The Effects achand Menisms of Electroacupuncture Combined with Exercise Training on T2DM Sarcopenia Based on the Omi/HtrA2 Autophagy Pathway

The Effects achand Menisms of Electroacupuncture Combined with Exercise Training on T2DM Sarcopenia Based on the Omi/HtrA2 Autophagy Pathway | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article The Effects achand Menisms of Electroacupuncture Combined with Exercise Training on T2DM Sarcopenia Based on the Omi/HtrA2 Autophagy Pathway Zhilin Xie, Qingxia Shen, Kai Yang, Yingmei Fu, Fang Liu, Tong Chen, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5762572/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Purpose: To explore the effects and mechanisms of electroacupuncture combined with exercise training on sarcopenia in mice with type 2 diabetes mellitus (T2DM). Methods: C57bcs-db mice were randomly divided into 5 groups: control group, model group, electroacupuncture group, exercise group, electroacupuncture + running group. Except for the control group, the other four groups were subcutaneously injected with dexamethasone solution to establish a T2DM sarcopenia model. After successful modeling, the mice in the running group were trained on the treadmill for 5 weeks. Mice in electroacupuncture group were connected to electrotherapy apparatus for 5 weeks. Mice in the electroacupuncture + running group were treated with both running and electroacupuncture. The grasping power, body weight and wet weight of both gastrocnemius of mice were measured. The morphology and structure of gastrocnemius were observed by HE staining. mRNA expressions of omi, Hax-1, LC3 and Becline were detected by RT-PCR, and the expression of related autophagy molecular proteins were detected by Western blot. Results: After the modeling, all the mice reached the successful criteria for modeling of sarcopenia. After intervention, the hind leg grip of mice in electroacupuncture group, running group and electroacupuncture + running group was significantly increased. The wet weight of gastrocnemius muscle in running + electroacupuncture group was significantly higher than that in model group, and the degree of muscle fiber atrophy of gastrocnemius muscle was significantly reduced. LC3mRNA level and omi/Htra2 protein expression level were significantly decreased in electroacupuncture + running group. Conclusion: The evidence provided by this study suggests that dexamethasone induces sarcopenia in T2DM mice. Electroacupuncture combined with exercise improves grip strength, skeletal muscle wet weight, and muscle fiber atrophy in mice, and reduces the expression of autophagy-related mRNA and proteins, indicating that electroacupuncture combined with exercise may be an effective treatment for T2DM-related sarcopenia. Type 2 diabetes mellitus (T2DM) sarcopenia autophagy exercise training electroacupuncture Omi/HtrA2 Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Introduction According to the latest data from the 10th edition of the IDF Global Diabetes Map released by the International Diabetes Federation (IDF)[ 1 ], the number of adults with diabetes worldwide was approximately 537 million in 2021, and this figure is projected to rise to 783 million by 2045. Type 2 diabetes mellitus is a group of chronic, progressive diseases characterized by impaired glucose metabolism. The disease has a high prevalence, and moderate exercise plays a crucial role in maintaining stable blood glucose levels. The currently accepted approach to managing T2DM includes adopting a healthy lifestyle, which involves dietary planning, physical activity, and weight control[ 1 ]. Sarcopenia is one of the common complications in patients with diabetes mellitus, severely impacting their quality of life and leading to disabilities such as falls, fractures, loss of independence, and increased risk of death[ 2 – 5 ]. Sarcopenia, primarily characterized by a decrease in skeletal muscle mass and function[ 2 ], is recognized as a common risk factor in diabetes mellitus[ 4 ]. Older adults with T2DM experience a faster decline in skeletal muscle mass and strength compared to non-diabetic individuals[ 6 ], and are more likely to develop sarcopenia. Given the high prevalence and detrimental effects of sarcopenia in diabetic patients, enhancing muscle mass and function is particularly important in the clinical management of this population[ 4 ]. Physical activity, particularly resistance training, has been shown to be effective in preventing muscle atrophy and improving muscle strength[ 7 ], benefiting both upper and lower extremity muscles[ 8 ]. Data from Roger[ 9 ]further suggest that cellular senescence contributes to age-related functional decline, and physical activity may mitigate this process. However, in clinical practice, some patients, particularly those who are elderly or obese, struggle to perform active resistance training, highlighting an urgent need for alternative treatments, such as passive training or medications, for sarcopenia. Acupuncture is a well-established traditional therapy for neuromuscular disorders. Electroacupuncture (EA) has been shown to improve oxidative stress and inflammation, maintaining mitochondrial function, though its effects and mechanisms on sarcopenia remain unclear. Studies have shown that EA promotes IGF-1 expression, inhibits myostatin, and stimulates the proliferation of skeletal muscle satellite cells, thereby repairing denervated skeletal muscle and delaying muscle atrophy[ 10 ]. Guo et al.[ 11 ] found that after EA intervention, oxidative stress, apoptosis, and levels of IL-6, TNF-α, Atrogin-1, and MuRF1 were suppressed, leading to reduced mitochondrial damage and improved muscle function. Peripheral nerve stimulation through acupuncture or EA has been shown to effectively promote recovery from neuromuscular dysfunction. Autophagy, a lysosome-dependent catabolic process, plays a key role in maintaining cellular homeostasis under stress. Impaired autophagy may contribute to various diabetes-related metabolic disorders, including sarcopenia[ 12 ]. It has been suggested that the pathogenesis of sarcopenia is related to autophagy and can be improved by pathways such as tea polyphenols and histone deacetylase, which enhance metabolism and mitochondrial dynamics[ 13 , 14 ]. Serine protease Omi, a member of the HtrA (High Temperature Requirement A) family, is a protein associated with heat stress, also known as HtrA2. Omi/HtrA2 protease regulates various cellular processes through interactions with multiple substrate proteins in the cytoplasm and mitochondria[ 15 ]. Recent studies[ 16 ]have demonstrated that Omi/HtrA2 regulates autophagy upstream of the core autophagy protein Beclin1 by cleaving Hax-1, a Bcl-2 family-associated protein, relieving its inhibition of Beclin1, and ultimately inducing autophagy. HtrA2/Omi protease deficiency has been linked to denervation-independent sarcopenia[ 17 ], while its absence disrupts mitochondrial protein homeostasis, induces mitochondrial stress responses (UPRmt), and inhibits myogenic differentiation of C2C12 myoblasts, leading to skeletal muscle degeneration in mnd2 mice[ 18 ]. Therefore, autophagy plays a critical role in maintaining skeletal muscle homeostasis. In this study, we employed interventions including exercise, electroacupuncture, and their combination to observe changes in muscle strength, muscle mass, and autophagy in skeletal muscle cells of T2DM mice with sarcopenia. We also analyzed the gene and protein expression of the Omi/HtrA2-induced autophagy pathway to provide a basis for further research and treatment of T2DM-related sarcopenia. Materials and Methods Experimental Animals and Grouping A total of 9 male 4-week-old C57BKS-DB mice, weighing 24.73 ± 1.95 g, were purchased from Jiangsu Jicui YaoKang Biotechnology Co., Ltd., with license number SCXK (Su) 2018-0008. The mice were housed under the national standard conditions for rodent laboratory animals: a constant room temperature of 23 ± 2°C, humidity of 40–45%, and a 12-hour light/dark cycle, with free access to water and food. After adaptive feeding, the mice were fed a normal diet for 3 weeks. They were weighed, their blood glucose levels and hindlimb grip strength were measured, and they were randomly divided into five groups: control group (n = 3), model group (n = 3), electroacupuncture group (n = 3), running group (n = 3), and electroacupuncture + running group (n = 3). Based on blood glucose levels, insulin was administered via intraperitoneal injection at a dose of 0.5-1 U/kg of short-acting insulin to maintain blood glucose at 20 ± 3 mmol/L. All animal experiments were conducted in accordance with Chinese legislation on animal management and general recommendations. The study was approved by the Ethics Committee of the First Affiliated Hospital of Hainan Medical University. Construction of a Mouse Model of Type 2 Diabetes Mellitus Combined with Sarcopenia Except for the control group, the other mice were used to construct a model of type 2 diabetes mellitus (T2DM) combined with sarcopenia. A fresh dexamethasone phosphate sodium solution (5 mg/mL) was diluted 10 times by adding 1 mL of the dexamethasone solution to 9 mL of 0.9% saline, resulting in a final concentration of 0.5 mg/mL. This freshly prepared dexamethasone solution was administered subcutaneously at a dose of 5 mg/kg for 8 weeks to induce the T2DM and sarcopenia model. The success of the model was confirmed when blood glucose levels reached ≥ 16.7 mmol/L, hindlimb grip strength decreased by 25%, and both muscle mass and function were significantly reduced. Intervention Methods After the successful establishment of the model, experimental interventions were carried out. Mice in the electroacupuncture group were fixed, and 0.25 mm × 13 mm disposable sterile acupuncture needles were used to puncture two acupoints: Pishu (BL20) and Zusanli (ST36). The needles were connected to a KWD-808II multifunctional pulse therapy instrument, with sparse-dense waves (frequency of 1 Hz) and the current intensity adjusted until slight muscle twitching occurred. This treatment was performed every other day, with each session lasting 20 minutes, for 3 days a week and resting 1 day for a total of 5 weeks. The running group was subjected to treadmill training, with the treadmill set at a 0° incline and a speed of 16 m/min for 20 minutes per session. The mice exercised for 3 days a week and rested for 1 day for 5 weeks. The electroacupuncture + running group underwent both electroacupuncture and exercise interventions, following the same protocol as the respective groups. Hindlimb Grip Strength Measurement The grip strength of the mice was measured once per week. Before testing, the mice were allowed to adapt to the grip strength meter (DS2-50N) for 30 seconds. The meter was set to measurement mode, and once the mice had fully grasped the grid with all four paws, their tails were gently pulled backward until the grip was released. The highest value of three measurements, expressed in newtons (N), was recorded as