Obesity is a major global health burden that is linked to type 2 diabetes, cardiovascular disease, and metabolic syndrome. Chitosan oligosaccharides (COS) are bioactive compounds that are derived from the depolymerization of the chitosan in crustacean shells and are promising candidates for natural anti-adipogenesis effects. However, there is incomplete understanding of the molecular mechanisms by which structurally defined low-molecular-weight COS modulates adipogenic transcription networks and global transcriptional reprogramming. MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) mass spectrometry and ¹³C NMR spectroscopy indicated a predominance of dimeric species (DP2) at m/z 344.79, which represents a lower molecular weight fraction and is proposed to improve the membrane permeability and intracellular bioavailability of COS. In a 3T3-L1 preadipocyte model, COS treatment at concentrations of 320–1280 µg/mL dose-dependently reduced intracellular lipid accumulation, triglyceride content, and adipocyte maturation while enhancing lipolysis and insulin-mediated glucose uptake. Western blot analysis indicated dose-dependent downregulation of PPARγ and C/EBPα. Transcriptomic RNA-seq analysis indicated large-scale transcriptional reprogramming with the altered expression of genes involved in PPAR signaling, PI3K-Akt, AMPK, insulin signaling, and fatty acid metabolism pathways among differentially expressed genes. These findings demonstrate that COS suppresses adipogenesis through the coordinated modulation of adipogenic transcription factors and multiple metabolic signaling pathways. The results support its potential as a promising natural compound but warrant preclinical investigation in the context of obesity and metabolic disorders.

Holothuria scabra has long been acknowledged in traditional medicine for its therapeutic properties. The transforming growth factor-beta (TGF-β) superfamily is crucial in regulating cellular processes, including differentiation, proliferation, and immune responses. This study marks the first exploration of the gene expression localization, sequence conservation, and functional roles of H. scabra TGF-β proteins, specifically activin (HolscActivin), inhibin (HolscInhibin), and the TGF-β receptor (HolscTGFBR), across various organs. In situ hybridization indicated that HolscActivin and HolscInhibin are expressed in the intestine, respiratory tree, ovary, testis, and inner body wall. This suggests their roles in nutrient absorption, gas exchange, reproduction, and extracellular matrix remodeling. Notably, HolscTGFBR demonstrated a similar tissue-specific expression pattern, except for its absence in the respiratory tree. Bioinformatics analysis reveals that HolscTGFBR shares significant sequence similarity with HomsaTGFBR, especially in regions essential for signal transduction and inhibition. Molecular docking results indicate that HolscActivin may promote receptor activation, while HolscInhibin functions as a natural antagonist, reflecting the signaling mechanisms of human TGF-β proteins. Interestingly, cross-species ternary complex docking with human TGF-β receptors further supports these findings, showing that HolscActivin moderately engages the receptors, whereas HolscInhibin exhibits strong binding, suggestive of competitive inhibition. These results indicate that H. scabra TGF-β proteins retain the structural and functional features of vertebrate TGF-β ligands, supporting their potential applications as natural modulators in therapeutic and functional food development.

Crustacean shell waste from the seafood industry represents an underutilized food waste with the potential to produce chitosan and chitosan oligosaccharides (COS), with promising applications in bone regeneration. However, the molecular mechanisms of COS during osteogenic differentiation remain uncharacterized. In this study, chitosan was extracted from mud crab (Scylla olivacea) shell waste and COS was prepared from chitosan by acid hydrolysis and then characterized by FT-IR and FESEM, confirming the chitosan backbone and a degree of deacetylation (DD) exceeding 80%. The COS treatment in the concentration range of 5–320 µg/mL significantly enhanced MC3T3-E1 pre-osteoblast proliferation without cytotoxicity, and it increased alkaline phosphatase (ALP) activity, extracellular matrix mineralization, and Runx2/osterix (OSX) expression. Transcriptomics of pre-osteoblasts treated with COS at 320 µg/mL for 21 days identified 19,643 differentially expressed genes, revealing coordinated activation of multiple osteogenesis-related signaling pathways, including ECM–integrin/focal adhesion, PI3K-Akt/MAPK, TGF-β/BMP, Wnt/β-catenin, and parathyroid hormone (PTH) synthesis. These findings demonstrate that marine-derived COS promotes osteoblast differentiation through the coordinated activation of multiple signaling pathways, providing the first comprehensive transcriptomic characterization of MC3T3-E1 pre-osteoblast directly treated with COS during osteogenic differentiation. Therefore, this study supports the use of COS as a promising bioactive oligosaccharide for bone regeneration and osteoporosis management, and further in vivo and clinical studies are recommended.

