Reduced serum levels of mitochondria-derived peptide MOTS-c in patients with obstructive sleep apnea

Kumagai H., Kim S., Miller B., Natsume T., Wan J., Kumagai M.E., Ramirez II R., Lee S.H., Sato A., Mehta H.H., Yen K., Cohen P.

MOTS-c, a mitochondrial microprotein, has been described as a novel regulator of glucose and lipid metabolism. In addition to its role as a metabolic regulator, MOTS-c prevents skeletal muscle atrophy in high-fat-fed mice. Here, we examined the preventive effect of MOTS-c on skeletal muscle mass using an immobilization-induced muscle atrophy model and explored its underlying mechanisms. Male C57BL/6J mice (10-week-old) were randomly assigned to one of the three experimental groups: non-immobilization control group (sterilized water injection), immobilization control group (sterilized water injection), and immobilization and MOTS-c treated group (15 mg/kg/day MOTS-c injection). We used casting tape for the immobilization experiment. After eight days of the experimental period, skeletal muscle samples were collected and used for the Western blotting, RNA sequencing, lipid, and collagen assays. Immobilization reduced ~15% of muscle mass, while MOTS-c treatment attenuated muscle loss with only a 5% reduction. MOTS-c treatment also normalized phospho-AKT, phospho-FOXO1, and phospho-FOXO3a expression levels, and reduced circulating inflammatory cytokines, such as interleukin-1b (IL-1β), interleukin-6 (IL-6), chemokine C-X-C motif ligand 1 (CXCL1), and monocyte chemoattractant protein 1 (MCP-1), in immobilized mice. An unbiased RNA sequencing and its downstream analyses demonstrated that MOTS-c modified adipogenic-modulating gene expression within the peroxisome proliferator-activated receptors (PPARs) pathway. Supporting this observation, muscle fatty acid levels were lower in the MOTS-c treated group than in the casted-controls. These results suggest that MOTS-c treatment inhibits skeletal muscle lipid infiltration by regulating adipogenesis-related genes and prevents immobilization-induced muscle atrophy.

Tang M., Su Q., Duan Y., Fu Y., Liang M., Pan Y., Yuan J., Wang M., Pang X., Ma J., Laher I., Li S.

AbstractMyocardial remodeling and dysfunction are commonly observed in type 2 diabetes mellitus (T2DM). Aerobic exercise can partly alleviate diabetes-induced myocardial dysfunction through its antioxidant actions. MOTS-c is a potential exercise mimic. This study aimed to investigate the effects of MOTS-c on improving diabetic heart function and its mechanism and to identify whether MOTS-c improved antioxidant defenses due to aerobic exercise. Herein, we established a rat model of T2DM induced by high-fat diet combined with a low-dose streptozotocin injection. Interventions were performed using intraperitoneal injections of MOTS-c (i.p. 0.5 mg/kg/day, 7 days/week) or aerobic exercise training (treadmill, 20 m/min, 60 min/day, 5 days/week) for 8 weeks. Myocardial ultrastructure was assessed using transmission electron microscopy (TEM), myocardial lipid peroxidation levels (MDA), superoxide dismutase (SOD), glutathione (GSH), and catalase (CAT) levels were assessed using colorimetric methods, and molecular analyses including MOTS-c, Kelch-like ECH-associated protein 1 (Keap1), Nuclear factor E2-related factor 2 (Nrf2), adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK)and phospho-AMPK (p-AMPK) were examined using Western blot. The results showed that MOTS-c, with or without exercise, reduced myocardial ultrastructural damage and improved glucolipid metabolism and cardiac function in T2DM. Furthermore, MOTS-c increased antioxidant markers such as SOD, CAT, and the protein expression of myocardial MOTS-c, Keap1, Nrf2, and p-AMPK. MOTS-c with exercise treatment reduced myocardial MDA and increased p-AMPK significantly comparing to only exercise or MOTS-c alone. Our findings suggest that MOTS-c may be a helpful supplement for overcoming exercise insufficiency and improving myocardial structure and function in diabetes.

Domin R., Pytka M., Żołyński M., Niziński J., Rucinski M., Guzik P., Zieliński J., Ruchała M.