the grip strength for each mouse. Body Weight and Blood Glucose Measurement During the experiment, the body weight and blood glucose of the mice were measured once a week at the same time. Blood samples were collected from the tail after disinfection, and blood glucose was measured using a glucometer (Abbott). No fasting or water restrictions were imposed before measurement. Animal Sacrifice and Tissue Collection At the end of the experiment, body weight, blood glucose, and grip strength were measured before the mice were sacrificed. The mice were euthanized by cervical dislocation, and sterile ophthalmic scissors and forceps were used to dissect the skin and tissue of both hindlimbs. The complete gastrocnemius muscle from one side was placed in a 1.5 mL cryotube and immediately frozen in liquid nitrogen. After all the tissues were collected, they were stored in liquid nitrogen. The gastrocnemius muscle from the other side was placed in a 50 mL centrifuge tube containing 30 mL of 4% paraformaldehyde and stored in a cool, dark place. Skeletal Muscle Cross-Section Observation The embedded blocks containing skeletal muscle tissues were dehydrated by immersion in 50% ethanol and 60% ethanol for 2 hours, then further dehydrated using a tissue processor. The tissues were cleared in xylene for 5 minutes and immersed in molten paraffin. The tissues were embedded using a tissue embedding machine, and liquid paraffin was slowly added. After solidification on an ice platform, the paraffin blocks were sectioned at a thickness of approximately 5 µm using a microtome. The sections were flattened in a water bath and transferred to adhesive slides. The slides were labeled and baked overnight. Standard H&E staining was performed, and the sections were observed under a microscope at 20x magnification. RNA Extraction and qRT-PCR Detection Total RNA from tissues was extracted using the Eastep Super Total RNA Extraction Kit (Promega LS1040). For each sample, total RNA was reverse transcribed into cDNA using the SuperMix for qPCR (gDNA digester plus) (YEASEN H7101160) reverse transcription kit, according to the manufacturer's instructions, for quantitative analysis. The denaturation temperature was set at 95°C for 10 seconds, and the annealing and extension temperature was 60°C for 30 seconds, for a total of 40 cycles. The gene expression was normalized to ACTB using the ΔΔCt method to analyze relative gene expression changes. The primers used were as follows: Omi/HtrA2 (5’-ATCTCAAACGGATCAGGATTCGT-3’, 5’-CAGCCTCACTCGTACTCGG-3’); Hax-1(5’-CGAGGCTTTTTCGGCTTTCC-3’,5’-GCATAGCTCTCTCGACCCCA-3’);Beclin1(5’-GGCGGCTCCTATTCCATCAA-3’,5’-GTCCACTGCTCCTCAGAGTTA-3’);LC3(5’-GACCGCTGTAAGGAGGTGC-3’,5’-CTTGACCAACTCGCTCATGTTA-3’);and ACTB(5’-GGCTGTATTCCCCTCCATCG-3’,5’-CCAGTTGGTAACAATGCCATGT-3’). The expression levels of the target genes were normalized to ACTB, and the data were presented as fold changes relative to the cycle threshold (Ct) values. The relative expression levels of Omi/Htr A2, Hax-1, Beclin1, and LC3 were analyzed. Western Blot Detection The tissues were homogenized in a mortar with lysis buffer (PMSF) and repeatedly frozen and thawed in liquid nitrogen, followed by lysis on ice for 15 minutes. The samples were then centrifuged at 12,000 rpm at 4°C for 30 minutes. The supernatant (2 µL) was taken to measure protein concentration using the BCA method. The remaining protein samples were separated on an SDS-PAGE gel and transferred to PVDF membranes. The membranes were blocked with 5% skim milk, then incubated overnight at 4°C with the following primary antibodies: Beclin 1 (Abcam, ab62557), HtrA2/Omi (Abcam, ab75982), Hax-1 (Abcam, ab137613), and LC3B (Abcam, ab48394), with the dilution ratio of LC3B at 1:2000, and the other primary antibodies at 1:1000. β-Actin (Beyotime, AF5003) was used as an internal control at a dilution ratio of 1:1000. All bands were visualized using a chemiluminescent ECL kit (Thermo Fisher), and band intensity was analyzed using the TanonImage software. Statistical Analysis Data were analyzed using SPSS 24.0 software. Mouse modeling data were presented as mean ± standard deviation (SD), and paired t-tests were used to compare data before and after treatments within the same group. For data with normal distribution, one-way analysis of variance (ANOVA) was used for between-group comparisons, while the Kruskal-Wallis test was applied for data that did not follow a normal distribution. P < 0.05 was considered a statistically significant difference. Results Construction of the Mouse Model of Type 2 Diabetes Mellitus Combined with Sarcopenia Before the dexamethasone intervention, the mice underwent 1–2 weeks of adaptive feeding, during which their body weight and grip strength increased. All groups of mice were in good condition, with shiny fur and responsive behaviors. From the 3rd week, the mice were administered dexamethasone at 5 mg/kg to induce the model. The model mice exhibited symptoms such as excessive drinking, reduced food intake, polyuria, and slow weight gain, and some even experienced weight loss. Gradually, their fur became dull and rough, and they displayed irritability, fatigue, and irregular bowel movements. From the 4th week onward, grip strength began to decline. Blood glucose levels were measured, and a glucose level ≥ 16.7 mmol/L was observed. By the 10th week, the muscle mass and function of the modeled mice had significantly decreased, with hindlimb grip strength reduced by 25%. Although grip strength decreased in the non-modeled mice, it did not reach the 25% threshold, indicating that the model of type 2 diabetes mellitus combined with sarcopenia had been successfully established. Comparison of Grip Strength, Body Weight, and Skeletal Muscle Wet Weight Among Different Groups of Mice After the model construction was completed, interventions were administered to the type 2 diabetes mellitus (T2DM) mice with sarcopenia in the 11th week. After 5 consecutive weeks of treatment, the grip strength of the mice gradually began to recover. There was no significant difference in grip strength in the control group. In contrast, the model group showed significantly lower grip strength at the 15th week compared to before the intervention (P < 0.05). However, in the running, electroacupuncture, and electroacupuncture combined with running groups, the grip strength at the 15th week was significantly higher than at the 10th week. The results indicated that the grip strength of sarcopenic mice further declined, but symptoms were alleviated after interventions such as exercise, electroacupuncture, or the combination of electroacupuncture and exercise. The electroacupuncture group showed a more pronounced effect compared to the running group. Before the intervention, there were no statistically significant differences in grip strength among the model, running, electroacupuncture, and running + electroacupuncture groups. However, there was a significant difference between the control and model groups, as well as between the model group and the other intervention groups (Table 1 , Fig. 1 a). Furthermore, it was observed that in the control and intervention-treated mice, body weight began to stabilize and increase slowly, while body weight in the model group continued to decline, though not significantly (Table 2 , Fig. 1 b). After the mice were sacrificed, the bilateral gastrocnemius muscles were weighed. Compared to the control group, the skeletal muscle wet weight in the model group was significantly lower. Compared with the model group, the running group, the electroacupuncture group, and the running + electroacupuncture group showed significantly increased wet weight of skeletal muscle. (Fig. 2 ). Table 1 Comparison of gripping power of mice before and after intervention Group (mean ± standard deviation) Mouse Grip Strength (N) Control group(n = 3) Model group (n = 3) Running group (n = 3) Electroacupuncture group (n = 3) Running + electroacupuncture group (n = 3) Pre-intervention 2.56 ± 0.18 1.20 ± 0.09 1.04 ± 0.22 1.02 ± 0.11 1.11 ± 0.03 Post-intervention 2.57 ± 0.10 1.13 ± 0.11*### 1.77 ± 0.66*^^^ 1.67 ± 0.10***^^^ 1.70 ± 0.04*^^^ Data are shown as means ± SEM. * indicates P < 0.05 for post-intervention compared to pre-intervention grip strength for each group, and ** indicates P < 0.01 for post-intervention compared to pre-intervention grip strength for each group, *** indicates P < 0.001 for post-intervention compared to pre-intervention grip strength for each group. ### indicates P < 0.001 for grip strength between control group and model group after intervention. ^^^represents P < 0.001 between the intervention groups and the model group after intervention. Table 2 Comparison of gripping power of mice before and after intervention Group (mean ± standard deviation) Mouse Weight (g) Control group(n = 3) Model group (n = 3) Running group (n = 3) Electroacupuncture group (n = 3) Running + electroacupuncture group (n = 3) Pre-intervention 54.78 ± 0.47 43.36 ± 12.65 41.13 ± 9.65 30.37 ± 1.54 38.17 ± 3.61 Post-intervention 58.92 ± 1.55 36.53 ± 9.50 46.85 ± 12.85 33.25 ± 1.68 39.85 ± 3.19 Data are shown as means ± SEM. Morphological Changes in Skeletal Muscle Tissue of Mice HE staining results of skeletal muscle in each group showed that in the control group, muscle fibers in the cross-sections appeared polygonal and uniformly sized. The muscle cell nuclei were distributed along the periphery of the fibers, with intact nuclear membranes, and the muscle fibers were arranged in an orderly manner. In the model group, the cross-sections revealed irregularly shaped, atrophied, and deformed muscle fibers with a substantial infiltration of inflammatory cells. Several muscle cells exhibited nuclear migration towards the interior, showing clear multinucleation, and the muscle fibers were arranged disorderly. The running and electroacupuncture groups exhibited similar patterns, with a reduction in inflammatory cell infiltration and nuclear migration compared to the model group. In the running + electroacupuncture group, the cross-sections showed a marked reduction in inflammatory cell infiltration and nuclear migration compared to the model group. Moreover, some muscle fibers showed improvement in morphology, approaching a normal state (Fig. 3 ). Comparison of Omi/HtrA2, Hax-1, Beclin1, and LC3 mRNA Levels in Skeletal Muscle Tissue of Each Group We used qPCR to evaluate the effect of exercise and electroacupuncture on autophagy levels in sarcopenic mice. Compared with the control group, the LC3 mRNA level was significantly increased in the model group. Compared with the model group, the LC3 mRNA level in the running group + electroacupuncture group showed a downward trend (P = 0.056) (Fig. 4 c). A similar trend was observed in the qPCR results for Beclin1, a gene that reflects autophagy levels (Fig. 4 d). In the results for Omi, the expression levels in all intervention groups showed a decreasing trend compared to the model group (Fig. 4 a). For Hax-1, the results showed a decreasing trend in Hax-1 gene expression