MFN2 overexpression in skeletal muscles of young and old mice causes a mild hypertrophy without altering mitochondrial respiration and H2O2 emission.

AIM: Sarcopenia, the aging-related loss of muscle mass and function, is a debilitating process negatively impacting the quality of life of affected individuals. Although the mechanisms underlying sarcopenia are incompletely understood, impairments in mitochondrial dynamics, including mitochondrial fusion, have been proposed as a contributing factor. However, the potential of upregulating mitochondrial fusion proteins to alleviate the effects of aging on skeletal muscles remains unexplored. We therefore hypothesized that overexpressing Mitofusin 2 (MFN2) in skeletal muscle in vivo would mitigate the effects of aging on muscle mass and improve mitochondrial function. METHODS: MFN2 was overexpressed in young (7 mo) and old (24 mo) male mice for 4 months through intramuscular injections of an adeno-associated viruses. The impacts of MFN2 overexpression on muscle mass and fiber size (histology), mitochondrial respiration, and H2O2 emission (Oroboros fluororespirometry), and various signaling pathways (qPCR and western blotting) were investigated. RESULTS: MFN2 overexpression increased muscle mass and fiber size in both young and old mice. No sign of fibrosis, necrosis, or inflammation was found upon MFN2 overexpression, indicating that the hypertrophy triggered by MFN2 overexpression was not pathological. MFN2 overexpression even reduced the proportion of fibers with central nuclei in old muscles. Importantly, MFN2 overexpression had no impact on muscle mitochondrial respiration and H2O2 emission in both young and old mice. MFN2 overexpression attenuated the increase in markers of impaired autophagy in old muscles. CONCLUSION: MFN2 overexpression may be a viable approach to mitigate aging-related muscle atrophy and may have applications for other muscle disorders.

Molecular mechanisms underlying physical exercise-induced brain BDNF overproduction.

Accumulating evidence supports that physical exercise (EX) is the most effective non-pharmacological strategy to improve brain health. EX prevents cognitive decline associated with age and decreases the risk of developing neurodegenerative diseases and psychiatric disorders. These positive effects of EX can be attributed to an increase in neurogenesis and neuroplastic processes, leading to learning and memory improvement. At the molecular level, there is a solid consensus to involve the neurotrophin brain-derived neurotrophic factor (BDNF) as the crucial molecule for positive EX effects on the brain. However, even though EX incontestably leads to beneficial processes through BDNF expression, cellular sources and molecular mechanisms underlying EX-induced cerebral BDNF overproduction are still being elucidated. In this context, the present review offers a summary of the different molecular mechanisms involved in brain's response to EX, with a specific focus on BDNF. It aims to provide a cohesive overview of the three main mechanisms leading to EX-induced brain BDNF production: the neuronal-dependent overexpression, the elevation of cerebral blood flow (hemodynamic hypothesis), and the exerkine signaling emanating from peripheral tissues (humoral response). By shedding light on these intricate pathways, this review seeks to contribute to the ongoing elucidation of the relationship between EX and cerebral BDNF expression, offering valuable insights into the potential therapeutic implications for brain health enhancement.

Autophagy ablation in skeletal muscles worsens sepsis-induced muscle wasting, impairs whole-body metabolism, and decreases survival.

Septic patients frequently develop skeletal muscle wasting and weakness, resulting in severe clinical consequences and adverse outcomes. Sepsis triggers sustained induction of autophagy, a key cellular degradative pathway, in skeletal muscles. However, the impact of enhanced autophagy on sepsis-induced muscle dysfunction remains unclear. Using an inducible and muscle-specific Atg7 knockout mouse model (Atg7iSkM-KO), we investigated the functional importance of skeletal muscle autophagy in sepsis using the cecal ligation and puncture model. Atg7iSkM-KO mice exhibited a more severe phenotype in response to sepsis, marked by severe muscle wasting, hypoglycemia, higher ketone levels, and a decreased in survival as compared to mice with intact Atg7. Sepsis and Atg7 deletion resulted in the accumulation of mitochondrial dysfunction, although sepsis did not further worsen mitochondrial dysfunction in Atg7iSkM-KO mice. Overall, our study demonstrates that autophagy inactivation in skeletal muscles triggers significant worsening of sepsis-induced muscle and metabolic dysfunctions and negatively impacts survival.

MYTHO is a novel regulator of skeletal muscle autophagy and integrity.

