Skip to main content
NCBI home page
Wiley Open Access Collection logoLink to Wiley Open Access Collection
. 2025 Apr 20;33(5):974–985. doi: 10.1002/oby.24274

Semaglutide‐induced weight loss improves mitochondrial energy efficiency in skeletal muscle

Ran Hee Choi 1,2,✉, Takuya Karasawa 1,2,3,✉, Cesar A Meza 1,2, J Alan Maschek 1,4, Allison M Manuel 4, Linda S Nikolova 5, Kelsey H Fisher‐Wellman 6, James E Cox 1,4,7, Amandine Chaix 1,2,8, Katsuhiko Funai 1,2,7,8,✉
PMCID: PMC12015655  PMID: 40254778

Abstract

Objective

Glucagon‐like peptide‐1 receptor agonists (e.g., semaglutide) potently induce weight loss, thereby reducing obesity‐related complications. However, weight regain occurs when treatment is discontinued. An increase in skeletal muscle oxidative phosphorylation (OXPHOS) efficiency upon diet‐mediated weight loss has been described, which may contribute to reduced systemic energy expenditure and weight regain. We set out to determine the unknown effect of semaglutide on muscle OXPHOS efficiency.

Methods

C57BL/6J mice were fed a high‐fat diet for 12 weeks before receiving semaglutide or vehicle for 1 or 3 weeks. The rates of ATP production and oxygen (O2) consumption were measured via high‐resolution respirometry and fluorometry to determine OXPHOS efficiency in muscle at these two time points.

Results

Semaglutide treatment led to significant reductions in fat and lean mass. Semaglutide improved skeletal muscle OXPHOS efficiency, measured as ATP produced per O2 consumed in permeabilized muscle fibers. Mitochondrial proteomic analysis revealed changes restricted to two proteins linked to complex III assembly (LYRM7 and TTC19; p < 0.05 without multiple corrections) without substantial changes in the abundance of OXPHOS subunits.

Conclusions

These data indicate that weight loss with semaglutide treatment increases skeletal muscle mitochondrial efficiency. Future studies could test whether it contributes to weight regain.

Study Importance.

What is already known?

  • Glucagon‐like peptide‐1 (GLP‐1) receptor agonists successfully reduce adiposity in mice and in humans.

  • In rodents, dietary intervention‐induced weight loss improves skeletal muscle oxidative phosphorylation (OXPHOS) energy efficiency.

What does this study add?

  • Semaglutide‐induced weight loss also increased muscle mitochondrial OXPHOS efficiency in mice.

  • Semaglutide‐induced increase in mitochondrial efficiency was observed in permeabilized muscle fibers, but not in isolated mitochondria.

How might these results change the direction of research or the focus of clinical practice?

  • GLP‐1 receptor agonists likely promote reduced energy expenditure in a similar fashion to dietary intervention‐induced weight loss. This may influence propensity for weight regain or beneficial effects of increasing energy expenditure.

INTRODUCTION

Obesity predisposes individuals to a broad spectrum of diseases, including type 2 diabetes and cancer. Modest (5%–10%) weight loss improves clinical parameters associated with obesity‐related diseases [1]. Lifestyle interventions can successfully induce ≥5% weight loss from an overweight state; however, maintaining a weight‐reduced state has been extremely challenging. Almost 80% of people who have undergone weight loss by various methods are expected to undergo a weight‐regain phase over the next 5 years [2]. Combating weight rebound for sustainable weight management remains a primary challenge and is compounded by environmental, behavioral, and physiological factors that promote regain.

Weight loss from an overweight state increases energy efficiency, thereby lowering the fuel cost during low levels of physical activity [3, 4, 5]. This metabolic adaptation persists for several years following weight loss and makes individuals susceptible to significant weight regain [3, 6]. It has been demonstrated that increased skeletal muscle work efficiency during, as well as after, weight loss contributes to decreased energy expenditure [4]. Potential mechanisms underlying metabolic adaptations in the weight‐reduced state include decreased glycolytic enzymatic activity such as phosphofructokinase, lower bioactive thyroid hormones, increased activity of sarcoplasmic/endoplasmic reticulum Ca2+ ATPase (SERCA), and shift in myosin heavy chain (MHC) isoforms toward MHC I, characterized as more efficient and oxidative, that may enhance skeletal muscle work efficiency by altering myofibrillar ATPases [4, 5, 7]. Similar changes in metabolic rates can be found in the context of exercise training and high‐altitude adaptation [8, 9].

Recently, we have shown that dietary intervention‐induced weight loss in overweight mice increased mitochondrial oxidative phosphorylation (OXPHOS) efficiency of skeletal muscle, quantified by ATP production per oxygen (O2) consumption (P/O ratio) [10]. Those changes were not associated with changes in SERCA abundance or efficiency compared to lean controls and overweight groups. These results suggest that increased energy efficiency in a weight‐reduced state may be due to alteration of mitochondrial OXPHOS efficiency, which requires less O2 to produce equimolar ATP production.

In recent years, glucagon‐like peptide‐1 (GLP‐1) receptor agonists such as semaglutide have shown striking efficacy for weight loss, inducing 10% to 15% reduction in body weight within 1 year. In the semaglutide treatment effect in people with obesity and semaglutide effects on cardiovascular outcomes in people with overweight or obesity trials, lasting 68 weeks and 208 weeks, respectively, weekly subcutaneous injection of semaglutide (2.4 mg) decreased weight by ~10% to 15% within 1 year, which was successfully sustained for the experimental duration [11, 12]. Importantly, individuals who stopped taking GLP‐1 receptor agonists regained 10% of their lost weight within a year [13]. Although changes in food intake likely largely contribute to weight regain, there have been limited studies on the contribution of energy efficiency/expenditure on the propensity for weight regain during or after GLP‐1 receptor agonist treatment. Furthermore, neither the duration of changed energy efficiency during and/or after weight loss nor the outcomes of the sustained increase in energy efficiency have been fully described. As mentioned earlier, increased energy efficiency from weight loss is similar to that observed with exercise training; therefore, increased energy efficiency in the weight‐reduced state could have potential benefits on health, such as by improving glycemic controls and reducing oxidative stress [14, 15].