in the model group compared to the control group. In contrast, the expression levels of Hax-1 in the running, electroacupuncture, and running + electroacupuncture groups showed an increasing trend compared to the model group (Fig. 4 b). These trends suggest that autophagy levels are higher in mice with muscle wasting syndrome compared to normal mice, and that exercise, acupuncture, or a combination of both can reduce autophagy levels in diabetic mice with muscle wasting syndrome. Comparison of Omi/HtrA2, Hax-1, Beclin1, and LC3Ⅱ/LC3Ⅰ Protein Levels in Skeletal Muscle Tissue of Each Group The results from the Western blot (WB) experiments showed that, compared to the control group, the HtrA2/Omi protein levels were significantly elevated in the model group. In contrast, the HtrA2/Omiprotein levels were significantly reduced in the electroacupuncture + running group compared to the model group (Fig. 5a). Regarding Hax-1protein, the results indicated a decreasing trend in the model group compared to the control group. However, the Hax-1 protein levels in all intervention groups showed an increasing trend compared to the model group (Fig. 5b). As for the Beclin1protein levels, there was an increasing trend in the model group compared to the control group (Fig. 5d). In contrast, the protein levels in the intervention groups showed a decreasing trend compared to the model group. Figure 5 Western blot analysis of Omi/HtrA2, Hax-1, LC3II/LC3Ⅰ, and Beclin1 in mouse muscle tissues(n = 3). Data are shown as means ± SEM. * P < 0.05 vs. model group, # P < 0.05 vs. control group. Discussion Currently, the treatment methods and mechanisms for sarcopenia are still under exploration. Using sarcopenic patients as test subjects presents severe ethical concerns[ 19 ], making it necessary to conduct treatment-related research using animal models. At present, there is no internationally established standard for constructing sarcopenia animal models[ 19 ]. To study the treatment and mechanisms of T2DM combined with sarcopenia, we induced muscle weakness in db/db mice using dexamethasone. The grip strength of the modeled mice decreased by 25%, meeting the criteria for sarcopenia modeling, while the control group showed no significant decrease in grip strength. In addition to reduced grip strength, HE staining under electron microscopy showed that the skeletal muscle fibers of dexamethasone-induced mice were atrophied and deformed compared to normal mice, with reduced cross-sectional areas of the muscle fibers. This suggests that dexamethasone can accelerate the onset of sarcopenia in diabetic mice, providing new data and ideas for constructing sarcopenia animal models. Exercise training, particularly resistance training, is considered an effective method to prevent muscle atrophy[ 20 ] and improve muscle strength[ 7 , 21 , 22 ], benefiting both upper and lower limb strength[ 8 ]. In this study, treadmill training was used to improve muscle strength in T2DM mice. The results showed that after exercise intervention, the grip strength of the mice increased significantly, while the grip strength of the untreated model group continued to decline. This suggests that exercise training has a positive therapeutic effect on T2DM combined with sarcopenia. Our experimental results are supported by the study of Serra-Rexach et al.[ 23 ], who conducted resistance training for 8 weeks in elderly individuals over 90 years old. The training intensity increased from 30–70% of one repetition maximum (1RM), resulting in a 10.6 kg increase in leg press 1RM, improved muscle strength, and reduced fall risk. High-intensity resistance training (80% 1RM) was more effective in improving sarcopenia-related muscle weakness compared to low-intensity resistance training (≤ 50% 1RM). Snijders [ 24 ] conducted 24 weeks of supervised resistance training for elderly individuals, significantly improving their skeletal muscle mass and strength. One year after stopping treatment, muscle mass and strength declined. A meta-analysis[ 25 ]suggested that supervised resistance training is effective in improving muscle strength in the elderly. However, many clinical T2DM sarcopenia patients cannot tolerate strict exercise training due to obesity or aging. Furthermore, the effectiveness of drugs for treating T2DM sarcopenia remains controversial, with some hypoglycemic drugs having adverse effects on muscles[ 4 ]. Therefore, passive treatments in traditional Chinese medicine, such as acupuncture, electroacupuncture, and electrical stimulation, combined with exercise training, are of great significance for treating sarcopenia patients. To explore whether acupuncture, as a passive treatment, has a positive effect on diabetic sarcopenia, we conducted interventions with exercise, electroacupuncture, and their combination in mice. The results showed significant improvements in grip strength in all intervention groups, particularly in the electroacupuncture and combined treatment groups. This indicates that exercise, electroacupuncture, and their combination can enhance muscle strength in T2DM sarcopenic mice. Moreover, skeletal muscle wet weight in the combined treatment group was significantly higher than in the model group, demonstrating that exercise and electroacupuncture interventions have a positive effect on muscle mass in T2DM sarcopenia. T. Brock Symons[ 26 ]used casting to induce muscle atrophy in rats, followed by treatment with acupuncture, electroacupuncture, and electrical stimulation. The results showed that these treatments reduced the negative effects of casting and slowed the degradation of muscle protein. The findings are consistent with our results. Animal models in mice indicate that the absence of mnd2 (the mouse homolog of HtrA2) results in neuronal degeneration and muscle atrophy, ultimately leading to shortened lifespan. This suggests that Omi/HtrA2 is essential for normal cell survival. Hax-1 is a substrate of Omi/HtrA2, involved in apoptosis, autophagy, cell migration, and mRNA transport[ 27 ]Under apoptotic stimuli, Omi/HtrA2strongly cleaves Hax-1, triggering early apoptosis. Recent studies show that Omi/HtrA2regulates autophagy upstream of Beclin1, by cleaving Hax-1 (HS1-associated protein X-1) and relieving its inhibition of Beclin1, ultimately inducing autophagy[ 16 ]. Studies have reported that the lack of HtrA2/Omiprotease activity induces non-denervation-dependent muscle atrophy[ 17 ]. Additionally, HtrA2 protease deficiency leads to mitochondrial protein imbalance, inducing UPRmt and inhibiting myogenic differentiation of C2C12 myoblasts. Mice deficient in HtrA2 protease activity showed a distinct muscular dystrophy phenotype[ 18 ]. In our study, HtrA2/Omi protein levels were significantly elevated in T2DM sarcopenic mice and decreased after exercise and electroacupuncture treatments. In our study, HtrA2/Omi protein levels were significantly elevated in T2DM sarcopenic mice and decreased after exercise and electroacupuncture treatments. The qPCR results for LC3 also showed a similar trend, with higher LC3 levels in T2DM sarcopenic mice, which significantly decreased after electroacupuncture combined with exercise intervention. This suggests that the autophagy level induced by the HtrA2/OMI autophagy pathway is elevated in T2DM sarcopenic mice, and the combination of electroacupuncture and exercise can reduce this autophagy. Some limitations exist in this study. Although certain autophagy proteins showed a decreasing trend after the intervention, there was no statistically significant difference. This may be due to factors such as obesity and reduced muscle mass in the T2DM sarcopenic mice, leading to poor performance in active training. Additionally, due to the death of some mice after model establishment, the sample size was reduced, potentially affecting the results for autophagy-related genes and proteins such as HtrA2/Omi, Hax-1, LC3, and Beclin1. Future studies should aim to increase the sample size and improve the representativeness of the data. Moreover, muscle mass and cross-sectional areas were not quantitatively analyzed in this study. Establishing uniform criteria for sarcopenia mouse models will also be crucial for future research. Conclusion The evidence provided by this study suggests that exercise and electroacupuncture therapy improve muscle function and mass in T2DM sarcopenic mice by modulating the HtrA2/Omi autophagy pathway. These therapies reduce the mRNA and protein expression levels of autophagy-related factors, alleviating autophagy-induced muscle damage. The combination of active exercise therapy and passive electroacupuncture therapy offers a new direction for treating diabetic sarcopenia, providing a solid theoretical and experimental foundation for the clinical application of integrative traditional Chinese and Western medicine, with broad potential for development. Declarations Funding This study was supported by Natural Science Foundation of Hainan Province of China (No. 821RC756) and The New Medical Technology Research and Transformation Seed Program of Shanghai Municipal Health Commission,.Study on multimodal rehabilitation detection and treatment of patients with hemiplegia based on portable intelligent devices (No.2024ZZ1030). Conflict of interest The authors have no relevant financial or non-financial interests to disclose. Author contributions Zhilin Xie: Collection and assembly of data, data analysis and interpretation, manuscript writing, final approval of manuscript. Jing Zhang: Conception and design, final approval of manuscript. Xiaoliu Li: Collect lower limb electromyographic signals, electromyographic stimulation is used to recruit muscles. Qingxia Shen, Kai Yang: Assist in literature reviewand, collection and assembly of data, final approval of manuscript. Yingmei Fu: Assist in literature reviewand, final approval of manuscript. Tong Chen: Design of experimental scheme of TCM acupuncture and moxibustion, final approval of manuscript. Fang Liu: Acupuncture point selection, final approval of manuscript. All authors reviewed the manuscript. Acknowledgements We thank Dr Man Xizo from the school of basic medicine and life sciences of Hainan Medical University for her guidance in the design of the experiment and her assistance in preparing this manuscript, and Professor Yiqiang Xie from the the College of Traditional Chinese Medicine of Hainan Medical University for his guidance preparing this the manuscript. Ethical approval The animal experiments were approved by the Ethics Committee of Hainan Medical University (NoHYLL-2021-173). References A. Kumar, R. Gangwar, A.A. Zargar, R. Kumar,A. Sharma, Prevalence of Diabetes in India: A Review of IDF Diabetes Atlas 10th Edition. Curr Diabetes Rev. 20 , e130423215752 (2024). https://doi.org/10.2174/1573399819666230413094200 A.J. Cruz-Jentoft, G. Bahat, J. Bauer, Y. Boirie, O. Bruyere, T. Cederholm, C. Cooper, F. Landi, Y. Rolland, A.A. Sayer, S.M. Schneider, C.C. Sieber, E. Topinkova, M. Vandewoude, M. Visser, M. Zamboni, P. 