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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...

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Autores principales: 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
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2012
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.
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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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