In this study, we set out to determine the influence of GLP‐1 receptor agonist‐induced weight loss on skeletal muscle mitochondrial energy efficiency. We hypothesized that semaglutide treatment will increase energy efficiency of skeletal muscle mitochondrial ATP production similarly to our observation with dietary intervention‐induced weight loss [10].

METHODS

Animals

Male C57BL/6J (RRID:IMSR_JAX:000664) mice were purchased from Jackson Laboratory at 20 weeks of age. Mice were maintained on a high‐fat diet (HFD; 42% of calories from fat consisting of 210 g/kg anhydrous milk fat, with a composition of 61.8% saturated fat, 27.3% monounsaturated fat, 4.7% polyunsaturated fat, Inotiv TD88137) for the duration of the experiments. Body composition was determined by Bruker Minispec nuclear magnetic resonance (NMR) (Bruker Corp.) before and after the interventions. At 12 weeks of HFD feeding, mice were divided into two groups: vehicle (phosphate‐buffered saline [PBS]); and semaglutide (Novo Nordisk A/S, subcutaneous, 3 nmol/kg body weight [bw]/day). The dose of 3 nmol/kg bw/day is relevant to the clinical doses of semaglutide (0.25 mg to 2.4 mg/week) [16]. Reagents were administered once daily for either 1 week or 3 weeks. Mice were housed in a 12‐h light/dark cycle in a temperature‐controlled room at 22°C and were provided access to food and water ad libitum. For terminal experiments, mice were injected intraperitoneally with ketamine (80 mg/kg) and xylazine (10 mg/kg), after which tissues were collected. All protocols were approved by Institutional Animal Care and Use Committee at the University of Utah.

Preparation of permeabilized muscle fiber bundles

A small portion of mouse red gastrocnemius muscle was dissected and placed in buffer X (7.23mM K2EGTA, 2.77mM CaK2EGTA, 20mM imidazole, 20mM taurine, 5.7mM ATP, 14.3mM phosphocreatine, 6.56mM MgCl2·6H2O, and 50mM 2‐[N‐Morpholino] ethanesulfonic acid potassium salt [K‐MES], pH 7.4). Fiber bundles were separated and permeabilized for 30 min at 4°C with saponin (30 μg/mL). Immediately afterward, permeabilized muscle fiber bundles (PmFB) were washed in buffer Z (105mM K‐MES, 30mM KCl, 10mM K2HPO4, 5mM MgCl2·6H2O, 0.5 mg/mL of bovine serum albumin [BSA], and 1mM EGTA, pH 7.4) with 0.5mM pyruvate and 0.2mM malate for 15 min. After washing, PmFB were placed in buffer Z and analyzed within 1 hour. All measurements from PmFB were performed in duplicate, but not those from isolated mitochondria.

Mitochondria isolation from mouse skeletal muscle

Mouse gastrocnemius muscle was freshly dissected and used to isolate mitochondria, as previously described [17]. Briefly, the tissue was minced in ice‐cold mitochondrial isolation medium (MIM) buffer (0.3M sucrose, 10mM HEPES, and 1mM EGTA, pH 7.1) with BSA (1 mg/mL) and homogenized using a TH‐01 homogenizer (Omni International, Inc.). The homogenate was then centrifuged at 800g for 10 min at 4°C. The supernatant was transferred to a new tube and then centrifuged at 12,000g for 10 min at 4°C. The supernatant was removed and the crude mitochondrial pellet was washed in 1 mL of MIM buffer. Subsequently, the pellet was centrifuged again at 12,000g for 10 min at 4°C, and the pellet was resuspended in 100 μL of MIM buffer.

High‐resolution respirometry and fluorometry

PmFB and isolated mitochondria were used to determine O2 consumption and ATP production, as previously described [10]. Respiration was measured using the Oxygraph‐2 K (Oroboros Instruments). ATP production was determined by enzymatically coupling ATP production to nicotinamide adenine dinucleotide phosphate (NADPH) synthesis using the Fluorolog‐QM (Horiba Scientific), as previously described [18]. Both experiments were performed in buffer Z in the presence of 0.5mM malate, 5mM pyruvate, 5mM glutamate, and 10mM succinate. Respiration and ATP production were recorded with a subsequent ADP titration (20, 200, and 2000 μM for PmFB; 2, 20, and 200 μM for isolated mitochondria). In PmFB experiments, 20mM creatine monohydrate and 10 μM blebbistatin were added to buffer Z to inhibit myosin ATPase. The P/O ratio was analyzed by dividing the rate of ATP production by the rate of atomic O2 consumption [18]. Mitochondrial respiration and ATP production were measured using independent Oroboros respirometers and Horiba fluorometers using different aliquots from the same samples. We find that this approach maximizes our ability to perform experiments on a greater number of mice, reducing experimental variability [10, 18].

Statistical analysis

Statistical analysis was performed using GraphPad Prism 10.2.3 software (GraphPad Software). Two‐way ANOVA followed by Tukey multiple comparisons test or unpaired t test was performed for group comparison. All data are presented as mean (SE), and statistical significance was set at p ≤ 0.05.

Additional experimental details can be found in the online Supporting Information Methods.

RESULTS

Semaglutide treatment induces weight loss in overweight mice

Wild‐type C57BL/6J mice were fed an HFD (42% of calories from fat) for 12 weeks before being treated with either vehicle (PBS) or semaglutide (3 nmol/kg bw/day, subcutaneous injection) for 1 week or 3 weeks (Figure 1A). During the intervention, mice were maintained on the HFD. Mice treated with semaglutide lost about 12% (1.7%) of weight after 1 week of treatment and about 23% (1.66%) after 3 weeks (Figure 1B,C). Weight loss was due to disproportionately more loss in fat mass than lean mass. Fat mass was significantly decreased by 26% (2.23%) after 1 week and by 42% (3.66%) after 3 weeks of semaglutide treatment compared to the vehicle groups (Figure 1D,E). Lean mass was significantly reduced by 9% (1.5%) both after 1 week and 3 weeks (Figure 1F,G), and this was similarly reflected in gastrocnemius muscle mass (Figure 1H). Thus, semaglutide intervention significantly reduced weight, fat mass, and lean mass.

