Search Results (316)

Diabetes mellitus severely impairs skeletal muscle regeneration after injury, limiting satellite cell activation and angiogenesis and disrupting barrier integrity while increasing fibrosis. Hypobaric hypoxia has been proposed to improve the regenerative microenvironment through hypoxia-responsive signaling, but its temporal effects and the coordination between vascular and myogenic programs in diabetic muscle remain unclear. To clarify these processes, adult male mice were divided into five groups: diabetes mellitus control (DM), cardiotoxin-injured (CTX) diabetes assessed on days 7 and 14 (CTX7, CTX14), and hypobaric-hypoxia-treated diabetic injury assessed on days 7 and 14 (H+CTX7, H+CTX14). Animals in the hypoxia groups were exposed to a hypobaric hypoxia chamber for 8 h per day for 14 days. Fibrosis, angiogenic and myogenic markers, and endothelial junctional genes were examined using histology, immunofluorescence, immunoblotting, and qRT-PCR (Quantitative Real-Time PCR). Hypobaric hypoxia on day 7 enhanced HIF-1α (hypoxia-inducible factor 1 alpha), VEGF (vascular endothelial growth factor), eNOS (endothelial nitric oxide synthas), Kdr (kinase insert domain receptor, VEGFR-2), and Angpt2 (angiopoietin-2) expression, accompanied by simultaneous endothelial sprouting and early myogenic stimulation compared to CTX7. Improvements were observed in Angpt1 (angiopoietin-1), Cdh5 (cadherin-5, VE-cadherin), Emcn (endomucin), the Angpt1/Angpt2 ratio, and CD31 density. Myogenin and MyHC (myosin heavy chain) were induced with a reduction in eMyHC (embryonic myosin heavy chain) in accordance with stabilization of endothelium and maturation of fibers, which occurred by day 14. A decrease in fibrosis and an increase in the myofiber cross-sectional area occurred. These findings suggest that hypobaric hypoxia modulates HIF-1α signaling, which in turn induces the VEGF-Kdr-eNOS pathway and the angiopoietin–Tie2–VE-cadherin pathway. Together, these pathways coordinate vascular remodeling and myogenic regeneration, ultimately improving the structural and functional recovery of diabetic muscle. Full article

Hypertrophic cardiomyopathy (HCM) is a common and deadly cardiac disease characterized by enlarged myocytes, increased myocardial wall thickening, and fibrosis. A majority of HCM cases are associated with mutations in the β-myosin heavy chain (MYH7) converter domain locus, which leads to varied pathophysiological and clinical manifestations. Using base-editing technology, we generated mutant human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) harboring HCM-causing myosin converter domain mutations (MYH7 c.2167C>T [R723C]; MYH6 c.2173C>T [R725C]) to define HCM pathogenesis in vitro. In this study, we integrated transcriptomic analysis with phenotypic and molecular analyses to dissect the HCM disease mechanisms using MYH6/7 myosin mutants. Our KEGG analysis of bulk RNA-sequencing data revealed significant upregulation of transcripts associated with HCM in the mutant hiPSC-CMs. Further, in-depth transcriptomic analysis using Gene-Ontology (GO-term) analysis for biological process showed upregulation of several transcripts associated with heart development and disease. Notably, our analysis showed robust upregulation of cytoskeletal transcripts, including actin-cytoskeleton networks, sarcomere components, and other structural proteins in the mutant CMs. Furthermore, cellular and nuclear morphological analysis showed that the MYH6/7 mutation induced cellular hypertrophy and increased aspect ratio compared to the isogenic control. Immunostaining experiments showed marked sarcomere disorganization with lower sarcomeric order and higher dispersion in the mutant hiPSC-CMs, highlighting the remodeling of the myofibril arrangement. Notably, the MYH6/7 mutant showed reduced cortical F-actin expression and increased central F-actin expression compared to the isogenic control, confirming the cytoskeletal remodeling and sarcomeric organization during HCM pathogenesis. These pathological changes accumulated progressively over time, underscoring the chronic and evolving nature of HCM driven by the MYH6/7 mutations. Together, our findings provide critical insights into the cellular and molecular underpinnings of MYH6/7-mutation-associated disease. These findings offer valuable insights into HCM pathogenesis, aiding in future therapies. Full article