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Yoo, Proportion and Characteristics of the Subjects with Low Muscle Mass and Abdominal Obesity among the Newly Diagnosed and Drug-Naive Type 2 Diabetes Mellitus Patients. Diabetes Metab J. 43 , 105-113 (2019). https://doi.org/10.4093/dmj.2018.0036 F. Giallauria, A. Cittadini, N.A. Smart,C. Vigorito, Resistance training and sarcopenia. Monaldi Arch Chest Dis. 84 , 738 (2016). https://doi.org/10.4081/monaldi.2015.738 R. Borde, T. Hortobagyi,U. Granacher, Dose-Response Relationships of Resistance Training in Healthy Old Adults: A Systematic Review and Meta-Analysis. Sports Med. 45 , 1693-1720 (2015). https://doi.org/10.1007/s40279-015-0385-9 R.A. Fielding, E.J. Atkinson, Z. Aversa, T.A. White, A.A. Heeren, M.M. Mielke, S.R. Cummings, M. Pahor, C. Leeuwenburgh,N.K. LeBrasseur, Biomarkers of Cellular Senescence Predict the Onset of Mobility Disability and Are Reduced by Physical Activity in Older Adults. J Gerontol A Biol Sci Med Sci. 79 , (2024). https://doi.org/10.1093/gerona/glad257 X.Q. Huang, J.S. Xu, X.R. Ye,X. Chen, Wide Pulse Width Electroacupuncture Ameliorates Denervation-Induced Skeletal Muscle Atrophy in Rats via IGF-1/PI3K/Akt Pathway. Chin J Integr Med. 27 , 446-454 (2021). https://doi.org/10.1007/s11655-021-2865-0 F. Guo, L. Fu,Z. Lu, Effect of electroacupuncture combined with sulforaphane in the treatment of sarcopenia in SAMP8 mice. Iran J Basic Med Sci. 27 , 560-566 (2024). https://doi.org/10.22038/IJBMS.2024.71345.15509 M. Kitada,D. Koya, Autophagy in metabolic disease and ageing. Nat Rev Endocrinol. 17 , 647-661 (2021). https://doi.org/10.1038/s41574-021-00551-9 S. Chen, X. Wang, C. Zhang, L. Xue, W. Feng, H. Xie, L. Cheng, C. Lyu, X. Li,H. Zhao, [Mechanism of tea polyphenols improving the sarcopenia in the aged type 2 diabetes model rats via mitochondrial quality control]. Wei Sheng Yan Jiu. 53 , 540-546 (2024). https://doi.org/10.19813/j.cnki.weishengyanjiu.2024.04.004 T.W. Lee, H.W. Liu, Y.F. Lin, T.I. Lee, Y.H. Kao,Y.J. Chen, Histone deacetylase inhibition improves metabolism and mitochondrial dynamics: A potential novel therapeutic strategy for sarcopenia coexisting with diabetes mellitus. Med Hypotheses. 158 , 110724 (2021). https://doi.org/10.1016/j.mehy.2021.110724 X.J. Su, L. Huang, Y. Qu,D. Mu, Progress in research on the role of Omi/HtrA2 in neurological diseases. Rev Neurosci. 30 , 279-287 (2019). https://doi.org/10.1515/revneuro-2018-0004 B. Li, Q. Hu, H. Wang, N. Man, H. Ren, L. Wen, N. Nukina, E. Fei,G. Wang, Omi/HtrA2 is a positive regulator of autophagy that facilitates the degradation of mutant proteins involved in neurodegenerative diseases. Cell Death Differ. 17 , 1773-1784 (2010). https://doi.org/10.1038/cdd.2010.55 H. Zhou, D. Yuan, W. Gao, J. Tian, H. Sun, S. Yu, J. Wang,L. Sun, Loss of high-temperature requirement protein A2 protease activity induces mitonuclear imbalance via differential regulation of mitochondrial biogenesis in sarcopenia. IUBMB Life. 72 , 1659-1679 (2020). https://doi.org/10.1002/iub.2289 H. Sun, L. Shen, P. Zhang, F. Lin, J. Ma, Y. Wu, H. Yu,L. Sun, Inhibition of High-Temperature Requirement Protein A2 Protease Activity Represses Myogenic Differentiation via UPRmt. Int J Mol Sci. 23 , (2022). https://doi.org/10.3390/ijms231911761 W.Q. Xie, M. He, D.J. Yu, Y.X. Wu, X.H. Wang, S. Lv, W.F. Xiao,Y.S. Li, Mouse models of sarcopenia: classification and evaluation. J Cachexia Sarcopenia Muscle. 12 , 538-554 (2021). https://doi.org/10.1002/jcsm.12709 M.S. Fragala, E.L. Cadore, S. Dorgo, M. Izquierdo, W.J. Kraemer, M.D. Peterson,E.D. Ryan, Resistance Training for Older Adults: Position Statement From the National Strength and Conditioning Association. J Strength Cond Res. 33 , 2019-2052 (2019). https://doi.org/10.1519/JSC.0000000000003230 S. Vikberg, N. Sorlen, L. Branden, J. Johansson, A. Nordstrom, A. Hult,P. Nordstrom, Effects of Resistance Training on Functional Strength and Muscle Mass in 70-Year-Old Individuals With Pre-sarcopenia: A Randomized Controlled Trial. J Am Med Dir Assoc. 20 , 28-34 (2019). https://doi.org/10.1016/j.jamda.2018.09.011 C. Hurst, S.M. Robinson, M.D. Witham, R.M. Dodds, A. Granic, C. Buckland, S. De Biase, S. Finnegan, L. Rochester, D.A. Skelton,A.A. Sayer, Resistance exercise as a treatment for sarcopenia: prescription and delivery. Age Ageing. 51 , (2022). https://doi.org/10.1093/ageing/afac003 J.A. Serra-Rexach, N. Bustamante-Ara, M. Hierro Villaran, P. Gonzalez Gil, M.J. Sanz Ibanez, N. Blanco Sanz, V. Ortega Santamaria, N. Gutierrez Sanz, A.B. Marin Prada, C. Gallardo, G. Rodriguez Romo, J.R. Ruiz,A. Lucia, Short-term, light- to moderate-intensity exercise training improves leg muscle strength in the oldest old: a randomized controlled trial. J Am Geriatr Soc. 59 , 594-602 (2011). https://doi.org/10.1111/j.1532-5415.2011.03356.x T. Snijders, M. Leenders, L. de Groot, L.J.C. van Loon,L.B. Verdijk, Muscle mass and strength gains following 6 months of resistance type exercise training are only partly preserved within one year with autonomous exercise continuation in older adults. Exp Gerontol. 121 , 71-78 (2019). https://doi.org/10.1016/j.exger.2019.04.002 A. Lacroix, T. Hortobagyi, R. Beurskens,U. Granacher, Effects of Supervised vs. Unsupervised Training Programs on Balance and Muscle Strength in Older Adults: A Systematic Review and Meta-Analysis. Sports Med. 47 , 2341-2361 (2017). https://doi.org/10.1007/s40279-017-0747-6 T. Brock Symons, J. Park, J.H. Kim, E.H. Kwon, J. Delacruz, J. Lee, Y. Park, E. Chung,S. Lee, Attenuation of skeletal muscle atrophy via acupuncture, electro-acupuncture, and electrical stimulation. Integr Med Res. 12 , 100949 (2023). https://doi.org/10.1016/j.imr.2023.100949 L. Cilenti, M.M. Soundarapandian, G.A. Kyriazis, V. Stratico, S. Singh, S. Gupta, J.V. Bonventre, E.S. Alnemri,A.S. Zervos, Regulation of HAX-1 anti-apoptotic protein by Omi/HtrA2 protease during cell death. J Biol Chem. 279 , 50295-50301 (2004). https://doi.org/10.1074/jbc.M406006200 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5762572","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":398882124,"identity":"c3e0a91c-89aa-4f20-ab16-24e68daf8c31","order_by":0,"name":"Zhilin Xie","email":"","orcid":"","institution":"The First Affiliated Hospital of Hainan Medical University","correspondingAuthor":false,"prefix":"","firstName":"Zhilin","middleName":"","lastName":"Xie","suffix":""},{"id":398882125,"identity":"3fe7408c-6458-45d4-b193-7211c68c91cf","order_by":1,"name":"Qingxia Shen","email":"","orcid":"","institution":"The First Affiliated Hospital of Hainan Medical University","correspondingAuthor":false,"prefix":"","firstName":"Qingxia","middleName":"","lastName":"Shen","suffix":""},{"id":398882126,"identity":"8dd76cb8-5fba-4653-b97a-15d4f60c9135","order_by":2,"name":"Kai Yang","email":"","orcid":"","institution":"The First Affiliated Hospital of Hainan Medical University","correspondingAuthor":false,"prefix":"","firstName":"Kai","middleName":"","lastName":"Yang","suffix":""},{"id":398882127,"identity":"cfcfc921-ca8c-4ddb-9d10-c28b960987fc","order_by":3,"name":"Yingmei Fu","email":"","orcid":"","institution":"The First Affiliated Hospital of Hainan Medical University","correspondingAuthor":false,"prefix":"","firstName":"Yingmei","middleName":"","lastName":"Fu","suffix":""},{"id":398882128,"identity":"9dae8217-da45-4ef5-83ce-817ecb8c9fef","order_by":4,"name":"Fang Liu","email":"","orcid":"","institution":"The First Affiliated Hospital of Hainan Medical University","correspondingAuthor":false,"prefix":"","firstName":"Fang","middleName":"","lastName":"Liu","suffix":""},{"id":398882129,"identity":"2c1255a5-2788-4290-abd9-96e9f97784fa","order_by":5,"name":"Tong Chen","email":"","orcid":"","institution":"The First Affiliated Hospital of Hainan Medical University","correspondingAuthor":false,"prefix":"","firstName":"Tong","middleName":"","lastName":"Chen","suffix":""},{"id":398882130,"identity":"cdb75d73-e91b-4200-a7f1-e144e3f2c3cf","order_by":6,"name":"Xiaoliu Li","email":"","orcid":"","institution":"Minhang Hospital，Fudan University","correspondingAuthor":false,"prefix":"","firstName":"Xiaoliu","middleName":"","lastName":"Li","suffix":""},{"id":398882131,"identity":"48fffff0-e698-4985-bb49-748040b6e9e5","order_by":7,"name":"Jing Zhang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA/UlEQVRIiWNgGAWjYDACCRDBAyTZGxsOSFRIyMkTr4Xn8MEHFmcsjA0biNICZqQlG1S2VSQyHCCgQ35287OHX2Qs8uQdcswkbs6TSGBsYH746AYeLYxzjpkby/BIFBseOGMmOXObRB47A5uxcQ4eLcwSCWbSEjwSiRsbe8ykJbdJFDM28LBJ49PCJpH+DaKlmcdM+u8cicSGAwS08EjkmEl+AGqZz8aWbCDZQIQWCYmcMmmgxsQNPMwHH0gckzA2bCbgF/kZ6dskf/bUJc6f/xAYlTV1cvLszQ8f49MCAsy8PQwMBgfgXALKQYDxxw+gdQ1EqBwFo2AUjIKRCQDRxUhPsvE+jQAAAABJRU5ErkJggg==","orcid":"","institution":"The First Affiliated Hospital of Hainan Medical University","correspondingAuthor":true,"prefix":"","firstName":"Jing","middleName":"","lastName":"Zhang","suffix":""}],"badges":[],"createdAt":"2025-01-04 09:08:15","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5762572/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5762572/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":73365724,"identity":"93ac94b0-52f6-4aeb-8dea-b4e6ff3d2791","added_by":"auto","created_at":"2025-01-09 09:31:13","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":141422,"visible":true,"origin":"","legend":"\u003cp\u003eTrends in grip strength and body weight of mice in intervention groups (a) Trends in grip strength of mice in each group (b) Trends in body weight of mice in each group\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-5762572/v1/d6160630f2b18665671db5bc.png"},{"id":73365726,"identity":"6406c79f-e262-4417-b66a-1820885f13eb","added_by":"auto","created_at":"2025-01-09 09:31:13","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":81594,"visible":true,"origin":"","legend":"\u003cp\u003eSkeletal muscle wet weight of mice in each group after intervention (n=3). Data are expressed as mean ± SEM. *** indicates P\u0026lt;0.001for each group compared with the model group, and #### indicates P\u0026lt;0.001 for the model group compared with the control group.\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-5762572/v1/ffaae83d836a79b6a8b5fde6.png"},{"id":73365728,"identity":"e66acb7d-8b61-4d45-8150-e439599f70ac","added_by":"auto","created_at":"2025-01-09 09:31:13","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":349721,"visible":true,"origin":"","legend":"\u003cp\u003eLight microscopic observation of skeletal muscle tissues of mice in each group (×100)\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-5762572/v1/059b26e02d4cfb7fe94e5159.png"},{"id":73365740,"identity":"28b4fc16-d00e-4afa-91fd-7454930fb0b3","added_by":"auto","created_at":"2025-01-09 09:31:14","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":234857,"visible":true,"origin":"","legend":"\u003cp\u003eqRT-PCR was used to detect Omi/Htr A2 (a), Hax-1 (b), LC3 (c), and Beclin1 (d) mRNA expression in mouse muscle tissues (n=3). Data are shown as means ± SEM.