FIGURE 1.

FIGURE 1

Open in a new tab

Semaglutide induces weight loss in overweight mice. (A) Schematic of the experimental design. Twenty‐week‐old C57BL/6J mice were fed a 42% HFD for 12 weeks before semaglutide (3 nmol/kg of body weight/day) intervention. Semaglutide was subcutaneously injected daily for 1 or 3 weeks. Vehicle groups received an equal volume of PBS. (B) Body weight was recorded every week. (C) Percent change in body weight after semaglutide treatment was calculated from the baseline of 12 weeks of an HFD. (D) Fat mass was determined by NMR before and after treatment. (E) Percentage change in fat mass was analyzed by NMR. (F) Lean mass was determined by NMR before and after treatment. (G) Percentage change in lean mass was calculated by NMR. (H) After the completion of semaglutide treatment, gastrocnemius muscle was harvested, and wet weight was measured. Data are represented as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, and **** p < 0.0001, significant difference vs. corresponding controls. ## p < 0.01 and ### p < 0.001, significant difference vs. vehicle, 3 weeks. n = 8 and 9 for 1‐week vehicle and 3‐week vehicle, respectively, and n = 11 for both 1‐week and 3‐week semaglutide. HFD, high‐fat diet.

Semaglutide treatment increases skeletal muscle OXPHOS energy efficiency

In order to test whether semaglutide‐induced weight loss increases energy efficiency for mitochondrial ATP synthesis, we first quantified the rates of ATP production and O2 consumption and P/O ratio in red gastrocnemius PmFB harvested from 1‐week and 3‐week vehicle and semaglutide‐treated groups. The red portions of gastrocnemius muscle were selected due to them providing sufficient tissue mass for a variety of mitochondrial assays. The rate of ATP production (JATP) showed a trend toward an increase after 1 week of semaglutide treatment, with a more pronounced effect observed after 3 weeks of treatment (Figure 2A,B). Additionally, although O2 consumption (JO2) did not change significantly after 1 week, it was markedly decreased after 3 weeks of semaglutide treatment (Figure 2C,D). When calculating P/O ratio to determine energy efficiency, semaglutide tended to increase P/O ratio after 1 week, with a significant improvement after 3 weeks (Figure 2E,F). This improved P/O ratio was not associated with changes in OXPHOS subunit levels (Figure 2G,H). Overall, these results indicate that 3 weeks of semaglutide treatment robustly improved mitochondrial OXPHOS efficiency (p < 0.01, vs. 3‐week vehicle) in gastrocnemius muscle by reducing JO2 without affecting OXPHOS subunits.

FIGURE 2.

FIGURE 2

Open in a new tab

Semaglutide treatment increases skeletal muscle OXPHOS energy efficiency. A small piece of red gastrocnemius was collected and permeabilized in the buffer with saponin (30 μg/mL). (A,B) JATP was determined by fluorometer. (C,D) JO2 was analyzed by high‐resolution respirometry. (E,F) OXPHOS efficiency was determined as P/O ratio. (G,H) Abundance of OXPHOS subunits in whole lysate of gastrocnemius muscle was determined by Western blot. Data are represented as mean ± SEM. *p < 0.05, significant difference vs. corresponding controls. n = 10 and 13 for 1‐week vehicle and 3‐week vehicle and n = 10 and 14 for 1‐week and 3‐week semaglutide, respectively. JATP, rate of ATP production; JO2, mitochondrial O2 consumption; OXPHOS, oxidative phosphorylation; P/O, ATP produced per oxygen consumed.

Semaglutide‐induced increase in mitochondrial OXPHOS efficiency is not observed when quantified in isolated mitochondria from skeletal muscle

Mitochondrial bioenergetic phenotyping in PmFB enables assessment of respiratory responses in situ, better preserving the native conformation of mitochondrial reticulum and membrane structures. However, these preparations also contain other subcellular structures, including other ATP synthases and ATPases. Although respiratory assay buffers contain inhibitors for some of these systems, we cannot rule out the possibility that nonmitochondrial mechanisms could influence ATP fluorometry (less concerning for O2 consumption) and thereby affect P/O ratio measurements.

In order to address this, we conducted the same bioenergetic phenotyping in isolated mitochondria from gastrocnemius muscle. In isolated mitochondria, neither JATP, JO2, nor P/O ratio was significantly changed by semaglutide (Figure 3A–F), and OXPHOS subunits remained unchanged (Figure 3G,H). We observed that JATP responses stopped after the second dose of ADP (20 μM; Figure 3A,B), unlike what was observed in PmFB (Figure 2A,B), whereas JO2 continued to respond to each ADP titration (Figure 3C,D). This led to a decrease in P/O ratio at the highest ADP compared to the second dose (Figure 3E,F). Overall, the semaglutide‐induced increase in muscle OXPHOS efficiency is only observed in PmFB and not in isolated mitochondria.

FIGURE 3.

FIGURE 3

Open in a new tab

Semaglutide‐induced increase in mitochondrial OXPHOS efficiency is not observed when quantified in isolated mitochondria from skeletal muscle. Mitochondria were isolated from whole gastrocnemius muscle. (A,B) JATP was determined by fluorometer. (C,D) JO2 was analyzed by high‐resolution respirometry. (E,F) OXPHOS efficiency was determined as P/O ratio. (G,H) Abundance of OXPHOS subunits in isolated mitochondria from gastrocnemius muscle was determined by Western blot. Data are represented as mean ± SEM. n = 12 and 13 for 1‐week vehicle and 3‐week vehicle, respectively, and n = 15 for both 1‐week and 3‐week semaglutide. JATP, rate of ATP production; JO2, mitochondrial O2 consumption; OXPHOS, oxidative phosphorylation; P/O, ATP produced per oxygen consumed.

