OBJECTIVES: To explore the mechanism by which Tougu Xiaotong Capsule (TXC) promotes chondrogenic differentiation and cartilage repair in mice with osteoarthritis (OA). METHODS: Fifty 8-week-old male C57BL mice were randomly divided into normal control group, cartilage damage (induced by subchondral ring-shaped drilling) model group and TXC treatment groups at low, moderate and high doses (184, 368 and 736 mg/kg, respectively). Saline (in normal control and model groups) and TXC were administered after modeling by daily gavage for 6 consecutive weeks. The changes of cartilage damage in the mice were assessed by measuring thermal withdrawal latency (TWL) and mechanical withdrawal threshold (MWT) and using micro-CT, modified safranine O and fast green staining, HE staining, and qPCR. Primary cultures of mouse synovial mesenchymal stem cells (SMSCs) with lentivirus vector transfection for interfering CXCL12, TXC treatment, or both for 24 h were examined for chondrogenic differentiation using immunofluorescence staining, scratch assay, immunocytochemistry, and Western blotting. RESULTS: In mouse models with cartilage damage, TXC treatment at the moderate dose significantly alleviated joint pain, promoted cartilage repair, and upregulated the mRNA expression levels of CXCL12, GDF5, collagen II, aggrecan, Comp and Sox9 in the cartilage tissue. In primary mouse SMSCs, CXCL12 knockdown resulted in significant reduction of GDF5 protein expression, migration ability and Sox9 protein expression, and these changes were obviously reversed by TXC treatment. CONCLUSIONS TXC promotes chondrogenic differentiation of mouse SMSCs to promote repair of cartilage damage in mice by activating the CXCL12/GDF5 pathway.
Animals ; Drugs, Chinese Herbal/therapeutic use* ; Osteoarthritis/metabolism* ; Male ; Growth Differentiation Factor 5/metabolism* ; Mice, Inbred C57BL ; Mice ; Chemokine CXCL12/metabolism* ; Signal Transduction/drug effects* ; Cell Differentiation/drug effects* ; Cartilage, Articular/drug effects* ; Mesenchymal Stem Cells/cytology*

The mandibular condyle is a critical growth center in craniofacial bone development, especially during postnatal stages. Postnatal condyle osteogenesis requires precise spatiotemporal coordination of growth factor signaling cascades and hierarchical gene regulatory networks. Plagl1, which encodes a zinc finger transcription factor, is a paternally expressed gene. We demonstrate that PLAGL1 is highly expressed in cranial neural crest cell (CNCC)-derived lineage cells in mouse condyles. Using the CNCC-derived lineage-specific Plagl1 knockout mouse model, we evaluate the function of PLAGL1 during postnatal mouse condyle development. Our findings show that PLAGL1 contributes significantly to osteoblast differentiation, and its deficiency impairs osteogenic lineage differentiation, which consequently disrupts mandibular condyle development. Mechanistically, insulin-like growth factor 2 (IGF2) in complex with IGF-binding proteins (IGFBPs) has been identified as the principal PLAGL1 effector responsible for osteogenic regulation during postnatal condyle morphogenesis. Plagl1 deficiency significantly downregulates the IGF2/IGFBP pathway, leading to disordered glucose metabolism, defective extracellular matrix organization, and impaired ossification. Exogenous IGF2 treatment rescues impaired osteoblast differentiation caused by Plagl1 deficiency. In conclusion, the PLAGL1-IGF2 axis is a critical regulator of osteogenesis during mandibular condyle development.
Animals ; Osteogenesis/genetics* ; Insulin-Like Growth Factor II/metabolism* ; Mice ; Transcription Factors/metabolism* ; Mice, Knockout ; Cell Differentiation ; DNA-Binding Proteins/genetics* ; Mandibular Condyle/growth & development* ; Osteoblasts/cytology* ; Signal Transduction ; Neural Crest/cytology*

