Myoblast Apoptosis Research Articles

Functional study of the ST6GAL2 gene regulating skeletal muscle growth and development

ST6GAL2, a member of the sialoglycosyltransferase family, primarily localizes within the cellular Golgi apparatus. However, the role of the ST6GAL2 gene in skeletal muscle growth and development remains elusive. In this study, the impact of the ST6GAL2 gene on the proliferation, differentiation, and apoptosis of primary chicken myoblasts at the cellular level was investigated. Quantitative fluorescent PCR was used to measure the expression levels of genes. Subsequently, using gene knockout mice, we assessed its effects on skeletal muscle growth and development in vivo. Our findings reveal that the ST6GAL2 gene promotes the expression of cell cycle and proliferation-related genes, including CCNB2 and PCNA, and apoptosis-related genes, such as Fas and Caspase-9. At the individual level, double knockout of ST6GAL2 inhibited the formation of both fast and slow muscle fibers in the quadriceps, extensor digitorum longus, and tibial anterior muscle, while promoting their formation in the gastrocnemius and soleus. These results collectively demonstrate that the ST6GAL2 gene facilitates the proliferation, apoptosis, and fusion processes of primary chicken myoblasts. Additionally, it promotes the enlargement of cross-sectional muscle fiber areas and regulates the formation of fast and slow muscle fibers at the individual level, albeit inhibiting muscle fusion. This study provides valuable insights into the role of the ST6GAL2 gene in promoting proliferation of skeletal muscle.

Delta-like noncanonical notch ligand 2 regulates the proliferation and differentiation of sheep myoblasts through the Wnt/β-catenin signaling pathway.

This study delved into the role ofdelta-like noncanonical notch ligand 2(DLK2) in the cell cycle, proliferation, apoptosis, and differentiation of myoblasts, as well as its interaction with the classical Wnt/β-catenin signaling pathway in regulating myoblast function. The research revealed that upregulation of DLK2 in myoblasts during the proliferation phase enhanced myoblast proliferation, facilitated cell cycle progression, and reduced apoptosis. Conversely, downregulation of DLK2 expression using siRNA during the differentiation phase promoted myoblast hypertrophy and fusion, suppressed the expression of muscle fiber degradation factors, and expedited the differentiation process. DLK2 regulates myoblasts function by influencing the expression of various factors associated with the Wnt/β-catenin signaling pathway, including CTNNB1, FZD1, FZD6, RSPO1, RSPO4, WNT4, WNT5A, andadenomatous polyposis coli. In essence, DLK2, with the involvement of the Wnt/β-catenin signaling pathway, plays a crucial regulatory role in the cell cycle, proliferation, apoptosis, and differentiation of myoblasts.

Maintaining mitochondrial NAD+ homeostasis is key for heat-induced skeletal muscle injury prevention despite presence of intracellular cation alterations.

Mitochondrial dysfunction is implicated in heat-induced skeletal muscle (SKM) injury and its underlying mechanisms remain unclear. Evidence suggests that cellular ions and molecules, including divalent cations and adenine nucleotides, are involved in the regulation of mitochondrial function. In this study, we examined Ca2+, Mg2+, and NAD+ levels in mouse C2C12 myoblasts and SKM in response to heat exposure. During heat exposure, mitochondrial Ca2+ levels increased significantly, whereas cytosolic Ca2+ levels remained unaltered. The mitochondrial Ca2+ levels in the SKM of heat-exposed mice were 28% higher compared to control mice. No changes in cytosolic Ca2+ were detected between the two groups. Following heat exposure, cytosolic and mitochondrial Mg2+ levels were reduced by 47% and 23% in C2C12 myoblasts, and by 51% and 44% in mouse SKMs, respectively. In addition, heat exposure decreased mitochondrial NAD+ levels by 32% and 26% in C2C12 myoblasts and mouse SKMs, respectively. Treatment with the NAD+ precursor nicotinamide riboside (NR) partially prevented heat-induced depletion of NAD+. Additionally, NR significantly reduced heat-increased mitochondrial fission, mitochondrial depolarization, and apoptosis in C2C12 myoblasts and mouse SKMs. No effects of NR on heat-induced changes in intracellular Ca2+ and Mg2+ levels were observed. This study provides in vitro and in vivo evidence that acute heat stress causes alterations in mitochondrial Ca2+, Mg2+, and NAD+ homeostasis. Our results suggest mitochondrial NAD+ homeostasis as a therapeutic target for the prevention of heat-induced SKM injury.