Even though blebbistatin and EGTA were included in the assay buffer to inhibit MHC and SERCA ATPase activities, we further examined the possibility that semaglutide might influence P/O ratio by changes in MHC or SERCA isoforms. Nevertheless, no differences were observed (Figure S1A–D). These results suggest that cellular compartments associated with mitochondrial network preserved in PmFB may play a role in the improvement of semaglutide‐induced OXPHOS efficiency in skeletal muscle.

Semaglutide treatment does not alter skeletal muscle mitochondrial supercomplex formation

Mitochondrial respiratory complexes are thought to have supercomplexes that have improved efficiency for electron transport compared to free complexes [19, 20]. However, semaglutide treatment did not appear to influence mitochondrial OXPHOS supercomplex assembly (Figures 4A–F and S2A–F). Mitochondrial morphology, including shape and size of individual mitochondria, as well as those of cristae, has a strong influence on OXPHOS [21]. Mitochondrial morphology analyzed by electron microscopy, however, did not show any noticeable differences between vehicle and semaglutide treatment (Figure 4G). Previously, we have shown that mitochondrial cardiolipin can also influence OXPHOS [10, 17, 22], but semaglutide did not influence cardiolipin, nor did it have any robust influence on other lipid classes in mitochondria (Figure 5A and Figure S3A–K). These findings indicate that alterations in mitochondrial supercomplex formation, morphology, and lipidome are unlikely to be responsible for the improvement in muscle P/O ratio induced by semaglutide.

FIGURE 4.

FIGURE 4

Open in a new tab

Semaglutide treatment does not alter skeletal muscle mitochondrial supercomplex formation. (A–F) Mitochondrial supercomplex formation was analyzed in isolated mitochondria from gastrocnemius muscle by native polyacrylamide gel electrophoresis (PAGE) and quantified. (G) Representative mitochondrial morphology analyzed by electron microscopy. All data are from 3‐week semaglutide treatment. n = 6 for both vehicle and semaglutide for native PAGE.

FIGURE 5.

FIGURE 5

Open in a new tab

Semaglutide‐induced weight loss does not robustly influence the abundance of OXPHOS subunits. (A) Relative abundance of mitochondrial lipids. (B) Principal component analysis of mitochondrial proteomic data. (C) Volcano plot of differentially abundant mitochondria proteins between vehicle and semaglutide groups. (D–H) Heat map of abundance of OXPHOS subunits determined by mass spectrometry. All data are from 3‐week semaglutide treatment. n = 6 for both vehicle and semaglutide. OXPHOS, oxidative phosphorylation.

Semaglutide‐induced weight loss does not robustly influence the abundance of OXPHOS subunits

The improvement in OXPHOS efficiency may result from changes in the abundance of respiratory subunits. Thus, we performed proteomic analyses on skeletal muscle mitochondria with or without semaglutide treatment. Consistent with our previous experiments with dietary intervention‐induced weight loss [10], semaglutide treatment did not appear to influence the muscle mitochondrial proteome by principal component analyses (Figure 5B), nor was there a single mitochondrial protein whose abundance was statistically significantly different (q < 0.05; Figure 5C–H). However, two proteins associated with complex III assembly were trending to be elevated (assembly factor LYRM7 increased by 41% and tetratricopeptide repeat domain 19 [TTC19] increased by 38%, both p < 0.05 without correcting for multiple comparison; Figure 5F). It is unclear whether changes in these complex III subunits are sufficient to explain the improvement in skeletal muscle OXPHOS efficiency with semaglutide treatment.

DISCUSSION

GLP‐1 receptor agonists, including semaglutide, have revolutionized obesity‐related health care by their efficacy in inducing weight loss, as well as improving other comorbidities, including diabetes, cardiovascular disease, and chronic kidney disease. Weight loss, including that induced by dietary intervention or by GLP‐1 receptor agonists, is known to induce a reduction in energy expenditure. The consequence of a perpetual reduction in resting and exercise‐induced energy expenditure extends beyond the propensity for weight regain, as it may attenuate all secondary benefits of elevated energy expenditure, such as glycemic control. With the emergence of GLP‐1 receptor agonists and other weight‐loss drugs, studying the biology of the weight‐reduced state has become increasingly relevant. In this study, we showed that semaglutide promotes an increase in skeletal muscle OXPHOS energy efficiency, in a similar fashion to our previous observation with weight loss induced by a dietary intervention [10]. The mechanism by which semaglutide treatment improved OXPHOS efficiency remains to be fully elucidated, but we found two components of complex III, i.e., LYRM7 and TTC19, to become elevated after semaglutide treatment.

We found that semaglutide treatment improved skeletal muscle mitochondrial efficiency when analyzed in red gastrocnemius PmFB. JATP tended to increase after 1 week of treatment without a change in JO2, and this effect became more pronounced after 3 weeks, with significantly attenuated JO2. These changes resulted in an ~50% increase in P/O ratio, indicating improved OXPHOS efficiency. These findings align with previous studies that have shown that weight loss, whether induced by diet or exercise, can improve mitochondrial efficiency in skeletal muscle [10, 23]. Consistent with our previous study, the higher P/O ratio was driven by a reduction in JO2 in response to the same ADP stimuli [10]. We also observed a trend toward increased JATP with semaglutide treatment. Activation of GLP‐1 receptor signaling enhances ATP synthesis in pancreatic β cells [24, 25]. Nevertheless, increased JATP was not observed in our previous paper with dietary intervention‐induced weight loss [10]. Our previous dietary intervention study induced weight loss by switching an HFD back to standard chow, whereas the mice remained on an HFD for our current study with semaglutide‐induced weight loss. Thus, these nuanced differences between dietary intervention and semaglutide treatment may be due to differences in their dietary compositions.