Magnolol, a compound extracted from Magnolia officinalis, demonstrates potential efficacy in addressing metabolic dysfunction and cardiovascular diseases. Its biological activities encompass anti-inflammatory, antioxidant, anticoagulant, and anti-diabetic effects. Growth/differentiation factor-15 (GDF-15), a member of the transforming growth factor β superfamily, is considered a potential therapeutic target for metabolic disorders. This study investigated the impact of magnolol on GDF-15 production and its underlying mechanism. The research examined the pharmacological effect of magnolol on GDF-15 expression in vitro and in vivo, and determined the involvement of endoplasmic reticulum (ER) stress signaling in this process. Luciferase reporter assays, chromatin immunoprecipitation, and in vitro DNA binding assays were employed to examine the regulation of GDF-15 by activating transcription factor 4 (ATF4), CCAAT enhancer binding protein γ (CEBPG), and CCCTC-binding factor (CTCF). The study also investigated the effect of magnolol and ATF4 on the activity of a putative enhancer located in the intron of the GDF-15 gene, as well as the influence of single nucleotide polymorphisms (SNPs) on magnolol and ATF4-induced transcription activity. Results demonstrated that magnolol triggers GDF-15 production in endothelial cells (ECs), hepatoma cell line G2 (HepG2) and hepatoma cell line 3B (Hep3B) cell lines, and primary mouse hepatocytes. The cooperative binding of ATF4 and CEBPG upstream of the GDF-15 gene or the E1944285 enhancer located in the intron led to full-power transcription of the GDF-15 gene. SNP alleles were found to impact the magnolol and ATF4-induced transcription activity of GDF-15. In high-fat diet ApoE^-/- mice, administration of magnolol induced GDF-15 production and partially suppressed appetite through GDF-15. These findings suggest that magnolol regulates GDF-15 expression through priming of promoter and enhancer activity, indicating its potential as a drug for the treatment of metabolic disorders.
Lignans/pharmacology* ; Growth Differentiation Factor 15/metabolism* ; Animals ; Biphenyl Compounds/pharmacology* ; Endoplasmic Reticulum Stress/drug effects* ; Activating Transcription Factor 4/genetics* ; Mice ; Humans ; Male ; Magnolia/chemistry* ; CCAAT-Enhancer-Binding Proteins/genetics* ; Mice, Inbred C57BL

Background and Objectives: Intraperitoneal injection (i.p.) of D-galactose (D-gal) accelerates aging and develops aging models. A low dose of long-term use and a high dose of short-term use of D-gal can induce natural aging in mice, like brain, cardiac, liver, renal, and skin aging, and erectile dysfunction. Our research aims to determine whether a high dose of short-term use of D-gal. i.p. in rats can induce natural aging and affect the following parameters: body weight (BW), Superoxide Dismutase (SOD), Vascular endothelial growth factor (VEGF), C-reactive protein (CRP), and myostatin. Methods: A daily D-gal i.p. dose of 300 mg/ml/kg for seven days was carried out to induce aging parameters in the rats. After seven days, the body and gastrocnemius circumference of the rats were weighed, and biochemical analysis for SOD, VEGF, CRP, and myostatin in the blood plasma was done. Results: The data obtained were analyzed using nonparametric statistics Friedman test and Mann-Whitney test. After the seven day-intervention, both the control (NaCl 0.9% i.p.) and the high dose of short-term use of D-gal i.p. groups showed no significant difference in the body weight and gastrocnemius circumference. However, D-gal administration could increase the blood plasma level of SOD, VEGF, CRP, and myostatin. Conclusion We conclude that a high dose of short-term intraperitoneal D-galactose can be administrated to induce aging in rat models. The SOD, VEGF, CRP and myostatin can be used as aging parameters.
Aging ; Galactose ; Myostatin ; Vascular Endothelial Growth Factor A

Background and Objectives: Intraperitoneal injection (i.p.) of D-galactose (D-gal) accelerates aging and develops aging models. A low dose of long-term use and a high dose of short-term use of D-gal can induce natural aging in mice, like brain, cardiac, liver, renal, and skin aging, and erectile dysfunction. Our research aims to determine whether a high dose of short-term use of D-gal. i.p. in rats can induce natural aging and affect the following parameters: body weight (BW), Superoxide Dismutase (SOD), Vascular endothelial growth factor (VEGF), C-reactive protein (CRP), and myostatin. Methods: A daily D-gal i.p. dose of 300 mg/ml/kg for seven days was carried out to induce aging parameters in the rats. After seven days, the body and gastrocnemius circumference of the rats were weighed, and biochemical analysis for SOD, VEGF, CRP, and myostatin in the blood plasma was done. Results: The data obtained were analyzed using nonparametric statistics Friedman test and Mann-Whitney test. After the seven day-intervention, both the control (NaCl 0.9% i.p.) and the high dose of short-term use of D-gal i.p. groups showed no significant difference in the body weight and gastrocnemius circumference. However, D-gal administration could increase the blood plasma level of SOD, VEGF, CRP, and myostatin. Conclusion We conclude that a high dose of short-term intraperitoneal D-galactose can be administrated to induce aging in rat models. The SOD, VEGF, CRP and myostatin can be used as aging parameters.
Aging ; D-Galactose ; Galactose ; Myostatin ; VEGF ; Vascular Endothelial Growth Factor A

