Search Results (1,438)

This study aimed to investigate the ameliorative effects of glucuronolactone (GL) as a dietary additive on high-fat diet (HFD)-induced growth suppression and metabolic disorders in turbot. A 10-week feeding trial was conducted using juvenile turbot (16.7 ± 0.03 g). Two diets with different protein (%)/lipid (%) levels were formulated: PC (54/12) and NC (47/17). Based on the NC diet, three experimental diets were prepared by supplementing 200 (G200), 400 (G400), and 600 (G600) mg/kg of GL. The present results show that compared to the PC group, HFDs significantly inhibited the growth performance of turbot and induced severe metabolic disorders, hepatointestinal damage, and gut microbiota dysbiosis. Dietary GL supplementation effectively reversed these adverse effects. Specifically, compared to the NC group, GL supplementation significantly restored growth performance, enhanced non-specific immunity, and systematically improved metabolic health. In the liver, GL notably ameliorated tissue damage and downregulated key lipogenic genes (SREBP1, ACC, FAS, PPARγ), while upregulating genes involved in lipid oxidation and catabolism (PPARα1, CPT1, ACOX1, HSL, LPL) and lipid transport (ApoB100, MTP), thereby alleviating hepatic lipid deposition. Furthermore, GL activated the Nrf2/Keap1 antioxidant pathway, up-regulating the expression of genes such as SOD, CAT, GPX, and HO-1. It also suppressed the NF-κB-mediated inflammatory response (downregulation of IL-1β, IFN-γ and TNF-α2; upregulation of IL-10 and TGF-β2) and the mitochondrial apoptosis pathway (increased Bcl-2/Bax ratio; downregulation of Caspase3/7/9), collectively mitigating oxidative damage and cellular apoptosis. Moreover, GL restored intestinal morphology, enhanced the expression of tight junction proteins (Claudin-3, Claudin-7, ZO-1, Occludin) and MUC2, and inhibited MLCK signaling. These improvements led to a reduction in serum D-LA levels, indicating strengthened intestinal barrier function. Concurrently, GL reshaped the gut microbiota composition by enriching beneficial bacteria such as Akkermansia and suppressing potential pathogens like Listeria. In summary, GL effectively alleviated HFD-induced growth suppression and metabolic damage in turbot by improving lipid metabolism and alleviating hepatic injury, while concurrently restoring intestinal barrier integrity and microbiota homeostasis. Full article

Background: To investigate the effects of probiotics and their postbiotics on mouse health, this study utilized healthy mice randomly assigned to a control group (CK, n = 6), a probiotic group (L, n = 6, oral gavage 200 μL Pediococcus lactis), and a postbiotic group (PL, n = 6, oral gavage 200 μL Pediococcus lactis postbiotic). Methods: Following 21 days of continuous intervention, changes in gut metabolic profiles, microbial community structure, tissue morphology, and tight junction protein expression were systematically analyzed using metabolomics, 16S rRNA sequencing, hematoxylin and eosin (HE) staining, and immunohistochemistry techniques. Results: The results revealed that screening for significantly altered endogenous metabolites identified core differences concentrated in metabolites related to intestinal barrier repair, anti-inflammation, and antioxidant activity (e.g., 3-indolepropionic acid, astaxanthin, hydroxybenzoic acid). 16S rRNA sequencing revealed that the overall community structure was relatively stable according to principal component analysis, although differences were detected in specific taxa. However, LEfSe analysis identified significantly enriched functional microbial groups at multiple taxonomic levels in the PL group: phylum: Actinomycetota; class: Coriobacteriia; order: Coriobacteriales, Erysipelotrichales; family: Erysipelotrichaceae, Eggerthellaceae; genus: norank_Erysipelotrichaceae, Intestinimonas. These results suggest that although the overall community structure remained relatively stable, specific taxa may have differed between groups. Hematoxylin and eosin staining revealed no pathological lesions in intestinal tissues from either group, with intact mucosal architecture. Immunohistochemistry demonstrated significantly elevated expression of intestinal tight junction proteins Claudin 1, MUC-2, Occludin, and ZO-1 in the PL group compared to the CK group (p < 0.001). Conclusions: In summary, this probiotic (Pediococcus lactis) and its postbiotic showed promising effects, which may be related to changes in specific microbiota taxa, intestinal metabolic profiles, and tight junction protein expression. Beyond maintaining gut microbiota and tissue homeostasis, it enhances intestinal barrier function, suppresses latent inflammation, and boosts antioxidant capacity. Postbiotics may exhibit superior efficacy compared to probiotics. This provides robust experimental evidence for its development and application in gut health products for healthy populations. However, these findings still require further validation in studies with longer intervention periods and in disease models. Full article