No significant improvements in mitochondrial efficiency were detected when measuring OXPHOS efficiency in isolated mitochondria. This may suggest the critical role of the intact cellular environment, which is preserved in PmFB but disrupted during mitochondrial isolation [26, 27]. PmFB maintain the integrity of key cellular components, including mitochondrial structural arrangement, intracellular organelles, and multienzyme systems essential for energy transfer [26, 28, 29]. Intact myofibrillar ATPases, including myosin ATPase, in PmFB potentially lead to this difference. We used blebbistatin, which inhibits specifically myosin II ATPase, to prevent spontaneous contraction during the measurement [18, 30]. There is a possibility that the addition of blebbistatin did not inhibit other myosin ATPases, such as myosin I ATPases, present in PmFB, as we precisely dissected red gastrocnemius (type I; oxidative) for PmFB. Moreover, it has been reported that increased skeletal muscle work efficiency during, as well as after, weight loss via dietary intervention has been linked to the switching myosin isoforms from more glycolytic (IIa, IIb and IIx) to oxidative (I) [4]. Other candidates for intact myofibrillar ATPases in PmFB, including Ca2+‐activated myosin ATPase and SERCA, can also be involved in generating this different observation between PmFB and isolated mitochondria, as they largely participate in ATP consumption. In order to avoid this possibility, we added a chelating agent, i.e., 1mM EGTA, to our experimental buffer. Different types of fiber or myofibrillar ATPases possibly result in distinct outcomes [27, 31, 32]. Moreover, we did not find that semaglutide treatment resulted in robust changes in MHC or SERCA isoforms. This likely explains why observed improvements were specific to PmFB, underscoring the importance of studying mitochondrial function in a more physiologically relevant context. These findings suggest that future studies should investigate how different fiber types and mitochondria sources contribute to OXPHOS efficiency under weight‐loss conditions.

Our previous study found that shifts in cardiolipin after diet‐induced weight loss potentially mediated increased OXPHOS efficiency [10]. In the current study, semaglutide‐induced improvement in OXPHOS efficiency did not appear to be related to such changes in mitochondrial cardiolipin. There were some nuanced changes in some other lipid species, and it is unclear whether these changes have influenced OXPHOS efficiency. It remains possible that semaglutide influences other aspects of skeletal muscle mitochondria to influence bioenergetics. One possibility is that GLP‐1 receptor agonists decrease mitochondrial electron leak. In leukocytes from patients with type 2 diabetes, GLP‐1 receptor agonists reduced the reactive oxygen species production [33], potentially suggesting that it might reduce mitochondrial electron leak to improve efficiency. However, electron leak usually accounts for a very small portion of total electron flux (often less than 1% when quantified) [34]. Therefore, it is difficult to imagine that a twofold increase in P/O ratio is completely accounted for by reduced electron leak. GLP‐1 receptor agonists are also known to affect mitochondrial dynamics in substantia nigra [35]. Even though we did not observe a robust change in mitochondrial morphology, there may be other subtle effects not detected by electron microscopy that contribute to improved OXPHOS efficiency.

Semaglutide treatment resulted in a 12% and 23% reduction in weight after 1 and 3 weeks of treatment, respectively. Fat mass was reduced by 26% and 42%, respectively, over the same time points. Additionally, lean mass decreased by ~10% at both time points. These findings are consistent with previous clinical trials and animal studies that have shown that GLP‐1 receptor agonists effectively promote weight loss [11, 36]. Skeletal muscle is a significant contributor to energy expenditure [3, 37]. A decrease in skeletal muscle mass during weight loss has been associated with a reduction in energy expenditure [3]. In addition to this effect, energy expenditure per unit of lean mass appears to be reduced, promoted by an increase in skeletal muscle work efficiency [4, 5, 10]. In contrast, our previous study on a dietary intervention‐induced weight loss resulted in minimal loss of lean mass [10]. Because we did not have a pair‐fed control group in the current study, it is unclear whether semaglutide has a direct effect on muscle mass through GLP‐1 receptor agonism. Alternatively, it is noteworthy that the previous study with dietary intervention‐induced weight loss involved changing mice from an HFD to standard chow, whereas mice remained on an HFD during the weight‐loss phase in the current study with semaglutide.

In the current study, semaglutide did not significantly alter the skeletal muscle mitochondrial proteome when corrected for multiple comparisons. However, we identified two proteins related to complex III assembly, i.e., LYRM7 and TTC19, that trended to be greater (p < 0.05 with t tests). LRYM7 is involved in the incorporation of the iron–sulfur cluster into Rieske (Fe‐S) protein (UQCRFS1) [38]. TTC19 is known to interact with ubiquinol‐cytochrome c reductase core protein 1 (UQCRC1) and UQCRFS1 [39], although semaglutide did not influence the abundance of these subunits. Defects in LYRM7 and TTC19 are linked to complex III deficiencies, i.e., in nuclear type 8 (MC3DN8) and nuclear type 2 (MC3DN2), respectively, which result in increased oxidative stress and inefficient electron transfer [40, 41, 42]. The association between weight loss and these proteins is unknown. Thus, semaglutide may influence LYRM7 and TTC19, potentially influencing complex III to enhance electron transport efficiency and reducing oxidative stress.

There are several limitations in the current study that should be addressed in future research. First, only male mice were used for the study, strictly from the standpoint that, in our hands, female mice take substantially longer to achieve weight gain. We are currently working on studies with a longer HFD regimen or other obesity model to study both sexes. Second, we did not include the measurements of whole‐body energy expenditure with indirect calorimetry, primarily because we found that daily injection of semaglutide/vehicle interfered with indirect calorimetry measurements. We are actively working on strategies to circumvent these limitations for our future studies.

In summary, our study demonstrates that semaglutide treatment significantly improves mitochondrial OXPHOS efficiency in skeletal muscle. Increased OXPHOS efficiency was only observed when quantified in permeabilized fibers, suggesting that intact subcellular structures may be necessary for this effect. In contrast to our previous findings with dietary intervention‐induced weight loss, we did not find semaglutide to influence the skeletal muscle mitochondrial proteome or lipidome. Alternatively, improved mitochondrial energy efficiency may be mediated by posttranslational modifications or by changes in mitochondria dynamics and ultrastructure. Although tools to study these changes in vivo are currently limited, future studies should explore alternative molecular mechanisms that drive the improved OXPHOS efficiency with semaglutide treatment. Greater understanding of these effects could help identify targets to mitigate risks for weight regain.

