UBE2C promotes myoblast differentiation and skeletal muscle regeneration through the Akt signaling pathway.

Ubiquitin-conjugation enzyme E2C (UBE2C) is a crucial component of the ubiquitin-proteasome system that is involved in numerous cancers. In this study, we find that UBE2C expression is significantly increased in mouse embryos, a critical stage during skeletal muscle development. We further investigate the function of UBE2C in myogenesis. Knockdown of UBE2C inhibits C2C12 cell differentiation and decreases the expressions of MyoG and MyHC, while overexpression of UBE2C promotes C2C12 cell differentiation. Additionally, knockdown of UBE2C, specifically in the tibialis anterior muscle (TA), severely impedes muscle regeneration in vivo. Mechanistically, we show that UBE2C knockdown reduces the level of phosphorylated protein kinase B (p-Akt) and promotes the degradation of Akt. These findings suggest that UBE2C plays a critical role in myoblast differentiation and muscle regeneration and that UBE2C regulates myogenesis through the Akt signaling pathway.

NIR Light-Fuse Drug-Free Photothermal Armor-Piercing Microcapsule for Femoral Vein Thrombosis Therapy.

Acute thrombosis and its complications are leading global causes of disability and death. Existing thrombolytic drugs, such as alteplase and urokinase (UK), carry a significant bleeding risk during clinical treatments. Thus, the development of a novel thrombolysis strategy is of utmost urgency. Based on the previous work, the hollow structure of microcapsules (MC) is fabricated. Subsequently, armor-piercing MC, known as Fucoidan/S-Nitrosoglutathione/Melanin@MC (FGM@MC) is obtained, using a layer-by-layer (LBL) self-assembly method. Utilizing near-infrared (NIR) light as a trigger, the FGM@MC demonstrated photothermal thrombolysis at the site of thrombus due to its stable and outstanding photothermal properties. Simultaneously, photothermal stimulation leads to the release of a significant amount of nitric oxide from the FGM@MC, resulting in cavitation effects for mechanical thrombolysis. In vivo experiments confirmed the stable release of nitric oxide under NIR light irradiation. Treatment of femoral vein thrombosis in rats revealed that the thrombolytic effectiveness of FGM@MC+NIR (53.71%) is comparable to that of UK (59.70%). Notably, FGM@MC does not interfere with the coagulation function of rats and exhibits a favorable safety profile. In conclusion, this study demonstrates that the drug-free armor-piercing microcapsule has significant potential in the treatment of thrombosis, offering a safe and effective alternative to traditional thrombolytic therapies.

Urbanization-induced soil organic carbon loss and microbial-enzymatic drivers: insights from aggregate size classes in Nanchang city, China.

Soil microorganisms and enzymes play crucial roles in soil organic carbon (SOC) sequestration by promoting soil aggregate formation and stability and by participating in SOC cycling and accumulation. However, the effects by which soil microorganisms and enzymes act as mediators driving dynamic changes in SOC during rapid urbanization remain unclear. Therefore, this study selected the built-up area of Nanchang City, China (505 km2), as the study area. Sampling surveys were conducted using 184 sample plots stratified based on the proportion of impermeable surface area to distinguish different urbanization levels. The driving factors of dynamic changes in SOC of different aggregates during the process of urbanization were analyzed using the soil microbial community and enzyme activities. The results demonstrated that with an increase in urbanization intensity, both SOC content and stock exhibited a significant decline (p < 0.05). The highest SOC stock and contribution rate were observed in the 0.25-1 mm aggregates, and they were significantly influenced by urbanization (p < 0.05). In addition, the biomass of gram-positive bacteria (G+) and actinomycetota, and the activities of N-acetylglucosaminidase and acid phosphatase (AP) were significantly higher in low-urbanization areas than in high-urbanization areas (p < 0.05). SOC of each aggregate was positively correlated with fungi, arbuscular mycorrhizal fungi, G+, gram-negative bacteria, actinomycetota, protozoa, ß-1,4-glucosidase, N-acetylglucosaminidase, AP, urease, and catalase. Compared to soil enzymes, soil microorganisms exhibited a greater role in SOC sequestration (22.7%). Additionally, a structural equation model indicated that urbanization can directly or indirectly lead to a decrease in SOC of aggregates by altering soil physicochemical properties and affecting microbial and enzyme dynamics. However, the larger vegetation characteristics index mitigate the negative impacts of urbanization on SOC. Overall, urbanization had a negative impact on soil carbon storage. In the future, it is important to consider strategies that focus on improving soil nutrients, maintaining soil structure, protecting existing urban trees, and enhancing plant diversity during the urbanization process. These measures can help increase soil microbial biomass and enzyme activity, thereby improving soil and aggregate-related SOC content. The study could contribute to enhancing carbon sequestration in urban greenspaces.