Deoxynivalenol triggers mitotic catastrophe and apoptosis in C2C12 myoblasts

Deoxynivalenol (DON), commonly known as vomitoxin, is a mycotoxin produced by fungi and is frequently found as a contaminant in various cereal-based food worldwide. While the harmful effects of DON have been extensively studied in different tissues, its specific impact on the proliferation of skeletal muscle cells remains unclear. In this study, we utilized murine C2C12 myoblasts as a model to explore the influence of DON on their proliferation. Our observations indicated that DON exhibits dose-dependent toxicity, significantly inhibiting the proliferation of C2C12 cells. Through the application of RNA-seq analysis combined with gene set enrichment analysis, we identified a noteworthy downregulation of genes linked to the extracellular matrix (ECM) and condensed chromosome. Concurrently with the reduced expression of ECM genes, immunostaining analysis revealed notable changes in the distribution of fibronectin, a vital ECM component, condensing into clusters and punctate formations. Remarkably, the exposure to DON induced the formation of multipolar spindles, leading to the disruption of the normal cell cycle. This, in turn, activated the p53-p21 signaling pathway and ultimately resulted in apoptosis. These findings contribute significant insights into the mechanisms through which DON induces toxicity within skeletal muscle cells.

Bta-miR-181d and Bta-miR-196a mediated proliferation, differentiation, and apoptosis in Bovine Myogenic Cells.

Skeletal muscle is an important component of livestock and poultry organisms. The proliferation and differentiation of myoblasts are highly coordinated processes, which rely on the regulation of miRNA. MiRNAs are widely present in organisms and play roles in various biological processes, including cell proliferation, differentiation, and apoptosis. MiR-181d and miR-196a, identified as tumor suppressors, have been found to be involved in cell proliferation, apoptosis, directed differentiation, and cancer cell invasion. However, their role in beef cattle skeletal muscle metabolism remains unclear. In this study, we discovered that overexpression of bta-miR-181d and bta-miR-196a in Qinchuan cattle myoblasts inhibited proliferation and apoptosis while promoting myogenic differentiation through EDU staining, flow cytometry analysis, immunofluorescence staining, and Western blotting. RNA-seq analysis of differential gene expression revealed that after overexpression of bta-miR-181d and bta-miR-196a, the differentially expressed genes were mainly enriched in the PI3K-Akt and MAPK signaling pathways. Furthermore, the phosphorylation levels of key proteins p-AKT in the PI3K signaling pathway and p-MAPK in the MAPK signaling pathway were significantly decreased after overexpression of bta-miR-181d and bta-miR-196a. Overall, this study provides preliminary evidence that bta-miR-181d and bta-miR-196a may regulate proliferation, apoptosis, and differentiation processes in Qinchuan cattle myoblasts by affecting the phosphorylation status of key proteins in PI3K-Akt and MAPK-ERK signaling pathways.