Autophagy is a critical process in the regulation of muscle mass, function and integrity. The molecular mechanisms regulating autophagy are complex and still partly understood. Here, we identify and characterize a novel FoxO-dependent gene, d230025d16rik which we named Mytho (Macroautophagy and YouTH Optimizer), as a regulator of autophagy and skeletal muscle integrity in vivo. Mytho is significantly up-regulated in various mouse models of skeletal muscle atrophy. Short term depletion of MYTHO in mice attenuates muscle atrophy caused by fasting, denervation, cancer cachexia and sepsis. While MYTHO overexpression is sufficient to trigger muscle atrophy, MYTHO knockdown results in a progressive increase in muscle mass associated with a sustained activation of the mTORC1 signaling pathway. Prolonged MYTHO knockdown is associated with severe myopathic features, including impaired autophagy, muscle weakness, myofiber degeneration, and extensive ultrastructural defects, such as accumulation of autophagic vacuoles and tubular aggregates. Inhibition of the mTORC1 signaling pathway in mice using rapamycin treatment attenuates the myopathic phenotype triggered by MYTHO knockdown. Skeletal muscles from human patients diagnosed with myotonic dystrophy type 1 (DM1) display reduced Mytho expression, activation of the mTORC1 signaling pathway and impaired autophagy, raising the possibility that low Mytho expression might contribute to the progression of the disease. We conclude that MYTHO is a key regulator of muscle autophagy and integrity.

Endothelial cells are an important source of BDNF in rat skeletal muscle.

BDNF (brain-derived neurotrophic factor) is present in skeletal muscle, controlling muscular metabolism, strength and regeneration processes. However, there is no consensus on BDNF cellular source. Furthermore, while endothelial tissue expresses BDNF in large amount, whether endothelial cells inside muscle expressed BDNF has never been explored. The aim of the present study was to provide a comprehensive analysis of BDNF localization in rat skeletal muscle. Cellular localization of BDNF and activated Tropomyosin-related kinase B (TrkB) receptors was studied by immunohistochemical analysis on soleus (SOL) and gastrocnemius (GAS). BDNF and activated TrkB levels were also measured in muscle homogenates using Western blot analysis and/or Elisa tests. The results revealed BDNF immunostaining in all cell types examined with a prominent staining in endothelial cells and a stronger staining in type II than type I muscular fibers. Endothelial cells but not other cells displayed easily detectable activated TrkB receptor expression. Levels of BDNF and activated TrkB receptors were higher in SOL than GAS. In conclusion, endothelial cells are an important and still unexplored source of BDNF present in skeletal muscle. Endothelial BDNF expression likely explains why oxidative muscle exhibits higher BDNF levels than glycolytic muscle despite higher the BDNF expression by type II fibers.

Tofacitinib improved peripheral endothelial dysfunction and brain-derived neurotrophic factor levels in the rat adjuvant-induced arthritis model.

This study aimed to explore the effect of Tofacitinib on endothelial dysfunction and cerebral levels of brain-derived neurotrophic factor (BDNF) in the adjuvant-induced arthritis (AIA) rat model. Tofacitinib (10 mg/kg twice a day) or vehicle was administered from the first signs of inflammation. Arthritis scores were daily monitored while other parameters including endothelial function assessed from aortic rings, radiographic scores, blood pressure, heart rate, circulating levels of triglycerides, cholesterol, and interleukin (IL)-1ß, tumor necrosis factor-α (TNF-α), IL-17A, and cerebral BDNF levels were determined after 3 weeks of treatment. A group of non-AIA rats served as controls. In AIA rats, as compared with vehicle, Tofacitinib significantly reduced arthritis and radiographic scores, decreased total cholesterol and low-density lipoprotein cholesterol (LDL-C), but changed neither blood pressure nor heart rate and proinflammatory cytokines levels. It also fully restored acetylcholine (Ach)-induced relaxation (p < 0.05) through increased nitric oxide (NO) synthase activity, reduced BH4 deficiency and O2 -° production, decreased cyclo-oxygenase-2 (COX-2)/arginase activities, and enhanced endothelium-derived hyperpolarizing factor (EDHF) production. These effects translated into a decrease in atherogenic index and an elevation of BDNF levels in the prefrontal cortex (p < 0.05) and hippocampus (p < 0.001). The present study identified Tofacitinib as an efficient therapeutic option to reduce cardiovascular risk and improve BDNF-dependent cognition in arthritis.

Potential mechanisms involved in regulating muscle protein turnover after acute exercise: A brief review.

