Effects of RNA interference-mediated silencing of toll-like receptor 4 gene on proliferation and apoptosis of human breast cancer MCF-7 and MDA-MB-231 cells: An in vitro study
Authors: Ya-Qiong Huang
Dai Liu
Jian-Nan Gong
Er-Jing Cheng
Yan-Ting Dong
Yan Chen
Xiao-Ling Gao
Shu-Juan Wang
Jiao-Jiao Yang
Xiang-Li Wang
Dong-Dong An
Bei Wang
Journal: Journal of Cellular Physiology (2018)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: Bcl-2; CDK4; caspase-3; cyclin D1; human breast cancer
Affiliations: Department of Respiratory and Critical Care Medicine, The Second Hospital of Shanxi Medical University, Taiyuan, P. R. China., Shanxi Medical University, Taiyuan, P. R. China.

Abstract:  Breast cancer is known as the most prevalent cancer in women worldwide, and has an undeniable negative impact on public health, both physically, and mentally. This study aims to investigate the effects of toll-like receptor 4 (TLR4) gene silencing on proliferation and apoptosis of human breast cancer cells to explore for a new theoretical basis for its treatment. TLR4 small interference RNA (siRNA) fragment recombinant plasmids were constructed, including TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3. Human breast cancer MCF-7 and MDA-MB-231 cells were assigned into blank, negative control (NC), TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3 groups. MCF-7 and MDA-MB-231 cell growth was detected by MTT assay. Apoptosis and cell cycle were determined by flow cytometry. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis were conducted to determine the expression of TLR4, CDK4, cyclin D1, Livin, Bcl-2, p53, c-FLIP, and caspase-3. In comparison with the NC and blank groups, the TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3 groups showed decreased the expression of TLR4, inhibited proliferation of MCF-7 and MDA-MB-231 cells and promoted MCF-7 and MDA-MB-231 cell apoptosis, and the cells were blocked in G1 phase. In comparison with the NC and blank groups, in the TLR4 siRNA-1, TLR4 siRNA-2, and TLR4 siRNA-3 groups, siRNA-TLR4 significantly increased expression of p53 and caspase-3 in MCF-7 and MDA-MB-231 cells, while it decreased the expressions of CDK4, cyclinD1, Livin, Bal-2, and c-FLIP. The study demonstrates that TLR4 gene silencing inhibits proliferation and induces apoptosis of MCF-7 and MDA-MB-231 cells. 

Retracted: Yes

PubPeer comments:
 FIGURE 4(a): mRNA expression of p5d3 and caspase-3 are increased, while CDK4, cyclin D1, Livin, Bcl-2, c-FLIP decreased in MCF-7 and MDA-MB-231 cells in the TLR4 siRNA-1 and TLR4 siRNA-2 groups. (a) Agarose gel electrophoresis of each gene by PCR amplification . I have concerns. ![file] , 
 FIGURE 5(a, c):  Protein expression of p53 and caspase-3 are increased, while CDK4, cyclin D1, Livin, Bcl-2, c-FLIP are decreased in MCF-7, and MDA-MB-231 cells in the TLR4 siRNA-1 and TLR4 siRNA-2 groups. (a) protein bands of MCF-7 related cells detected by Western blot analysis ... (c) protein bands of MDA-MB-231 related cells detected by Western blot analysis . ![file] Compare the loading band of 5(a) with Figure 7C from [Lu et al (2018)] and Fig. 3C from [Jiang et al (2018)] : ![] The loading band of 5(c) has similarities to that of 5C in [Wu et al (2018)] , though resolution limitations make it hard to be sure. We can say, though, that the protein blobs come from the same restricted gamut. ![file] UPDATE: Another sighting. Fig 4A of [Wang et al (2018)] ,  protein expression levels of EGFR, p-EGFR, Akt and p-Akt as determined using western blotting . ![] , 
 #2 Fig 5(a) again, at right, compared with Fig. 3A from [Huang et al (2017)] at left.  Western blotting images for NLRP3 and caspase-1. ![file] The highlighted protein blob is distinctive. The papers are otherwise unrelated. , 
 Retracted [20 July 2021] . ---------------------------------------------- The above article, published online on 22 June 2018 in Wiley Online Library has been retracted by agreement between the journal s Editor in Chief, Prof. Dr. Gregg Fields, and Wiley Periodicals LLC. The retraction has been agreed following an investigation based on allegations raised by a third party. A detailed investigation revealed that several image elements of the experimental data were published elsewhere in a different scientific context. Thus, the editors consider the conclusions of this article to be invalid. ,