The role of LAP proteins expressed in skeletal muscles (ERBIN, LANO, and SCRIBBLE) and at neuromuscular junctions (NMJs) remains largely unknown. Our previous data demonstrate that LAP proteins are differentially expressed in muscle cells, nerve endings, and terminal Schwann cells, though they are all expressed in myofibers and accumulate at NMJs. ERBIN and SCRIBBLE align with acetylcholine receptor clusters (CHRNs) at the NMJ. In vivo ablation of Erbin is associated with smaller CHRN and upregulation of Lano and Scribble. However, SCRIBBLE was also shown to influence the fate decision of muscle stem cells. Here, we investigated how the absence of SCRIBBLE in skeletal muscle cells might impair skeletal muscle fibers or NMJs. Although conditional Scribble knockout mice did not exhibit changes in weight or viability, force per weight decreased slightly. This was supported by compromised neuromuscular transmission and increased NMJ fragmentation. Moreover, Scribble knockout muscles transcribe less myosin heavy chain genes. Here, we also showed that RAB5, an effector of endocytic recycling, interacts with all LAP proteins, but in Scribble knockout muscles, reduced interaction was detected with ERBIN and LANO. These data suggest that a delicate signaling network employing LAP proteins is necessary for skeletal muscle fibers and NMJs. Full article

This study employed a multi-technique approach to investigate the structural and conformational changes in proteins in Meretrix lyrata (M. lyrata) adductor, foot, and siphon tissues during boiling. Data-independent acquisition (DIA) quantitative proteomics was utilized to identify differentially expressed proteins (DEPs) in six temporal comparison groups (20–0 s, 40–20 s, 60–40 s, 80–60 s, 100–80 s, and 120–100 s). The results showed that key myofibrillar proteins, including myosin heavy chain, paramyosin, and actin, exhibited tissue-specific expression patterns, while low-molecular-weight degradation fragments (<17 kDa) appeared with prolonged heating. Turbidity measurements peaked in adductor and siphon tissues at 60 s and in foot tissue at 80 s. Heating resulted in a narrowed particle size distribution (100–1000 nm), and a decreased zeta potential, indicating a reduction in protein surface charge. Fourier transform infrared spectroscopy revealed hydrogen bond disruption and secondary structure transitions, marked by a reduction in α-helix content with a corresponding increase in β-sheet and random coil structures. In total, 6527 proteins were identified, and Gene Ontology (GO) enrichment analysis highlighted the DEPs’ involvement in biological regulation and metabolic processes. Collectively, these results provide comprehensive characterization of protein denaturation, degradation, and structural reorganization in M. lyrata tissues during the boiling process. Full article

Background/Objectives: Premature births below 32 weeks of gestation generally require respiratory oxygen support, leading to a relatively hyperoxic environment compared to in utero conditions. Transient hyperoxia exposure has been linked to an elevated risk of chronic lung disease and retinopathy of prematurity; however, its effects on skeletal muscles remain elusive. This study aimed to investigate the effects of hyperoxic exposure in rats as a model of premature infants receiving supplemental oxygen (30–60% O₂ for several weeks). We hypothesized that rats exposed to postnatal hyperoxia would exhibit muscle fiber atrophy and alterations in fiber type. Methods: We used a rat model in which newborns were exposed to 80% oxygen from birth until postnatal day 12. We assessed the gastrocnemius muscles of rat legs at 12 weeks. Results: Rats exposed to hyperoxia showed substantially increased protein expression of Atrogin-1, along with elevated levels of adipophilin, myogenic differentiation factor 1, and myogenin. No significant changes were observed in the expression of slow or fast myosin heavy chain proteins. However, myofiber size in the gastrocnemius muscle was reduced in the hyperoxia-exposed group compared to the control group. Conclusions: Thus, transient hyperoxia exposure during early life can impede skeletal muscle development, potentially extending into adulthood. Full article