The maintenance of skeletal muscle quality involves various signal pathways that interact with each other. Under normal physiological conditions, these intersecting signal pathways regulate and coordinate the hypertrophy and atrophy of skeletal muscles, balancing the protein synthesis and degradation of muscle. When the total rate of protein synthesis exceeds that of protein degradation, the muscle gradually becomes enlarged, while when the total rate of protein synthesis is lower than that of protein degradation, the muscle shrinks. Myocyte atrophy mainly involves two protein degradation pathways, namely ubiquitin-proteasome and autophagy-lysosome. Protein degradation pathway is activated during muscle atrophy, resulting in the loss of muscle mass. Muscle atrophy can occur under various conditions such as malnutrition, aging and cachexia. Skeletal muscle atrophy caused by orthopedic diseases mainly includes disuse muscular atrophy caused by fracture and denervation muscular atrophy. The signal pathways that control and coordinate protein synthesis and degradation in skeletal muscle include insulin-like growth factor 1 (IGF1)-Akt-mammalian target of rapamycin (mTOR), myostatin-activin A-Smad, G protein α inhibitory peptide 2 (Gαi2)-PKC, nuclear factor κB (NF-κB), ectodysplasin A2 receptor (EDA2R)-NF-κB inducing kinase (NIK) and mitogen-activated protein kinase (MAPK) pathways. This paper provides a comprehensive review of the protein degradation pathways in skeletal muscle atrophy and the associated signal pathways regulating protein degradation in muscular atrophy.
Humans ; Muscular Atrophy/etiology* ; Muscle, Skeletal/pathology* ; Signal Transduction ; Animals ; Insulin-Like Growth Factor I/metabolism* ; Myostatin/physiology* ; TOR Serine-Threonine Kinases/metabolism* ; Autophagy/physiology* ; NF-kappa B/metabolism* ; Proteolysis ; Proteasome Endopeptidase Complex/physiology*

The fiber type transition of skeletal muscle is an intricate and essential physiological process in the body, significantly influencing both the function and metabolism of skeletal muscle. This phenomenon is not only affected by external environmental changes but also intricately regulated by internal physiological mechanisms. Therefore, exploring the physiological process of muscle fiber type transition holds considerable significance for the treatment of human neuromuscular disorders and the improvement of meat quality in livestock and poultry. It has been discovered that the cytokines secreted by skeletal muscle, i.e., myokines, play a role in the fiber type transition of skeletal muscle. Myokines mainly act on skeletal muscle in autocrine and paracrine forms to participate in signal transduction and regulate the fiber type transition of skeletal muscle. This paper reviews the functional differences among various muscle fiber types, expounds the effects and mechanisms of myokines in regulating the transition processes of these fiber types, and prospects the future research directions in this field. This review is expected to provide theoretical support for enhancing the meat quality of livestock and poultry and treating skeletal muscle-related diseases.
Humans ; Animals ; Cytokines/metabolism* ; Muscle Fibers, Skeletal/metabolism* ; Muscle, Skeletal/metabolism* ; Signal Transduction ; Muscle Fibers, Slow-Twitch/metabolism* ; Muscle Fibers, Fast-Twitch/metabolism* ; Myostatin/metabolism* ; Myokines

OBJECTIVE: To explore the effect of basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), and the combination of bFGF and EGF in the neural differentiation of human bone marrow mesenchymal stem cells (hBMSCs), and the role of Wnt/β-catenin signaling pathway in this process. METHODS: The identified 4th-generation hBMSCs were divided into five groups according to different induction conditions, namely control group (group A), EGF induction group (group B), bFGF induction group (group C), EGF and bFGF combined induction group (group D), and EGF, bFGF, and Dickkopf-related protein 1 (DKK-1) combined induction group (group E). After 7 days of continuous induction, the cell morphology was observed by inverted fluorescence phase contrast microscopy, levels of genes that were related to neural cells [Nestin, neuron-specific enolase (NSE), microtubule-associated protein 2 (MAP-2), and glial fibrillary acidic protein (GFAP)] and key components of the Wnt/β-catenin signaling pathway (β-catenin and Cyclin D1) were detected by RT-PCR, and the levels of proteins that were related to neural cells (Nestin and GFAP) as well as genes that were involved in Wnt/β-catenin signaling pathway [β-catenin, phosphorylation β-catenin (P-β-catenin), Cytoplasmic β-catenin, and Nuclear β-catenin] were explored by cellular immunofluorescence staining and Western blot. RESULTS: When compared to groups A and B, the typical neuro-like cell changes were observed in groups C-E, and most obviously in group D. RT-PCR showed that the relative expressions of Nestin, NSE, and MAP-2 genes in groups C-E, the relative expressions of GFAP gene in groups D and E, the relative expression of NSE gene in group B, the relative expressions of β-catenin gene in groups C and D, and the relative expressions of Cyclin D1 gene in groups B-D significantly increased when compared with group A ( P<0.05). Compared with group E, the relative expressions of Nestin, NSE, MAP-2, GFAP, β-catenin, and CyclinD1 genes significantly increased in group D ( P<0.05); compared with group C, the relative expression of Nestin gene in group D significantly decreased ( P<0.05), while NSE, MAP-2, and GFAP genes significantly increased ( P<0.05). The cellular immunofluorescence staining showed that the ratio of NSE- and GFAP-positive cells significantly increased in groups C-E than in group A, in group D than in groups C and E ( P<0.05). Western blot assay showed that the relative expression of NSE protein was significantly higher in groups C and D than in group A and in group D than in groups C and E ( P<0.05). In addition, the relative expression of GFAP protein was significantly higher in groups C-E than in group A and in group D than in group E ( P<0.05). Besides, the relative expressions of β-catenin, Cytoplasmic β-catenin, Nuclear β-catenin, and the ratio of Nuclear β-catenin to Cytoplasmic β-catenin were significantly higher in groups C and D than in group A and in group D than in group E ( P<0.05), whereas the relative expression of P-β-catenin protein was significantly lower in groups C and D than in group A and in group D than in group E ( P<0.05). CONCLUSION Different from EGF, bFGF can induce neural differentiation of hBMSCs. In addition, EGF can enhance the hBMSCs neural differentiation of bFGF, while the Wnt/β-catenin signaling pathway may play a positive regulatory role in these processes.
Humans ; beta Catenin/metabolism* ; Bone Marrow Cells ; Cell Differentiation ; Cells, Cultured ; Epidermal Growth Factor/metabolism* ; Mesenchymal Stem Cells ; Wnt Signaling Pathway ; Neurons ; Fibroblast Growth Factor 2/metabolism*

