The necessity of iodinated contrast agents in radiographic diagnosis comes with an increased risk of kidney injury. The sclerotium of Poria cocos (Schw.) Wolf, a traditional Chinese medicine, exhibits anti-inflammatory and diuretic pharmacological properties. Its extracted polysaccharides demonstrate anti-apoptotic, antioxidant, and anti-inflammatory activities. This study aimed to investigate the effects and potential mechanisms of Poria cocos polysaccharides (PCP) on contrast-induced acute kidney injury (CI-AKI). The purified PCP was characterized using FTIR and monosaccharide analysis. A CI-AKI mouse model was established to evaluate its preventive effects via oral gavage. Mitochondrial structure, function, and biogenesis were assessed by measuring mitochondrial membrane potential (MMP), ROS, ATP, and mitochondrial DNA copy number. Antioxidant capacity and apoptosis levels were evaluated by detecting 8-OHdG, caspase 3/7 activity, cleaved-PARP, MDA, and GSH. Finally, the mechanism of action of PCP was analyzed using qPCR and immunoblotting. The results showed that PCP significantly mitigated CI-AKI by inhibiting contrast-induced apoptosis and oxidative stress in kidney tissue. Furthermore, PCP maintained mitochondrial homeostasis in the kidneys by enhancing mitochondrial biogenesis and antioxidant capacity. These findings were consistent with in vitro data. Notably, PGC-1α was identified as a key factor in the nephroprotective effects of PCP, with its activation attributed to the CaMKK2-AMPK signaling pathway. These findings suggest that PCP offers significant benefits in mitigating CI-AKI by enhancing antioxidant defenses and promoting mitochondrial biogenesis through activation of the CaMKK2/AMPK/PGC-1α signaling pathway in the kidney. Therefore, PCP could be a potential preventive agent for CI-AKI.
Malvidin and its derivatives exhibit potent anti-inflammatory and antioxidant properties, yet their roles and mechanisms in infection-related acute lung injury (ALI) remain unclear. This study aimed to investigate the protective effects of malvidin-3-O-rhamnoside (Mv3rh) in Klebsiella pneumoniae (KP)-induced ALI and to elucidate the underlying mechanisms. A mouse model of KP-induced ALI was employed to assess the effects of Mv3rh on lung pathology, bacterial burden, myeloperoxidase (MPO) activity, alveolar macrophage proportion, cytokine levels, and survival. Mechanistic studies in alveolar macrophages included RNA sequencing, detection of reactive oxygen species (ROS), and evaluation of inflammasome activation, pyroptosis, and antioxidant responses using quantitative PCR (qPCR), Western blotting, flow cytometry, immunoprecipitation, and immunofluorescence. Potential interactions between Mv3rh with NOD-like receptor pyrin domain-containing 3 (NLRP3) or the Toll-like receptor 4/MD-2 complex were examined by molecular docking, cellular thermal shift assay (CETSA), and the interaction with TLR4 was further assessed by surface plasmon resonance (SPR). In vivo, Mv3rh pretreatment significantly mitigated KP-induced ALI, as evidenced by improved lung histopathology, reduced alveolar septal thickening, edema, and neutrophil infiltration, together with improved survival. These protective effects were accompanied by reduced systemic cytokine levels, lower bacterial burdens in the lung and blood, and partial restoration of alveolar macrophage-like cells in bronchoalveolar lavage fluid (BALF). In alveolar macrophages, Mv3rh suppressed TLR4/NF-κB signaling, reduced NLRP3 inflammasome priming and activation, inhibited cleavage of caspase-1 and GSDMD-N, and attenuated macrophage pyroptosis. In addition, Mv3rh activated the NRF2/HO-1 pathway, promoted NRF2 nuclear translocation, and restored antioxidant enzyme activities, including glutathione peroxidase (GPX), superoxide dismutase (SOD), and catalase (CAT). Mv3rh attenuates inflammation, oxidative stress, and macrophage pyroptosis in KP-induced ALI by regulating the TLR4/NF-κB/NLRP3 inflammasome axis and the NRF2/HO-1 antioxidant pathway. These findings suggest that Mv3rh possesses immunomodulatory activity and may be beneficial in KP-associated ALI.
Organic anion-transporting polypeptide (OATP) transporters have a significant influence on the disposition of drugs, significantly influencing their efficacy and toxicity. Humans express two transporters (OATP1B1 and OATP1B3), while murine possess only a single ortholog (Oatp1b2). To elucidate the impact of OATP1B1 and 1B3 on the pharmacokinetics of the hepatoprotective agent glycyrrhizin and its potential drug-drug interactions, we conducted a study utilizing wild-type, murine Oatp1b2 knockout, and newly generated humanized OATP1B1 and OATP1B3 transgenic mice (Slco1b2 -/- ; 1B1/1B3 tg ). These models were characterized physiologically and biochemically, followed by intravenous administration of glycyrrhizin for pharmacokinetic analysis. Compared to wild-type controls, Oatp1b2 knockout mice exhibited a 2-fold decrease in liver AUC0-7h and up to 26.9-fold reduction in liver-to-plasma ratios, indicating impaired hepatic uptake of glycyrrhizin. Additionally, the absence of Oatp1b2 resulted in a 2.8-fold increase in plasma AUC0-7h and enhanced renal distribution of glycyrrhizin. However, the knock-in of human OATP1B1 and OATP1B3 partially or fully restored the impaired hepatic uptake and reduced plasma levels of glycyrrhizin in Oatp1b2 knockout mice. Furthermore, we evaluated drug-drug interactions between glycyrrhizin and glecaprevir/pibrentasvir, a clinically relevant combination. Glecaprevir/pibrentasvir inhibited OATP1B1/1B3-mediated hepatic uptake of glycyrrhizin at therapeutic concentrations, increasing systemic exposure and highlighting drug-drug interaction risks. Our study provides the first in vivo evidence of glycyrrhizin hepatic transport by human OATP1B1 and OATP1B3, emphasizing the need for careful dosing in patients with variable OATP1B/1B3 activity due to transporter-mediated drug-drug interactions. Additionally, this humanized mouse model can serve as a robust in vivo tool for the prediction of human OATP1B1 and 1B3 function.