Oxidative stress-induced macrophage activation and vascular injury are major contributors to atherosclerosis, a chronic inflammatory disease associated with approximately 20 million deaths worldwide. This study investigated the antioxidant and anti-inflammatory activities of structurally characterized low-molecular-weight chitosan oligosaccharides (COS) derived from mud crab (Scylla olivacea) shell waste by hydrochloric acid hydrolysis and explored their molecular interactions with inflammation-related targets. Structural characterization by ¹³C-NMR and MALDI-TOF confirmed the identity of COS, while DPPH and ABTS analysis demonstrated concentration-dependent antioxidant activity. In LPS-induced RAW 264.7 macrophages, COS showed no cytotoxicity and significantly reduced nitric oxide production at 80 and 160 µg/mL. Molecular docking predicted favorable interactions of COS with several inflammation-associated proteins, with iNOS and COX-2 exhibiting the most favorable docking scores and extensive hydrogen-bonding and polar interactions. Transcriptomic profiling further revealed broad transcriptional remodeling and enrichment of pathways related to inflammation and atherosclerosis, including TNF, NF-κB, MAPK, Toll-like receptor, and lipid-and-atherosclerosis signaling. COS markedly suppressed inflammatory mediators, particularly Nos2 (iNOS) and Ptgs2 (COX-2) together with multiple cytokines and chemokines, consistent with reduced nitric oxide production and modulation of macrophage activation and foam cell-associated processes. Integration of docking and transcriptomic analyses identified iNOS and COX-2 as convergent candidate anti-inflammatory targets of COS, supporting its ability to attenuate inflammatory signaling through relevant pathways. These findings suggest that COS may modulate macrophage inflammatory responses through coordinated regulation of oxidative stress and inflammation-related signaling pathways.

The reliance on synthetic repellents such as N,N-diethyl-meta-toluamide (DEET) has raised health and environmental concerns, prompting the search for safer, plant-derived alternatives. Catnip (Nepeta cataria L.) is a rich source of iridoid monoterpenes, particularly nepetalactones, which are well known for their strong insect-repellent properties. However, the efficient extraction of nepetalactones remains challenging, and their precise mechanisms of action in insect inhibition are not yet fully understood. Thus, this study investigated the chemical composition from various methods, protein–ligand interactions, and pharmacokinetic safety profiles of catnip-derived compounds compared to DEET, with a focus on their interactions with odorant-binding proteins (OBPs) from Anopheles gambiae (AgamOBP), Culex quinquefasciatus (CquiOBP), and Aedes aegypti (AaegOBP). Gas chromatography–tandem mass spectrometry (GC–MS/MS) confirmed the presence of nepetalactone isomers as the major constituents in catnip extracts obtained through stream distillation and dried leaves extracted in olive oil fractions. Molecular docking revealed that cis,cis- and cis,trans-nepetalactones and nepetalactone exhibited high binding affinities, surpassing those of DEET. Molecular dynamics simulations demonstrated that all OBP–ligand complexes achieved stable conformations. Notably, cis,trans-nepetalactone formed a more stable complex with AgamOBP than DEET. These findings suggest that nepetalactones stabilize OBP–ligand interactions while inducing subtle conformational flexibility, potentially disrupting mosquito odorant recognition in a manner distinct from DEET. ADMET predictions indicated that nepetalactones exhibit favorable absorption, distribution, and safety profiles with reduced predicted toxicity compared to DEET. Collectively, these results establish nepetalactones as promising candidates for the development of effective, safe, and sustainable plant-based repellents.

The reliance on synthetic repellents such as N,N-diethyl-meta-toluamide (DEET) has raised health and environmental concerns, prompting the search for safer, plant-based alternatives. Catnip (Nepeta cataria L.), a rich source of iridoid monoterpenes, particularly nepetalactones, known for strong insect-repellent activity. However, their efficient extraction and molecular mechanisms in insect inhibition remains challenging. This study examined the chemical composition, protein–ligand interactions, and safety profiles of nepetalactones in comparison with DEET, with particular focus on mosquito odorant-binding proteins (OBPs) from Anopheles gambiae (AgamOBP), Culex quinquefasciatus (CquiOBP), and Aedes aegypti (AaegOBP). GC–MS/MS analysis identified nepetalactone isomers as the predominant constituents in catnip extracts obtained via steam distillation and olive oil extraction from dried leaves. Molecular docking results indicated that cis,cis-, cis,trans-, and nepetalactone isomers exhibited higher binding affinities toward the target OBPs than DEET. Furthermore, molecular dynamics simulations confirmed that all nepetalactone–OBP complexes exhibited stable conformations characterized by low average RMSD values and persistent hydrogen bond formation. Notably, cis,trans-NL–AaegOBP, NL–AaegOBP, and cis,cis-NL–AgamOBP complexes displayed lower binding free energies (ΔG_MM-PBSA) compared to DEET. These findings suggest that nepetalactones stabilize OBP–ligand interactions while inducing subtle conformational flexibility, potentially disrupting mosquito odorant recognition in a manner distinct from DEET. ADMET predictions indicated that nepetalactones exhibit favorable absorption, distribution, and safety profiles with reduced predicted toxicity compared to DEET. Collectively, these results establish nepetalactones as promising candidates for the development of effective, safe, and sustainable plant-based repellents.