\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-5762572/v1/ee92e2de024f9838e176bbaf.png"},{"id":73365736,"identity":"f5ce3524-ce87-4998-b63e-1cea66772f41","added_by":"auto","created_at":"2025-01-09 09:31:13","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":182939,"visible":true,"origin":"","legend":"\u003cp\u003eWestern blot analysis of Omi/HtrA2, Hax-1, LC3II/LC3Ⅰ, and Beclin1 in mouse muscle tissues(n=3). Data are shown as means ± SEM. * P\u0026lt;0.05 vs. model group, # P\u0026lt;0.05 vs. control group.\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-5762572/v1/d813c5b369cac3c7a3feca70.png"},{"id":74773659,"identity":"0a1cec09-f6f0-46a6-b27f-dc7e625617ce","added_by":"auto","created_at":"2025-01-26 12:08:30","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1845648,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5762572/v1/2ee535ee-1662-4d2c-a2c6-7a0da4ecd486.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"The Effects achand Menisms of Electroacupuncture Combined with Exercise Training on T2DM Sarcopenia Based on the Omi/HtrA2 Autophagy Pathway","fulltext":[{"header":"Introduction","content":"\u003cp\u003eAccording to the latest data from the 10th edition of the IDF Global Diabetes Map released by the International Diabetes Federation (IDF)[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e], the number of adults with diabetes worldwide was approximately 537\u0026nbsp;million in 2021, and this figure is projected to rise to 783\u0026nbsp;million by 2045. Type 2 diabetes mellitus is a group of chronic, progressive diseases characterized by impaired glucose metabolism. The disease has a high prevalence, and moderate exercise plays a crucial role in maintaining stable blood glucose levels. The currently accepted approach to managing T2DM includes adopting a healthy lifestyle, which involves dietary planning, physical activity, and weight control[\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Sarcopenia is one of the common complications in patients with diabetes mellitus, severely impacting their quality of life and leading to disabilities such as falls, fractures, loss of independence, and increased risk of death[\u003cspan additionalcitationids=\"CR3 CR4\" citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e–\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Sarcopenia, primarily characterized by a decrease in skeletal muscle mass and function[\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e], is recognized as a common risk factor in diabetes mellitus[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Older adults with T2DM experience a faster decline in skeletal muscle mass and strength compared to non-diabetic individuals[\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e], and are more likely to develop sarcopenia. Given the high prevalence and detrimental effects of sarcopenia in diabetic patients, enhancing muscle mass and function is particularly important in the clinical management of this population[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e].\u003c/p\u003e \u003cp\u003ePhysical activity, particularly resistance training, has been shown to be effective in preventing muscle atrophy and improving muscle strength[\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e], benefiting both upper and lower extremity muscles[\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Data from Roger[\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]further suggest that cellular senescence contributes to age-related functional decline, and physical activity may mitigate this process. However, in clinical practice, some patients, particularly those who are elderly or obese, struggle to perform active resistance training, highlighting an urgent need for alternative treatments, such as passive training or medications, for sarcopenia.\u003c/p\u003e \u003cp\u003eAcupuncture is a well-established traditional therapy for neuromuscular disorders. Electroacupuncture (EA) has been shown to improve oxidative stress and inflammation, maintaining mitochondrial function, though its effects and mechanisms on sarcopenia remain unclear. Studies have shown that EA promotes IGF-1 expression, inhibits myostatin, and stimulates the proliferation of skeletal muscle satellite cells, thereby repairing denervated skeletal muscle and delaying muscle atrophy[\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Guo et al.[\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e] found that after EA intervention, oxidative stress, apoptosis, and levels of IL-6, TNF-α, Atrogin-1, and MuRF1 were suppressed, leading to reduced mitochondrial damage and improved muscle function. Peripheral nerve stimulation through acupuncture or EA has been shown to effectively promote recovery from neuromuscular dysfunction.\u003c/p\u003e \u003cp\u003eAutophagy, a lysosome-dependent catabolic process, plays a key role in maintaining cellular homeostasis under stress. Impaired autophagy may contribute to various diabetes-related metabolic disorders, including sarcopenia[\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. It has been suggested that the pathogenesis of sarcopenia is related to autophagy and can be improved by pathways such as tea polyphenols and histone deacetylase, which enhance metabolism and mitochondrial dynamics[\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Serine protease Omi, a member of the HtrA (High Temperature Requirement A) family, is a protein associated with heat stress, also known as HtrA2. Omi/HtrA2 protease regulates various cellular processes through interactions with multiple substrate proteins in the cytoplasm and mitochondria[\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. Recent studies[\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]have demonstrated that Omi/HtrA2 regulates autophagy upstream of the core autophagy protein Beclin1 by cleaving Hax-1, a Bcl-2 family-associated protein, relieving its inhibition of Beclin1, and ultimately inducing autophagy. HtrA2/Omi protease deficiency has been linked to denervation-independent sarcopenia[\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e], while its absence disrupts mitochondrial protein homeostasis, induces mitochondrial stress responses (UPRmt), and inhibits myogenic differentiation of C2C12 myoblasts, leading to skeletal muscle degeneration in mnd2 mice[\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. Therefore, autophagy plays a critical role in maintaining skeletal muscle homeostasis.\u003c/p\u003e \u003cp\u003eIn this study, we employed interventions including exercise, electroacupuncture, and their combination to observe changes in muscle strength, muscle mass, and autophagy in skeletal muscle cells of T2DM mice with sarcopenia. We also analyzed the gene and protein expression of the Omi/HtrA2-induced autophagy pathway to provide a basis for further research and treatment of T2DM-related sarcopenia.\u003c/p\u003e "},{"header":"Materials and Methods","content":"\u003cp\u003e \u003cb\u003eExperimental Animals and Grouping\u003c/b\u003e \u003c/p\u003e\u003cp\u003eA total of 9 male 4-week-old C57BKS-DB mice, weighing 24.73 ± 1.95 g, were purchased from Jiangsu Jicui YaoKang Biotechnology Co., Ltd., with license number SCXK (Su) 2018-0008. The mice were housed under the national standard conditions for rodent laboratory animals: a constant room temperature of 23 ± 2°C, humidity of 40–45%, and a 12-hour light/dark cycle, with free access to water and food. After adaptive feeding, the mice were fed a normal diet for 3 weeks. They were weighed, their blood glucose levels and hindlimb grip strength were measured, and they were randomly divided into five groups: control group (n = 3), model group (n = 3), electroacupuncture group (n = 3), running group (n = 3), and electroacupuncture + running group (n = 3). Based on blood glucose levels, insulin was administered via intraperitoneal injection at a dose of 0.5-1 U/kg of short-acting insulin to maintain blood glucose at 20 ± 3 mmol/L. All animal experiments were conducted in accordance with Chinese legislation on animal management and general recommendations. The study was approved by the Ethics Committee of the First Affiliated Hospital of Hainan Medical University.\u003c/p\u003e\u003cp\u003e \u003cb\u003eConstruction of a Mouse Model of Type 2 Diabetes Mellitus Combined with Sarcopenia\u003c/b\u003e \u003c/p\u003e\u003cp\u003eExcept for the control group, the other mice were used to construct a model of type 2 diabetes mellitus (T2DM) combined with sarcopenia. A fresh dexamethasone phosphate sodium solution (5 mg/mL) was diluted 10 times by adding 1 mL of the dexamethasone solution to 9 mL of 0.9% saline, resulting in a final concentration of 0.5 mg/mL. This freshly prepared dexamethasone solution was administered subcutaneously at a dose of 5 mg/kg for 8 weeks to induce the T2DM and sarcopenia model. The success of the model was confirmed when blood glucose levels reached ≥ 16.7 mmol/L, hindlimb grip strength decreased by 25%, and both muscle mass and function were significantly reduced.\u003c/p\u003e\u003cp\u003e \u003cb\u003eIntervention Methods\u003c/b\u003e \u003c/p\u003e\u003cp\u003eAfter the successful establishment of the model, experimental interventions were carried out. Mice in the electroacupuncture group were fixed, and 0.25 mm × 13 mm disposable sterile acupuncture needles were used to puncture two acupoints: Pishu (BL20) and Zusanli (ST36). The needles were connected to a KWD-808II multifunctional pulse therapy instrument, with sparse-dense waves (frequency of 1 Hz) and the current intensity adjusted until slight muscle twitching occurred. This treatment was performed every other day, with each session lasting 20 minutes, for 3 days a week and resting 1 day for a total of 5 weeks. The running group was subjected to treadmill training, with the treadmill set at a 0° incline and a speed of 16 m/min for 20 minutes per session. The mice exercised for 3 days a week and rested for 1 day for 5 weeks. The electroacupuncture + running group underwent both electroacupuncture and exercise interventions, following the same protocol as the respective groups.\u003c/p\u003e\u003cp\u003e \u003cb\u003eHindlimb Grip Strength Measurement\u003c/b\u003e \u003c/p\u003e\u003cp\u003eThe grip strength of the mice was measured once per week. Before testing, the mice were allowed to adapt to the grip strength meter (DS2-50N) for 30 seconds. The meter was set to measurement mode, and once the mice had fully grasped the grid with all four paws, their tails were gently pulled backward until the grip was released. The highest value of three measurements, expressed in newtons (N), was recorded as the grip strength for each mouse.\u003c/p\u003e\u003cp\u003e \u003cb\u003eBody Weight and Blood Glucose Measurement\u003c/b\u003e \u003c/p\u003e\u003cp\u003eDuring the experiment, the body weight and blood glucose of the mice were measured once a week at the same time. Blood samples were collected from the tail after disinfection, and blood glucose was measured using a glucometer (Abbott). No fasting or water restrictions were imposed before measurement.