The myometrium is the smooth muscle layer of the uterus, whose dysfunctions are involved in various pathologies leading to infertility, such as adenomyosis and uterine fibroids. Developing relevant in vitro models of the myometrium is crucial for investigating the pathogenesis of these diseases. In this study, we propose a novel approach for cultivating mouse myometrial smooth muscle cells (SMCs) using plant-derived cellulose scaffolds. The scaffolds were obtained through the decellularization of green onion leaf, celery stalk, or bluegrass leaf, subsequently coated with collagen type I. We found that the structure of the green onion leaf scaffold provides unidirectional orientation of cultured cells, mimicking the tissue-specific organization of mouse myometrial SMCs in vivo. The mouse myometrial SMCs, cultured on this scaffold, proliferated, maintained viability up to 2.5 months, and retained the expression of the main markers of smooth muscle contractility (α-smooth muscle actin, transgelin, calponin, smooth muscle myosin heavy chains, connexin-43). To reproduce the native myometrium structure, a multilayered cultivation system was created. In a system of two overlaying scaffolds, cells also retained the viability and expression of smooth muscle contractility markers. The developed approach can be used for three-dimensional myometrium modeling to study the pathogenesis of myometrium-associated diseases. Full article

Background: We recently generated T cell receptor (TCR) transgenic (Tg) mice specific to cardiac myosin heavy chain-α (Myhc-α 334–352) on both myocarditis-resistant (C57BL/6) and susceptible (A/J) genetic backgrounds. We noted that the antigen-specific TCRs were expressed in CD4⁺ and CD8⁺ T cells in both strains, but their responses differed. While the T cells from naïve Tg C57BL/6 mice do not respond to Myhc-α 334–352, whereas those from A/J mice spontaneously respond to the antigen, suggesting their underlying molecular mechanisms might differ. Methods: To investigate the mechanisms of differences in the antigen-responsiveness between the Tg C57BL/6 and A/J mice, we performed bulk RNA sequencing on CD4⁺ and CD8⁺ T cells sorted by flow cytometry. Differentially expressed genes, Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, gene set enrichment analysis (GSEA) of GO and KEGG, and transcription factor (TF) network analyses were performed to identify pathways and regulators of immune responses. Results: First, the principal component analysis of the transcriptomic profiles distinguished CD4⁺ from CD8⁺ T cells, which also differed between the two strains. Second, the differentially expressed cytokine and cytotoxicity genes revealed similar patterns between CD4⁺ and CD8⁺ T cells. Importantly, KEGG enrichment analysis revealed downregulated pathways in both CD4⁺ and CD8⁺ T cells that are associated with viral myocarditis, and various autoimmune conditions in C57BL/6 as compared to A/J mice. Similarly, the GSEA of GO revealed negative regulation of heart contraction and positive regulation of cardiac muscle hypertrophy processes were negatively enriched in CD4⁺ T cells of C57BL/6 mice. Finally, by generating the transcription factor (TF) networks, 22 TFs were found common to both CD4⁺ and CD8⁺ T cells, whereas eight TFs were unique to CD4⁺ or CD8⁺ T cells that have a role in T cell activation, tolerance, and T regulatory cells. Conclusions: Our data provide new insights into the transcriptomic profiles that may contribute to the genetic resistance mechanisms for developing cardiac autoimmunity. Full article

In this report, we showed that oral administration of Dendrobium Taiseed Tosnobile (DTT, also known as Taiwan Emperor No.1) allowed Lewis Lung Carcinoma (LLC) tumor-bearing mice to maintain body weight and grip strength in a dose-dependent manner. Histological analysis showed that treatment with DTT water extract significantly reduced muscle fiber damage by inducing muscle regeneration and improved the cross-sectional area of the rectus femoris, soleus, and gastrocnemius of LLC tumor-bearing C57BL/6 female mice. Further studies revealed that DTT water extract also reduced the expression of inflammatory cytokines such as IL-6 and TNF-α, both in vitro and in vivo. Other analyses showed that DTT water extract promoted the differentiation of C2C12 myoblasts with or without IL-6 by maintaining Myosin Heavy Chain (MyHC) levels. This suggests that DTT water extract acts against muscle wasting via multiple mechanisms. Interestingly, vitamin B1 was identified as an ingredient in DTT water extract through an HPLC analysis. Vitamin B1 was shown to ameliorate IL-6 but not TNF-α generation in active THP-1 cells and protected C2C12 myotubes against IL-6. Further studies showed that DTT and vitamin B1 promoted the multi-nucleus fusion step of C2C12 differentiation by inducing E-cadherin-β-catenin expression with or without IL-6 treatment. In summary, DTT water extract protects muscle cells under cancer conditions through direct and indirect mechanisms, with vitamin B1 being a key functional ingredient that reduces IL-6 generation and aids muscle cell fusion against IL-6 treatment. Full article