Exosomes (EXs) mediate intercellular communication in the tissue microenvironment. We previously demonstrated that endothelial progenitor cell-derived exosomes (EPC-EXs) from exercised mice protect neurons and cerebral endothelial cells from hypoxia- and hypertension- induced injury ex vivo, suggesting their therapeutic potential in hypertensive ischemic injury. Here, we investigated whether exercise-conditioned EPC-EXs (ET-EPC-EXs) confer protection against acute ischemic injury. Hypertensive transgenic mice were divided into donor and recipient groups. Donor mice underwent treadmill exercise to generate ET-EPC-EXs. Recipient mice was subjected to middle cerebral artery occlusion and received ET-EPC-EXs via tail vein injection (2 × 10⁸/100 μL saline) two hours after stroke onset. Cerebral blood flow (CBF) was assessed, and brains were collected on day two for histological and molecular analyses. Our data showed that ET-EPC-EXs were robustly taken up by cerebral cells, predominantly in the penumbra in the ipsilateral hemisphere. ET-EPC-EXs reduced cell death and microglia activation and restored tight-junction proteins. Moreover, ET-EPC-EX treatment preserved CBF and improved sensorimotor function on day two post-stroke. Mechanistically, ET-EPC-EXs suppressed p38 activation, accompanied by reduced matrix metalloproteinase-3 and cytochrome c levels in the ipsilateral brain. Collectively, these findings demonstrate that EPC-EXs from exercise mice improve sensorimotor functions and confer protection in hypertensive ischemic brain injury, likely through attenuation of neuroinflammation and preservation of vascular integrity via modulation of the p38 signaling. Full article

Background/Objectives: Ulcerative colitis (UC) involves inflammatory response, oxidative stress, changes in metabolites, and the gut microbiota. Jingangteng capsule (JGTC) has been utilized clinically for the treatment of inflammatory diseases for many years. However, the efficacy of JGTC in ameliorating UC remains unclear, and the underlying mechanisms have not yet been elucidated. This study aims to investigate the effect and mechanism of JGTC on UC. Methods: The chemical compositions of JGTC were examined using ultra-high-performance liquid chromatography with quadrupole time-of-fight mass spectrometry. The anti-UC effect of JGTC was evaluated by assessing the disease activity index (DAI), colon length, intestinal barrier recovery, and inflammatory factors in a dextran sulfate sodium (DSS)-induced colitis model. Mechanisms were investigated through fecal 16S rDNA sequencing, metabolomics analysis, enzyme-linked immunosorbent assay (ELISA), Western blotting, and network pharmacology analysis. Results: JGTC significantly reduced the DAI scores in UC mice, increased their body weight and colon length (p < 0.001), repairing damaged intestinal tissue. It decreased the levels of inflammatory cytokines TNF-α, IL-6, IL-1β, and LPS (p < 0.01, p < 0.001), alleviating intestinal inflammation. It also raised the expression of tight junction proteins ZO-1, Claudin-1, and Occludin (p < 0.05, p < 0.001), thereby enhancing intestinal barrier function. Fecal metabolomic analysis revealed that the favorable alterations in amino acid and lipid metabolites were more pronounced. Heat maps showed strong correlations between pharmacological indicators and gut microbiota, as well as between the main differential metabolites and gut microbial communities. UPLC-QTOF-MS detection yielded 33 components of JGTC, and network pharmacology analysis based on these components predicted pathways of action of JGTC in UC. Functional pathways closely associated with significantly differential metabolites and metabolic pathways were also investigated. The PI3K-AKT-mTOR pathway was one of them, which is consistent with the conclusions drawn from network pharmacology. JGTC significantly modulated key factors in this pathway, inhibiting the expression of PI3K, Akt, PDK1, and mTOR, while augmenting the expression of PTEN (p < 0.05, p < 0.01, p < 0.001). It also mitigated the levels of related oxidative stress factors MDA, MPO, and D-LA, and raised SOD levels (p < 0.01, p < 0.001). Conclusions: JGTC improved the excessive inflammatory response in UC by regulating intestinal flora and metabolic disorders, affecting the PI3K-AKT-mTOR signaling pathway, restoring intestinal tissue damage and intestinal barrier, and inhibiting inflammatory and oxidative stress factors. Full article

