Antiproteolytic effects of plasma from hibernating bears: A new approach for muscle wasting therapy?

doi:10.1016/j.clnu.2007.07.003

Clinical Nutrition

Volume 26, Issue 5, October 2007, Pages 658-661

https://doi.org/10.1016/j.clnu.2007.07.003 Get rights and content

Summary

Background & aims

In rodents and humans, inactivity or starvation leads to atrophy of skeletal muscle including a decrease in the number and size of muscle cells and in the myofibrillar protein content. It has previously been described that in overwintering bears the inactivity does not provoke any loss of skeletal muscle cell number or size. Taking all these into account, the aim of this study is to test if hibernating bear plasma has any antiproteolytic effect on incubated rat skeletal muscle.

Methods

Rat skeletal extensor digitorum longus (EDL) muscles were incubated in the presence of hibernating, non-hibernating and control bear plasma. After that, proteolytic rate was evaluated as levels of tyrosine released to the medium and muscle mRNA content for different proteolytic systems were measured by Northern blot.

Results

Rat skeletal EDL muscles incubation in the presence of hibernating bear plasma resulted in a 40% decrease of the net proteolytic rate. This inhibition of proteolysis was accompanied by decreases in the expression of both lysosomal (cathepsin B) and ubiquitin-dependent (ubiquitin) proteolytic systems.

Conclusions

The results suggest that during hibernation the bear is able to produce a powerful proteolytic inhibitor which is released to the circulation and blocks muscle wasting associated with immobilization.

Introduction

The precise mechanism by which intracellular proteins are degraded is largely unknown, although it is accepted that proteolysis may occur inside and outside the lysosomes. The ATP-ubiquitin-dependent proteolytic system has been shown to be involved in the alterations of protein metabolism related to several pathophysiological conditions such as cancer, chronic infection and chronic heart failure.1, 2 During the pathological conditions commented on, muscle wasting leads to cachexia, a syndrome characterized by weight loss and profound metabolic abnormalities. Unfortunately therapeutic approaches to stop muscle wasting have not been very satisfactory, partly because of the toxicity of inhibitors of the ubiquitin-dependent proteolytic system. In addition, this proteolytic system is involved in many biological processes, such as cell proliferation and intracellular signaling.³ Therefore, a non-muscle specific inhibition would result in dramatic toxic effects for the organism. Interestingly, muscle wasting is also observed in non-pathological conditions, such as immobilization or microgravity, where an important muscle atrophy develops. In these conditions, an activation of the ubiquitin-dependent proteolytic system has also been described.⁴

Hibernating animals, such as bears, stay inside their winter dens for periods of at least 4 months. During this “sleeping” period they do not eat or have any physiological activity. In spite of this starvation and immobilization period, they do not develop muscle atrophy (their weight loss only includes adipose tissue mobilization), unlike humans, who would be weakened by 90% strength loss over the same period.⁵

In view of this observation, we hypothesized that a type of inhibitory activity on skeletal muscle proteolysis must be induced during hibernation. Bearing this in mind, we decided to test if hibernating bear plasma has any antiproteolytic effect on incubated rat skeletal muscle.

Section snippets

Bear plasma

Bear plasma was obtained from Ursus arctos. Sample collection was carried out in their natural habitat (Pirinees, Artíes, Vall d’Aran) using anesthesia.

Animals

Male Wistar rats (Interfauna, Barcelona, Spain) weighing about 60 g were used. The animals were maintained on a regular light–dark cycle (light on from 08:00 a.m. to 08:00 p.m.) at an ambient temperature of 22±2 °C and had free access to food and water. The diet (B.K. Universal, Sant Vicents dels Horts, Barcelona, Spain) consisted of 45.5–48.5%

Results

The results presented in Table 1 clearly show that the presence of hibernating plasma resulted in a 40% decrease of the proteolytic rate, as compared with the muscles incubated in the presence of non-hibernating bear plasma. It is therefore clear from these results that a material present in the plasma during hibernation is able to block total proteolysis. Bearing all this in mind, we decided to assess the expression of genes involved in protein breakdown. As observed in Table 2, both cathepsin

Discussion

Hibernating animals are subject to both extreme starvation and immobilization. In non-hibernating mammals, such as humans, these two conditions led to a massive and dangerous muscle wasting associated with cachexia. However, during hibernation, the animals do not experience muscle wasting,⁵ therefore indicating that neither starvation nor immobilization can trigger the proteolysis induction that takes place in skeletal muscle during wasting. The decrease of net proteolytic rate in incubated rat