Untangling the Impacts of Ethanol on Myoblast Proliferation

Alcohol-mediated myopathy frequently accompanies chronic alcohol misuse. This decreased skeletal muscle mass and function increases risk for frailty and falls, decreases quality of life, and worsens prognosis for other comorbid conditions. After muscle injury or atrophy, satellite cells are activated as myoblasts, which proliferate, differentiate, and fuse with myofibers to aid in repairing and rebuilding skeletal muscle. While decreased myoblast differentiation with ethanol (EtOH) is well-documented, we have also observed fewer myoblasts with in vitro EtOH after the proliferative phase of myogenesis. The extent of this decrease and whether this is due primarily to increased cell death, decreased cell division, or a combination thereof remains unanswered. Therefore, the objective of this study is to quantify effects of EtOH on myoblast cell division and death during the proliferative phase. C2C12 myoblasts were proliferated in the presence or absence of 50 mM EtOH for 24, 48, and 72 hrs. Nuclei were stained with DAPI to quantify differences in cell count at each time point. Lactate dehydrogenase (LDH) activity was measured in cell culture supernatant at 48 and 72 hrs and normalized to cell count to assess overall cytotoxicity. T-tests were used to determine differences between conditions at each time point. There were no significant differences in cell counts between cells grown with 0 and 50 mM EtOH at 24 or 48 hrs. At 72 hrs, EtOH significantly decreased cell count (p=0.03). There were no significant differences for LDH activity at 48 hrs. At 72 hrs, LDH activity was significantly lower in the 50 mM group (p=0.05). These preliminary results suggest that EtOH decreases cell count in a time-dependent manner. Cytotoxicity appeared lower with EtOH, although this is likely due to lower intracellular LDH activity given our previously observed adverse impacts of EtOH on myoblast glycolytic function. Regardless, the present data do not support increased cytotoxicity with 50 mM EtOH. Next steps will be to assess alterations in myoblast apoptosis and cell division with EtOH. Future work based on the results of these experiments will examine possible nutraceutical supplements and exercise-like stimulation as interventions to rescue alcohol-mediated effects on myoblast proliferation, which may improve subsequent differentiation and therefore regenerative capacity. This work was supported by Texas Tech University (TTU) startup funds and a TTU College of Arts & Sciences Undergraduate Research Experiences grant (DL). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Adiponectin attenuates H2O2-induced apoptosis in chicken skeletal myoblasts through the lysosomal-mitochondrial axis.

Adiponectin has previously been investigated for exerting its protective effect against myocardial injury through anti-apoptotic and anti-oxidative actions. Therefore, the present study aimed to investigate the nature and mechanism of adiponectin inhibition of H2O2-induced apoptosis in chicken skeletal myoblasts. Skeletal muscle satellite cells were differentiated and assigned into three groups. Group C was on the blank control group, group H was stimulated with the H2O2 (500μmol/L, 4h) alone group, group A + H was pre-treated with adiponectin (10μg/mL, 24h) and stimulated with the H2O2 (500μmol/L, 4h) group. Cytotoxicity inhibited by adiponectin was evaluated by the CCK-8 assay. The degree of apoptosis and oxidative damage was investigated by the TdT-mediated dUTP nick end labeling (TUNEL) and reactive oxygen species (ROS) staining assays. Oxidative stress was assessed by evaluating lipid peroxidation, superoxide dismutase, and reduced glutathione. Acridine orange (AO) staining detected lysosomal membrane permeability. The changes in mitochondrial membrane potential (MMP) were analyzed using 5,5,6,6'-tetrachloro-1,1,3,3-tetraethylimidacarbocyanine iodide (JC-1) dye under a fluorescence microscope. The lysosomal function, mitochondrial function, and apoptosis-related mRNA and protein expression levels were quantified by real-time quantitative PCR and western blot, respectively. The results suggested that adiponectin treatment attenuated H2O2-induced cytotoxicity and oxidative stress in skeletal myoblasts. Compared with H2O2 treatment, TUNEL and ROS staining demonstrated lower apoptosis upon adiponectin treatment. AO staining confirmed the amelioration of lysosomal membrane damage, and JC-1 staining revealed an increase in mitochondrial membrane potential after adiponectin treatment. At the molecular level, adiponectin treatment inhibited the expression of the lysosomal apoptotic factors cathepsin B, chymotrypsin B, and the mitochondrial apoptotic pathway cytochrome-c (cyt-c) and caspase-8; decreased the apoptotic marker gene Bax; and increased the expression of the anti-apoptotic marker gene Bcl-2. Adiponectin treatment attenuated H2O2-induced apoptosis in skeletal myoblasts, possibly by inhibiting oxidative stress and apoptosis through the lysosomal-mitochondrial axis.

CircUBE3C modulates myoblast development by binding to miR-191 and upregulating the expression of p27.

