Profound skeletal muscle loss can lead to severe disability and cosmetic deformities. Mesenchymal stem cell (MSC)-derived exosomes have shown potential as an effective therapeutic tool for tissue regeneration. This study aimed to determine the regenerative capacity of MSC-derived exosomes for skeletal muscle regeneration. Exosomes were isolated from human adipose tissue-derived MSCs (AD-MSCs). The effects of MSC-derived exosomes on satellite cells were investigated using cell viability, relevant genes, and protein analyses. Moreover, NOD-SCID mice were used and randomly assigned to the healthy control (n = 4), muscle defect (n = 6), and muscle defect + exosome (n = 6) groups. Muscle defects were created using a biopsy punch on the quadriceps of the hind limb. Four weeks after the surgery, the quadriceps muscles were harvested, weighed, and histologically analyzed. MSC-derived exosome treatment increased the proliferation and expression of myocyte-related genes, and immunofluorescence analysis for myogenin revealed a similar trend. Histologically, MSC-derived exosome-treated mice showed relatively preserved shapes and sizes of the muscle bundles. Immunohistochemical staining revealed greater expression of myogenin and myoblast determination protein 1 in the MSC-derived exosome-treated group. These results indicate that exosomes extracted from AD-MSCs have the therapeutic potential for skeletal muscle regeneration.

Intervertebral discs (IVDs) have poor nutrient diffusion, because the nucleus pulposus (NP) lacks direct vascular supply and likely generates adenosine triphosphate by anaerobic glycolysis. Regulation of glycolysis is mediated by hypoxia-inducible factor-1α (HIF-1α), a transcription factor that responds to local oxygen tension. Constitutively active HIF-1α (CA HIF-1α) was created by point mutation and determined the protective role of HIF-1α in IVD degeneration. Under fluoroscopy, rat caudal IVD segments were stabbed by a needle puncture, and pcDNA3- HIF-1α wild-type (WT) or pcDNA3-CA HIF-1α was transfected into NP cell lines. The constitutive activity of CA HIF-1α was analyzed using a luciferase assay after cell lysis. Next, IVD tissue samples were retrieved from five patients with degenerative lumbar spinal stenosis at the time of surgery, and NP cells were cultured. NP cells were transfected with CA HIF-1α, and relevant gene expression was measured. HIF-1α protein levels in the nucleus were significantly higher, and transcriptional activity was 10.3-fold higher in NP cells with CA HIF-1α than in those with HIF-1α WT. Gene transfer of CA HIF-1α into NP cells enhanced the expression of Glut-1, Glut-3, aggrecan, type II collagen, and Sox9. Moreover, CA HIF-1α reduced the apoptosis of NP cells induced by the Fas ligand. The HIF-1α and collagen 2 expression levels were notably increased in the NP cells of the CA HIF-1α transfected segments in histology and immunohistochemistry study. Collectively, these results suggest that activation of HIF-1α signaling pathway may play a protective role against IVD degeneration and could be used as a future therapeutic agent.

Recently, neuromediators such as substance P (SP) have been found to be important factors in tendon homeostasis. Some studies have found SP to be the cause of inflammation and tendinopathy, whereas others have determined it to be a critical component of tendon healing. As demonstrated by these conflicting findings, the effects of SP on tendinopathy remain unclear. In this study, we hypothesized that the duration of SP exposure determines its effect on the tendons, with repetitive long-term exposure leading to the development of tendinopathy. First, we verified the changes in gene and protein expression using in vitro tenocytes with 10-day exposure to SP. SP and SP + Run groups were injected with SP in their Achilles tendon every other day for 14 days. Achilles tendons were then harvested for biomechanical testing and histological processing. Notably, tendinopathic changes with decreased tensile strength, as observed in the Positive Control, were observed in the Achilles in the SP group compared to the Negative Control. Subsequent histological analysis, including Alcian blue staining, also revealed alterations in the Achilles tendon, which were generally consistent with the findings of tendinopathy in SP and SP + Run groups. Immunohistochemical analysis revealed increased expression of SP in the SP group, similar to the Positive Control. In general, the SP + Run group showed worse tendinopathic changes. These results suggest that sustained exposure to SP may be involved in the development of tendinopathy. Future research on inhibiting SP is warranted to target SP in the treatment of tendinopathy and may be beneficial to patients with tendinopathy.

