Nitrosative Redox Homeostasis and Antioxidant Response Defense in Disused Vastus lateralis Muscle in Long-Term Bedrest (Toulouse Cocktail Study)

Abstract

Increased oxidative stress by reactive oxygen species (ROS) and reactive nitrogen species (RNS) is a major determinant of disuse-induced muscle atrophy. Muscle biopsies (thigh vastus lateralis, VL) obtained from healthy male subjects enrolled in the Toulouse Cocktail bedrest (BR) study were used to assess efficacy of an antioxidant cocktail (polyphenols, omega-3, vitamin E, and selenium) to counteract the increased redox homeostasis and enhance the antioxidant defense response by using label-free LC–MS/MS and NITRO-DIGE (nitrosated proteins), qPCR, and laser confocal microscopy. Label-free LC–MS/MS indicated that treatment prevented the redox homeostasis dysregulation and promoted structural remodeling (TPM3, MYH7, MYBPC, MYH1, MYL1, HRC, and LUM), increment of RyR1, myogenesis (CSRP3), and skeletal muscle development (MUSTN1, LMNA, AHNAK). These changes were absent in the Placebo group. Glycolysis, tricarboxylic acid cycle (TCA), oxidative phosphorylation, fatty acid beta-oxidation, and mitochondrial transmembrane transport were normalized in treated subjects. Proteins involved in protein folding were also normalized, whereas protein entailed in ion homeostasis decreased. NITRO-DIGE analysis showed significant protein nitrosylation changes for CAT, CA3, SDHA, and VDAC2 in Treatment vs. Placebo. Similarly, the nuclear factor erythroid 2-related factor 2 (Nrf-2) antioxidant response element (Nrf-2 ARE) signaling pathway showed an enhanced response in the Treatment group. Increased nitrosative redox homeostasis and decreased antioxidant defense response were found in post-BR control (Placebo, n = 10) vs. the antioxidant cocktail treated group (Treatment, n = 10). Taken together, increased nitrosative redox homeostasis and muscle deterioration during BR-driven physical inactivity were prevented, whereas decreased antioxidant nitrosative stress defense response was attenuated by Treatment suggesting positive effects of the nutritional intervention protocol in bedrest.

Effects of Omega-3 and Antioxidant Cocktail Supplement on Prolonged Bed Rest: Results from Serum Proteome and Sphingolipids Analysis

“…Our previous proteomic study on muscle sample extracts of the same Cocktail study subjects identified prevention of nitrosative stress and muscle deterioration in cocktailtreated subjects, highlighting a reduction of inflammatory pathways and development of skeletal muscle/muscle contraction [20]. Specifically, muscle proteins related to inflammatory pathways (HP, C4A, C3 and IGHG3) decreased, while proteins associated with the development of skeletal muscle/muscle contraction prelaminin A/C (LMNA), neuroblast differentiation-associated protein (AHNAK), tropomyosin alpha-3 chain (TPM3), myosin-1(MYH1), myosin light chain 1/3 (MYL1), myosin-7 (MYH7), myosin-binding protein C (MYBPC1), ryanodine receptor 1 (RYR1) and sarcoplasmic reticulum histidine-rich calcium-binding protein (HRC) increased upon cocktail treatment [20]. In serum, increased levels of HP, C4A and C3 can suggest their cellular release into the blood stream and can be potential biomarkers to monitor the cocktail treatment.…”

“…Recent observations from human and animal studies [16][17][18] demonstrated that the additive and synergistic effects of a combined use of molecules such as polyphenols, vitamins and essential fatty acids can influence some metabolic features associated with physical inactivity [19]. In muscle, previous studies from the Toulouse (Cocktail) BR study indicated that the combined use of ω-3, polyphenols and vitamins influences protein post-translational modifications targeting the redox homeostasis balance [20]. Results from nitrosoproteome, protein ubiquitination and protein carbonylation on protein extracts from vastus lateralis muscle biopsies indicated increased nitrosative redox homeostasis, lower protein ubiquitination and lower carbonylation, suggesting protection from oxidative damage in subjects under cocktail treatment.…”

“…Results from nitrosoproteome, protein ubiquitination and protein carbonylation on protein extracts from vastus lateralis muscle biopsies indicated increased nitrosative redox homeostasis, lower protein ubiquitination and lower carbonylation, suggesting protection from oxidative damage in subjects under cocktail treatment. However, the preservation of lipemic homeostasis at the systemic level was not addressed [20,21].…”

Exercise and mitochondrial mechanisms in patients with sarcopenia

“…The accumulation of unfolded proteins in the mitochondrial matrix also stimulates the uptake of PRKN into the mitochondria (in addition to changes in the mitochondrial membrane potential) ( Yan et al, 2020 ). PHB1 and PHB2 stimulate a wide range of cellular functions, including mitophagy, cellular signaling, mitochondrial biogenesis, aging, and apoptosis ( Blottner et al, 2021 ).…”

Oxidative Stress in Skeletal Muscle

“…Another effect induced by dietary supplements is described in the paper by Dieter Blottner et al [ 4 ] who, as part of a long-term bed-rest project (BR) to study muscular atrophy in humans (Toulouse Cocktail Study), used a diet enriched with polyphenols, omega-3, vitamin E, and selenium (Toulouse Cocktail) to assess its effectiveness in the antioxidant defense response and to improve the functional status of muscle.…”