Inflammatory bowel disease (IBD) is a chronic gastrointestinal disorder with a high incidence of anxiety and depression. However, the underlying mechanisms of these symptoms remain to be fully elucidated. This study investigated the effects and mechanisms of a 20% ethanolic extract of Petasites japonicus leaves (EPJ) on dextran sulfate sodium (DSS)-induced colitis and depression-like behaviors. The physiological compounds identified in the EPJ were citric acid, chlorogenic acid, caffeic acid, fukinolic acid, 3,5-dicaffeoylquinic acid, quercetin 3-O-β-D-glucose-6″-acetate, 4,5-dicaffeoylquinic acid, kaempferol-3-O-(6″-acetyl)-β-glucopyranoside, and pedunculoside. EPJ significantly alleviated DSS-induced colitis, as evidenced by improvements in body weight loss (87.41% vs. 76.02% in the DSS group), colon length (5.75 vs. 4.34 cm), intestinal permeability (52.80 vs. 163.01 μg/mL), and myeloperoxidase (MPO) activity (0.24 vs. 0.67 U/mg) (p < 0.05). Histological analysis further confirmed recovery of goblet cells and attenuation of muscle layer thickening. EPJ also reversed DSS-induced gut microbiota dysbiosis and contributed to the restoration of microbial homeostasis. Behavioral assessments showed that EPJ effectively ameliorated depression-like behaviors. EPJ improved antioxidant systems in colon and brain tissues by modulating malondialdehyde (MDA) levels and reduced glutathione (GSH) and superoxide dismutase (SOD) activity. EPJ further upregulated tight junction protein expression and suppressed TLR4/NF-κB inflammatory pathway activation in both colon and brain tissues. Moreover, EPJ modulated serum stress-related hormones, normalized hypothalamic–pituitary–adrenal (HPA) axis dysregulation, regulated the BDNF/TrkB signaling pathway, and modulated tryptophan–kynurenine metabolism. Collectively, these findings suggest that EPJ exerts protective effects against DSS-induced colitis and depression-like behaviors. Full article

This study investigated the synergistic effects of liquid fermentation of rapeseed meal (RSM) on feed microbiota, growth performance, and muscle development in growing pigs. RSM was fermented using four compound probiotics and eleven enzyme preparations, and microbial changes were analyzed using 16S rRNA sequencing. Seventy-two Duroc × Jingfen White pigs were randomly assigned to three groups: soybean meal (Ctrl), RSM, and fermented RSM (FRSM). FRSM showed higher trichloroacetic acid-soluble protein (TCA-sp) content and significantly lower neutral detergent fiber (NDF), acid detergent fiber (ADF), anti-nutritional factors (ANFs), and toxins (TS) (p < 0.01). Fermentation increased microbial diversity, with higher abundances of Lactobacillus and Pediococcus. Compared with Ctrl and RSM, the feed-to-gain ratio (F/G) decreased in the FRSM group (p < 0.01). FRSM also improved serum antioxidant capacity, enhanced intestinal villus height (VH)and villus height/crypt depth ratio (VH/CD), and upregulated the expression of tight junction proteins (ZO-1, occludin) and the anti-inflammatory factor IL-10 (p < 0.01). FRSM group also increased myofiber diameter and cross-sectional area in the longissimus dorsi and elevated MyoD, MyoG and Myf5 expression (p < 0.01). RNA-seq revealed 2094 differentially expressed genes enriched in metabolic pathways. Overall, FRSM improved growth performance, intestinal health, and muscle development in growing pigs, which may guide the development of protein resource utilization technologies. Full article

Taurine is abundant in seminal plasma and is involved in redox balance, osmoregulation, and sperm membrane stability. However, its role in protecting dairy goat bucks against heat stress-associated declines in sperm quality remains unclear. In this study, eighteen Guanzhong dairy goat bucks were assigned to three groups: control (NC), field heat stress (HS), and HS with taurine supplementation (HS + Tau). Heat stress reduced seminal plasma taurine abundance and was associated with metabolic reprogramming, impaired sperm quality, disturbed redox homeostasis, and decreased LH and testosterone levels. Specifically, HS reduced sperm motility, viability, membrane integrity, and kinematic performance, increased sperm abnormalities, and shortened in vitro sperm survival time. Taurine supplementation alleviated these adverse changes and shifted the seminal plasma metabolome toward a more homeostatic profile. Metabolomic analysis indicated that HS was associated with the accumulation of long-chain acylcarnitines in seminal plasma. Complementary mouse and TM4 Sertoli cell experiments provided preliminary mechanistic support, suggesting that taurine may partially protect Sertoli cell tight-junction proteins, particularly ZO-1, under heat- and acylcarnitine-related stress, and may be associated with the modulation of p38/AKT signaling. Collectively, these findings suggest that taurine alleviates heat stress-induced declines in sperm quality in dairy goat bucks, at least in part, by modulating seminal plasma metabolism. Full article

