRNA can inhibit the proliferation of VSMCs in vivo and in vitro and attenuate neointimal formation. |
format | Online Article Text |
id | pubmed-3286375 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-32863752012-02-25 Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA Sun, Jiangbin Zheng, Jinhua Ling, Kaitelynne H Zhao, Keyan Xie, Zhongshang Li, Bo Wang, Tiance Zhu, Zhicheng Patel, Amit N Min, Weiping Liu, Kexiang Zheng, Xiufen J Transl Med Research BACKGROUND: Proliferation and migration of vascular smooth muscle cells (VSMCs) play a key role in neointimal formation which leads to restenosis of vein graft in venous bypass. STAT-3 is a transcription factor associated with cell proliferation. We hypothesized that silencing of STAT-3 by siRNA will inhibit proliferation of VSMCs and attenuate intimal thickening. METHODS: Rat VSMCs were isolated and cultured in vitro by applying tissue piece inoculation methods. VSMCs were transfected with STAT 3 siRNA using lipofectamine 2000. In vitro proliferation of VSMC was quantified by the MTT assay, while in vivo assessment was performed in a venous transplantation model. In vivo delivery of STAT-3 siRNA plasmid or scramble plasmid was performed by admixing with liposomes 2000 and transfected into the vein graft by bioprotein gel applied onto the adventitia. Rat jugular vein-carotid artery bypass was performed. On day 3 and7 after grafting, the vein grafts were extracted, and analyzed morphologically by haematoxylin eosin (H&E), and assessed by immunohistochemistry for expression of Ki-67 and proliferating cell nuclear antigen (PCNA). Western-blot and reverse transcriptase polymerase chain reaction (RT-PCR) were used to detect the protein and mRNA expression in vivo and in vitro. Cell apoptosis in vein grafts was detected by TUNEL assay. RESULTS: MTT assay shows that the proliferation of VSMCs in the STAT-3 siRNA treated group was inhibited. On day 7 after operation, a reduced number of Ki-67 and PCNA positive cells were observed in the neointima of the vein graft in the STAT-3 siRNA treated group as compared to the scramble control. The PCNA index in the control group (31.3 ± 4.7) was higher than that in the STAT-3 siRNA treated group (23.3 ± 2.8) (P < 0.05) on 7d. The neointima in the experimental group(0.45 ± 0.04 μm) was thinner than that in the control group(0.86 ± 0.05 μm) (P < 0.05).Compared with the control group, the protein and mRNA levels in the experimental group in vivo and in vitro decreased significantly. Down regulation of STAT-3 with siRNA resulted in a reduced expression of Bcl-2 and cyclin D1. However, apoptotic cells were not obviously found in all grafts on day 3 and 7 post surgery. CONCLUSIONS: The STAT-3 siRNA can inhibit the proliferation of VSMCs in vivo and in vitro and attenuate neointimal formation. BioMed Central 2012-01-04 /pmc/articles/PMC3286375/ /pubmed/22216901 http://dx.doi.org/10.1186/1479-5876-10-2 Text en Copyright ©2012 Sun et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Sun, Jiangbin Zheng, Jinhua Ling, Kaitelynne H Zhao, Keyan Xie, Zhongshang Li, Bo Wang, Tiance Zhu, Zhicheng Patel, Amit N Min, Weiping Liu, Kexiang Zheng, Xiufen Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA |
title | Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA |
title_full | Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA |
title_fullStr | Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA |
title_full_unstemmed | Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA |
title_short | Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA |
title_sort | preventing intimal thickening of vein grafts in vein artery bypass using stat-3 sirna |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3286375/ https://www.ncbi.nlm.nih.gov/pubmed/22216901 http://dx.doi.org/10.1186/1479-5876-10-2 |
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