Transformable Magnetic Liquid-Metal Nanoplatform for Intracellular Drug Delivery and MR Imaging-Guided Microwave Thermochemotherapy.

Improved techniques for the administration of chemotherapeutic drugs are required to enhance tumor therapy efficacy and reduce the side effects of chemotherapy due to insufficient targeting and limited intratumoral drug release. Controlled drug delivery systems combined with thermotherapy are expected to play an important role in personalized tumor therapy. Herein, a novel microwave-responsive transformable magnetic liquid-metal (MLM) nanoplatform is designed for effective endosomal escape that facilitates intracellular drug delivery and enhanced anticancer therapy. The MLM nanoplatform exhibits a sensitive magnetic resonance imaging function for imaging-guided therapy and brilliant synergistic effects of chemotherapy with microwave thermal therapy to kill tumor cells. Once endocytosed by targeted tumor cells, the deep penetration of microwave energy can be absorbed by the MLM nanoplatform to convert heat and reactive oxygen species, which induces the shape transformation from nanospheres to large rods, resulting in the physical disruption of the endosomal membrane for intracellular drug release. Furthermore, the MLM nanoplatform synergistic therapy could activate immunomodulatory effects by M1 macrophage polarization and T cell infiltration, thus inhibiting tumor growth and lung metastasis. This work based on microwave-driven transformable magnetic liquid-metal nanoplatform provides novel ways to precisely control drug delivery and high-efficiency cancer therapy.

All-in-One Self-Powered Microneedle Device for Accelerating Infected Diabetic Wound Repair.

Diabetic wound healing remains a significant clinical challenge due to the complex microenvironment and attenuated endogenous electric field. Herein, a novel all-in-one self-powered microneedle device (termed TZ@mMN-TENG) is developed by combining the multifunctional microneedle carried tannin@ZnO microparticles (TZ@mMN) with the self-powered triboelectric nanogenerator (TENG). In addition to the delivery of tannin and Zn2+ , TZ@mMN also effectively conducts electrical stimulation (ES) to infected diabetic wounds. As a self-powered device, the TENG can convert biomechanical motion into exogenous ES to accelerate the infected diabetic wound healing. In vitro experiment demonstrated that TZ@mMN shows excellent conductive, high antioxidant ability, and effective antibacterial properties against both Staphylococcus aureus and Escherichia coli (>99% antibacterial rates). Besides, the TZ@mMN-TENG can effectively promote cell proliferation and migration. In the diabetic rat full-thickness skin wound model infected with Staphylococcus aureus, the TZ@mMN-TENG can eliminate bacteria, accelerate epidermal growth (regenerative epidermis: ≈303.3 ± 19.1 µm), enhance collagen deposition, inhibit inflammation (lower TNF-α and IL-6 expression), and promote angiogenesis (higher CD31 and VEGF expression) to accelerate infected wound repair. Overall, the TZ@mMN-TENG provides a promising strategy for clinical application in diabetic wound repair.

Assembly of Metal-Phenolic Networks onto Microbubbles for One-Step Generation of Functional Microcapsules.

The one-step assembly of metal-phenolic networks (MPNs) onto particle templates can enable the facile, rapid, and robust construction of hollow microcapsules. However, the required template removal step may affect the refilling of functional species in the hollow interior space or the in situ encapsulation of guest molecules during the formation of the shells. Herein, a simple strategy for the one-step generation of functional MPNs microcapsules is proposed. This method uses bovine serum albumin microbubbles (BSA MBs) as soft templates and carriers, enabling the efficient pre-encapsulation of guest species by leveraging the coordination assembly of tannic acid (TA) and FeIII ions. The addition of TA and FeIII induces a change in the protein conformation of BSA MBs and produces semipermeable capsule shells, which allow gas to escape from the MBs without template removal. The MBs-templated strategy can produce highly biocompatible capsules with controllable structure and size, and it is applicable to produce other MPNs systems like BSA-TA-CuII and BSA-TA-NiII . Finally, those MBs-templated MPNs capsules can be further functionalized or modified for the loading of magnetic nanoparticles and the pre-encapsulation of model molecules through covalence or physical adsorption, exhibiting great promise in biomedical applications.