The mitochondrial open reading frame of 12S rRNA-c (MOTS-c) is a mitochondrial-derived peptide that regulates the nuclear genome during stressful conditions such as hypoxia, which is typical of exercise and training. We aim to mainly investigate the relationship between serum MOTS-c concentration and muscle strength parameters measured during the countermovement jump test with oxygen consumption (VO2) measured during the cardiopulmonary exercise test to exhaustion. Physically active healthy volunteers (17 male, three female, median age 30 years), not involved in any regular exercise program or participating in any sports competitions, performed five consecutive countermovement jump tests and cardiopulmonary exercise tests until maximal exhaustion and underwent a body composition assessment by means of bioelectrical impedance analysis, and had serum MOTS-c concentration measured at rest. Serum MOTS-c concentration was positively correlated with the average power and average and maximal force of the jumps, both overall muscle mass and leg muscle mass, but not with body fat percentage. There was no correlation with peak VO2. A higher serum MOTS-c concentration is associated with greater muscle mass, force, and power generated during jumping in healthy individuals but not exercise capacity reflected by peak VO2. More studies are needed to better understand the physiological and clinical values of these findings and why MOTS-c is better associated with measures of muscle strength and not endurance in physically active people.

Kumagai H., Kim S., Miller B., Natsume T., Lee S.H., Sato A., Ramirez R., Wan J., Mehta H.H., Yen K., Cohen P.

MOTS-c is a novel mitochondrial DNA-encoded microprotein and is expressed in several tissues including skeletal muscle. Since the primary function of MOTS-c is to improve aging- and high-fat diet-induced impaired glucose uptake in the skeletal muscle, MOTS-c has been described as a metabolic regulator and exercise-mimetic microprotein. However, the direct and functional target has not been identified yet. We have previously observed increased casein kinase 2 (CK2) activity in MOTS-c-treated mouse skeletal muscle. Therefore, based on this observation, we hypothesized that CK2 is a direct target of MOTS-c in the skeletal muscle and performed a series of experiments to test our hypothesis. First, dot blot and kinase activity assays showed that MOTS-c directly bound and activated CK2 in vitro. Then, since CK2 is a complex of alpha (CK2α) and beta (CK2β) subunits, we performed a dot blot assay using CK2α and CK2β subunits and observed that MOTS-c bound to the CK2α subunit but not the CK2β subunit. This binding was confidently confirmed using a surface plasmon resonance assay (also called a Biacore assay). On the MOTS-c coding region, there is a naturally occurring variation causing a K14Q amino acid replacement that has been suggested to be a bio-inactive form of MOTS-c and increases type 2 diabetes risk. Interestingly, the Biacore assay demonstrated that the binding affinity of K14Q MOTS-c to CK2α was 16 times less than that of wild type (WT) MOTS-c, and K14Q MOTS-c did not increase CK2 activity in an in vitro kinase assay. Then, we conducted mouse experiments to validate these observations in vivo. A single administration of WT MOTS-c (7.5 mg/kg) into young male C57BL/6J mice (n = 3/group) significantly increased CK2 activity in the skeletal muscle, but not K14Q MOTS-c. Additionally, a western blot following CK2α immunoprecipitation (IP) confirmed the binding between MOTS-c and CK2α in MOTS-c administered mouse skeletal muscle. Importantly, skeletal muscle proteomics analysis following CK2α IP showed that MOTS-c changed the CK2 interactome by recruiting over 30 additional proteins and building a chaperone complex network, which was important for myofibrillogenesis and maintenance of muscle structure and function. Additionally, CK2α no longer binds to PP2A, a protein phosphatase that dephosphorylates AKT, in MOTS-c treated skeletal muscle. Next, we investigated whether CK2 mediates the MOTS-c induced skeletal muscle glucose uptake. A single administration of WT MOTS-c (7.5 mg/kg) into young male C57BL/6J mice (n = 10-11/group) significantly increased skeletal muscle 2-Deoxy-D-glucose (2DG) uptake, but not K14Q MOTS-c. As expected, administration of a CK2 inhibitor (CX-4945, 25 mg/kg) with WT MOTS-c eliminates the MOTS-c effect, suggesting that MOTS-c promoted 2DG uptake by activating CK2. Altogether, our study demonstrated that MOTS-c directly binds to and activates CK2, resulting in physiologically crucial effects on skeletal muscle. NIH grants R01AG061834, R01AG069698, P30AG068345, R01AG068405, and P01AG034906. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Chang J.L., Goldberg A.N., Alt J.A., Ashbrook L., Auckley D., Ayappa I., Bakhtiar H., Barrera J.E., Bartley B.L., Billings M.E., Boon M.S., Bosschieter P., Braverman I., Brodie K., Cabrera‐Muffly C., et. al.

