Transfer of siRNA against XIAP induces apoptosis and reduces tumor cells growth potential in human breast cancer in vitro and in vivo
Authors: Yu Wang
Ruhui Zhang
Yingchao Zhang
Mingku Jia
Wenzeng Guan
Wenxin Gao
Xichun Han
Journal: Breast cancer research and treatment (2006)
Discipline: CLINICAL MEDICINE
Keywords: Gene targeting; short interfering RNA; siRNA; gene therapy; X-linked inhibitor of apoptosis
Affiliations: Department of Surgery, The Second Hospital of Jilin University, Changchun, PR China.

Abstract:  Background: Gene targeting using short interfering RNA(siRNA) has become a common strategy to explore gene function because of its prominent efficacy and specificity. It is proven that the application of siRNA technology to gene therapy is effective. In this study, we constructed a siRNA expression plasmid against gene X-linked inhibitor of apoptosis (XIAP), and then used breast cancer cells MCF-7 to assess its functions. Materials and methods: XIAP siRNA plasmid was constructed using an U6pro vector contained U6 promoter, After the plasmid had been transfected into MCF-7 cells and effected on the cell cycle, the expression change of XIAP was examined by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. The apoptosis of the transfected cells was analyzed by flow cytometry, and TUNEL method. The in vitro cellular growth activities were assayed by MTT incorporation. Twenty-four nude mice were randomly divided into 3 equal groups and were inoculated with electroinjection of blank plasmid, scrambled nucleotide control (control siRNA), or siRNA against XIAP subcutaneously respectively, then the appearance and size of tumors were observed. Four weeks later the mice were killed and the volumes of tumor were calculated so as to evaluate the therapeutic effects of siRNA against XIAP. Results: The successful construction of siRNA against XIAP plasmid was identified with sequencing. After the siRNA expression vector was transfected into the MCF-7 cells, the expression of XIAP gene was inhibited significantly (by 90%). The cellular growth activities in the MCF-7 cells transfected with siRNA against XIAP plasmid decreased obviously. The siRNA against XIAP plasmid knocked down XIAP expression in MCF-7 cells obviously, arrested the cell cycle in G1 phase, inhibited cell proliferation significantly, and promoted cell apoptosis in a tendency. TUNEL assay and flow cytometry showed that the classic apoptosis characters of the MCF-7 cells transfected with siRNA against XIAP plasmid manifested an apoptosis rate of 77.2%, significantly higher than those in the control siRNA group and in the blank plasmid group (both p &lt; 0.01). The growth speed and formation rate of xenograft tumor in mice transfected with siRNA against XIAP transfected mice slowed down significantly. By HE staining, a lot of necrotic tissues could be observed in the siRNA against XIAP transfected group, however, there was no similar inhibitive effect in the control siRNA or blank plasmid group. Conclusion: This study represents that MCF-7 transfected cells with siRNA against XIAP remarkably suppress tumor growth and induces apoptosis, both in vitro and in vivo. This novel modality may be a promising tool for cancer therapy. 

Retracted: No

PubPeer comments:
 The Western blots and other figures in this paper have an interesting layout that appears to be shared with many other papers, as discussed in a January 2020 post on Leonid Schneider s blog: https: forbetterscience.com 2020 01 24 the-full-service-paper-mill-and-its-chinese-customers The list of these papers is available here: https: docs.google.com spreadsheets d 1KXqTAyl4j-jVorFPMD2XRpr76LcIKJ0CVyIvRj0exYQ edit?usp=sharing In almost all papers, Western blot panels within the same figure, and across figures and papers, appear to share the same background, while the bands are regularly spaced, all have similar rounded edges without the usual smudges and specks, and with some bands showing a recognizable  jumping sardine shape. Also in almost all papers, bar graphs show a characteristic layout with bars in solid shades of grey, and the left-most bar in black, as well as double-edged error bars. Many papers have  cell viability and  apoptotic cells graphs. In some of these papers, including this one, flow cytometry plot show a unnaturally dense and well defined  hairball of events in Q3 with a few regularly spaced events in Q1 and Q2, and a rectangular or crab-shaped block of events in Q4. To illustrate the recognizable layout of the graphs in this paper, here is Figure 1.  , 
 Figure 3. * Red boxes: Panel E (migration) III appears to show an overlap with panel G (invasion) III.  , 
 Retracted [10 May 2021] . -------------------------------------------- The above article, published online on 23 Oct 2017 in Wiley Online Library , has been retracted by agreement between the journal s Editor in Chief, Prof. Dr. Christian Behl, and Wiley Periodicals LLC. The retraction has been agreed following an investigation based on allegations raised by a third party. Several flaws and inconsistencies between results presented and experimental methods described were found, thus the editors consider the conclusions of this article to be invalid. The authors do not agree on the retraction. ,