Motility-related microRNAs identified in pig seminal plasma exosomes by high-throughput small RNA sequencing.

Boar fertility is a key determinant of the production efficiency of the whole pig breeding industry and boar sperm motility is the seminal parameter with the greatest impact on the fecundity of a sow. Exosomes are small, extracellular vesicles found in many body fluids. Seminal plasma exosomes, which are secreted by the epididymis, prostate, seminal vesicles, and testes, contain a large number of miRNAs, the types and levels of which can reflect the physiological state of source cells. It has been shown that the expression profile of seminal plasma exosomal miRNA differs between low-motility semen and normal semen. The aim of this study was to investigate the relationship between semen motility and exosomal miRNA profiles to obtain information that would allow to predict boar fertility, as well as contribute to the understanding of the mechanisms by which exosomal miRNAs regulate semen motility. Three high-motility (semen motility >90 %) and three low-motility (semen motility <80 %) semen samples were collected from Landrace and Yorkshire boars, respectively, and seminal plasma exosomes were extracted by ultracentrifugation. Exosome characterization was performed using transmission electron microscopy, NTA, and Western blot. The expression profiles of exosomal miRNAs associated with semen motility in the two boar breeds were subsequently determined by small RNA sequencing. The results showed that 297 known miRNAs and 295 novel RNAs were co-expressed in the four groups. Notably, six miRNAs (ssc-miR-122-5p, ssc-miR-486, ssc-miR-451, ssc-miR-345-3p, ssc-miR-362, and ssc-miR-500-5p) were found to be differentially expressed in both boar breeds. Enrichment analysis of the target genes of the differentially expressed miRNAs showed that they were mainly involved in biological processes such as regulation of transcription from RNA polymerase II promoter, regulation of gene expression, and intracellular signal transduction and signaling pathways such as the PI3K-Akt, MAPK, and Ras signaling pathways. The six differentially expressed miRNAs identified in this study have significant potential as noninvasive markers of boar semen motility. Meanwhile, the results of the enrichment analysis provide novel insights into the mechanisms underlying the regulation of semen motility.

Layer-by-layer microneedle patch with antibacterial and antioxidant dual activities for accelerating bacterial-infected wound healing.

Bacterial-infected wound healing has always been a huge challenge to humans. Owing to the appearance of antibiotic resistance, there is an emergency need to design antibiotic-free wound dressings to treat such wounds. Herein, a novel antibiotic-free microneedle patch was designed, which its backing layer with antioxidant effect was coated with sodium carboxymethyl cellulose, 2-O-α-D-glucopyranosyl-L-ascorbic acid (GLAA), and 2-hydroxypropyltrimethyl ammonium chloride chitosan through electrostatic interaction based on layer-by-layer self-assembly technique, and its tips consisted of gelatin and tannic acid (TA) via hydrogen bonding interaction (CGH/GTA MN patch). The obtained CGH/GTA MN patch could effectively puncture the skin, and exhibit properties of pH-responsive TA and GLAA release. In vitro experiments showed that the obtained CGH/GTA MN patch has excellent antioxidative (scavenging DPPH efficacy is above 80 %, and scavenging ABTS efficiency reaches about 100 %), antibacterial (antibacterial rates of nearly 100 % for both Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli)), biodegradable, and biocompatible properties. In the S. aureus-infected rat wounds, the CGH/GTA MN patch could efficiently accelerate infected-wound healing by eliminating S. aureus infection, inhibiting inflammation, promoting angiogenesis, and accelerating epidermal regeneration. Thus, this study will provide a promising strategy to heal bacterial-infected wounds.

Layer-by-Layer Microneedle-Mediated rhEGF Transdermal Delivery for Enhanced Wound Epidermal Regeneration and Angiogenesis.