Noncoding RNAs, including miRNAs (microRNAs) and circRNAs (circular RNA), are crucial regulators of myoblast proliferation and differentiation during muscle development. However, the specific roles and molecular mechanisms of circRNAs in muscle development remain poorly understood. Based on the existing circRNA-miRNA-mRNA network, our study focuses on circUBE3C, exploring its differential expression in fetal and adult muscle tissue of the cattle and investigating its impact on myoblast proliferation, apoptosis, and differentiation. The functional analysis of overexpression plasmids and siRNAs (small interfering RNAs) targeting circUBE3C was comprehensively evaluated by employing an array of advanced assays, encompassing CCK-8 (cell counting kit-8), EdU (5-ethynyl-20-deoxyuridine), flow cytometry, western blot analysis, and RT-qPCR. In vivo investigations indicated that overexpression of circUBE3C impedes the process of skeletal muscle regeneration. Mechanistically, we demonstrated that circUBE3C interacts with miR-191 and alleviates the suppression of p27 through cytoplasmic separation, bioinformatics prediction, dual-luciferase reporter assay, and RIP (RNA immunoprecipitation). Our findings indicate that the novel circRNA circUBE3C competitively binds to miR-191, thereby inhibiting proliferation and promoting apoptosis in bovine primary myoblasts and unveiling a regulatory pathway in bovine skeletal muscle development. These findings expand our understanding of circRNA functions in mammals and provide a basis for further exploration of their role in myogenesis and muscle diseases.

Functional identification of PGM1 in the regulating development and depositing of inosine monophosphate specific for myoblasts.

Inosine monophosphate (IMP) is naturally present in poultry muscle and plays a key role in improving meat flavour. However, IMP deposition is regulated by numerous genes and complex molecular networks. In order to excavate key candidate genes that may regulate IMP synthesis, we performed proteome and metabolome analyses on the leg muscle, compared to the breast muscle control of 180-day-old Jingyuan chickens (hens), which had different IMP content. The key candidate genes identified by a differential analysis were verified to be associated with regulation of IMP-specific deposition. The results showed that the differentially expressed (DE) proteins and metabolites jointly involve 14 metabolic pathways, among which the purine metabolic pathway closely related to IMP synthesis and metabolism is enriched with four DE proteins downregulated (with higher expression in breast muscles than in leg muscles), including adenylate kinase 1 (AK1), adenosine monophosphate deaminase 1 (AMPD1), pyruvate kinase muscle isoenzyme 2 (PKM2) and phosphoglucomutase 1 (PGM1), six DE metabolites, Hypoxanthine, Guanosine, L-Glutamine, AICAR, AMP and Adenylsuccinic acid. Analysis of PGM1 gene showed that the high expression of PGM1 promoted the proliferation and differentiation of myoblasts and inhibited the apoptosis of myoblasts. ELISA tests have shown that PGM1 reduced adenosine triphosphate (ATP) and IMP and uric acid (UA), while enhancing the biosynthesis of hypoxanthine (HX). In addition, up-regulation of PGM1 inhibited the expression of purine metabolism pathway related genes, and promoted the IMP de novo and salvage synthesis pathways. This study preliminarily explored the mechanism of action of PGM1 in regulating the growth and development of myoblasts and specific IMP deposition in Jingyuan chickens, which provided certain theoretical basis for the development and utilization of excellent traits in Jingyuan chickens.

LMNA-related muscular dystrophy involving myoblast proliferation and apoptosis through the FOXO1/GADD45A pathway