AUTHOR CONTRIBUTIONS

Ran Hee Choi, Takuya Karasawa, Amandine Chaix, and Katsuhiko Funai designed the study. Ran Hee Choi, Takuya Karasawa, Cesar A. Meza, and Kelsey H. Fisher‐Wellman performed mouse and mitochondrial experiments and analyzed data. J. Alan Maschek and James E. Cox conducted lipidomic analyses. Allison M. Manuel, James E. Cox, and Kelsey H. Fisher‐Wellman conducted the proteomic analysis. Linda S. Nikolova assisted with electron microscopic images. Ran Hee Choi and Katsuhiko Funai wrote the manuscript with edits from all authors.

CONFLICT OF INTEREST STATEMENT

The authors declared no conflicts of interest.

Supporting information

Figure S1. Analysis of MHC and SERCA isoforms. (A) Tibialis anterior muscle embedded in OCT at 3‐week intervention was analyzed for MHC immunofluorescence to determine fiber types. (B) SERCA 1 and SERCA 2 protein expression levels were analyzed in gastrocnemius muscle by Western blotting and (C and D) quantified.

Figure S2. Mitochondrial supercomplexes in mouse gastrocnemius muscles after 1 week of the intervention. (A–F) Supercomplexes formations were analyzed in isolated mitochondria from gastrocnemius muscle by native PAGE and quantified. All data are from 1‐week semaglutide treatment. n = 5‐6 for both vehicle and semaglutide.

Figure S3. Mitochondrial lipidome in mouse gastrocnemius muscles. Mitochondrial lipidome was analyzed in isolated mitochondrial from gastrocnemius muscle of 3 weeks vehicle and semaglutide groups. (A) Heatmap of abundance of each mitochondrial phospholipidome (B) Total phospholipids from vehicle and semaglutide group (C) Absolute abundance of mitochondrial lipids (D–K) Absolute abundance of individual phospholipids species (D) Phosphocholine (PC), (E) Phosphatidylethanolamine (PE), (F) Phosphatidylinositol (PI), (G) Phosphatidylserine (PS), (H) Cardiolipin (CL), (I) Phosphatidylglycerol (PG), (J) Lysophosphatidylcholine (LPC), and (K) Lysophosphatidylethanolamine (LPE). n = 6 for both vehicle and semaglutide. **p < 0.01 significant difference vs. Veh.

OBY-33-974-s001.pdf (569.7KB, pdf)

Data S1. Supporting Information.

OBY-33-974-s002.docx (25.9KB, docx)

ACKNOWLEDGMENTS

This research is supported by National Institutes of Health (NIH) grants (DK107397, DK127979, GM144613, and AG074535 to Katsuhiko Funai; CA278826 to Kelsey H. Fisher‐Wellman; and AG065993 to Amandine Chaix) and the Grant‐in‐aid for Japan Society for the Promotion of Science (JSPS) Fellows (24KJ2039 to Takuya Karasawa). The University of Utah Metabolomics Core Facility is supported by DK110858.

Choi RH, Karasawa T, Meza CA, et al. Semaglutide‐induced weight loss improves mitochondrial energy efficiency in skeletal muscle. Obesity (Silver Spring). 2025;33(5):974‐985. doi: 10.1002/oby.24274

Ran Hee Choi and Takuya Karasawa contributed equally.

Contributor Information

Ran Hee Choi, Email: ranhee.choi@utah.edu.

Takuya Karasawa, Email: takuya.karasawa@utah.edu.

Katsuhiko Funai, Email: kfunai@utah.edu.

DATA AVAILABILITY STATEMENT

The data that support the findings of this study are available from the corresponding author upon reasonable request.