Appropriate treatments for acute traumas tend to avoid hemorrhages, vascular damage, and infections. However, in the homeostasis-imbalanced wound microenvironment, currently developed therapies could not precisely and controllably deliver biomacromolecular drugs, which are confronted with challenges due to large molecular weight, poor biomembrane permeability, low dosage, rapid degradation, and bioactivity loss. To conquer this, we construct a simple and effective layer-by-layer (LBL) self-assembly transdermal delivery patch, bearing microneedles (MN) coated with recombinant human epidermal growth factor (LBL MN-rhEGF) for a sustained release to wound bed driven by typical electrostatic force. Pyramidal LBL MN-rhEGF patches hold so enough mechanical strength to penetrate the stratum corneum, and generated microchannels allow rhEGF direct delivery in situ. The administrable delivery of biomacromolecular rhEGF through hierarchically coated MN arrays follows the diffusion mechanism of Fick's second law. Numerous efforts further have illustrated that finger-pressing LBL MN-rhEGF patches could not only promote cell proliferation of normal human dermal fibroblasts (NHDF) and human umbilical vein endothelial cells (HUVEC) in vitro but also take significant effects (regenerative epidermis: ∼144 µm; pro-angiogenesis: higher CD31 expression) in accelerating wound healing of mechanically injured rats, compared to the traditional dressing, which relies on passive diffusion. Our proof-of-concept features novel LBL biomacromolecular drug-delivery systems and self-administrated precision medicine modes at the point of care.

LBL Noninvasively Peelable Biointerfacial Adhesives for Cutaneo-Inspired pH/Tactility Artificial Receptors.

Besides barrier functions, skin possesses multiple sentiences to external stimuli (e.g., temperature, force, and humidity) for human-outside interaction. Thus, skincare should be taken very seriously, especially by patients with sensory disorders. However, currently available skin-mimicking devices are always limited by so much insufficient response functions and nontunable interface behaviors so as not to realize precise health monitoring and self-defense against injury. Herein, a bioinspired cutaneous receptor-perceptual system (CRPS) patch is presented, integrating hybrid pH indicators and triboelectric nanogenerators into biointerface film-adhesives that are fabricated through facile layer-by-layer (LBL) self-assembly of amide and Schiff-base linkages between alginate grafted with N-hydroxysuccinimide ester (AN), tannic acid (TA), and polyethylenimine (PEI). This CRPS patch is adhered robustly to the soft-curved skin surface without failure via "molecular suturing," and amino acid enables its benign peel-on-demand from tissue interfaces. Postdamage self-healing brings it without surgical reoperation, avoiding extra cost, pain, as well as infection risks. Significantly, CRPS patches as artificial chemo/mechanoreceptors can remotely visualize skin physiological status by pH-induced chromism using smartphones and prevent skin contact injury by tactility-driven self-powered electrical signals. Overall, the LBL-based strategy to create controllably biointerface-adhesive CRPS patches will usher in a new era of the mobihealth care platform supporting smart diagnosis and self-protection.

Notch signaling leads to a slower progression of embryonic myogenic differentiation in Landrace than in Langtang pigs.

Delving into porcine embryonic myogenesis is the key to elucidate the complex regulation of breed-specific differences in growth performance and meat production. Increasing evidence proves that pigs with less meat production show earlier embryonic myogenesis, but little is known about the underlying mechanisms. In this study, we examine the longissimus dorsi muscle (LDM) by immunohistochemistry and confirm that the differentiation of myogenic progenitors is increased ( P<0.05) in Lantang (LT, fatty) pigs compared with that in Landrace (LR, lean) pigs, which results in more ( P<0.001) differentiated myoblasts (Pax7 -/MyoD +) and less ( P<0.001) myogenic progenitors (Pax7 +/MyoD -) in LT pigs at 35 days post-conception (35dpc). Additionally, embryonic myogenic progenitors isolated from LT pigs show greater ( P<0.001) differentiation capacity with earlier expression of MyoD compared with those from LR pigs. Moreover, Notch signaling is more active ( P<0.05) in LR pig myogenic progenitors than in LT pig myogenic progenitors. Inhibition of Notch signaling in LR myogenic progenitors suppresses Pax7 expression and increases MyoD expression, thus promoting myogenic differentiation. Consistently, the process of myogenic progenitors differentiating into myoblasts in ex vivo embryo limbs is accelerated when Notch signaling is inhibited. These results indicate that Notch signaling facilitates the maintenance of myogenic progenitors and antagonizes myogenic differentiation by promoting Pax7 expression and preventing MyoD expression in LR pigs.