Background: Total glycosides of peony (TGP) have therapeutic potential for immune-related and inflammatory skin diseases, but their skin absorption is not satisfactory. This study aims to investigate how Evodia rutaecarpa volatile oil (VO-ER) enhances the permeability of TGP. Methods: Safety assessment was conducted through cell delivery and skin erythema tests. The chemical composition of VO-ER was identified via GC-MS. The study was conducted using modified Franz diffusion cells, microdialysis, confocal laser scanning microscopy (CLSM), attenuated total reflection–Fourier transform infrared spectroscopy (ATR-FTIR), molecular docking and molecular dynamics simulations (MD), laser Doppler flowmetry (LDF), and the western blotting method. Results: The study found that VO-ER promotes the permeation of total glycosides of peony in a concentration-dependent manner by disrupting the intercellular lipid tissue structure, downregulating the expression of claudin-1, claudin-7, and occludin, and improving local microcirculation, thereby promoting the absorption of TGP. Conclusions: VO-ER enhances the transdermal absorption of TGP through multiple mechanisms, such as disrupting the skin lipid barrier, downregulating tight junction proteins, and improving local skin microcirculation. This study provides a theoretical basis for VO-ER as a safe and effective new transdermal penetration enhancer, offering support for the development of topical preparations containing Evodia rutaecarpa and Paeonia lactiflora. Full article

The intestinal epithelial barrier regulates paracellular transport, and its dysfunction is associated with inflammatory and metabolic diseases. Among dietary fibers, inulin has attracted considerable attention due to its beneficial effects on intestinal health. Inulin’s actions have been attributed to protecting the structure and function of gut barrier components against inflammatory-associated damage. This review integrates preclinical and clinical studies evaluating the impact of inulin on intestinal permeability. Evidence from in vitro and in vivo models shows that inulin regulates the expression of tight junction proteins (TJPs), Paneth cell proliferation, and antimicrobial peptides, and modulates inflammatory signaling pathways. In addition, inulin prebiotic activity, via microbiota, stimulates the production of short-chain fatty acids (SCFAs) as butyrate that reinforces the barrier function. Understanding these pathways highlights the therapeutic potential of inulin as a nutritional strategy for treating barrier dysfunction. Clinical studies in obesity, metabolic disorders and inflammatory intestinal disease have associated inulin supplementation with improvements in biomarkers of intestinal permeability. Future studies are needed to test inulin’s safety in order to prevent potential risks and hazards. Full article

Tumor metastasis is closely associated with coagulation and inflammation, particularly via thrombin–PAR1 signaling. However, the potential of natural polysaccharides such as rhamnan sulfate (RS) to modulate these pathways and suppress metastasis remains unclear. We aimed to investigate the effects of orally administered RS derived from Monostroma nitidum on melanoma metastasis and its underlying mechanisms. Male C57BL/6J mice were orally administered water or RS daily. On day 8, saline or B16 melanoma cells were injected intravenously. Mice were treated for 21 days and divided into four groups (control, RS-only, M + W, and M + RS; n = 5/group). Metastasis and related molecular factors were analyzed in plasma, lung, and aortic tissues. Significant lung and aortic metastases were observed in the M + W group but were markedly suppressed in the M + RS group. RS reduced the expression of inflammatory factors (e.g., IL-6, PAR1), proteases, leukocyte activation markers, complement factors, angiogenic factors, and EMT-related factors. Conversely, thrombin, thrombomodulin, plasmin, TAFIa, and tight junction proteins were increased in RS-treated mice. RS suppresses melanoma metastasis by modulating thrombin–PAR1-mediated inflammation and associated pathways. These findings suggest RS as a potential therapeutic agent, although further mechanistic and clinical studies are required. Full article