\u003c/p\u003e\u003cp\u003e \u003cb\u003eAnimal Sacrifice and Tissue Collection\u003c/b\u003e \u003c/p\u003e\u003cp\u003eAt the end of the experiment, body weight, blood glucose, and grip strength were measured before the mice were sacrificed. The mice were euthanized by cervical dislocation, and sterile ophthalmic scissors and forceps were used to dissect the skin and tissue of both hindlimbs. The complete gastrocnemius muscle from one side was placed in a 1.5 mL cryotube and immediately frozen in liquid nitrogen. After all the tissues were collected, they were stored in liquid nitrogen. The gastrocnemius muscle from the other side was placed in a 50 mL centrifuge tube containing 30 mL of 4% paraformaldehyde and stored in a cool, dark place.\u003c/p\u003e\u003cp\u003e \u003cb\u003eSkeletal Muscle Cross-Section Observation\u003c/b\u003e \u003c/p\u003e\u003cp\u003eThe embedded blocks containing skeletal muscle tissues were dehydrated by immersion in 50% ethanol and 60% ethanol for 2 hours, then further dehydrated using a tissue processor. The tissues were cleared in xylene for 5 minutes and immersed in molten paraffin. The tissues were embedded using a tissue embedding machine, and liquid paraffin was slowly added. After solidification on an ice platform, the paraffin blocks were sectioned at a thickness of approximately 5 µm using a microtome. The sections were flattened in a water bath and transferred to adhesive slides. The slides were labeled and baked overnight. Standard H\u0026amp;E staining was performed, and the sections were observed under a microscope at 20x magnification.\u003c/p\u003e\u003cp\u003e \u003cb\u003eRNA Extraction and qRT-PCR Detection\u003c/b\u003e \u003c/p\u003e\u003cp\u003eTotal RNA from tissues was extracted using the Eastep Super Total RNA Extraction Kit (Promega LS1040). For each sample, total RNA was reverse transcribed into cDNA using the SuperMix for qPCR (gDNA digester plus) (YEASEN H7101160) reverse transcription kit, according to the manufacturer's instructions, for quantitative analysis. The denaturation temperature was set at 95°C for 10 seconds, and the annealing and extension temperature was 60°C for 30 seconds, for a total of 40 cycles. The gene expression was normalized to ACTB using the ΔΔCt method to analyze relative gene expression changes. The primers used were as follows: Omi/HtrA2 (5’-ATCTCAAACGGATCAGGATTCGT-3’, 5’-CAGCCTCACTCGTACTCGG-3’); Hax-1(5’-CGAGGCTTTTTCGGCTTTCC-3’,5’-GCATAGCTCTCTCGACCCCA-3’);Beclin1(5’-GGCGGCTCCTATTCCATCAA-3’,5’-GTCCACTGCTCCTCAGAGTTA-3’);LC3(5’-GACCGCTGTAAGGAGGTGC-3’,5’-CTTGACCAACTCGCTCATGTTA-3’);and ACTB(5’-GGCTGTATTCCCCTCCATCG-3’,5’-CCAGTTGGTAACAATGCCATGT-3’). The expression levels of the target genes were normalized to ACTB, and the data were presented as fold changes relative to the cycle threshold (Ct) values. The relative expression levels of Omi/Htr A2, Hax-1, Beclin1, and LC3 were analyzed.\u003c/p\u003e\u003cp\u003e \u003cb\u003eWestern Blot Detection\u003c/b\u003e \u003c/p\u003e\u003cp\u003eThe tissues were homogenized in a mortar with lysis buffer (PMSF) and repeatedly frozen and thawed in liquid nitrogen, followed by lysis on ice for 15 minutes. The samples were then centrifuged at 12,000 rpm at 4°C for 30 minutes. The supernatant (2 µL) was taken to measure protein concentration using the BCA method. The remaining protein samples were separated on an SDS-PAGE gel and transferred to PVDF membranes. The membranes were blocked with 5% skim milk, then incubated overnight at 4°C with the following primary antibodies: Beclin 1 (Abcam, ab62557), HtrA2/Omi (Abcam, ab75982), Hax-1 (Abcam, ab137613), and LC3B (Abcam, ab48394), with the dilution ratio of LC3B at 1:2000, and the other primary antibodies at 1:1000. β-Actin (Beyotime, AF5003) was used as an internal control at a dilution ratio of 1:1000. All bands were visualized using a chemiluminescent ECL kit (Thermo Fisher), and band intensity was analyzed using the TanonImage software.\u003c/p\u003e\u003cp\u003e \u003cb\u003eStatistical Analysis\u003c/b\u003e \u003c/p\u003e\u003cp\u003eData were analyzed using SPSS 24.0 software. Mouse modeling data were presented as mean ± standard deviation (SD), and paired t-tests were used to compare data before and after treatments within the same group. For data with normal distribution, one-way analysis of variance (ANOVA) was used for between-group comparisons, while the Kruskal-Wallis test was applied for data that did not follow a normal distribution. P \u0026lt; 0.05 was considered a statistically significant difference.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e \u003cb\u003eConstruction of the Mouse Model of Type 2 Diabetes Mellitus Combined with Sarcopenia\u003c/b\u003e \u003c/p\u003e\u003cp\u003eBefore the dexamethasone intervention, the mice underwent 1–2 weeks of adaptive feeding, during which their body weight and grip strength increased. All groups of mice were in good condition, with shiny fur and responsive behaviors. From the 3rd week, the mice were administered dexamethasone at 5 mg/kg to induce the model. The model mice exhibited symptoms such as excessive drinking, reduced food intake, polyuria, and slow weight gain, and some even experienced weight loss. Gradually, their fur became dull and rough, and they displayed irritability, fatigue, and irregular bowel movements. From the 4th week onward, grip strength began to decline. Blood glucose levels were measured, and a glucose level ≥ 16.7 mmol/L was observed. By the 10th week, the muscle mass and function of the modeled mice had significantly decreased, with hindlimb grip strength reduced by 25%. Although grip strength decreased in the non-modeled mice, it did not reach the 25% threshold, indicating that the model of type 2 diabetes mellitus combined with sarcopenia had been successfully established.\u003c/p\u003e\u003cp\u003e \u003cb\u003eComparison of Grip Strength, Body Weight, and Skeletal Muscle Wet Weight Among Different Groups of Mice\u003c/b\u003e \u003c/p\u003e\u003cp\u003eAfter the model construction was completed, interventions were administered to the type 2 diabetes mellitus (T2DM) mice with sarcopenia in the 11th week. After 5 consecutive weeks of treatment, the grip strength of the mice gradually began to recover. There was no significant difference in grip strength in the control group. In contrast, the model group showed significantly lower grip strength at the 15th week compared to before the intervention (P \u0026lt; 0.05). However, in the running, electroacupuncture, and electroacupuncture combined with running groups, the grip strength at the 15th week was significantly higher than at the 10th week. The results indicated that the grip strength of sarcopenic mice further declined, but symptoms were alleviated after interventions such as exercise, electroacupuncture, or the combination of electroacupuncture and exercise. The electroacupuncture group showed a more pronounced effect compared to the running group. Before the intervention, there were no statistically significant differences in grip strength among the model, running, electroacupuncture, and running + electroacupuncture groups. However, there was a significant difference between the control and model groups, as well as between the model group and the other intervention groups (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e, Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea). Furthermore, it was observed that in the control and intervention-treated mice, body weight began to stabilize and increase slowly, while body weight in the model group continued to decline, though not significantly (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb).\u003c/p\u003e\u003cp\u003eAfter the mice were sacrificed, the bilateral gastrocnemius muscles were weighed. Compared to the control group, the skeletal muscle wet weight in the model group was significantly lower. Compared with the model group, the running group, the electroacupuncture group, and the running + electroacupuncture group showed significantly increased wet weight of skeletal muscle. (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComparison of gripping power of mice before and after intervention\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colspan=\"5\" nameend=\"c6\" namest=\"c2\"\u003e \u003cp\u003eGroup (mean ± standard deviation)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMouse Grip Strength (N)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eControl group(n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eModel group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRunning group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eElectroacupuncture group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eRunning + electroacupuncture group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePre-intervention\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.56 ± 0.18\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.20 ± 0.09\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.04 ± 0.22\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.02 ± 0.11\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.11 ± 0.03\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePost-intervention\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.57 ± 0.10\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1.13 ± 0.11*###\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1.77 ± 0.66*^^^\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.67 ± 0.10***^^^\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.70 ± 0.04*^^^\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cp\u003eData are shown as means ± SEM. * indicates P \u0026lt; 0.05 for post-intervention compared to pre-intervention grip strength for each group, and ** indicates P \u0026lt; 0.01 for post-intervention compared to pre-intervention grip strength for each group, *** indicates P \u0026lt; 0.001 for post-intervention compared to pre-intervention grip strength for each group. ### indicates P \u0026lt; 0.001 for grip strength between control group and model group after intervention. ^^^represents P \u0026lt; 0.001 between the intervention groups and the model group after intervention.