Dynamic 3D genome reorganization during development and metabolic stress of the porcine liver.

Liver development is a complex process that is regulated by a series of signaling pathways. Three-dimensional (3D) chromatin architecture plays an important role in transcriptional regulation; nonetheless, its dynamics and role in the rapid transition of core liver functions during development and obesity-induced metabolic stress remain largely unexplored. To investigate the dynamic chromatin architecture during liver development and under metabolic stress, we generated high-resolution maps of chromatin architecture for porcine livers across six major developmental stages (from embryonic day 38 to the adult stage) and under a high-fat diet-induced obesity. The characteristically loose chromatin architecture supports a highly plastic genome organization during early liver development, which fundamentally contributes to the rapid functional transitions in the liver after birth. We reveal the multi-scale reorganization of chromatin architecture and its influence on transcriptional regulation of critical signaling processes during liver development, and show its close association with transition in hepatic functions (i.e., from hematopoiesis in the fetus to metabolism and immunity after birth). The limited changes in chromatin structure help explain the observed metabolic adaptation to excessive energy intake in pigs. These results provide a global overview of chromatin architecture dynamics associated with the transition of physiological liver functions between prenatal development and postnatal maturation, and a foundational resource that allows for future in-depth functional characterization.

Biomechanical Motion-Activated Endogenous Wound Healing through LBL Self-Powered Nanocomposite Repairer with pH-Responsive Anti-Inflammatory Effect.

Wound care is still worthy of concern, and effective measures such as electrical stimulating therapy (EST) have sparked compellingly for wound repair. Especially, portable and point-of-care EST devices get extremely desired but these are often limited by inevitable external power sources, lack of biological functions, and mechanical properties conforming to skin tissue. Herein, a dress-on-person self-powered nanocomposite bioactive repairer of wound is designed. As such, the cooperation of the film prepared by layer-by-layer self-assembling 2-hydroxypropyltrimethyl ammonium chloride chitosan (HTCC), alginate (ALG), and poly-dopamine/Fe3+ nanoparticles (PFNs), with a self-powered nanogenerator (SN) driven by motion into a nanocomposite repairer (HAP/SN-NR) is conducted. The HAP/SN-NR not only guides cell behavior (proliferation and migration rate ≈61.7%, ≈52.3%), but also facilitates neovascularization (enhanced CD31 expression >4-fold) through its self-powered EST, and the endogenous wound closure with no inflammatory in rats owing to reactive oxygen species (ROS)-clearance of HAP/SN-NR in vitro/vivo through responsively releasing poly-dopamine nanoparticles at wound pH. Enormous efforts illustrate that the repairer is endowed with high self-adhesion to tissue, self-healing, and biodegradation, accelerating wound healing (50% closure ≈5 days). This strategy sheds light on novel multifunctional portable sensor-type dressings and propels the development of intelligent medical devices.

Reorganization of chromatin architecture during prenatal development of porcine skeletal muscle.

Myofibres (primary and secondary myofibre) are the basic structure of muscle and the determinant of muscle mass. To explore the skeletal muscle developmental processes from primary myofibres to secondary myofibres in pigs, we conducted an integrative three-dimensional structure of genome and transcriptomic characterization of longissimus dorsi muscle of pig from primary myofibre formation stage [embryonic Day 35 (E35)] to secondary myofibre formation stage (E80). In the hierarchical genomic structure, we found that 11.43% of genome switched compartment A/B status, 14.53% of topologically associating domains are changed intradomain interactions (D-scores) and 2,730 genes with differential promoter-enhancer interactions and (or) enhancer activity from E35 to E80. The alterations of genome architecture were found to correlate with expression of genes that play significant roles in neuromuscular junction, embryonic morphogenesis, skeletal muscle development or metabolism, typically, NEFL, MuSK, SLN, Mef2D and GCK. Significantly, Sox6 and MATN2 play important roles in the process of primary to secondary myofibres formation and increase the regulatory potential score and genes expression in it. In brief, we reveal the genomic reorganization from E35 to E80 and construct genome-wide high-resolution interaction maps that provide a resource for studying long-range control of gene expression from E35 to E80.