Background/Objective: Cellular senescence is increasingly recognized as a key mechanism underlying sarcopenia, an age-related muscle disorder with no effective therapeutic. 6,4′-Dihydroxy-7-methoxyflavanone (DMF), a flavonoid isolated from Dalbergia odorifera T. Chen, has shown anti-senescence potential. This study aimed to investigate the protective effects of DMF against myoblasts senescence and elucidate the underlying molecular mechanisms. Method: A cellular model of senescence was established in C2C12 myoblasts using D-galactose (D-gal). The effects of DMF pretreatment were evaluated by assessing senescence phenotypes, myogenic differentiation, and mitochondrial function. The role of Sirtuin3 (SIRT3) was confirmed using siRNA-mediated knockdown. Results: DMF Pre-treatment effectively attenuated D-gal-induced senescence, as indicated by restored proliferation, reduced senescence-associated β-galactosidase activity, decreased DNA damage, and the downregulation of p53, p21^Cip1/WAF1 and p16^INK4a. Furthermore, DMF rescued myogenic differentiation capacity, enhancing the expression of Myoblast determination protein 1, Myogenin, Myosin heavy chain and Muscle-specific regulatory factor 4, and promoting myotube formation. Mechanistically, DMF was identified as a SIRT3 activator. It enhanced SIRT3 expression and activity, leading to the deacetylation and activation of the mitochondrial antioxidant enzyme superoxide dismutase 2. This consequently reduced mitochondrial reactive oxygen species, improved mitochondrial membrane potential and ATP production, and suppressed the NF-κB pathway by inhibiting IκBα phosphorylation and p65 acetylation/nuclear translocation. Crucially, all the beneficial effects of DMF—including oxidative stress reduction, mitochondrial functional recovery, anti-inflammatory action, and ultimately, the attenuation of senescence and improvement of myogenesis—were abolished upon SIRT3 knockdown. Conclusions: Our findings demonstrate that DMF alleviates myoblasts senescence and promotes myogenic differentiation by activating the SIRT3-SOD2 pathway, thereby reducing oxidative stress and NF-κB-driven inflammation responses. DMF emerges as a promising therapeutic candidate for sarcopenia. Full article

The study presents the potential of milkfish frame, a by-product of milkfish processing, as a source of dipeptidyl peptidase IV (DPP-IV) inhibitory and antioxidant peptides with potential applications in type 2 diabetes management. Proteomic analysis identified key proteins, including 65 kDa warm temperature acclimation protein 1 and myosin heavy chain. In silico prediction (BIOPEP-UWM) guided the selection of proteases for generating DPP-IV inhibitory peptides. Enzymatic hydrolysates were produced and evaluated for bioactivity. Among the treatments, pepsin hydrolysis (2% v/v, 8 h) yielded the highest peptide content (283.64 mg/g), soluble protein (86.46%), and DPP-IV inhibitory activity (68.47%). The resulting milkfish frame pepsin hydrolysate (MFH) was further enhanced through ultrafiltration and simulated gastrointestinal digestion, which improved the DPP-IV inhibitory and antioxidant capacities. Cytotoxicity assays confirmed that MFH (0–100 μg/mL) was non-toxic to FL83B hepatocytes after 24 h. Moreover, treating TNF-α-induced FL83B cells with 10 μg/mL MFHs improved cell viability, reducing the toxicity induced by TNF-α in cells. These findings show that MFHs exhibit promising antidiabetic potential and could serve as natural alternatives to synthetic drugs for type 2 diabetes management. This also demonstrates the valorization of fish processing by-products into functional food ingredients, advancing sustainable approaches in food innovation. Full article