LMNA-related muscular dystrophy is a major disease phenotype causing mortality and morbidity in laminopathies, but its pathogenesis is still unclear. To explore the molecular pathogenesis, a knock-in mouse harbouring the Lmna-W520R mutation was modelled. Morphological and motor functional analyses showed that homozygous mutant mice revealed severe muscular atrophy, profound motor dysfunction, and shortened lifespan, while heterozygotes showed a variant arrangement of muscle bundles and mildly reduced motor capacity. Mechanistically, the FOXO1/GADD45A pathway involving muscle atrophy processes was found to be altered in vitro and in vivo assays. The expression levels of FOXO1 and its downstream regulatory molecule GADD45A significantly increased in atrophic muscle tissue. The elevated expression of FOXO1 was associated with decreased H3K27me3 in its gene promotor region. Overexpression of GADD45A induced apoptosis and cell cycle arrest of myoblasts in vitro, and it could be partially restored by the FOXO1 inhibitor AS1842856, which also slowed the muscle atrophy process with improved motor function and prolonged survival time of homozygous mutant mice in vivo. Notably, the inhibitor also partly rescued the apoptosis and cell cycle arrest of hiPSC-derived myoblasts harbouring the LMNA-W520R mutation. Together, these data suggest that the activation of the FOXO1/GADD45A pathway contributes to the pathogenesis of LMNA-related muscle atrophy, and it might serve as a potential therapeutic target for laminopathies.

Synergistic effects of MFG-E8 and whey protein on mitigating d-galactose-induced sarcopenia through PI3K/AKT/PGC-1α and MAPK/ERK signaling pathways

Milk fat globule epidermal growth factor 8 (MFG-E8) and whey protein have emerged as promising bionutrient supplements for enhancing skeletal muscle mass and function. In the present study, aging-related sarcopenia rat model was employed to elucidate the effects of the combined administration of MFG-E8 and whey protein on the catabolism and anabolism of gastrocnemius protein. Combined intervention led to notable enhancements in the antioxidative stress status and mitochondrial biogenesis capacity of gastrocnemius muscle fibers in the aging rats, concomitant with a significant inhibition of lipid accumulation. Moreover, the synergistic effect of MFG-E8 and whey protein was found to exert modulatory effects on key signaling pathways, including PI3K/Akt/PGC-1α pathway and MAPK/ERK signaling pathways in the gastrocnemius muscle of the aging rats. Specifically, this combined intervention was observed to promote mitochondrial biogenesis and regulate the expression of protein anabolism and catabolism-related regulators, thereby facilitating the alleviation of mitochondrial oxidative stress and enhancing biogenesis in gastrocnemius tissues. The findings of our study provide compelling evidence for the potential of MFG-E8 as a promising dietary supplement with antisarcopenic properties to ameliorate muscle protein metabolism disorders and mitigate mitochondrial-mediated myoblast apoptosis induced by oxidative stress.

Effect of Bovine MEF2A Gene Expression on Proliferation and Apoptosis of Myoblast Cells.

Myocyte enhancer factor 2A (MEF2A) is a member of the myocyte enhancer factor 2 family. MEF2A is widely distributed in various tissues and organs and participates in various physiological processes. This study aimed to investigate the effect of MEF2A expression on the proliferation and apoptosis of bovine myoblasts. CCK8, ELISA, cell cycle, and apoptosis analyses were conducted to assess cell status. In addition, the mRNA expression levels of genes associated with bovine myoblast proliferation and apoptosis were evaluated using RT-qPCR. The results showed that the upregulation of MEF2A mRNA promoted the proliferation rate of myoblasts, shortened the cycle process, and increased the anti-apoptotic rate. Furthermore, the RT-qPCR results showed that the upregulation of MEF2A mRNA significantly increased the cell proliferation factors MyoD1 and IGF1, cell cycle factors CDK2 and CCNA2, and the apoptotic factors Bcl2 and BAD (p < 0.01). These results show that the MEF2A gene can positively regulate myoblast proliferation and anti-apoptosis, providing a basis for the analysis of the regulatory mechanism of the MEF2A gene on bovine growth and development.

Transcriptome-Based Identification of the Muscle Tissue-Specific Expression Gene CKM and Its Regulation of Proliferation, Apoptosis and Differentiation in Chicken Primary Myoblasts.