It is well established that resistance training increases muscle mass. Indeed, there is evidence to suggest that a single session of resistance training is associated with an increase in muscle protein synthesis in young adults. However, the fundamental mechanisms that are involved in regulating muscle protein turnover rates after an acute bout of physical exercise are unclear. Therefore, this review will briefly focus on summarizing the potential mechanisms behind the growth of skeletal muscle after physical exercise. We also present mechanistic differences that may exist between young and older individuals during muscle protein synthesis and breakdown after physical exercise. Pathways leading to the activation of AKT/mTOR signals after resistance exercise and the activation of AMPK signaling pathway following a HIIT (High intensity interval training) are discussed.

Regulation of muscle and mitochondrial health by the mitochondrial fission protein Drp1 in aged mice.

KEY POINTS: The maintenance of mitochondrial integrity is critical for skeletal muscle health. Mitochondrial dynamics play key roles in mitochondrial quality control; however, the exact role that mitochondrial fission plays in the muscle ageing process remains unclear. Here we report that both Drp1 knockdown and Drp1 overexpression late in life in mice is detrimental to skeletal muscle function and mitochondrial health. Drp1 knockdown in 18-month-old mice resulted in severe skeletal muscle atrophy, mitochondrial dysfunction, muscle degeneration/regeneration, oxidative stress and impaired autophagy. Overexpressing Drp1 in 18-month-old mice resulted in mild skeletal muscle atrophy and decreased mitochondrial quality. Our data indicate that silencing or overexpressing Drp1 late in life is detrimental to skeletal muscle integrity. We conclude that modulating Drp1 expression is unlikely to be a viable approach to counter the muscle ageing process. ABSTRACT: Sarcopenia, the ageing-related loss of skeletal muscle mass and function, is a debilitating process negatively impacting the quality of life of afflicted individuals. Although the mechanisms underlying sarcopenia are still only partly understood, impairments in mitochondrial dynamics, and specifically mitochondrial fission, have been proposed as an underlying mechanism. Importantly, conflicting data exist in the field and both excessive and insufficient mitochondrial fission were proposed to contribute to sarcopenia. In Drosophila melanogaster, enhancing mitochondrial fission in midlife through overexpression of dynamin-1-like protein (Drp1) extended lifespan and attenuated several key hallmarks of muscle ageing. Whether a similar outcome of Drp1 overexpression is observed in mammalian muscles remains unknown. In this study, we investigated the impact of knocking down and overexpressing Drp1 protein for 4 months in skeletal muscles of late middle-aged (18 months) mice using intra-muscular injections of adeno-associated viruses expressing shRNA targeting Drp1 or full Drp1 cDNA. We report that knocking down Drp1 expression late in life triggers severe muscle atrophy, mitochondrial dysfunctions, degeneration/regeneration, oxidative stress and impaired autophagy. Drp1 overexpression late in life triggered mild muscle atrophy and decreased mitochondrial quality. Taken altogether, our results indicate that both overexpression and silencing of Drp1 in late middle-aged mice negatively impact skeletal muscle mass and mitochondrial health. These data suggest that Drp1 content must remain within a narrow physiological range to preserve muscle and mitochondrial integrity during ageing. Altering Drp1 expression is therefore unlikely to be a viable target to counter sarcopenia.

Brain-derived neurotrophic factor is a full endothelium-derived factor in rats.

Most of what is known on vascular brain-derived neurotrophic factor (BDNF) derived from experiments on cultured endothelial cells. Therefore, the present study compared BDNF levels/localization in artery (aorta) vs vein (vena cava) from a same territory in rats either sedentary (SED) or exposed to treadmill exercise (EX) as a mean to stimulate endogenous endothelial nitric oxide (NO) production. In SED rats, for both artery and vein, BDNF was strongly expressed by endothelial cells, while only a faint and scattered expression was observed throughout the media. Endothelial and muscular BDNF staining as vascular BDNF protein levels were however higher in artery than in vein, while BDNF mRNA levels did not differ between vessels. Irrespective of the vessels, EX resulted in an increase (+50%) in BDNF protein levels with no change in BDNF mRNA levels, a selective endothelial BDNF overexpression (x4) and an increase in vascular levels of tropomyosin related kinase B receptors (TrkB) phosphorylated at tyrosine 816 (p-TrkBTyr816). Endothelial expressions of BDNF and p-TrkBTyr816 were positively associated when SED and EX rats were simultaneously examined. The results incite to consider endothelial BDNF as a full and NO-dependent endothelium-derived factor that exerts autocrine effects.

The effect of exercise on memory and BDNF signaling is dependent on intensity.

