Cheng Wang

Deer Horn Glue is rich in collagen- and tissue-derived proteins, but the link between its experimentally observed proteome, the theoretical peptide sequence space derived from that proteome, and bone-related molecular targets remains poorly defined. This study established a proteomics-guided multiscale computational workflow to prioritize putative receptor activator of nuclear factor-κB ligand (RANKL)-binding peptide candidates generated from proteins identified in one Deer Horn Glue sample. The Deer Horn Glue proteome was characterized by liquid chromatography–tandem mass spectrometry (LC-MS/MS). Experimentally identified proteins were subjected to in silico tryptic digestion and stepwise activity, safety, and physicochemical screening. All 49 retained candidates underwent global and interface-focused docking. Five prioritized peptides were examined by AlphaFold 3, three independently seeded 200 ns molecular dynamics simulations per complex, entropy-omitted molecular mechanics/generalized Born surface area (MM/GBSA) analysis, residue decomposition, and locally relaxed computational alanine substitution analysis. Proteomic analysis retained 707 target protein groups and generated 24,575 nonredundant theoretical peptide sequences. Stepwise screening retained 49 candidates. Global and interface-focused docking rankings showed modest agreement, and the expanded interface analysis identified additional candidates while retaining GASLQDWDFGK as the top-ranked sequence. Across three independent simulations, GASLQDWDFGK showed consistently low peptide root-mean-square deviation (RMSD), whereas HEFSVDMTCEGCSNAVTR formed the largest mean number of interfacial hydrogen bonds. Their entropy-omitted MM/GBSA estimates were generally the most favorable, although the order varied among simulations. Locally relaxed alanine substitutions highlighted reproducible energy-sensitive positions for experimental testing. By linking an experimentally observed Deer Horn Glue proteome to multiscale structural analysis, this workflow provides a traceable and reproducible strategy for prioritizing testable peptide–RANKL interaction hypotheses. The five candidates remain theoretical products of in silico digestion and require targeted detection, direct binding, and functional validation.

Atopic dermatitis (AD), a common chronic inflammatory skin disease, has been extensively studied using single-cell genomics. However, keratinocytes, as key effector cells in AD, have underlying mechanisms remain incompletely understood and require further investigation. We integrated single-cell transcriptomic data from skin tissues of healthy controls, chronic active AD patients, spontaneously healed AD (SHAD) patients, and an ovalbumin-induced AD mouse model. The study particularly emphasized the gene expression and cellular dynamics of keratinocytes across the different groups, as well as their interactions with immune cells. Compared to healthy controls, we observed significant changes in the keratinocyte transcriptome, cellular state, and keratinocyte-immune cell ligand-receptor interactions in AD skin, particularly the marked activation of genes involved in antigen processing and presentation. Interestingly, such gene activation was not observed in keratinocytes from the ovalbumin-induced AD mouse model, despite its phenotype closely resembling human AD. Furthermore, in SHAD, we identified a recovery of both the ligand-receptor interaction patterns and antigen processing and presentation genes, accompanied by a notable shift in the transcriptome. This involved a significant downregulation of genes related to cytoplasmic transcription and oxidative phosphorylation. Notably, this pattern was not observed in the self-healing mouse model following the removal of ovalbumin stimulation. Our results suggest that the persistent activation of antigen processing and presentation pathways in keratinocytes may be a key driver of chronic inflammation in AD. Therefore, redirecting anti-allergic therapeutic strategies from solely targeting immune cells to targeting of keratinocyte-mediated antigen presentation may offer a more effective approach. Furthermore, we raise concerns about the use of ovalbumin-induced mouse models to recapitulate human chronic AD, as the underlying mechanisms may differ significantly.