As a crucial regulatory molecule in the context of vascular stenosis, transforming growth factor-β (TGF-β), plays a pivotal role in its initiation and progression. TGF-β, a member of the TGF-β superfamily, can bind to the TGF-β receptor and transduce extracellular to intracellular signals through canonical Smad dependent or noncanonical signaling pathways to regulate cell growth, proliferation, differentiation, and apoptosis. Restenosis remains one of the most challenging problems in cardiac, cerebral, and peripheral vascular disease worldwide. The mechanisms for occurrence and development of restenosis are diverse and complex. The TGF-β pathway exhibits diversity across various cell types. Hence, clarifying the specific roles of TGF-β within different cell types and its precise impact on vascular stenosis provides strategies for future research in the field of stenosis.
Humans ; Transforming Growth Factor beta/metabolism* ; Constriction, Pathologic ; Signal Transduction ; Cell Differentiation ; Vascular Diseases ; Transforming Growth Factors ; Transforming Growth Factor beta1

The failure rate of dental implantation in patients with well-controlled type 2 diabetes mellitus (T2DM) is higher than that in non-diabetic patients. This due, in part, to the impaired function of bone marrow mesenchymal stem cells (BMSCs) from the jawbone marrow of T2DM patients (DM-BMSCs), limiting implant osseointegration. RNA N6-methyladenine (m6A) is important for BMSC function and diabetes regulation. However, it remains unclear how to best regulate m6A modifications in DM-BMSCs to enhance function. Based on the "m6A site methylation stoichiometry" of m6A single nucleotide arrays, we identified 834 differential m6A-methylated genes in DM-BMSCs compared with normal-BMSCs (N-BMSCs), including 43 and 790 m6A hypermethylated and hypomethylated genes, respectively, and 1 gene containing hyper- and hypomethylated m6A sites. Differential m6A hypermethylated sites were primarily distributed in the coding sequence, while hypomethylated sites were mainly in the 3'-untranslated region. The largest and smallest proportions of m6A-methylated genes were on chromosome 1 and 21, respectively. MazF-PCR and real-time RT-PCR results for the validation of erythrocyte membrane protein band 4.1 like 3, activity-dependent neuroprotector homeobox (ADNP), growth differentiation factor 11 (GDF11), and regulator of G protein signalling 2 agree with m6A single nucleotide array results; ADNP and GDF11 mRNA expression decreased in DM-BMSCs. Furthermore, gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses suggested that most of these genes were enriched in metabolic processes. This study reveals the differential m6A sites of DM-BMSCs compared with N-BMSCs and identifies candidate target genes to enhance BMSC function and improve implantation success in T2DM patients.
Humans ; Bone Marrow/metabolism* ; Bone Morphogenetic Proteins/metabolism* ; Dental Implants/adverse effects* ; Diabetes Mellitus, Type 2/metabolism* ; Growth Differentiation Factors/metabolism* ; Mesenchymal Stem Cells/metabolism* ; RNA/metabolism* ; RNA Processing, Post-Transcriptional