Triterpenoid Saponin W3 from Anemone flaccida Suppresses Osteoclast Differentiation through Inhibiting Activation of MAPKs and NF-κB Pathways
Authors: Xiangying Kong
Hongye Wan
Wenbin Wu
Xiaomin Li
Michun Zhong
Na Lin
Shiwei Jia
Xiaohui Su
Yue Yang
Journal: International journal of biological sciences (2015)
Discipline: BIOLOGY & BIOCHEMISTRY
Keywords: Anemone flaccida; Bone resorption; Osteoclasts; RANKL.
Affiliations: 1. Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, 100700 Beijing, China., 2. Guangzhou Kanghe Pharmaceutical Limited Company, 511440 Guangzhou, China.

Abstract:  Excessive bone resorption by osteoclasts within inflamed joints is the most specific hallmark of rheumatoid arthritis. A. flaccida has long been used for the treatment of arthritis in folk medicine of China; however, the active ingredients responsible for the anti-arthritis effects of A. flaccida are still elusive. In this study, W3, a saponin isolated from the extract of A. flaccida was identified as the major active ingredient by using an osteoclast formation model induced by receptor activator of nuclear factor kappa-B ligand (RANKL). W3 dose-dependently suppressed the actin ring formation and lacunar resorption. Mechanistic investigation revealed that W3 inhibited the RANKL-induced TRAF6 expression, decreased phosphorylation of mitogen-activated protein kinases (MAPKs) and IκB-α, and suppressed NF-κB p65 DNA binding activity. Furthermore, W3 almost abrogated the expression of c-Fos and nuclear factor of activated T cells (NFATc1). Therefore, our results suggest that W3 is a potential agent for treating lytic bone diseases although further evaluation in vivo and in clinical trials is needed. 

Retracted: No

PubPeer comments:
 Figures 2 and 3: * Green boxes: the control panels appear to show an overlap; however this is ok since they appear to be the same experiments * Red boxes: The TS 0.5 panel appears to show an overlap with the W3 0.5 panel. These appear to be different experiments. ![file] , 
 Figure 7A and 8A. * Blue boxes: Four lanes in the GAPDH panels of Figure 7A and 8A appear to show an overlap. The figures are showing the same experiment (W3 concentration series) but the lanes are shifted so the individual lanes no longer represent the same conditions. ![file] , 
 The same GAPDH blot appears to be visible in another paper by the same research group that was submitted about a year earlier, i.e. Kong X et al. J. Trans. Med. 2015, 13:91, DOI: 10.1186 s12967-015-0440-1. See: https: pubpeer.com publications 6D505D1A0ECB922DB1B3FE76DEB670 In the older paper, the blot represents a TS concentration series, while in this newer paper, the blot represents a W3 concentration series. Here are Figures 7A and 8A from this paper (on the right) compared to Figures 5A and 6A from the older Kong et al. paper (left). ![file] , 
 Figure 2A from this paper appears to look very similar to Figure 1A of the older paper, Kong X et al. J. Trans. Med. 2015, 13:91, DOI: 10.1186 s12967-015-0440-1. The experiments are the same, but the TS 2.5 image is rotated 180 degrees. ![file] , 
 #3 Fig 6B of this paper also seems to share a GAPDH loading control with 4B from [Kong et al (2015)] . The two figures represent increasing doses of W3 and of TS respectively. ![file] , 
 One more for the sake of completeness. Fig 8A:  W3 suppresses NF-u03baB activation in RANKL-induced RAW 264.7 cells. (A) Confluent RAW 264.7 cells were pre-treated with various concentrations of **W3** (0.1-2.5 u00b5g mL) or vehicle for 2 h followed by RANKL (50 ng mL) for 0.5 h . Below, Fig 6A from [Kong et al (2015)] :  TS suppresses RANKL-induced NF-u03baB activation in RAW 264.7 cells. Confluent RAW 264.7 cells were pre-treated with various concentrations of **TS** (0.1-2.5 u03bcg mL) for 2 h followed by RANKL (50 ng mL) for 30 min. Different sets of conditions but same GAPDH loading control. ![file] , 
 Dear Dr.Bik, I just got the message from the administration department of Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, and informed me that you raised some inquiries on my research papers via the Pubpeer. Many thanks for your kind concern and sorry for later response due to not checking the Pubpeer website recently. Regarding to the inquiries and concerns in the papers mentioned, I will take the highest priority and immediately start investigations, including carefully checking the manuscripts, raw data and experimental records. And also I will contact with the editorial offices of the relevant journals as soon as possible, if there is any risk to the highest degree of accuracy of our scientific findings. After a quick check on some inquiries and concerns (the careful check is still ongoing), I am confident in the validity of our scientific findings published in those articles. Nevertheless, there is no excuse for any mistakes in the papers as the corresponding author, a lab leader and supervisor. I will be more than happy to make the responses accordingly with careful double-checking via the Pubpeer. Again, many thanks for your concerns which definitely alert me and help me in my future research. Also I must sincerely apologize for any mistakes, including oversight and inconvenience caused, to my students, colleagues and collaborators. I will take this as an invaluable learning opportunity, and keep the accuracy and integrity of science as my first priority in my research. Best regards! Yours, sincerely, Na LIN ,