\u003c/p\u003e\u003cdiv class=\"gridtable\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eComparison of gripping power of mice before and after intervention\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e\u003ccolgroup cols=\"6\"\u003e\u003c/colgroup\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colspan=\"5\" nameend=\"c6\" namest=\"c2\"\u003e \u003cp\u003eGroup (mean ± standard deviation)\u003c/p\u003e \u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMouse Weight (g)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eControl group(n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eModel group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eRunning group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eElectroacupuncture group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eRunning + electroacupuncture group (n = 3)\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePre-intervention\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e54.78 ± 0.47\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e43.36 ± 12.65\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e41.13 ± 9.65\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e30.37 ± 1.54\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e38.17 ± 3.61\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePost-intervention\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e58.92 ± 1.55\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e36.53 ± 9.50\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e46.85 ± 12.85\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e33.25 ± 1.68\u003c/p\u003e \u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e39.85 ± 3.19\u003c/p\u003e \u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/table\u003e\u003c/div\u003e\u003cp\u003eData are shown as means ± SEM.\u003c/p\u003e\u003cp\u003e \u003cb\u003eMorphological Changes in Skeletal Muscle Tissue of Mice\u003c/b\u003e \u003c/p\u003e\u003cp\u003eHE staining results of skeletal muscle in each group showed that in the control group, muscle fibers in the cross-sections appeared polygonal and uniformly sized. The muscle cell nuclei were distributed along the periphery of the fibers, with intact nuclear membranes, and the muscle fibers were arranged in an orderly manner. In the model group, the cross-sections revealed irregularly shaped, atrophied, and deformed muscle fibers with a substantial infiltration of inflammatory cells. Several muscle cells exhibited nuclear migration towards the interior, showing clear multinucleation, and the muscle fibers were arranged disorderly. The running and electroacupuncture groups exhibited similar patterns, with a reduction in inflammatory cell infiltration and nuclear migration compared to the model group. In the running + electroacupuncture group, the cross-sections showed a marked reduction in inflammatory cell infiltration and nuclear migration compared to the model group. Moreover, some muscle fibers showed improvement in morphology, approaching a normal state (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e \u003cb\u003eComparison of Omi/HtrA2, Hax-1, Beclin1, and LC3 mRNA Levels in Skeletal Muscle Tissue of Each Group\u003c/b\u003e \u003c/p\u003e\u003cp\u003eWe used qPCR to evaluate the effect of exercise and electroacupuncture on autophagy levels in sarcopenic mice. Compared with the control group, the LC3 mRNA level was significantly increased in the model group. Compared with the model group, the LC3 mRNA level in the running group + electroacupuncture group showed a downward trend (P = 0.056) (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec). A similar trend was observed in the qPCR results for Beclin1, a gene that reflects autophagy levels (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ed). In the results for Omi, the expression levels in all intervention groups showed a decreasing trend compared to the model group (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea). For Hax-1, the results showed a decreasing trend in Hax-1 gene expression in the model group compared to the control group. In contrast, the expression levels of Hax-1 in the running, electroacupuncture, and running + electroacupuncture groups showed an increasing trend compared to the model group (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb). These trends suggest that autophagy levels are higher in mice with muscle wasting syndrome compared to normal mice, and that exercise, acupuncture, or a combination of both can reduce autophagy levels in diabetic mice with muscle wasting syndrome.\u003c/p\u003e\u003cp\u003e \u003cb\u003eComparison of Omi/HtrA2, Hax-1, Beclin1, and LC3Ⅱ/LC3Ⅰ Protein Levels in Skeletal Muscle Tissue of Each Group\u003c/b\u003e \u003c/p\u003e\u003cp\u003eThe results from the Western blot (WB) experiments showed that, compared to the control group, the HtrA2/Omi protein levels were significantly elevated in the model group. In contrast, the HtrA2/Omiprotein levels were significantly reduced in the electroacupuncture + running group compared to the model group (Fig.\u0026nbsp;5a). Regarding Hax-1protein, the results indicated a decreasing trend in the model group compared to the control group. However, the Hax-1 protein levels in all intervention groups showed an increasing trend compared to the model group (Fig.\u0026nbsp;5b). As for the Beclin1protein levels, there was an increasing trend in the model group compared to the control group (Fig.\u0026nbsp;5d). In contrast, the protein levels in the intervention groups showed a decreasing trend compared to the model group.\u003c/p\u003e\u003cp\u003e \u003cb\u003eFigure\u0026nbsp;5\u003c/b\u003e Western blot analysis of Omi/HtrA2, Hax-1, LC3II/LC3Ⅰ, and Beclin1 in mouse muscle tissues(n = 3). Data are shown as means ± SEM. * P \u0026lt; 0.05 vs. model group, # P \u0026lt; 0.05 vs. control group.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eCurrently, the treatment methods and mechanisms for sarcopenia are still under exploration. Using sarcopenic patients as test subjects presents severe ethical concerns[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e], making it necessary to conduct treatment-related research using animal models. At present, there is no internationally established standard for constructing sarcopenia animal models[\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. To study the treatment and mechanisms of T2DM combined with sarcopenia, we induced muscle weakness in db/db mice using dexamethasone. The grip strength of the modeled mice decreased by 25%, meeting the criteria for sarcopenia modeling, while the control group showed no significant decrease in grip strength. In addition to reduced grip strength, HE staining under electron microscopy showed that the skeletal muscle fibers of dexamethasone-induced mice were atrophied and deformed compared to normal mice, with reduced cross-sectional areas of the muscle fibers. This suggests that dexamethasone can accelerate the onset of sarcopenia in diabetic mice, providing new data and ideas for constructing sarcopenia animal models.\u003c/p\u003e\u003cp\u003eExercise training, particularly resistance training, is considered an effective method to prevent muscle atrophy[\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e] and improve muscle strength[\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e], benefiting both upper and lower limb strength[\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. In this study, treadmill training was used to improve muscle strength in T2DM mice. The results showed that after exercise intervention, the grip strength of the mice increased significantly, while the grip strength of the untreated model group continued to decline. This suggests that exercise training has a positive therapeutic effect on T2DM combined with sarcopenia. Our experimental results are supported by the study of Serra-Rexach et al.[\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e], who conducted resistance training for 8 weeks in elderly individuals over 90 years old. The training intensity increased from 30–70% of one repetition maximum (1RM), resulting in a 10.6 kg increase in leg press 1RM, improved muscle strength, and reduced fall risk. High-intensity resistance training (80% 1RM) was more effective in improving sarcopenia-related muscle weakness compared to low-intensity resistance training (≤ 50% 1RM). Snijders [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e] conducted 24 weeks of supervised resistance training for elderly individuals, significantly improving their skeletal muscle mass and strength. One year after stopping treatment, muscle mass and strength declined. A meta-analysis[\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]suggested that supervised resistance training is effective in improving muscle strength in the elderly.\u003c/p\u003e\u003cp\u003eHowever, many clinical T2DM sarcopenia patients cannot tolerate strict exercise training due to obesity or aging. Furthermore, the effectiveness of drugs for treating T2DM sarcopenia remains controversial, with some hypoglycemic drugs having adverse effects on muscles[\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Therefore, passive treatments in traditional Chinese medicine, such as acupuncture, electroacupuncture, and electrical stimulation, combined with exercise training, are of great significance for treating sarcopenia patients. To explore whether acupuncture, as a passive treatment, has a positive effect on diabetic sarcopenia, we conducted interventions with exercise, electroacupuncture, and their combination in mice. The results showed significant improvements in grip strength in all intervention groups, particularly in the electroacupuncture and combined treatment groups. This indicates that exercise, electroacupuncture, and their combination can enhance muscle strength in T2DM sarcopenic mice. Moreover, skeletal muscle wet weight in the combined treatment group was significantly higher than in the model group, demonstrating that exercise and electroacupuncture interventions have a positive effect on muscle mass in T2DM sarcopenia. T. Brock Symons[\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]used casting to induce muscle atrophy in rats, followed by treatment with acupuncture, electroacupuncture, and electrical stimulation. The results showed that these treatments reduced the negative effects of casting and slowed the degradation of muscle protein. The findings are consistent with our results.\u003c/p\u003e\u003cp\u003eAnimal models in mice indicate that the absence of mnd2 (the mouse homolog of HtrA2) results in neuronal degeneration and muscle atrophy, ultimately leading to shortened lifespan. This suggests that Omi/HtrA2 is essential for normal cell survival. Hax-1 is a substrate of Omi/HtrA2, involved in apoptosis, autophagy, cell migration, and mRNA transport[\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]Under apoptotic stimuli, Omi/HtrA2strongly cleaves Hax-1, triggering early apoptosis. Recent studies show that Omi/HtrA2regulates autophagy upstream of Beclin1, by cleaving Hax-1 (HS1-associated protein X-1) and relieving its inhibition of Beclin1, ultimately inducing autophagy[\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Studies have reported that the lack of HtrA2/Omiprotease activity induces non-denervation-dependent muscle atrophy[\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. Additionally, HtrA2 protease deficiency leads to mitochondrial protein imbalance, inducing UPRmt and inhibiting myogenic differentiation of C2C12 myoblasts. Mice deficient in HtrA2 protease activity showed a distinct muscular dystrophy phenotype[\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. In our study, HtrA2/Omi protein levels were significantly elevated in T2DM sarcopenic mice and decreased after exercise and electroacupuncture treatments. In our study, HtrA2/Omi protein levels were significantly elevated in T2DM sarcopenic mice and decreased after exercise and electroacupuncture treatments. The qPCR results for LC3 also showed a similar trend, with higher LC3 levels in T2DM sarcopenic mice, which significantly decreased after electroacupuncture combined with exercise intervention. This suggests that the autophagy level induced by the HtrA2/OMI autophagy pathway is elevated in T2DM sarcopenic mice, and the combination of electroacupuncture and exercise can reduce this autophagy.