Methionine down-regulates TLR4/MyD88/NF-Œ∫B signalling in osteoclast precursors to reduce bone loss during osteoporosis
Authors: A Surolia
K Manglani
V Vijayan
S Gupta
M Khandelwal
Journal: British Journal of Pharmacology (2014)
Discipline: PHARMACOLOGY & TOXICOLOGY
Keywords: MyD88; RANKL; alendronate; methionine; osteoclast
Affiliations: Molecular Sciences Laboratory, National Institute of Immunology, New Delhi, India., Molecular Biophysics Unit; Indian Institute of Science; Bangalore Karnataka India, Molecular Sciences Laboratory; National Institute of Immunology; New Delhi India

Abstract:  Background and purpose: Studies have demonstrated that a moderate intake of amino acids is associated with development of bone health. Methionine, a sulphur-containing essential amino acid, has been largely implicated for improving cartilage formation, however its physiological significance on bone integrity and functionality have not been elucidated. We investigated whether methionine can prevent osteoporotic bone loss. Experimental approach: The anti-resorptive effect of methionine, (250 mg kg(-1) body wt administered in drinking water for 10 weeks), was evaluated in ovariectomized (OVX) rats by monitoring changes in bone turnover, formation of osteoclasts from blood-derived mononuclear cells and changes in the synthesis of pro-osteoclastogenic cytokines. Key results: Methionine improved bone density and significantly decreased the degree of osteoclast development from blood mononuclear cells in OVX rats, as indicated by decreased production of osteoclast markers tartarate resistant acid phosphatase b (TRAP5b) and MIP-1Œ±. siRNA-mediated knockdown of myeloid differentiation primary response 88 [MyD88], a signalling molecule in the toll-like receptor (TLR) signalling cascade, abolished the synthesis of both TRAP5b and MIP-1Œ± in developing osteoclasts. Methionine supplementation disrupted osteoclast development by inhibiting TLR-4/MyD88/NF-Œ∫B pathway. Conclusions and implications: TLR-4/MyD88/NF-Œ∫B signalling pathway is integral for osteoclast development and this is down-regulated in osteoporotic system on methionine treatment. Methionine treatment could be beneficial for the treatment of postmenopausal osteoporosis. 

Retracted: No

PubPeer comments:
 Could the authors clarify the image duplication, where the treatment conditions are different???  , 
 #1 Thanks for pointing it out. we have noted it. we will look into the raw data and respond as soon as possible. , 
 The fold times in the two publications are different in the same conditions (same authors). Could the authors please clarify?  , 
 #3 Normal total homocysteine blood levels range from 5-12 u03bcmol l, and hyperhomocysteinemia is classified into three categories: mild (12u201330 u03bcmol l), moderate (30u2013100 u03bcmol l), and severe ( 100 u03bcmol l). In the normal range fold difference is 2.4 Hyperhomocysteinemia (mild), range of fold difference is 2.5 Hyperhomocysteinemia (moderate), range of fold difference is 3.3 Met 500 in two independent studies caused hyperhomocysteinemia in OVX rats and the fold difference between the two is ~3.6. Is that a cause for a major concern? , 
 Correction for the paper has been published, available at https: bpspubs.onlinelibrary.wiley.com doi 10.1111 bph.15568 , 
 Concerns on the images provided in the recently published erratum . - The immunoblots for IkB and its actin control have some background issues (blue and green arrows). - For the lanes for NFkB p65 and its Histone H1, the backgrounds are clear white. It seemed to be not concurrent with the other lanes. Could the authors provide their uncropped original immunoblot image for more clarifications?  , 
 Concerns on the images provided in the recently published erratum . The bands for the p65 NFkB for different conditions look the same (red box). There is no band for the Sham, suspecting that the bands were diminished because of the adjustments in the background (Yellow Arrow). Could the authors clarify with their uncropped original images?  , 
 Concerns on the images provided in the recently published erratum . In figure 14, the bands for IkB in the older and corrected version were shown. In the corrected version, the authors stated about the changes in the immunoblot images of IkB expression, which did not affect the conclusion. However, the results are not conclusive with the corrected images provided. In detail, the older version shows a lesser expression of IkB in the OVX and ALD+Met, though in the corrected version which is similar to that of other groups. The texts in the older and corrected version of the publications are also provided with the image file to show the statements of the authors. In the older version, there is a black line in the 2nd and 5th bands (OVX and ALD+Met) - Represented with BLUE ARROWS. These two bands were cropped from some other blots to show the reduction in the IkB expression and unfortunately, the image outline has appeared (in the red box). Suspected image manipulation in the older version and the results provided by the authors in the corrected version affect the conclusion of the publication. The serious question is: Since the authors represented the same immunoblot for IkB with changes in band 2 and 5 and the actin control were also provided. - How did bands 1, 3, and 4 of IkB remain the same in the corrected version? - How did only bands 2 and 5 changed in the corrected version? - How the background of the bands in the corrected version is different from the older version (clear white)? The authors are kindly requested to provide their raw data (original uncropped image) for the clarification.  , 
 Concerns: In the text, the authors explained phosphorylated ERK1 2 and Akt. However, the authors did not provide images for these in their erratum. If the authors think the same p-ERK and p-Akt in the older version is to be applicable, then the authors failed to show the respective loading controls. Since there is no bands appeared in the case of both p-ERK1 2 and p-Akt, loading control for the same is necessary to make sure that the samples were loaded.  ,