Background Evaluation and interpretation of the literature on obstructive sleep apnea is needed to consolidate and summarize key factors important for clinical management of the OSA adult patient. Toward this goal, an international collaborative of multidisciplinary experts in sleep apnea evaluation and treatment have produced the International Consensus statement on Obstructive Sleep Apnea (ICS:OSA). Methods Using previously defined methodology, focal topics in OSA were assigned as literature review (LR), evidence-based review (EBR), or evidence-based review with recommendations (EBR-R) formats. Each topic incorporated the available and relevant evidence which was summarized and graded on study quality. Each topic and section underwent iterative review and the ICS:OSA was created and reviewed by all authors for consensus. Results The ICS:OSA addresses OSA syndrome definitions, pathophysiology, epidemiology, risk factors for disease, screening methods, diagnostic testing types, multiple treatment modalities, and effects of OSA and treatment on the multiple comorbidities. Specific focus on outcomes with positive airway pressure (PAP) and surgical treatments were evaluated. Conclusion This review of the literature in OSA consolidates the available knowledge and identifies the limitations of the current evidence. This effort aims to highlight the basis of OSA evidence-based practice and identify future research needs. Knowledge gaps and opportunities for improvement include improving the metrics of OSA disease, determining the optimal OSA screening paradigms, developing strategies for PAP adherence and longitudinal care, enhancing selection of PAP alternatives and surgery, understanding health risk outcomes, and translating evidence into individualized approaches to therapy. This article is protected by copyright. All rights reserved

Park S., Kim Y., Bang H.I., Park Y.

Amado C.A., Martín-Audera P., Agüero J., Lavín B.A., Guerra A.R., Boucle D., Ferrer-Pargada D., Berja A., Martín F., Casanova C., García-Unzueta M.

BackgroundMOTS-c and Romo1 are mitochondrial peptides that are modulated by oxidative stress. No previous studies have explored circulating levels of MOTS-c in patients with chronic obstructive pulmonary disease (COPD).MethodsWe enrolled 142 patients with stable COPD and 47 smokers with normal lung function in an observational cross-sectional study. We assessed serum levels of both MOTS-c and Romo1 and associated these findings with clinical characteristics of COPD.ResultsCompared with smokers with normal lung function, patients with COPD had lower levels of MOTS-c (p = 0.02) and higher levels of Romo1 (p = 0.01). A multivariate logistic regression analysis revealed that above-median MOTS-c levels were positively associated with Romo1 levels (OR 1.075, 95% CI 1.005–1.150, p = 0.036), but no association was found with other COPD characteristics. Below-median levels of circulating MOTS-c were associated with oxygen desaturation (OR 3.25 95% CI 1.456–8.522, p = 0.005) and walking <350 meters (OR 3.246 95% CI 1.229–8.577, p = 0.018) in six-minute walk test. Above-median levels of Romo1 were positively associated with current smoking (OR 2.756, 95% CI 1.133–6.704, p = 0.025) and negatively associated with baseline oxygen saturation (OR 0.776 95% CI 0.641–0.939, p = 0.009).ConclusionsReduced levels of circulating MOTS-c and increased levels of Romo1 were detected in patients diagnosed with COPD. Low levels of MOTS-c were associated with oxygen desaturation and poorer exercise capacity using 6 min walk test. Romo1 was associated with current smoking and baseline oxygen saturation.Trial registrationwww.clinicaltrials.gov; No.: NCT04449419; URL: www.clinicaltrials.gov. Date of registration: June 26, 2020.

Clemente-Suárez V.J., Martín-Rodríguez A., Redondo-Flórez L., Ruisoto P., Navarro-Jiménez E., Ramos-Campo D.J., Tornero-Aguilera J.F.