\u003c/p\u003e\u003cp\u003eSome limitations exist in this study. Although certain autophagy proteins showed a decreasing trend after the intervention, there was no statistically significant difference. This may be due to factors such as obesity and reduced muscle mass in the T2DM sarcopenic mice, leading to poor performance in active training. Additionally, due to the death of some mice after model establishment, the sample size was reduced, potentially affecting the results for autophagy-related genes and proteins such as HtrA2/Omi, Hax-1, LC3, and Beclin1. Future studies should aim to increase the sample size and improve the representativeness of the data. Moreover, muscle mass and cross-sectional areas were not quantitatively analyzed in this study. Establishing uniform criteria for sarcopenia mouse models will also be crucial for future research.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThe evidence provided by this study suggests that exercise and electroacupuncture therapy improve muscle function and mass in T2DM sarcopenic mice by modulating the HtrA2/Omi autophagy pathway. These therapies reduce the mRNA and protein expression levels of autophagy-related factors, alleviating autophagy-induced muscle damage. The combination of active exercise therapy and passive electroacupuncture therapy offers a new direction for treating diabetic sarcopenia, providing a solid theoretical and experimental foundation for the clinical application of integrative traditional Chinese and Western medicine, with broad potential for development.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003eThis study was supported by Natural Science Foundation of Hainan Province of China (No. 821RC756) and The New Medical Technology Research and Transformation Seed Program of Shanghai Municipal Health Commission,.Study on multimodal rehabilitation detection and treatment of patients with hemiplegia based on portable intelligent devices (No.2024ZZ1030).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest\u0026nbsp;\u003c/strong\u003eThe authors have no relevant financial or non-financial interests to disclose.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions\u0026nbsp;\u003c/strong\u003eZhilin Xie: Collection and assembly of data, data analysis and interpretation, manuscript writing, \u0026nbsp;final approval of manuscript. Jing Zhang: Conception and design, final approval of manuscript.\u0026nbsp;Xiaoliu Li: Collect lower limb electromyographic signals, electromyographic stimulation is used to recruit muscles. Qingxia Shen, Kai Yang: Assist in literature reviewand, collection and assembly of data, final approval of manuscript. Yingmei Fu: Assist in literature reviewand, final approval of manuscript. Tong Chen: Design of experimental scheme of TCM acupuncture and moxibustion, final approval of manuscript. Fang Liu: Acupuncture point selection, final approval of manuscript. All authors reviewed the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u0026nbsp;\u003c/strong\u003eWe thank Dr Man Xizo from the school of basic medicine and life sciences of Hainan Medical University for her guidance in the design of the experiment and her assistance in preparing this manuscript, and Professor Yiqiang Xie from the the College of Traditional Chinese Medicine of Hainan Medical University for his guidance preparing this the manuscript.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthical approval\u0026nbsp;\u003c/strong\u003eThe animal experiments were approved by the Ethics Committee of Hainan Medical University (NoHYLL-2021-173).\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eA. Kumar, R. Gangwar, A.A. Zargar, R. Kumar,A. Sharma, Prevalence of Diabetes in India: A Review of IDF Diabetes Atlas 10th Edition. 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Wei Sheng Yan Jiu. \u003cstrong\u003e53\u003c/strong\u003e, 540-546 (2024). https://doi.org/10.19813/j.cnki.weishengyanjiu.2024.04.004\u003c/li\u003e\n\u003cli\u003eT.W. Lee, H.W. Liu, Y.F. Lin, T.I. Lee, Y.H. Kao,Y.J. Chen, Histone deacetylase inhibition improves metabolism and mitochondrial dynamics: A potential novel therapeutic strategy for sarcopenia coexisting with diabetes mellitus. Med Hypotheses. \u003cstrong\u003e158\u003c/strong\u003e, 110724 (2021). https://doi.org/10.1016/j.mehy.2021.110724\u003c/li\u003e\n\u003cli\u003eX.J. Su, L. Huang, Y. Qu,D. Mu, Progress in research on the role of Omi/HtrA2 in neurological diseases. Rev Neurosci. \u003cstrong\u003e30\u003c/strong\u003e, 279-287 (2019). https://doi.org/10.1515/revneuro-2018-0004\u003c/li\u003e\n\u003cli\u003eB. Li, Q. Hu, H. Wang, N. Man, H. Ren, L. Wen, N. Nukina, E. Fei,G. Wang, Omi/HtrA2 is a positive regulator of autophagy that facilitates the degradation of mutant proteins involved in neurodegenerative diseases. 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Rochester, D.A. Skelton,A.A. Sayer, Resistance exercise as a treatment for sarcopenia: prescription and delivery. Age Ageing. \u003cstrong\u003e51\u003c/strong\u003e, (2022). https://doi.org/10.1093/ageing/afac003\u003c/li\u003e\n\u003cli\u003eJ.A. Serra-Rexach, N. Bustamante-Ara, M. Hierro Villaran, P. Gonzalez Gil, M.J. Sanz Ibanez, N. Blanco Sanz, V. Ortega Santamaria, N. Gutierrez Sanz, A.B. Marin Prada, C. Gallardo, G. Rodriguez Romo, J.R. Ruiz,A. Lucia, Short-term, light- to moderate-intensity exercise training improves leg muscle strength in the oldest old: a randomized controlled trial. J Am Geriatr Soc. \u003cstrong\u003e59\u003c/strong\u003e, 594-602 (2011). https://doi.org/10.1111/j.1532-5415.2011.03356.x\u003c/li\u003e\n\u003cli\u003eT. Snijders, M. Leenders, L. de Groot, L.J.C. van Loon,L.B. Verdijk, Muscle mass and strength gains following 6 months of resistance type exercise training are only partly preserved within one year with autonomous exercise continuation in older adults. Exp Gerontol. \u003cstrong\u003e121\u003c/strong\u003e, 71-78 (2019). https://doi.org/10.1016/j.exger.2019.04.002\u003c/li\u003e\n\u003cli\u003eA. Lacroix, T. Hortobagyi, R. Beurskens,U. Granacher, Effects of Supervised vs. Unsupervised Training Programs on Balance and Muscle Strength in Older Adults: A Systematic Review and Meta-Analysis. Sports Med. \u003cstrong\u003e47\u003c/strong\u003e, 2341-2361 (2017). https://doi.org/10.1007/s40279-017-0747-6\u003c/li\u003e\n\u003cli\u003eT. Brock Symons, J. Park, J.H. Kim, E.H. Kwon, J. Delacruz, J. Lee, Y. Park, E. Chung,S. Lee, Attenuation of skeletal muscle atrophy via acupuncture, electro-acupuncture, and electrical stimulation. Integr Med Res. \u003cstrong\u003e12\u003c/strong\u003e, 100949 (2023). https://doi.org/10.1016/j.imr.2023.100949\u003c/li\u003e\n\u003cli\u003eL. Cilenti, M.M. Soundarapandian, G.A. Kyriazis, V. Stratico, S. Singh, S. Gupta, J.V. Bonventre, E.S. Alnemri,A.S. Zervos, Regulation of HAX-1 anti-apoptotic protein by Omi/HtrA2 protease during cell death. J Biol Chem. \u003cstrong\u003e279\u003c/strong\u003e, 50295-50301 (2004). https://doi.org/10.1074/jbc.M406006200\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Type 2 diabetes mellitus (T2DM), sarcopenia, autophagy, exercise training, electroacupuncture, Omi/HtrA2","lastPublishedDoi":"10.21203/rs.3.rs-5762572/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5762572/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003ePurpose:\u003c/strong\u003e To explore the effects and mechanisms of electroacupuncture combined with exercise training on sarcopenia in mice with type 2 diabetes mellitus (T2DM).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods: \u003c/strong\u003eC57bcs-db mice were randomly divided into 5 groups: control group, model group, electroacupuncture group, exercise group, electroacupuncture + running group. Except for the control group, the other four groups were subcutaneously injected with dexamethasone solution to establish a T2DM sarcopenia model. After successful modeling, the mice in the running group were trained on the treadmill for 5 weeks. Mice in electroacupuncture group were connected to electrotherapy apparatus for 5 weeks. Mice in the electroacupuncture + running group were treated with both running and electroacupuncture. The grasping power, body weight and wet weight of both gastrocnemius of mice were measured. The morphology and structure of gastrocnemius were observed by HE staining. mRNA expressions of omi, Hax-1, LC3 and Becline were detected by RT-PCR, and the expression of related autophagy molecular proteins were detected by Western blot.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults: \u003c/strong\u003eAfter the modeling, all the mice reached the successful criteria for modeling of sarcopenia. After intervention, the hind leg grip of mice in electroacupuncture group, running group and electroacupuncture + running group was significantly increased. The wet weight of gastrocnemius muscle in running + electroacupuncture group was significantly higher than that in model group, and the degree of muscle fiber atrophy of gastrocnemius muscle was significantly reduced. LC3mRNA level and omi/Htra2 protein expression level were significantly decreased in electroacupuncture + running group.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion: \u003c/strong\u003eThe evidence provided by this study suggests that dexamethasone induces sarcopenia in T2DM mice. Electroacupuncture combined with exercise improves grip strength, skeletal muscle wet weight, and muscle fiber atrophy in mice, and reduces the expression of autophagy-related mRNA and proteins, indicating that electroacupuncture combined with exercise may be an effective treatment for T2DM-related sarcopenia.\u003c/p\u003e","manuscriptTitle":"The Effects achand Menisms of Electroacupuncture Combined with Exercise Training on T2DM Sarcopenia Based on the Omi/HtrA2 Autophagy Pathway","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-01-09 09:31:08","doi":"10.21203/rs.3.rs-5762572/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"17ef8ece-5d2f-4e92-b498-7e88f93fed8b","owner":[],"postedDate":"January 9th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-01-26T12:08:13+00:00","versionOfRecord":[],"versionCreatedAt":"2025-01-09 09:31:08","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-5762572","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5762572","identity":"rs-5762572","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}