This study investigated the effects of dietary lysophospholipids on growth performance, hepatic lipid metabolism, intestinal health, and dietary lipid levels of largemouth bass. The 56-day experiment included five groups: CON (0% lysophospholipids), LL50 (0.05% lysophospholipids), LP50 (0.05% lysophospholipids—0.5% oil), LP100 (0.1% lysophospholipids—1.0% oil), and LP200 (0.1% lysophospholipids—2.0% oil), with 3 replicates (30 fish/replicate) per group. The results showed that compared with the CON group, dietary supplementation of 0.05% lysophospholipid had no significant effect on the growth performance of largemouth bass, but increased the crude protein content and decreased the crude lipid content in the whole body. An amount of 0.05% lysophospholipid improved hepatic lipid utilization efficiency. Specifically, this supplementation level promoted serum lipid transport (increased serum HDL-C content and decreased triglyceride and LDL-C contents), and enhanced hepatic lipid metabolism by regulating the expression of lipid metabolism-related genes (fas, hsl, and acc) and the levels of lipid metabolites (phosphatidylcholine and fatty acids), thereby reducing hepatic triglyceride content. In addition, 0.05% lysophospholipid improved intestinal health by increasing lipase activity and intestinal villus height, up-regulating the expression of the anti-inflammatory gene (tgf-β1) and tight junction protein genes (claudin-1, claudin-4, and zo-1), and down-regulating the expression of the pro-inflammatory gene (tnf-α). In terms of dietary lipid reduction, supplementation with 0.1% lysophospholipid allowed a 1% reduction in dietary lipid level without affecting the growth performance of largemouth bass, whereas at the same level of lysophospholipid supplementation, a 2% reduction in dietary lipid level resulted in decreased growth performance of largemouth bass. These findings provide theoretical support for the practical application of lysophospholipids, and demonstrate that reducing dietary lipid inclusion by adding lysophospholipids helps to reduce feed costs and improve aquaculture economic benefits. Full article

The purpose of our study was to assess if spinacetin (SPC), a flavonoid found in spinach, can alleviate the cyclophosphamide (CYP)-induced changes in cystometric and inflammatory parameters indicative of the development of hemorrhagic cystitis. The animal experiments were conducted in female Wistar rats. The cohort of 60 animals was grouped as follows: I—control, II—CYP group, III—SPC group, and IV—CYP + SPC group. The cystometry and biochemical analyses were performed after a fortnight of SPC administration. SPC was found to restore normal cystometric parameters in CYP-induced cystitis and, similarly, it normalized c-Fos expression changes in the central micturition regions. SPC further prevented a massive increase in the bladder wall thickness/permeability due to exposition to CYP administration. CYP instillation resulted in the elevation of biomarkers found in urine (brain-derived neurotrophic factor, BDNF, and nerve growth factor, NGF), and in the bladder detrusor muscle (Rho kinase and vesicular acetylcholine transporter, VAChT), which were successfully restored after administration of SPC. As for the biomarkers in the bladder urothelium, the CYP-induced increases in TNF-α, IL-1β, IL-6, calcitonin gene-related peptide (CGRP), malondialdehyde, 3-nitrotyrosine, insulin-like growth factor-binding protein 3 (IGFBP-3), occludin, organic cation transporter 3 (OCT-3), orosomucoid-1 (ORM1), pituitary adenylate cyclase receptor 1 (PAC1), synaptosomal-associated protein 23 (SNAP23), SNAP25, and synaptic vesicle glycoprotein (SV2A) levels were attenuated by SPC. Finally, CYP administration resulted in a decrease in the heparin-binding EGF-like growth factor (HB-EGF), hemopexin (HPX), T-H protein, and tight junction protein (Z01), and we noted the successful restoration of all these changes in concentrations after application of SPC. In summary, SPC robustly mitigated cyclophosphamide (CYP)-induced cystometric dysfunction and biochemical alterations characteristic of iatrogenic hemorrhagic cystitis. These findings position SPC as a compelling therapeutic candidate and warrant further translational investigation for the management of CYP-induced bladder injury. Full article