Di(2-ethylhexyl) phthalate (DEHP), a widely employed exogenous plasticizer, has become pervasive in the environment and living organisms due to its extensive use in food packaging, medical devices, and daily consumer products, and is established as a typical endocrine-disrupting chemical. Growing evidence indicates a strong association between DEHP exposure and the incidence of chronic bone disorders, including osteoporosis (OP), osteoarthritis (OA), and osteonecrosis of the femoral head (ONFH). However, the molecular mechanisms underlying its pathogenic effects across these diseases remain poorly defined. In this study, we applied an environmental network toxicology approach to integrate predicted protein targets of DEHP with known disease-associated targets of the three bone disorders using multiple databases. Through Venn analysis, protein–protein interaction (PPI) network construction, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, we identified core targets and key signaling pathways. Molecular docking and molecular dynamics (MD) simulations were further employed to validate the binding modes and stability between DEHP and the core targets, thereby elucidating common and distinct mechanisms of DEHP across these bone diseases. A total of 109 overlapping targets of DEHP and the three bone diseases were identified, among which 7 core targets—AKT1, SRC, ESR1, CASP3, MMP9, BCL2, and BCL2L1—were common to all three disorders. These are implicated in critical biological processes such as apoptosis regulation, inflammation, extracellular matrix degradation, and estrogen signaling. KEGG enrichment analysis revealed significant involvement of the PI3K-Akt, MAPK, Ras, TNF, and estrogen signaling pathways across all three diseases. Molecular docking and MD simulations confirmed stable binding of DEHP to key targets including AKT1, ESR1, and MMP9, supporting its potential to disrupt bone metabolic homeostasis via multi-target and multi-pathway mechanisms. Further analysis indicated that DEHP exerts both shared and disease-specific effects: it disrupts osteoblast/osteoclast balance in OP, amplifies inflammatory responses and matrix degradation in OA, and contributes to impaired angiogenesis and osteocyte necrosis in ONFH. This study systematically reveals how DEHP disrupts bone homeostasis through a multi-target and multi-pathway network, constructing a cross-disease osteotoxicity framework. It is the first to delineate the common and distinct molecular mechanisms of DEHP in OP, OA, and ONFH. Although these insights are derived from computational models and require further experimental validation, they provide a novel theoretical basis for combined intervention strategies targeting multiple bone diseases and for environmental health risk assessment.

Introduction: Bone, as both a crucial organ with mechanical support and immune regulatory functions, profoundly influencing the growth and development of fish in aquaculture systems. Deer antler and its aqueous extract Cervi Cornus Colla (CCC), are natural medicinal substances known for their bone-protective properties.Methods: This study investigated the osteoprotective effects of CCC against lipopolysaccharide (LPS)-induced injury to fish bones. An LPS-mediated zebrafish skeletal injury model was established and evaluated using Alcian Blue/Alizarin Red staining, fluorescence tracing, ELISA, and qPCR.Results: The median lethal concentration (LC₅₀) of CCC in zebrafish was 206.3 mg/mL, and staining revealed that CCC reversed LPS-induced reduction in cartilage and bone formation in a dose-dependent manner. In Tg (mpeg1: EGFP) zebrafish, CCC treatment significantly decreased macrophages-associated green fluorescence, suggesting that the extract suppressed the activation of macrophages caused by LPS. CCC also reduced the production of several inflammatory factors (IL-6, IL- 1β, and NO). Furthermore, qRT-PCR confirmed CCC restored COL2α1 and Sox9a mRNA levels. Additional studies revealed CCC inhibits LPS-induced activation of the TGF-β/Smad3 signaling pathway.Discussion: This study demonstrates that CCC can alleviate zebrafish bone injury, thereby providing an experimental basis for its application in the prevention and treatment of bone diseases in farmed fish.

Objectives: Atopic dermatitis (AD) is a long-term, recurring inflammatory skin condition characterized by impaired epidermal barrier function and abnormal immune system regulation. Pine pollen has traditionally been used for dermatological treatments, though its active components remain unclear. The primary objective of this study was to pinpoint the active constituents of pine pollen and elucidate its therapeutic effects against AD. Methods: The safety concentration ranges and protective efficacy of nine pine pollen constituents against 2,4-dinitrochlorobenzene (DNCB)-induced HaCaT cell damage were evaluated using the CCK-8 assay. Furthermore, models of DNCB-induced damage were established both in vitro (HaCaT cells) and in vivo (BALB/c mice) to explore the protective effects of the key functional component. Results: Our findings identified pine pollen polysaccharides (PPPS) as the principal bioactive constituent, characterized by a unique infrared absorption spectral profile and a sponge-like architecture with three-dimensional interconnected porous networks. In vitro, PPPS inhibited DNCB-induced decreases in cell viability, morphological abnormalities, oxidative stress, and apoptosis. In vivo, PPPS alleviated DNCB-induced skin lesions by attenuating epidermal hyperplasia, suppressing mast cell infiltration, inhibiting cell apoptosis, and downregulating the expression of IL-4 and IL-17A. Conclusions: This study provides evidence that PPPS from pine pollen can alleviate epidermal damage in AD, offering a novel therapeutic strategy for AD treatment.