Skeletal muscle is an essential tissue in meat-producing animals, and meat-producing traits have been a hot topic in chicken genetic breeding research. Current research shows that creatine kinase M-type-like (CKM) is one of the most abundant proteins in skeletal muscle and plays an important role in the growth and development of skeletal muscle, but its role in the development of chicken skeletal muscle is still unclear. Via RNA sequencing (RNA-seq), we found that CKM was highly expressed in chicken breast muscle tissue. In this study, the expression profile of CKM was examined by quantitative real-time PCR (qPCR), and overexpression and RNA interference techniques were used to explore the functions of CKM in the proliferation, apoptosis and differentiation of chicken primary myoblasts (CPMs). It was shown that CKM was specifically highly expressed in breast muscle and leg muscle and was highly expressed in stage 16 embryonic muscle, while CKM inhibited proliferation, promoted the apoptosis and differentiation of CPMs and was involved in regulating chicken myogenesis. Transcriptome sequencing was used to identify genes that were differentially expressed in CPMs after CKM disruption, and bioinformatics analysis showed that CKM was involved in regulating chicken myogenesis. In summary, CKM plays an important role in skeletal muscle development during chicken growth and development.

The Mouse CircGHR Regulates Proliferation, Differentiation and Apoptosis of Hepatocytes and Myoblasts.

The anterior pituitary gland of animals secretes growth hormone (GH) to bind to the growth hormone receptor (GHR) on the liver cell membrane through the blood circulation, thereby promoting the downstream gene insulin-like growth factor-1 (IGF1) expression, which is the canonical GH-GHR-IGF1 signaling pathway. Therefore, the amount of GHR and the integrity of its structure will affect animal growth and development. In the previous study, we found that the mouse GHR gene can transcribe a circular transcript named circGHR. Our group cloned the full-length of the mouse circGHR and analyzed its spatiotemporal expression profile. In this study, we further predicted the open reading frame of circGHR with bioinformatics, subsequently constructed a Flag-tagged protein vector and preliminarily verified its coding potential with western blot. Additionally, we found that circGHR could inhibit the proliferation of NCTC469 cells and has a tendency to inhibit cell apoptosis, while for C2C12 cells, it showed a tendency to inhibit cell proliferation and promote its differentiation. Overall, these results suggested that the mouse circGHR had the potential to encode proteins and affect cell proliferation, differentiation and apoptosis.

Mitochondrial Ca2+ overload due to altered proteostasis amplifies apoptosis in C2C12 myoblasts under hypoxia: Protective role of nanocurcumin formulation.

Severe hypoxia triggers apoptosis leads to myofibers loss and is attributable to impaired intracellular calcium (iCa2+ ) homeostasis, resulting in reduced muscle activity. Hypoxia increases intracellular Ca2+ by activating the release of Ca2+ from iCa2+ stores, however, the effect of increased [iCa2+ ] on the mitochondria of muscle cells at high-altitude hypoxia is largely unexplored. This study examined mitochondrial Ca2+ overload due to altered expression of mitochondrial calcium uptake 1 (MICU1), that is, a gatekeeper of the mitochondrial Ca2+ uniporter, impaired mitochondrial membrane potential (ΔΨm). p53 stabilization and its translocation to the mitochondria were observed following disrupted mitochondrial membrane integrity in myoblasts under hypoxia. Furthermore, the downstream effects of p53 led to the upregulation of proapoptotic proteins (Bax, Caspase-3, and cytochrome C) in myoblasts under hypoxia. Nanocurcumin-pyrroloquinoline quinone formulation (NCF; Indian patent no. 302877), developed to address hypoxia-induced consequences, was found to be beneficial in maintaining mitochondrial Ca2+ homeostasis and limiting p53 translocation into mitochondria under hypoxia in muscle myoblasts. NCF treatment also modulates heat shock proteins and apoptosis-regulating protein expression in myoblasts. Conclusively, we proposed that mitochondrial Ca2+ overload due to altered MICU1 expression intensifies apoptosis and mitochondrial dysfunctionality. The study also reported that NCF could improve mitochondrial [Ca2+ ] homeostasis and antiapoptotic ability in C2C12 myoblasts under hypoxia.

Dual role of endoplasmic reticulum stress-ATF-6 activation in autophagy and apoptosis induced by cyclic stretch in myoblast.