Oncogenic FGFR4 signaling represents a potential therapeutic target in various cancer types, including triple-negative breast cancer and hepatocellular carcinoma. However, resistance to FGFR4 single-agent therapy remains a major challenge, emphasizing the need for effective combinatorial treatments. Our study sought to develop a comprehensive computational model of FGFR4 signaling and to provide network-level insights into resistance mechanisms driven by signaling dynamics. An integrated approach, combining computational network modeling with experimental validation, uncovered potent AKT reactivation following FGFR4 targeting in triple-negative breast cancer cells. Analyzing the effects of cotargeting specific network nodes by systematically simulating the model predicted synergy of cotargeting FGFR4 and AKT or specific ErbB kinases, which was subsequently confirmed through experimental validation; however, cotargeting FGFR4 and PI3K was not synergistic. Protein expression data from hundreds of cancer cell lines was incorporated to adapt the model to diverse cellular contexts. This revealed that although AKT rebound was common, it was not a general phenomenon. For example, ERK reactivation occurred in certain cell types, including an FGFR4-driven hepatocellular carcinoma cell line, in which there is a synergistic effect of cotargeting FGFR4 and MEK but not AKT. In summary, this study offers key insights into drug-induced network remodeling and the role of protein expression heterogeneity in targeted therapy responses. These findings underscore the utility of computational network modeling for designing cell type–selective combination therapies and enhancing precision cancer treatment. Significance: Computational predictive modeling of signaling networks can decipher mechanisms of cancer cell resistance to targeted therapies and enable identification of more effective cancer type–specific combination treatment strategies.

An important objective of vascularized tissue regeneration is to develop agents for osteonecrosis. We aimed to identify the pro-angiogenic and osteogenic efficacy of adipose tissue-derived (AD) pericytes combined with Nel-like protein-1 (NELL-1) to investigate the therapeutic effects on osteonecrosis. Tube formation and cell migration were assessed to determine the pro-angiogenic efficacy. Vessel formation was evaluated in vivo using the chorioallantoic membrane assay. A mouse model with a 2.5 mm necrotic bone fragment in the femoral shaft was used as a substitute for osteonecrosis in humans. Bone formation was assessed radiographically (plain radiographs, three-dimensional images, and quantitative analyses), and histomorphometric analyses were performed. To identify factors related to the effects of NELL-1, analysis using microarrays, qRT-PCR, and Western blotting was performed. The results for pro-angiogenic efficacy evaluation identified synergistic effects of pericytes and NELL-1 on tube formation, cell migration, and vessel formation. For osteogenic efficacy analysis, the mouse model for osteonecrosis was treated in combination with pericytes and NELL-1, and the results showed maximum bone formation using radiographic images and quantitative analyses, compared with other treatment groups and showed robust bone and vessel formation using histomorphometric analysis. We identified an association between FGF2 and the effects of NELL-1 using array-based analysis. Thus, combinatorial therapy using AD pericytes and NELL-1 may have potential as a novel treatment for osteonecrosis.

Inadequate vessel maintenance or growth causes ischemia in diseases such as myocardial infarction, stroke, and neurodegenerative disorders. Therefore, developing an effective strategy to salvage ischemic tissues using a novel compound is urgent. Drug repurposing has become a widely used method that can make drug discovery more efficient and less expensive. Additionally, computational virtual screening tools make drug discovery faster and more accurate. This study found a novel drug candidate for pro-angiogenesis by in silico virtual screening. Using Gene Expression Omnibus (GEO) microarray datasets related to angiogenesis studies, differentially expressed genes were identified and characteristic direction signatures extracted from GEO2EnrichR were used as input data on L1000CDS² to screen pro-angiogenic molecules. After a thorough review of the candidates, a list of compounds structurally similar to TWS-119 was generated using ChemMine Tools and its clustering toolbox. ChemMine Tools and ChemminR structural similarity search tools for small-molecule analysis and clustering were used for second screening. A molecular docking simulation was conducted using AutoDock v.4 to evaluate the physicochemical effect of secondary-screened chemicals. A cell viability or toxicity test was performed to determine the proper dose of the final candidate, ellipticine. As a result, we found ellipticine, which has pro-angiogenic effects, using virtual computational methods. The noncytotoxic concentration of ellipticine was 156.25 nM. The phosphorylation of glycogen synthase kinase-3β was decreased, whereas the β-catenin expression was increased in human endothelial cells treated with ellipticine. We concluded that ellipticine at sublethal dosage could be successfully repositioned as a pro-angiogenic substance by in silico virtual screening.