Preventing intimal thickening of vein grafts in vein artery bypass using STAT-3 siRNA
Authors: Xiufen Zheng
Zhicheng Zhu
Bo Li
Kaitelynne H Ling
Keyan Zhao
Jiangbin Sun
Amit N Patel
Jinhua Zheng
Tiance Wang
Zhongshang Xie
Kexiang Liu
Weiping Min
Journal: Journal of Translational Medicine (2012)
Discipline: CLINICAL MEDICINE
Keywords: Vascular smooth muscle cells; STAT-3; siRNA; proliferation; migration
Affiliations: Department of Cardiovascular Surgery, The Second Hospital, Jilin University, Chang Chun, China.

Abstract:  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&amp;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 &lt; 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 &lt; 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. 

Retracted: No

PubPeer comments:
 image compressed ![file] Disclaimer: These potential image duplications have been detected automatically using ImageTwin http: dx.doi.org 10.1145 3019612.3019875 Although results have been curated, the automatic algorithm may have generated false positive results and cannot take into account all scientific explanations for apparent duplications like stripped and reprobed blots, time series, serial sections and reused controls. , 
 The finding above is from Figure 4, described as follows: u201cSTAT-3 siRNA prevent VMSCs proliferation in vivo. Left jugular vein was interposed into the left common carotid artery and treated with either STAT-3 siRNA or scramble siRNA as described in the Material Methods. The interposed veins were collected and applied to immunohistochemistry Staining with PCNA (A) and Ki-67 (B) on day 3 and 7 after operation as described in the Material Methods. (a b): Scramble siRNA treatment; (c d): STAT-3 siRNA. (a) (c): day 3; (b) (d): day 7. Data are representative of five mice in each group.u201d This suggests that the tissues were stained differently so I believe the similarities seen in #1 are unexpected. Could the authors comment? , 
 Figure 1B appears to have a repeated image. *** ![file] , 
 Figure 4 appears to have another set of images with similarities (one area shown within green boxes). Again, I think this is unexpected, but maybe I am misunderstanding. *** ![file] , 
 Thanks for your careful check. We will look up our original images to see if we uploaded them by mistake. ,