REFERENCES

  • 1. Ryan DH, Yockey SR. Weight loss and improvement in comorbidity: differences at 5%, 10%, 15%, and over. Curr Obes Rep. 2017;6(2):187‐194. doi: 10.1007/s13679-017-0262-y [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 2. Anderson JW, Konz EC, Frederich RC, Wood CL. Long‐term weight‐loss maintenance: a meta‐analysis of US studies. Am J Clin Nutr. 2001;74(5):579‐584. doi: 10.1093/ajcn/74.5.579 [DOI] [PubMed] [Google Scholar]
  • 3. Leibel RL, Rosenbaum M, Hirsch J. Changes in energy expenditure resulting from altered body weight. N Engl J Med. 1995;332(10):621‐628. doi: 10.1056/NEJM199503093321001 [DOI] [PubMed] [Google Scholar]
  • 4. Rosenbaum M, Vandenborne K, Goldsmith R, et al. Effects of experimental weight perturbation on skeletal muscle work efficiency in human subjects. Am J Physiol Regul Integr Comp Physiol. 2003;285(1):R183‐R192. doi: 10.1152/ajpregu.00474.2002 [DOI] [PubMed] [Google Scholar]
  • 5. Goldsmith R, Joanisse DR, Gallagher D, et al. Effects of experimental weight perturbation on skeletal muscle work efficiency, fuel utilization, and biochemistry in human subjects. Am J Physiol Regul Integr Comp Physiol. 2010;298(1):R79‐R88. doi: 10.1152/ajpregu.00053.2009 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6. Fothergill E, Guo J, Howard L, et al. Persistent metabolic adaptation 6 years after “The Biggest Loser” competition. Obesity (Silver Spring). 2016;24(8):1612‐1619. doi: 10.1002/oby.21538 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 7. Baldwin KM, Joanisse DR, Haddad F, et al. Effects of weight loss and leptin on skeletal muscle in human subjects. Am J Physiol Regul Integr Comp Physiol. 2011;301(5):R1259‐R1266. doi: 10.1152/ajpregu.00397.2011 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8. Joyner MJ, Coyle EF. Endurance exercise performance: the physiology of champions. J Physiol. 2008;586(1):35‐44. doi: 10.1113/jphysiol.2007.143834 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9. Murray AJ, Horscroft JA. Mitochondrial function at extreme high altitude. J Physiol. 2016;594(5):1137‐1149. doi: 10.1113/JP270079 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10. Ferrara PJ, Lang MJ, Johnson JM, et al. Weight loss increases skeletal muscle mitochondrial energy efficiency in obese mice. Life Metab. 2023;2(2):load014. doi: 10.1093/lifemeta/load014 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11. Wilding JPH, Batterham RL, Calanna S, et al. Early remdesivir to prevent progression to severe Covid‐19 in outpatients. N Engl J Med. 2022;386(4):989‐1002. doi: 10.1056/NEJMoa2032183 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12. Garvey WT, Batterham RL, Bhatta M, et al. Two‐year effects of semaglutide in adults with overweight or obesity: the STEP 5 trial. Nat Med. 2022;28(10):2083‐2091. doi: 10.1038/s41591-022-02026-4 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13. Wilding JPH, Batterham RL, Davies M, et al. Weight regain and cardiometabolic effects after withdrawal of semaglutide: the STEP 1 trial extension. Diabetes Obes Metab. 2022;24(8):1553‐1564. doi: 10.1111/dom.14725 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14. Toledo FG, Menshikova EV, Ritov VB, et al. Effects of physical activity and weight loss on skeletal muscle mitochondria and relationship with glucose control in type 2 diabetes. Diabetes. 2007;56(8):2142‐2147. doi: 10.2337/db07-0141 [DOI] [PubMed] [Google Scholar]
  • 15. Wang D, Jiang DM, Yu RR, et al. The effect of aerobic exercise on the oxidative capacity of skeletal muscle mitochondria in mice with impaired glucose tolerance. J Diabetes Res. 2022;2022:3780156. doi: 10.1155/2022/3780156 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16. Gabery S, Salinas CG, Paulsen SJ, et al. Semaglutide lowers body weight in rodents via distributed neural pathways. JCI Insight. 2020;5(6):e133429. doi: 10.1172/jci.insight.133429 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 17. Heden TD, Johnson JM, Ferrara PJ, et al. Mitochondrial PE potentiates respiratory enzymes to amplify skeletal muscle aerobic capacity. Sci Adv. 2019;5(9):eaax8352. doi: 10.1126/sciadv.aax8352 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 18. Lark DS, Torres MJ, Lin CT, Ryan TE, Anderson EJ, Neufer PD. Direct real‐time quantification of mitochondrial oxidative phosphorylation efficiency in permeabilized skeletal muscle myofibers. Am J Physiol Cell Physiol. 2016;311(2):C239‐C245. doi: 10.1152/ajpcell.00124.2016 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19. Bianchi C, Genova ML, Parenti Castelli G, Lenaz G. The mitochondrial respiratory chain is partially organized in a supercomplex assembly: kinetic evidence using flux control analysis. J Biol Chem. 2004;279(35):36562‐36569. doi: 10.1074/jbc.M405135200 [DOI] [PubMed] [Google Scholar]
  • 20. Greggio C, Jha P, Kulkarni SS, et al. Enhanced respiratory chain supercomplex formation in response to exercise in human skeletal muscle. Cell Metab. 2017;25(2):301‐311. doi: 10.1016/j.cmet.2016.11.004 [DOI] [PubMed] [Google Scholar]
  • 21. Frey TG, Mannella CA. The internal structure of mitochondria. Trends Biochem Sci. 2000;25(7):319‐324. doi: 10.1016/s0968-0004(00)01609-1 [DOI] [PubMed] [Google Scholar]
  • 22. Siripoksup P, Cao G, Cluntun AA, et al. Sedentary behavior in mice induces metabolic inflexibility by suppressing skeletal muscle pyruvate metabolism. J Clin Invest. 2024;134(11):e167371. doi: 10.1172/JCI167371 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23. Sparks LM, Redman LM, Conley KE, et al. Effects of 12 months of caloric restriction on muscle mitochondrial function in healthy individuals. J Clin Endocrinol Metab. 2017;102(1):111‐121. doi: 10.1210/jc.2016-3211 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24. Carlessi R, Chen Y, Rowlands J, et al. GLP‐1 receptor signalling promotes β‐cell glucose metabolism via mTOR‐dependent HIF‐1α activation. Sci Rep. 2017;7(1):2661. doi: 10.1038/s41598-017-02838-2 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25. Tsuboi T, da Silva Xavier G, Holz GG, Jouaville LS, Thomas AP, Rutter GA. Glucagon‐like peptide‐1 mobilizes intracellular Ca2+ and stimulates mitochondrial ATP synthesis in pancreatic MIN6 beta‐cells. Biochem J. 2003;369(Pt 2):287‐299. doi: 10.1042/BJ20021288 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26. Kuznetsov AV, Veksler V, Gellerich FN, Saks V, Margreiter R, Kunz WS. Analysis of mitochondrial function in situ in permeabilized muscle fibers, tissues and cells. Nat Protoc. 2008;3(6):965‐976. doi: 10.1038/nprot.2008.61 [DOI] [PubMed] [Google Scholar]
  • 27. Picard M, Taivassalo T, Gouspillou G, Hepple RT. Mitochondria: isolation, structure and function. J Physiol. 2011;589(Pt 18):4413‐4421. doi: 10.1113/jphysiol.2011.212712 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 28. Saks VA, Veksler VI, Kuznetsov AV, et al. Permeabilized cell and skinned fiber techniques in studies of mitochondrial function in vivo. Mol Cell Biochem. 1998;184(1–2):81‐100. [PubMed] [Google Scholar]
  • 29. Kay L, Nicolay K, Wieringa B, Saks V, Wallimann T. Direct evidence for the control of mitochondrial respiration by mitochondrial creatine kinase in oxidative muscle cells in situ. J Biol Chem. 2000;275(10):6937‐6944. doi: 10.1074/jbc.275.10.6937 [DOI] [PubMed] [Google Scholar]
  • 30. Perry CG, Kane DA, Lin CT, et al. Inhibiting myosin‐ATPase reveals a dynamic range of mitochondrial respiratory control in skeletal muscle. Biochem J. 2011;437(2):215‐222. doi: 10.1042/BJ20110366 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31. He ZH, Bottinelli R, Pellegrino MA, Ferenczi MA, Reggiani C. ATP consumption and efficiency of human single muscle fibers with different myosin isoform composition. Biophys J. 2000;79(2):945‐961. doi: 10.1016/S0006-3495(00)76349-1 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 32. Edman S, Flockhart M, Larsen FJ, Apro W. Need for speed: human fast‐twitch mitochondria favor power over efficiency. Mol Metab. 2024;79:101854. doi: 10.1016/j.molmet.2023.101854 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 33. Luna‐Marco C, de Maranon AM, Hermo‐Argibay A, et al. Effects of GLP‐1 receptor agonists on mitochondrial function, inflammatory markers and leukocyte‐endothelium interactions in type 2 diabetes. Redox Biol. 2023;66:102849. doi: 10.1016/j.redox.2023.102849 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 34. Jastroch M, Divakaruni AS, Mookerjee S, Treberg JR, Brand MD. Mitochondrial proton and electron leaks. Essays Biochem. 2010;47:53‐67. doi: 10.1042/bse0470053 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35. Lin TK, Lin KJ, Lin HY, et al. Glucagon‐like peptide‐1 receptor agonist ameliorates 1‐methyl‐4‐phenyl‐1,2,3,6‐tetrahydropyridine (MPTP) neurotoxicity through enhancing mitophagy flux and reducing α‐synuclein and oxidative stress. Front Mol Neurosci. 2021;14:697440. doi: 10.3389/fnmol.2021.697440 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36. Nunn E, Jaiswal N, Gavin M, et al. Antibody blockade of activin type II receptors preserves skeletal muscle mass and enhances fat loss during GLP‐1 receptor agonism. Mol Metab. 2024;80:101880. doi: 10.1016/j.molmet.2024.101880 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37. Zurlo F, Larson K, Bogardus C, Ravussin E. Skeletal muscle metabolism is a major determinant of resting energy expenditure. J Clin Invest. 1990;86(5):1423‐1427. doi: 10.1172/JCI114857 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 38. Sanchez E, Lobo T, Fox JL, Zeviani M, Winge DR, Fernandez‐Vizarra E. LYRM7/MZM1L is a UQCRFS1 chaperone involved in the last steps of mitochondrial complex III assembly in human cells. Biochim Biophys Acta. 2013;1827(3):285‐293. doi: 10.1016/j.bbabio.2012.11.003 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39. Bottani E, Cerutti R, Harbour ME, et al. TTC19 plays a husbandry role on UQCRFS1 turnover in the biogenesis of mitochondrial respiratory complex III. Mol Cell. 2017;67(1):96‐105. doi: 10.1016/j.molcel.2017.06.001 [DOI] [PubMed] [Google Scholar]
  • 40. Cherian A, Divya KP, Jose J, Thomas B. Multifocal cavitating leukodystrophy‐a distinct image in mitochondrial LYRM7 mutations. Mult Scler Relat Disord. 2021;47:102615. doi: 10.1016/j.msard.2020.102615 [DOI] [PubMed] [Google Scholar]
  • 41. Ghezzi D, Arzuffi P, Zordan M, et al. Mutations in TTC19 cause mitochondrial complex III deficiency and neurological impairment in humans and flies. Nat Genet. 2011;43(3):259‐263. doi: 10.1038/ng.761 [DOI] [PubMed] [Google Scholar]
  • 42. Invernizzi F, Tigano M, Dallabona C, et al. A homozygous mutation in LYRM7/MZM1L associated with early onset encephalopathy, lactic acidosis, and severe reduction of mitochondrial complex III activity. Hum Mutat. 2013;34(12):1619‐1622. doi: 10.1002/humu.22441 [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Figure S1. Analysis of MHC and SERCA isoforms. (A) Tibialis anterior muscle embedded in OCT at 3‐week intervention was analyzed for MHC immunofluorescence to determine fiber types. (B) SERCA 1 and SERCA 2 protein expression levels were analyzed in gastrocnemius muscle by Western blotting and (C and D) quantified.