Acknowledgments

This work was supported by grants from the Instituto de Salud Carlos III (03/0100) of the Ministerio de Sanidad y Consumo, from the Dirección General de Investigación Científica y Técnica (SAF4744-2005) of the Ministerio de Educación y Ciencia, from the Generalitat de Catalunya (SGR/00108), from Ministerio de Ciencia y Tecnologia (VIN03/021), and by grant SAF-2003-04223 from the Ministerio de Ciencia y Tecnología (MCyT). Special thanks to Jesús Fernández, (Zoologic de Barcelona) and Santiago

References (12)

J.M. Argilés et al.
The ubiquitin-dependent proteolytic pathway in skeletal muscle: its role in pathological states
Trends Pharmacol Sci
(1996)
D. Taillandier et al.
Regulation of proteolysis during reloading of the unweighted soleus muscle
Cell Biol
(2003)
P. Chomczynski et al.
Single-step method of RNA isolation by acid guanidinium thiocyanate phenol chloroform extraction
Anal Biochem
(1987)
D. Attaix et al.
The ubiquitin-proteasome system and skeletal muscle wasting
Essays Biochem
(2005)
A. Ciechanover et al.
The ubiquitin system: from basic mechanisms to the patient bed
IUBMB Life
(2004)
H.J. Harlow et al.
Muscle strength in overwintering bears
Nature
(2001)

There are more references available in the full text version of this article.

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The current findings identifying eight serum proteins that are uniquely expressed provide a starting point in the elucidation of their unique actions. Together with this information, we can begin to explore the applications of proteins identified in bears to human pathologies, including muscle wasting and osteoporosis (Donahue et al., 2006; Fuster et al., 2007; Harlow et al., 2001; Hershey et al., 2008; Lin et al., 2012; Lohuis et al., 2007), kidney disease (Stenvinkel et al., 2013), and metabolic syndrome (Berg von Linde et al., 2015; Martin, 2008; Rigano et al., 2017; Wu et al., 2013). It is possible that metabolites and/or additional serum proteins that were not detected in this study could play an important role in the regulation of hibernation phenotypes.
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Brown bears (Ursus arctos) hibernate for 5–7 months without eating, drinking, urinating, and defecating at a metabolic rate of only 25% of the summer activity rate. Nonetheless, they emerge healthy and alert in spring. We quantified the biochemical adaptations for hibernation by comparing the proteome, metabolome, and hematological features of blood from hibernating and active free-ranging subadult brown bears with a focus on conservation of health and energy. We found that total plasma protein concentration increased during hibernation, even though the concentrations of most individual plasma proteins decreased, as did the white blood cell types. Strikingly, antimicrobial defense proteins increased in concentration. Central functions in hibernation involving the coagulation response and protease inhibition, as well as lipid transport and metabolism, were upheld by increased levels of very few key or broad specificity proteins. The changes in coagulation factor levels matched the changes in activity measurements. A dramatic 45-fold increase in sex hormone-binding globulin levels during hibernation draws, for the first time, attention to its significant but unknown role in maintaining hibernation physiology. We propose that energy for the costly protein synthesis is reduced by three mechanisms as follows: (i) dehydration, which increases protein concentration without de novo synthesis; (ii) reduced protein degradation rates due to a 6 °C reduction in body temperature and decreased protease activity; and (iii) a marked redistribution of energy resources only increasing de novo synthesis of a few key proteins. The comprehensive global data identified novel biochemical strategies for bear adaptations to the extreme condition of hibernation and have implications for our understanding of physiology in general.
Hibernating bears (Ursidae): Metabolic magicians of definite interest for the nephrologist
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Two novel observations have helped to further elucidate this fascinating metabolic adaption in bears. Fuster et al.39 demonstrated that hibernating bears produce a powerful proteolytic inhibitor that blocks muscle wasting associated with immobilization. If this factor could be identified, it could serve as a treatment strategy for conditions in which protein energy wasting is a common feature.
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^☆: Conference presentation: The work was presented by Prof. Josep M. Argilés in the 11th world congress on advances in oncology, Hersonissos, Creta, 2006/10/12.

^a: These authors contributed equally to this work.

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ORIGINAL ARTICLEAntiproteolytic effects of plasma from hibernating bears: A new approach for muscle wasting therapy?☆

Summary

Background & aims

Methods

Results

Conclusions

Introduction

Section snippets

Bear plasma

Animals

Results

Discussion

Acknowledgments

Trends Pharmacol Sci

Cell Biol

Anal Biochem

The ubiquitin-proteasome system and skeletal muscle wasting

Essays Biochem

The ubiquitin system: from basic mechanisms to the patient bed

IUBMB Life

Muscle strength in overwintering bears

Nature

ORIGINAL ARTICLE
Antiproteolytic effects of plasma from hibernating bears: A new approach for muscle wasting therapy?☆