Cancer continues to be a significant global health issue. Traditional genetic-based approaches to understanding and treating cancer have had limited success. Researchers are increasingly exploring the impact of the environment, specifically inflammation and metabolism, on cancer development. Examining the role of mitochondria in this context is crucial for understanding the connections between metabolic health, physical activity, and cancer. This study aimed to review the literature on this topic through a comprehensive narrative review of various databases including MedLine (PubMed), Cochrane (Wiley), Embase, PsychINFO, and CinAhl. The review highlighted the importance of mitochondrial function in overall health and in regulating key events in cancer development, such as apoptosis. The concept of “mitochondrial fitness” emphasizes the crucial role of mitochondria in cell metabolism, particularly their oxidative functions, and how proper function can prevent replication errors and regulate apoptosis. Engaging in high-energy-demanding movement, such as exercise, is a powerful intervention for improving mitochondrial function and increasing resistance to environmental stressors. These findings support the significance of considering the role of the environment, specifically inflammation and metabolism, in cancer development and treatment. Further research is required to fully understand the mechanisms by which physical activity improves mitochondrial function and potentially reduces the risk of cancer.

Li S., Wang M., Ma J., Pang X., Yuan J., Pan Y., Li X., Liang Y., Laher I.

García-Benlloch S., Revert-Ros F., Blesa J.R., Alis R.

MOTS-c (mitochondrial open reading frame of the 12 S rRNA-c) is a newly discovered peptide that has been shown to have a protective role in whole-body metabolic homeostasis. This could be a consequence of the effect of MOTS-c on muscle tissue. Here, we investigated the role of MOTS-c in the differentiation of human (LHCN-M2) and murine (C2C12) muscle progenitor cells. Cells were treated with peptides at the onset of differentiation or after myotubes had been formed. We identified in silico a putative Src Homology 2 (SH2) binding motif in the YIFY region of the MOTS-c sequence, and created a Y8F mutant MOTS-c peptide to explore the role of this region. In both cellular models, treatment with wild-type MOTS-c peptide increased myotube formation whereas treatment with the Y8F peptide did not. MOTS-c wild-type, but not Y8F peptide, also protected against interleukin-6 (IL-6)-induced reduction of nuclear myogenin staining in myocytes. Thus, we investigated whether MOTS-c interacts with the IL-6/Janus kinase/ Signal transducer and activator of transcription 3 (STAT3) pathway, and found that MOTS-c, but not the Y8F peptide, blocked the transcriptional activity of STAT3 induced by IL-6. Altogether, our findings suggest that, in muscle cells, MOTS-c interacts with STAT3 via the putative SH2 binding motif in the YIFY region to reduce STAT3 transcriptional activity, which enhances myotube formation. This newly discovered mechanism of action highlights MOTS-c as a potential therapeutic target against muscle-wasting in several diseases. • MOTS-c has a myogenic effect on myoblast differentiation in vitro. • The internal hydrophobic YIFY motif of MOTS-c is responsible for this effect. • MOTS-c seems to interact with the IL-6/JAK/STAT3 pathway to enhance muscle formation.

Laher I., Wei Y., Liu Y., Ayas N.

Shen C., Wang J., Feng M., Peng J., Du X., Chu H., Chen X.

Oxidative stress and the inflammatory response contribute to the progression of cardiovascular disease. The present study aimed to investigate whether the mitochondrial-derived peptide MOTS-c could alleviate H2O2-induced oxidative stress and inflammatory status in H9c2 cells through activation of nuclear factor erythroid 2-related Factor 2 (Nrf2)/antioxidative response element (ARE) and inhibition of the NF-κB pathway. Rat H9c2 cardiomyocytes were obtained, and 10, 20 or 50 μM MOTS-c was pretreated for 24 h before treatment with H2O2. Then, the cell was treated with 100 μM H2O2 for 1 h to induce oxidative stress. An inhibition model of sh-Nrf2 was constructed via a lentivirus expression system, and an activation model of NF-κB was achieved using phorbol 12-myristate-13-acetate (PMA). Cell viability was determined using a Cell Counting kit-8 assay. Relative measurement of relative protein and mRNA expression used western blotting and qRT-PCR, respectively. Intracellular reactive oxygen species (ROS) levels were detected using dichlorodihydrofluorescein diacetate, and malondialdehyde (MDA) and superoxide dismutase (SOD) levels were determined via commercial kits. The protein expression and distribution in the cells were visualized by immunofluorescence analysis. Enzyme-linked immunosorbent assay was used to detect the levels of inflammatory cytokines, including TNF-α, IL-6 and IL-1β. We found that H2O2 treatment significantly decreased cell viability and the level of SOD, increased the levels of ROS and MDA, and upregulated the expression of inflammatory cytokines, including TNF-α, IL-6 and IL-1β, in H9c2 cells. The expression levels of Nrf2, HO-1 and NQO-1 were significantly downregulated in the H2O2, while the phosphorylation of NF-κBp65 was promoted by H2O2. However, pretreatment with MOTS-c significantly reversed H2O2-induced damage in H9c2 cells. Moreover, both inhibition of the Nrf2/ARE pathway and activation of the NF-κB pathway significantly decreased the effects of MOTS-c, suggesting that MOTS-c might play a role in alleviating oxidative damage via the Nrf2/ARE and NF-κB pathways. Our investigation indicated that MOTS-c could protect against H2O2-induced inflammation and oxidative stress in H9c2 cells by inhibiting NF-κB and activating the Nrf2/ARE pathways.