Exposure of healthy tissue to ionizing radiation (IR) occurs due to nuclear accidents and terrorism, as well as radiotherapy. The vascular endothelium is a key target of IR, and microvascular endothelial cells (ECs) are particularly vulnerable to radiation. IR induces EC activation leading to endothelial cell injury. Human ghrelin is a stomach-derived peptide with pleiotropic effects, including protection against inflammation. We hypothesize that human ghrelin improves survival in total body irradiation (TBI) and that ghrelin’s protective effect could be mediated by attenuating endothelial cell injury. To test this, mice were exposed to TBI and after 24 h were treated subcutaneously with human ghrelin once daily for 4 days and monitored for 30 days. The survival rate of the human ghrelin-treated group was significantly higher than that of the vehicle group. Subsequently, human ghrelin treatment showed an effective dose modification factor of 1.0681. On day 4 after TBI, human ghrelin significantly attenuated EC permeability in the lungs and improved tight junction protein ZO-1 expression. Human ghrelin also improved ZO-1 and Claudin5 expression in primary mouse lung vascular endothelial cells. Taken together, these results indicate that human ghrelin improves survival after TBI, and its survival benefit is in part due to the attenuation of EC permeability and microvascular barrier dysfunction. Full article

Background and Objectives: Pro-inflammatory diet and high-fat diet (HFD) often coexist in real-world, but their combined impact on the gut–liver axis and potential nutritional countermeasures remain insufficiently studied. This study aimed to evaluate a pro-inflammatory–high-fat composite dietary pattern on the intestine and liver in the population, and to further evaluate the protective potential of astaxanthin (ATX) in complementary experimental systems. Methods: Data from the NHANES 2005–2010 were used to construct four composite exposure groups based on the dietary inflammation index (DII) and energy from fat. Survey-weighted regression analyses were performed to examine associations with systemic inflammation and liver injury. Interaction and C-reactive protein (CRP)-mediated effect analyses were conducted. Fifty SD rats were randomly divided into control group, model group induced by HFD combined with inflammatory factors, and low-, medium-, and high-dose Haematococcus pluvialis (HP) intervention groups. Serum lipids, liver enzymes, liver and colon pathology, and inflammatory and oxidative markers were measured in rats. In an in vitro organ-on-chip barrier model, the effect of ATX was observed when colonic barrier damage was induced using palmitic acid and lipopolysaccharides. Results: The high DII combined with HFD showed the largest increases in CRP, liver enzymes, and fatty liver index. A synergistic interaction was observed between DII and HFD, with CRP mediating approximately 20% of the effect. In rat model, HP-derived ATX improved the lipid profile, attenuated hepatic steatosis and oxidative damage, and reduced colonic pro-inflammatory cytokines, while restoration of tight junction proteins was limited. In colon organoid model, ATX showed limited efficacy in improving inflammation and barrier function. Conclusions: The pro-inflammatory–high-fat dietary pattern synergistically exacerbates gut–liver dysfunction. HP-derived ATX alleviates metabolic and inflammation-induced enterohepatic comorbidity, but its effect on repairing barrier structure is limited. Full article

Alterations in tight junction (TJ) organization and dysregulation of cancer stem cell (CSC)-associated markers are increasingly recognized as molecular features linked to colorectal cancer (CRC) progression, heterogeneity and clinical outcome. Bioactive dietary compounds such as spermidine (SPD) and eugenol (EUG) have been proposed as modulators of cancer-related molecular pathways; however, their combined effects on CRC spheroid models relevant to molecular characterization remain insufficiently defined. In the present study, the molecular impact of SPD and EUG, administered individually or in combination, was evaluated in primary and metastatic CRC spheroids. First-generation spheroids derived from Caco-2 and SW620 cells were exposed to SPD, EUG, or SPD+EUG at the time of seeding, and spheroid growth and self-renewal capacity were monitored across successive generations. The expression of TJ- and CSC-associated markers was assessed at both the transcript and protein levels using reverse transcription–quantitative polymerase chain reaction (RT-qPCR), Western blotting and immunohistochemistry. The combined SPD+EUG treatment was associated with a marked reduction in spheroid area and self-renewal capacity in both CRC models. Baseline molecular profiling revealed higher TJ marker expression in Caco-2 spheroids and enrichment of CSC-associated markers in SW620 spheroids. Treatment-induced modulation of CSC- and TJ-related transcripts was observed; however, transcript-level changes were not consistently mirrored at the protein level, indicating the involvement of post-transcriptional regulatory mechanisms. In particular, Occludin (OCLN), Zonula occludens-1 (ZO-1), CD133, ALDH1A1, SOX2 and VE-cadherin exhibited divergent RNA and protein expression patterns depending on cell type and treatment condition. Collectively, these findings underscore the relevance of three-dimensional CRC spheroid models for molecular profiling studies and highlight the importance of integrating transcript- and protein-level analyses when evaluating bioactive compounds with potential diagnostic and translational relevance in colorectal cancer. Full article