Association of lithocholic acid with skeletal muscle hypertrophy through TGR5-IGF-1 and skeletal muscle mass in cultured mouse myotubes, chronic liver disease rats and humans
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Curated by eLife
Evaluation Summary:
This paper, of interest to both basic scientists and clinicians, addresses the clinically important condition of reduced muscle mass in human chronic liver disease. It seeks causative mechanisms under these conditions. It uses in vivo and in vitro techniques to draw associations between bile acid concentrations and muscle disease. They implicate a specific bile acid for the first time.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Abstract
Hepatic sarcopenia is one of many complications associated with chronic liver disease (CLD) and has a high mortality rate; however, the liver-muscle axis is not fully understood. Therefore, few effective treatments exist for hepatic sarcopenia, the best of which being branched-chain amino acid (BCAA) supplementation to help increase muscle mass. Our aim was to investigate the molecular mechanism(s) of hepatic sarcopenia focused on bile acid (BA) composition.
Methods:
The correlation between serum BA levels and psoas muscle mass index (PMI) was examined in 73 CLD patients. Gastrocnemius muscle phenotype and serum BA levels were assessed in CLD rats treated with BCAA. Mouse skeletal muscle cells (C2C12) were incubated with lithocholic acid (LCA), G-protein-coupled receptor 5 (TGR5) agonist or TGR5 antagonist to assess skeletal muscle hypertrophy.
Results:
In human CLD, serum LCA levels were the sole factor positively correlated with PMI and were significantly decreased in both the low muscle mass group and the deceased group. Serum LCA levels were also shown to predict patient survival. Gastrocnemius muscle weight significantly increased in CLD rats treated with BCAA via suppression of protein degradation pathways, coupled with a significant increase in serum LCA levels. LCA treated C2C12 hypertrophy occurred in a concentration-dependent manner linked with TGR5-Akt pathways based upon inhibition results via a TGR5 antagonist.
Conclusions:
Our results indicate LCA-mediated skeletal muscle hypertrophy via activation of TGR5-IGF1-Akt signaling pathways. In addition, serum LCA levels were associated with skeletal muscle mass in cirrhotic rats, as well as CLD patients, and predicted overall patient survival.
Funding:
This research was supported by JSPS KAKENHI Grant Number 22K08011 and 21H02892, and AMED under Grant Number JP21fk0210090 and JP22fk0210115. Maintaining cirrhotic rats were partially supported by Otsuka Pharmaceutical Company.
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Evaluation Summary:
This paper, of interest to both basic scientists and clinicians, addresses the clinically important condition of reduced muscle mass in human chronic liver disease. It seeks causative mechanisms under these conditions. It uses in vivo and in vitro techniques to draw associations between bile acid concentrations and muscle disease. They implicate a specific bile acid for the first time.
(This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)
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Reviewer #1 (Public Review):
My impression is of a careful and thorough study, that potentially could provide a paradigm for future studies in this direction. A potential causative pathway for hepatic-related sarcopenia is identified. Parallel studies are made in both experimental and human clinical systems.
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Reviewer #2 (Public Review):
Tamai et al aimed to investigate the association between bile acid concentration (lithocholic acid) in both humans, and an in vivo, rat model of chronic liver disease induced by CCl4 treatment. They show that a decline in bile acid concentrations in CLD rats treated with BCAA, and an increase in concentrations in CLD rats, without BCAA treatment, compared to controls. This decline in LCA concentration was negatively associated with gastrocnemius muscle mass (i.e., muscle mass increased). However, lithocholic acid was associated with an increase in gastrocnemius muscle mass in the CLD rats + BCAA group. The authors also highlight a significant, positive correlation between psoas muscle index in CLD patients and lithocholic acid concentrations. Of interest, the authors treated C2C12 skeletal muscle cells, an …
Reviewer #2 (Public Review):
Tamai et al aimed to investigate the association between bile acid concentration (lithocholic acid) in both humans, and an in vivo, rat model of chronic liver disease induced by CCl4 treatment. They show that a decline in bile acid concentrations in CLD rats treated with BCAA, and an increase in concentrations in CLD rats, without BCAA treatment, compared to controls. This decline in LCA concentration was negatively associated with gastrocnemius muscle mass (i.e., muscle mass increased). However, lithocholic acid was associated with an increase in gastrocnemius muscle mass in the CLD rats + BCAA group. The authors also highlight a significant, positive correlation between psoas muscle index in CLD patients and lithocholic acid concentrations. Of interest, the authors treated C2C12 skeletal muscle cells, an immortalised rat cell line with lithocholic acid in order to investigate a mechanistic link between the associations observed within in vivo data. The authors suggest that the TGF5-IGF-1-Akt3 pathway may be linked to muscle hypertrophy. However, these suggestions are not fully supported by the data within the manuscript and need to be extended further to downstream targets related to mTOR, and ultimately muscle protein synthesis.
The multi-model approach is a clear strength of this manuscript. Both animal and human work were used to investigate associations in LCA concentration and muscle mass. The authors also use an in vitro model to investigate potential mechanistic links which may explain the associations identified within in vivo models. However, the manuscript does have a number of flaws. Firstly, the authors do not set out specific and detailed aims in relation to their in vitro work. Secondly, the rationale for the inclusion of BCAA treatment in rats does not come across in the manuscript. Additionally, the flow throughout the manuscript, in particular the results section, needs improvement.
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Reviewer #3 (Public Review):
In this study, Dr Tamai et al. investigated the association between bile acid level and skeletal muscle mass using a rat model and patients with HCCs. The authors found that LCA level was closely associated with skeletal muscle mass in both CLD rats and human patients with HCCs.
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