Woodhead J.S., Merry T.L.

Acute exercise, and in particular aerobic exercise, increases skeletal muscle energy demand causing mitochondrial stress, and mitochondrial-related adaptations which are a hallmark of exercise training. Given that mitochondria are central players in the exercise response, it is imperative that they have networks that can communicate their status both intra- and inter-cellularly. Peptides encoded by short open-reading frames within mitochondrial DNA, mitochondrial-derived peptides (MDPs), have been suggested to form a newly recognised branch of this retrograde signalling cascade that contribute to coordinating the adaptive response to regular exercise. Here we summarise the recent evidence that acute high intensity exercise in humans can increase concentrations of the MDPs humanin and MOTS-c in skeletal muscle and plasma, and speculate on the mechanisms controlling MDP responses to exercise stress. Evidence that exercise training results in chronic changes in MDP expression within tissues and the circulation is conflicting and may depend on the mode, duration, intensity of training plan and participant characteristics. Further research is required to define the effect of these variables on MDPs and to determine whether MDPs other than MOTS-c have exercise mimetic properties. MOTS-c treatment of young and aged mice improves exercise capacity/performance and leads to adaptions that are similar to that of being physically active (weight loss, increased antioxidant capacity and improved insulin sensitivity), however, studies utilising a MOTS-c inactivating genetic variant or combination of exercise + MOTS-c treatment in mice suggest that there are distinct and overlapping pathways through which exercise and MOTS-c evoke metabolic benefits. Overall, MOTS-c, and potentially other MDPs, may be exercise-sensitive myokines and further work is required to define inter- and intra-tissue targets in an exercise context.

Osorio R.S., Martínez-García M.Á., Rapoport D.M.

Due in part to overall improvements in health, the population of elderly individuals is increasing rapidly. Similarly, obstructive sleep apnoea (OSA) is both gaining increased recognition and also increasing due to the worldwide obesity epidemic. The overlap of OSA and ageing is large, but there is strong plausibility for causation in both directions: OSA is associated with pathological processes that may accelerate ageing and ageing-related processes; ageing may cause physical and neurological changes that predispose to obstructive (and central) apnoea. In addition, the common symptoms (e.g. excessive daytime sleepiness, and defects in memory and cognition), possible physiological consequences of OSA (e.g. accelerated cardiovascular and cerebrovascular atherosclerosis), and changes in metabolic and inflammatory markers overlap with the symptoms and associated conditions seen in ageing. There is also the possibility of synergy in the effects of these symptoms and conditions on quality of life, as well as a need to separate treatable consequences of OSA from age-related complaints. Taken together, the aforementioned considerations make it essential to review the interaction of OSA and ageing, both proven and suspected. The present review examines some aspects of what is known and points to the need for further investigation of the relationships, given the large number of potentially affected subjects.

von Walden F., Fernandez-Gonzalo R., Norrbom J., Emanuelsson E.B., Figueiredo V.C., Gidlund E., Norrbrand L., Liu C., Sandström P., Hansson B., Wan J., Cohen P., Alkner B.

In this manuscript, we report for the first time, to our knowledge, the response of circulating levels of mitochondrial-derived peptides humanin and MOTS-c to acute resistance and endurance exercise. Our data support that acute endurance exercise stimulates MDP levels in plasma, whereas acute resistance exercise does not.