High mobility group box 2 regulates skeletal muscle development through ribosomal protein S6 kinase 1.

HMGB2, a DNA-binding protein, highly expresses during embryogenesis and plays an important role in development of some organs and tissues. However, it remains to be further investigated weather HMGB2 influences muscle development. In this work, we identified HMGB2 as an essential factor in myogenesis. Compared to wild type (WT) mice, body weights of systemic hmgb2 homozygous knockout (hmgb2-/- ) mice especially males were reduced. Diameter and cross-section area of tibialis anterior (TA) muscle fibers as well as expression of Myogenin and MyHC were all decreased in hmgb2-/- mice. CTX injury model revealed that HMGB2 was required for satellite cell proliferation and muscle regeneration. Moreover, HMGB2 interacted with S6K1 and regulated the kinase activity of S6K1 during cell proliferation. Knockdown and inactivation of S6K1 in C2C12 cells both resulted in impaired proliferation and differentiation. Furthermore, expression of cyclin D1 and Myf5 were both decreased when HMGB2 or S6K1 were knocked down and kinase activity of S6K1 was inhibited. These results indicate that HMGB2 is required for skeletal muscle development and regeneration, and HMGB2 maintains proliferation of myoblasts through regulating kinase activity of S6K1.

Biocompatible Self-Healing Coating Based on Schiff Base for Promoting Adhesion of Coral Cells.

Layer-by-layer self-assembly (LBL) technique is a very efficient and convenient method to modify the substrate surface. In this study, we report a self-repairing surface coating that can promote cell adhesion, especially for enhancing the adhesion of coral cells on the basal surface. The results confirmed that the modified chitosan-dialdehyde starch film based on Schiff base has good biocompatibility for common mammalian cells, such as normal human dermal fibroblasts (NHDFs) and relatively special cells (coral cells). The cytotoxicity test indicated that the optical density values of the experimental group films at 490 nm were higher than those of the control group in this study. In addition, the self-repairing coating modified by phase transition lysozyme can maintain its adhesion ability underwater for a period of time. Therefore, they have great application on substrates requiring underwater adhesion. Our results confirmed that the modified chitosan-dialdehyde starch self-healing films could provide a biocompatible coating material to promote the adhesion of normal human epidermal fibroblasts or coral cells.

MLL1 promotes myogenesis by epigenetically regulating Myf5.

OBJECTIVES: Mixed lineage leukaemia protein-1 (MLL1) mediates histone 3 lysine 4 (H3K4) trimethylation (me3) and plays vital roles during early embryonic development and hematopoiesis. In our previous study, we found its expression was positively correlated with embryonic myogenic ability in pigs, indicating its potential roles in mammalian muscle development. The present work aimed to explore the roles and regulation mechanisms of MLL1 in myogenesis. MATERIALS AND METHODS: The expression of MLL1 in C2C12 cells was experimentally manipulated using small interfering RNAs (siRNA). 5-ethynyl-2'-deoxyuridine (EdU) assay, cell cycle assay, immunofluorescence, qRT-PCR and Western blot were performed to assess myoblast proliferation and differentiation. Chromatin immunoprecipitation assay was conducted to detect H3K4me3 enrichment on myogenic factor 5 (Myf5) promoter. A cardiotoxin (CTX)-mediated muscle regeneration model was used to investigate the effects of MLL1 on myogenesis in vivo. RESULTS: MLL1 was highly expressed in proliferating C2C12 cells, and expression decreased after differentiation. Knocking down MLL1 suppressed myoblast proliferation and impaired myoblast differentiation. Furthermore, knockdown of MLL1 resulted in the arrest of cell cycle in G1 phase, with decreased expressions of Myf5 and Cyclin D1. Mechanically, MLL1 transcriptionally regulated Myf5 by mediating H3K4me3 on its promoter. In vivo data implied that MLL1 was required for Pax7-positive satellite cell proliferation and muscle repair. CONCLUSION: MLL1 facilitates proliferation of myoblasts and Pax7-positive satellite cells by epigenetically regulating Myf5 via mediating H3K4me3 on its promoter.

Zfp422 promotes skeletal muscle differentiation by regulating EphA7 to induce appropriate myoblast apoptosis.