Figure S2. Mitochondrial supercomplexes in mouse gastrocnemius muscles after 1 week of the intervention. (A–F) Supercomplexes formations were analyzed in isolated mitochondria from gastrocnemius muscle by native PAGE and quantified. All data are from 1‐week semaglutide treatment. n = 5‐6 for both vehicle and semaglutide.

Figure S3. Mitochondrial lipidome in mouse gastrocnemius muscles. Mitochondrial lipidome was analyzed in isolated mitochondrial from gastrocnemius muscle of 3 weeks vehicle and semaglutide groups. (A) Heatmap of abundance of each mitochondrial phospholipidome (B) Total phospholipids from vehicle and semaglutide group (C) Absolute abundance of mitochondrial lipids (D–K) Absolute abundance of individual phospholipids species (D) Phosphocholine (PC), (E) Phosphatidylethanolamine (PE), (F) Phosphatidylinositol (PI), (G) Phosphatidylserine (PS), (H) Cardiolipin (CL), (I) Phosphatidylglycerol (PG), (J) Lysophosphatidylcholine (LPC), and (K) Lysophosphatidylethanolamine (LPE). n = 6 for both vehicle and semaglutide. **p < 0.01 significant difference vs. Veh.

OBY-33-974-s001.pdf (569.7KB, pdf)

Data S1. Supporting Information.

OBY-33-974-s002.docx (25.9KB, docx)

Data Availability Statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Articles from Obesity (Silver Spring, Md.) are provided here courtesy of Wiley

RESOURCES