Mandibular growth that is induced by functional appliances is closely associated with skeletal and neuromuscular adaptation. Accumulating evidence has proved that apoptosis and autophagy have a vital role in adaptation process. However, little is known about the underlying mechanisms. This study sought to determine whether ATF-6 is involved in stretch-induced apoptosis and autophagy in myoblast. The study also sought to uncover the potential molecular mechanism. Apoptosis was assessed by TUNEL and Annexin V and PI staining. Autophagy was detected by transmission electron microscopy (TEM) analysis and immunofluorescent staining for autophagy-related protein light chain 3 (LC3). Real time-PCR and western blot were performed to evaluate the expression level of mRNA and proteins that were associated with endoplasmic reticulum stress (ERS), autophagy and apoptosis. Cyclic stretch significantly decreased the cell viability and induced apoptosis and autophagy of myoblasts time-dependently. Stretching stimuli activated ATF-6 pathway and induced ERS-mediated apoptosis. Moreover, using 4-PBA significantly inhibited ERS-related apoptosis, as well as partially decreasing autophagy. In addition, inhibition of autophagy by 3-MA enhanced apoptosis by affecting the expression of CHOP and Bcl-2. However, it had no obvious effects on ERS-related proteins of GRP78 and ATF-6. More importantly, knockdown ATF-6 effectively weakened apoptosis and autophagy. It did so by regulating the expression of Bcl-2, Beclin1 and CHOP, but not cleaved Caspase-12, LC3II and p62 in stretched myoblast. ATF-6 pathway was activated in myoblast by mechanical stretch. ATF-6 may regulate the process of stretch-induced myoblast apoptosis and autophagy via CHOP, Bcl-2 and Beclin1 signaling.

The cytoprotection of milk-derived MFG-E8 on mitochondria-injured L6 cell via mediation of Akt/bcl-2/bax-caspase-3 signaling pathway

Mitochondria-dependent myoblast apoptosis induced by oxidative stress and decrease in successful protein synthesis play a crucial role in loss of muscle mass and functionality further to result in sarcopenia. Here, we investigated the relationship between milk fat globule-EGF factor Ⅷ (MFG-E8) and mitochondrial-dependent apoptosis pathway. MFG-E8 exert cytoprotection through increasing L6 cells survival/apoptotic ratio, increasing mitochondrial membrane potential and regulating S and G2/M phases. By observing cell ultrastructure, MFG-E8 improved mitochondrial homeostasis mainly through decreasing cytochrome-c release, expression of caspase-9 and caspase-3, mitochondrial vacuolation and mitophagy thereby inhibiting cell apoptosis. From molecular perspective, MFG-E8 repaired mitochondria fragmentation by increasing mitochondrial DNA replication and regulated expression of key mitochondrial-apoptotic factors (upregulation: B-cell lymphoma-2 like 1 (bcl2l1), bcl2l2, cyclooxygenase-1 and mitochondrial uncoupling protein-2; Downregulation: p21 and p53) further to improve mitochondrial function and inhibit apoptosis. Moreover, MFG-E8 inhibited mitochondrial-dependent apoptosis via Akt/bcl-2/bax/caspase-3 signaling cascades. Taken together, our research provided a promising approach for deep exploration of MFG-E8 on cytoprotection against mitochondrial injury and mitochondrial-mediated apoptosis and application of anti-apoptosis in alleviating sarcopenia.

Ginsenoside Rb1 Prevents Oxidative Stress-Induced Apoptosis and Mitochondrial Dysfunction in Muscle Stem Cells via NF-κB Pathway.