Potato protein (PP) at concentrations of 0.025–0.3% was added to tropical fish surimi, including lizardfish (LZ) and threadfin bream (TB), and cold-water fish, namely Alaska pollock (AP) and Pacific whiting (PW), to examine its effect on proteolytic inhibition and surimi gel texture. Tropical fish surimi, particularly LZ, exhibited the highest degree of autolysis induced by endogenous proteases (p < 0.05), as evidenced by degradation of myosin heavy chain and tropomyosin. PP demonstrated a broad range of proteolytic inhibition activities against chymotrypsin, trypsin, papain, and cathepsin L, with chymotrypsin being the most susceptible. At a PP concentration of 0.3%, the highest autolytic inhibition was obtained in AP (72.24%), followed by LZ surimi (60.44%, p < 0.05). Egg white protein (EW) showed autolytic inhibitory activity at 14.50–50.52% in all species at 0.3%. Surimi gels with only 0.025% PP exhibited breaking forces and distance comparable to those with added 0.3% EW, regardless of the cooking regimes. In tropical surimi, PP at 0.3% increased the breaking force by 4.13–5.38-fold under a setting condition (as the best heating regime) compared with the control. At this concentration, PP decreased the whiteness of LZ and AP in the set surimi gels by 7.03% and 6.42% (p < 0.05), respectively, whereas its effect on TB and PW surimi was negligible. The study demonstrates that PP can be a promising alternative to EW to control proteolytic degradation and improve textural properties of cold-water and tropical surimi. Full article

This study examined the effect of long-term physical activity during the finishing period on meat and fat quality, and metabolic gene expression in obese Alentejano (AL) pigs. From 87.3 to 161.6 kg BW and for 130 days, eighteen pigs were assigned to either individual pens without an exercise area (NE, n = 9) or an outdoor park with an exercise area (WE, n = 9). Both groups received identical commercial diets at 85% ad libitum intake. Loin (Longissimus lumborum—LL), tenderloin (Psoas major—PM), and dorsal subcutaneous fat samples were obtained at slaughter, and analyzed for fatty acid composition and gene expression. Physical activity modulated the fatty acid profile and key metabolic genes in muscle and fat tissues. WE pigs showed higher palmitoleic (p = 0.031) and linolenic (p = 0.022) acids in LL, while Fatty acid synthase and Leptin in LL were downregulated (p = 0.071 and p = 0.018, respectively); Fatty acid binding protein 4 was downregulated (p = 0.003) and Stearoyl-CoA desaturase upregulated (p = 0.020) in the PM of WE pigs, indicating changes in lipid metabolism. Also, Myosin heavy chain 7 was upregulated (p = 0.016) in LL, suggesting oxidative muscle remodeling. These findings suggest that moderate, long-term physical activity during finishing induces modest but favorable metabolic adaptations in muscle and fat tissues without compromising meat quality in AL pigs, supporting its use in traditional rearing systems aimed at balancing animal welfare and product quality in local breeds. Full article

Muscle atrophy is defined as reductions in muscle size and function and represents a critical concern affecting elderly populations, immobilized patients, and individuals following specific dietary regimens, such as fasting and low-protein diets. This study investigated the protective effects of Stellaria dichotoma root extract and its isolated bioactive compounds during muscle atrophy using both in vitro and in vivo experimental models. First, S. dichotoma root extract prevented dexamethasone (DEX)-induced atrophy in C2C12 myotubes. Through systematic solvent partitioning and resin chromatography, five compounds (1–5) were successfully isolated from the n-butanol fraction. Dichotomine B (2) was identified as the most abundant and bioactive constituent. Treatment with dichotomine B significantly preserved the myotube diameter, enhanced the fusion index, and maintained the myosin heavy chain protein level while suppressing key atrophic biomarkers, including FoxO3a, MuRF-1, and Atrogin-1, in DEX-treated myotubes. Furthermore, dichotomine B (2) reduced proteolysis in serum-free cultured C2C12 myotubes and in mice subjected to 48 h of fasting, preserving muscle mass and strength. These findings suggest that S. dichotoma root extract and its principal compound, dichotomine B (2), have promising therapeutic potential and provide an opportunity to develop novel pharmacological interventions against muscle wasting through suppression of proteolysis pathways. Full article