Zinc finger protein 422 (Zfp422) is a widely expressed zinc finger protein that serves as a transcriptional factor to regulate downstream gene expression, but until now, little is known about its roles in myogenesis. We found here that Zfp422 plays a critical role in skeletal muscle development and regeneration. It highly expresses in mouse skeletal muscle during embryonic development. Specific knockout of Zfp422 in skeletal muscle impaired embryonic muscle formation. Satellite cell-specific Zfp422 deletion severely inhibited muscle regeneration. Myoblast differentiation and myotube formation were suppressed in Zfp422-deleted C2C12 cells, isolated primary myoblasts, and satellite cells. Chromatin Immunoprecipitation Sequencing (ChIP-Seq) revealed that Zfp422 regulated ephrin type-A receptor 7 (EphA7) expression by binding an upstream 169-bp DNA sequence, which was proved to be an enhancer of EphA7. Knocking EphA7 down in C2C12 cells or deleting Zfp422 in myoblasts will inhibit cell apoptosis which is required for myoblast differentiation. These results indicate that Zfp422 is essential for skeletal muscle differentiation and fusion, through regulating EphA7 expression to maintain proper apoptosis.

The SNPs in myoD gene from normal muscle developing individuals have no effect on muscle mass.

BACKGROUND: Myogenic Differentiation 1 (MyoD) is a crucial master switch in regulating muscle-specific gene transcription. Forced expression of myoD is equipped to induce several cell lineages into myoblast, which then differentiate and fuse into myotube. Pig is one of the most significant livestock supplying meat, and has been classified into lean, fat and miniature pig breeds. However, the mechanisms underlying muscle mass variation among different pig breeds have remained unclear. Considering the important effect of MyoD on muscle development, it remains to be investigated whether the difference in muscle mass is caused by its single nucleotide polymorphisms (SNPs) which are the major differences among pig breeds at DNA level. RESULTS: In this study, we identified the locations of porcine myoD regulatory regions including proximal regulatory region (PRR), distal regulatory region (DRR), and core enhancer (CE) region. There are 8 SNPs in the regulatory regions and 6 SNPs in gene body region, which were identified from lean, fat and miniature pig populations. However, these SNPs have no effects on its temporal expression and transcriptional activity which might lead to the distinction in postnatal muscle mass. In addition, overexpression of myoD clones across from amphibious to mammals including xenopus tropicalis, chicken, mouse and pig whose gene identities vary from 68 to 84%, could promote myogenesis in NIH3T3 fibroblasts cells. CONCLUSIONS: These results proved that myoD nucleotide variations from different pig populations have no effect on muscle mass, suggesting that the function of myoD is highly conserved not only among different pig breeds, but also across different species. Thus, it would be futile to discover SNPs affecting muscle mass in pig populations with normal muscle development.

Integrated miRNA-mRNA transcriptomic analysis reveals epigenetic-mediated embryonic muscle growth differences between Wuzhishan and Landrace pigs1.

Pig is one of the major dietary protein sources for human consumption, from which muscle is the largest protein origin. However, molecular mechanisms concerning early porcine embryonic muscle development distinctions between pig breeds are still unclear. In this study, an integrated analysis of transcriptome and miRNAome was conducted using longissimus dorsi muscle of 4 early embryonic stages around the primary myofiber formation time (18-, 21-, 28-, and 35-d post coitus) from 2 pig breeds (Landrace [LR] and Wuzhishan [WZS]) differing in meat mass. The global miRNA/mRNA expression profile showed that WZS prepared for myogenic developmental processes earlier than LR. After identifying and analyzing the interaction network of top 100 up-/down-regulated miRNA and their target genes, we were able to find 3 gene clusters: chromatin modification-related (Chd2, H3f3a, Chd6, and Mll1), myogenesis-related (Pax3, Pbx1, Mef2a, and Znf423), and myosin component-related (Mylk, Myo5a, Mylk4, Myh9, and Mylk2) gene clusters. These genes may involve in miRNA-gene myogenic regulatory network that plays vital role in regulating distinct early porcine embryonic myogenic processes between LR and WZS. In summary, our study reveals an epigenetic-mediated myogenic regulatory axial that will help us to decipher molecular mechanisms concerning early porcine embryonic muscle development distinctions between pig breeds.