Sarcopenia, featured by the progressive loss of skeletal muscle function and mass, is associated with the impaired function of muscle stem cells (MuSCs) caused by increasing oxidative stress in senescent skeletal muscle tissue during aging. Intact function of MuSCs maintains the regenerative potential as well as the homeostasis of skeletal muscle tissues during aging. Ginsenoside Rb1, a natural compound from ginseng, exhibited the effects of antioxidation and against apoptosis. However, its effects of restoring MuSC function during aging and improving age-related sarcopenia remained unknown. In this study, we investigated the role of Rb1 in improving MuSC function and inhibiting apoptosis by reducing oxidative stress levels. We found that Rb1 inhibited the accumulation of reactive oxygen species (ROS) and protected the cells from oxidative stress to attenuate the H2O2-induced cytotoxicity. Rb1 also blocked oxidative stress-induced apoptosis by inhibiting the activation of caspase-3/9, which antagonized the decrease in mitochondrial content and the increase in mitochondrial abnormalities caused by oxidative stress via promoting the protein expression of genes involved in mitochondrial biogenesis. Mechanistically, it was proven that Rb1 exerted its antioxidant effects and avoided the apoptosis of myoblasts by targeting the core regulator of the nuclear factor-kappa B (NF-κB) signal pathway. Therefore, these findings suggest that Rb1 may have a beneficial role in the prevention and treatment of MuSC exhaustion-related diseases like sarcopenia.

Impaired autophagy and mitochondrial dynamics are involved in Sorafenib-induced cardiomyocyte apoptosis

Sorafenib (Sor), a novel multi-target anticancer drug also induces severe toxicity in heart, while the mechanism of its cardiotoxicity remains to be fully elucidated. Dysregulation of autophagy and mitochondrial dynamics imbalance have been implicated in cardiomyocyte death. The aim of this study is to test the hypothesis that Sor disrupts autophagy and mitochondrial dynamics, thereby aggravating Sor-induced oxidative stress damage to cardiomyocytes. Our results revealed that Sor (≥ 5 μM) concentration- and time-dependently reduced cell viability and induced apoptosis in H9c2 myoblasts. Sor treatment promoted intracellular reactive oxygen species (ROS) generation, and subsequent Ca2+ overload as well as apoptosis, which were abolished by the ROS scavenger MPG. Sor inhibited the basal autophagy activity of cells, as supported by the fact that ERK1/2 inhibition-dependent decreases of autophagosomes and autolysosomes, and p62 accumulation in a concentration- and time-dependent manner. Improving autophagy with rapamycin abrogated Sor-induced ROS and Ca2+ overloads, and cell apoptosis. Furthermore, Sor compromised mitochondrial morphology and caused excessive mitochondrial fragmentation in cells. The imbalance of mitochondrial dynamics was attributed to ROS-mediated CaMKII overactivity, and increased phosphorylation of dynamin-related protein 1 (phosph-Drp1). Suppression of CaMKII with KN-93 or mitochondrial fission with mitochondrial division inhibitor-1 (Mdivi-1) attenuated Sor-induced ROS and Ca2+ overloads as well as apoptosis. In conclusion, these results provide the first evidence that impairments in autophagy and mitochondrial dynamics are involved in Sor-induced cardiomyocyte apoptosis. The present study may provide a potential strategy for preventing or reducing cardiotoxicity of Sor.

MiRNA-10a-5p Targeting the BCL6 Gene Regulates Proliferation, Differentiation and Apoptosis of Chicken Myoblasts.

sProliferation, differentiation, and apoptosis are three essential stages in cell development, and miRNAs can achieve extensive regulation of cellular developmental processes by repressing the expression of target genes. According to our previous RNA-seq results, miRNA-10a-5p was differentially expressed at different periods in chicken myoblasts, revealing a possible association with muscle development. In this study, we concluded that miRNA-10a-5p inhibited chicken myoblasts’ proliferation and differentiation and promoted chicken myoblasts’ apoptosis by directly targeting BCL6, a critical transcription factor involved in muscle development and regeneration. Overexpression of BCL6 significantly facilitated myoblasts’ proliferation and differentiation and suppressed myoblasts’ apoptosis. On the contrary, knockdown of BCL6 significantly repressed myoblasts’ proliferation and differentiation and induced myoblasts’ apoptosis. The results above suggest that miRNA-10a-5p plays a potential role in skeletal muscle growth, development and autophagy by targeting the BCL6 gene. We first revealed the functions of miRNA-10a-5p and BCL6 in the proliferation, differentiation, and apoptosis of chicken myoblasts.