In this study, we aimed to characterize intramuscular fat (IMF) tissue in fattening steers through a comparison with subcutaneous fat (SCF) tissue. The IMF of the longissimus thoracis et lumborum and the SCF of the back fat from three fattening steers (mean body weight of 703.50 ± 11.45 kg) were collected, and the muscle tissue, connective tissue, and fascia were carefully removed. Gene and protein expressions and the lipid contents were assessed via transcriptomic, proteomic, and lipidomic analyses, respectively. Subsequently, tissue-specific factors were identified using integrated analysis. The results revealed that the expressions of sarcoplasmic/endoplasmic reticulum Ca²⁺ transporting 2 (ATP2A2), enolase 3 (ENO3), fructose-bisphosphatase 2 (FBP2), myosin heavy chain 7 (MYH7), myosin light chain 3 (MYL3), myosin light chain kinase (MYLK), glycogen phosphorylase (PYGM), troponin C1 (TNNC1), and tropomyosin 2 (TPM2) significantly increased in IMF at both the mRNA and protein levels, whereas those of fatty acid-binding protein 4 (FABP4), stearoyl-CoA desaturase (SCD), and apolipoprotein E (APOE) were reduced. The abundances of both phosphatidylinositol (PI) (18:1/20:4) and phosphatidylcholine (PC) (15:0/18:2) were positively correlated with APOE. Conversely, that of PI (18:1/20:4) was negatively correlated with ENO3 and PYGM, whereas PC (15:0/18:2) was negatively correlated with TNNC1 and MYLK. In conclusion, we identified calcium signaling and glycolysis as key IMF-regulating pathways. ATP2A2, ENO3, FBP2, MYH7, MYL3, MYLK, PYGM, TNNC1, TPM2, and LPE 18:0 were negatively associated with IMF deposition, whereas FABP4, SCD, APOE, PI (18:1/20:4), and PC (15:0/18:2) were positively associated with it. These findings offer underlying IMF-related targets to promote IMF deposition in cattle. Full article

Duck enteritis virus (DEV) is responsible for duck viral enteritis, a contagious and lethal disease in waterfowls. The host proteins targeted by DEV are unknown. In this study, we developed a recombinant DEV rVP26-Flag and identified 17 host proteins that interact with VP26 in infected chicken embryo fibroblast cells using co-immunoprecipitation in conjunction with liquid chromatography–tandem mass spectrometry (Co-IP-MS/MS). The 17 potential targets of VP26 proteins include Xirp1, TMOD3, DCN, ATP5PD, AP3M1, MYO5A, MYH10, MYH9 (non-muscle myosin IIA heavy chain), and GSN. Most of these proteins are microfilament or cytoskeletal proteins with functions such as cytoskeletal protein binding, actin filament interaction, microfilament motor activity, and myosin II interaction. Using the Search Tool for the Retrieval of Interacting Genes analysis, we predicted a functional network of microfilament cytoskeletal proteins interacting with VP26. Interaction between DEV VP26 and the carboxyl-terminus domain of MYH9 (1651–1960 aa) was verified via co-localization and Co-IP assays. We also demonstrated that the inhibition of actin polymerization with cytochalasin D and latrunculin A reduced the DEV titer. Furthermore, siRNA-mediated knockdown of MYH9, which has intrinsic ATPase activity, also resulted in a reduced viral titer. A targeted inhibitor of myosin II ATPase, (-)-Blebbistatin, significantly suppressed DEV infection both in vitro and in vivo. These results suggest that the actin–myosin II network plays a crucial role in DEV proliferation, with MYH9 being an important host factor influencing DEV infection. Full article