To investigate the therapeutic impact of Forsythia suspensa extract (FS) on RSV-infected mice and explore its antiviral pharmacodynamic foundations. Methods: An integrated analytical approach, combining UPLC-Q-TOF/MS with network pharmacology, was employed to analyze and identify the chemical constituents in FS, particularly those exhibiting antiviral properties against RSV. The study integrated network pharmacology and metabolomics for further analysis, and molecular docking and interaction experiments were conducted to validate the pharmacodynamic mechanisms. Finally, an RSV pneumonia mouse model was employed to evaluate the therapeutic influence of FS, including pathological and immunohistochemistry assessments. Twenty-five components in FS were identified through UPLC-Q-TOF/MS analysis. Integrated network pharmacology data revealed 43 effective components and predicted 113 potential targets of FS for anti-RSV activity. Metabolomics analysis identified 14 metabolite biomarkers closely linked to RSV-induced metabolic disruptions involving pathways. Moreover, molecular docking and Biacore experiments provided additional confirmation that FS primarily exerts its effects through compounds such as rutin, quercetin, and kaempferol. Immunohistochemistry experiments demonstrated a significant reduction in the expression of relevant proteins following FS administration, affirming its capacity to ameliorate lung inflammation induced by RSV infection through the modulation of Toll-like receptor signaling pathways. The data presented in this study illustrate that FS exerts its anti-RSV effects by regulating the Toll-like receptor signaling pathway and the arachidonic acid metabolism pathway via rutin, quercetin, and kaempferol. Furthermore, the approach of combining network pharmacology with metabolomics proves to be an effective research strategy for investigating the bioactive constituents of medicinal plants and elucidating their pharmacological effects.

Objectives: PI3K plays a pivotal role in the inflammatory response by modulating the production and release of inflammatory factors. Pik3cb is one of the subunits of PI3K, and its specific role in myocardium inflammation remains unelucidated. This study aimed to investigate the role of Pik3cb in the inflammatory response and to elucidate the underlying mechanism. Methods: An inflammation model was established using H9c2 cells treated with LPS and ATP, and Pik3cb expression was evaluated in this model system. Subsequently, an overexpression model was constructed by transfecting cells with a Pik3cb overexpression plasmid, after which the effects of Pik3cb overexpression on the PI3K/AKT and NF-κB/NLRP3 inflammatory signaling pathways were assessed. Results: These analyses revealed that the expression and distribution of Pik3cb were significantly reduced in the LPS/ATP-induced cellular inflammation model group, whereas plasmid-mediated overexpression of Pik3cb significantly inhibited the activation of the PI3K/AKT signaling pathway in response to LPS/ATP stimulation. Additionally, the LPS/ATP-induced activation of the NF-κB/NLRP3 axis was significantly inhibited following Pik3cb overexpression. Conclusion: This study demonstrates that Pik3cb acts as a negative regulator of LPS/ATP-induced inflammation in cardiomyocytes, exerting anti-inflammatory effects by inhibiting the PI3K/AKT/NF-κB/NLRP3 signaling axis. These findings provide a potential therapeutic target for the treatment of myocardial inflammation.

Cardiomyocyte death caused by hypoxia is one of the main causes of myocardial infarction or heart failure, and mitochondria play an important role in this process. Agrimonolide (AM) is a monomeric component extracted from Agrimonia pilosa L. and has antioxidant, antitumor, and anti‐inflammatory effects. This study aimed to investigate the role and mechanism of AM in improving hypoxia‐induced H9c2 cell damage. The results showed that low AM concentrations promote H9c2 cell proliferation and increase cellular ATP content. Transcriptome sequencing showed that AM induces differential expression of genes in H9c2 cells. Gene ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses revealed that these genes were concentrated in mitochondrial function. Subsequent experiments confirmed that AM regulates hypoxia‐induced cell cycle arrest. AM inhibited the rate of apoptosis by regulating the expression of apoptosis‐related proteins, reducing the level of cleaved Caspase 3 and Bax, and increasing the level of Bcl2, thereby protecting H9c2 cells from hypoxia‐induced apoptosis. AM restored the mitochondrial membrane potential, inhibited the generation of ROS, maintained the normal shape of the mitochondria, improved the level of the mitochondrial functional proteins OPA1, MFN1, MFN2, Tom20, and increased the level of ATP. In conclusion, AM protects H9c2 cells from hypoxia‐induced cell damage.