The aims of the present study were to investigate in brain of adult rats (1) whether exercise-induced activation of brain-derived neurotrophic factor (BDNF)/tropomyosin-related kinase B (TrkB) pathway is dependent on exercise intensity modality and (2) whether exercise-induced improvement of memory is proportional to this pathway activation. Wistar rats were subjected to low (12 m/min) or high (18 m/min) exercise intensity on horizontal treadmill (30 min/day, 7 consecutive days) that corresponds to ~ 40 and 70% of maximal aerobic speed, respectively. Animals treated with scopolamine to induce memory impairment were subjected to novel object recognition test to assess potential improvement in cognitive function. Expressions of BDNF, phosphorylated TrkB receptors, synaptophysin (a marker of synaptogenesis), c-fos (a neuronal activity marker) and phosphorylated endothelial nitric oxide synthase (a cerebral blood flow marker) were measured in prefrontal cortex and hippocampus of different groups of rats. In terms of cognition, our data reported that only the most intense exercise improves memory performance. Our data also revealed that BDNF pathway is dependent on intensity modality of exercise with a gradual effect in hippocampus whereas only the highest intensity leads to this pathway activation in prefrontal cortex. Our study revealed that memory improvement through BDNF pathway activation is dependent on exercise intensity. While reporting that our protocol is sufficient to improve cognition in animals with impaired memory, our data suggest that prefrontal cortex is possibly a more suitable structure than hippocampus when neuroplastic markers are used to mirror potential improvement in memory performance.

Brain-derived Neurotrophic Factor Pathway after Downhill and Uphill Training in Rats.

INTRODUCTION: The elevation of brain-derived neurotrophic factor (BDNF) levels in the brain and the subsequent phosphorylation of its cognate tropomyosin-related kinase B (TrkB) receptors at tyrosine 816 (pTrkB) are largely involved in the positive effect of aerobic exercise on brain functioning. Although BDNF levels were reported to increase in proportion with exercise intensity, the effect of the type of contraction is unknown. Therefore, the cerebral BDNF/TrkB pathway was investigated after uphill and downhill treadmill activities at equivalent intensity to preferentially induce eccentric and concentric contractions, respectively. METHODS: A treadmill activity (30 min·d for seven consecutive days) either in a horizontal position at two different speeds to modulate intensity (experiment 1) or at three different inclinations (null, -10%, and +5%) but at equivalent intensity to modulate the type of contraction (experiment 2) was induced in rats. Both experiments included sedentary rats. Levels of BDNF, pTrkB, synaptophysin (marker of synaptogenesis), endothelial nitric oxide synthase phosphorylated at serine 1177 (peNOS), and c-fos levels (indicators of elevation in blood flow in the cerebrovasculature and neuronal activity, respectively) were measured in motor- and cognition-related brain regions using Western blotting analysis. RESULTS: Experiment 1 indicated that treadmill activity induces an intensity-dependent increase in peNOS, c-fos, and BDNF levels. Experiment 2 showed that intensity of exercise as well as activation of the cerebral BDNF pathway, and synaptogenesis did not differ among horizontal, uphill, and downhill treadmill activities. CONCLUSION: The cerebral response of the BDNF pathway to a treadmill activity is dependent on exercise intensity, but not on the type of contraction (eccentric vs concentric).

Plasma cholesterol level determines in vivo prion propagation.

Transmissible spongiform encephalopathies are fatal neurodegenerative diseases with an urgent need for therapeutic and prophylactic strategies. At the time when the blood-mediated transmission of prions was demonstrated, in vitro studies indicated a high binding affinity of the scrapie prion protein (PrPSc) with apoB-containing lipoproteins, i.e., the main carriers of cholesterol in human blood. The aim of the present study was to explore the relationship between circulating cholesterol-containing lipoproteins and the pathogenicity of prions in vivo. We showed that, in mice with a genetically engineered deficiency for the plasma lipid transporter, phospholipid transfer protein (PLTP), abnormally low circulating cholesterol concentrations were associated with a significant prolongation of survival time after intraperitoneal inoculation of the 22L prion strain. Moreover, when circulating cholesterol levels rose after feeding PLTP-deficient mice a lipid-enriched diet, a significant reduction in survival time of mice together with a marked increase in the accumulation rate of PrPSc deposits in their brain were observed. Our results suggest that the circulating cholesterol level is a determinant of prion propagation in vivo and that cholesterol-lowering strategies might be a successful therapeutic approach for patients suffering from prion diseases.