Cryptotanshinone (CPT) exerts inhibitory effects on breast cancers (BC). However, its effect and potential mechanism on breast cancer cell migration and invasion are not well understood. This study aims to investigate the potential mechanism of CPT on migration and invasion by network pharmacology and experiments. Wound healing and Transwell Matrigel invasion assays were performed to analyze the effects of CPT on migration and invasion in breast cancer cell lines MCF-7 and 4 T1. Western blot and immunofluorescence analyses were performed to verify the effect of CPT on epithelial-mesenchymal transition (EMT). Network pharmacology and experiments were used to explore the potential mechanism of CPT on EMT in breast cancer. Molecular docking, molecular dynamics (MD) simulation, cellular thermal shift assays (CETSA), drug affinity responsive target stability (DARTS), transfection, and mutagenesis were performed to confirm the direct binding effect of CPT on MEK1. Finally, BC nude mouse xenografts were established to explore the anti-metastasis effect of CPT in vivo. We found that CPT inhibited the EMT process to attenuate the migration and invasion of BC cells in a dose-dependent manner. Subsequently, network pharmacology, CETSA, DARTS, and enzyme activity assays confirmed that CPT directly binds MEK1 to inhibit its phosphorylation and kinase activity. Moreover, the molecular docking, MD, and mutagenesis results showed that CPT bound to MEK1 at Gly80 and Asp208. Moreover, MEK inhibitor (AZD-6244) intervention studies confirmed that CPT inhibits EMT by targeting the MEK pathway. Finally, the mouse tail vein injection model showed that CPT can effectively inhibit BC metastasis in vivo. Our results demonstrated that CPT modulates breast cancer migration and invasion via the MEK/ERK/EMT axis by directly targeting MEK1. Our study provides evidence suggesting that CPT may serve as a potential agent for suppressing breast cancer metastasis.
To elucidate the mechanism by which Resveratrol (Res) ameliorates hypertrophic scar (HS) formation by targeting acid-sensing ion channel 3 (ASIC3) to modulate macrophage-fibroblast (FB) crosstalk. A rabbit-ear HS model was established in vivo. Hematoxylin-eosin (H&E) staining, Masson staining, immunofluorescence (IF), Western blot (WB), and quantitative real-time PCR (RT-qPCR) were used to assess the effects of Res on scar hyperplasia, collagen deposition, FB activation, and macrophage polarization. In vitro, FB activation was stimulated by combined treatment with TGF-β1 and lactic acid, and a Transwell co-culture system comprising FB and human monocyte-derived M0 macrophages was established. Scratch assay, FCM, IF, and WB were performed to assess the impacts of Res on FB activation, migration, and macrophage polarization. Additionally, ASIC3 gene knockout experiments were conducted both in vivo and in vitro to confirm the mechanism underlying Res-mediated HS improvement. In vitro, Res significantly inhibited FB migration in a dose-dependent manner and downregulated the protein expression of α-SMA, COL1A1, COL3A1, reduced M-CSF secretion, suppressed macrophage polarization toward the M2 phenotype, and decreased TGF-β1 mRNA expression. It also blocked activation of the PI3K/Akt signaling pathway downstream of ASIC3. These effects were completely abolished after ASIC3 gene knockdown. In vivo, Res significantly reduced the scar elevation index (SEI) in rabbit-ear HS. It improved collagen fiber arrangement and decreased collagen deposition. It markedly inhibited M2 macrophage polarization and TGF-β1 mRNA expression. After ASIC3 knockout, the anti-HS effects of Res, as well as its regulatory effects on macrophage polarization and fibrotic factors, were abrogated. Res ameliorates HS by inhibiting ASIC3 expression. This disrupts the ASIC3-M-CSF-TGF-β1 positive feedback loop. It restores the balance of macrophage polarization, inhibits FB activation, reduces abnormal collagen deposition, and ultimately attenuates HS formation.
Parkinson's disease (PD) is a progressive neurodegenerative disorder characterized by dopaminergic neuron loss and α-synuclein (α-syn) aggregation, often linked to lysosomal dysfunction. Cathepsin D (CTSD), a lysosomal hydrolase essential for α-syn clearance, becomes functionally impaired when its maturation is disrupted, exacerbating proteostatic stress. This study investigated whether ginsenoside Rg1(Rg1) restores CTSD maturation and lysosomal function to mitigate PD pathology. MPTP-induced zebrafish and mouse PD models, as well as MPP+-treated SH-SY5Y cells, animals and cells were treated with Rg1 at different concentrations. Motor behavior, dopaminergic neuron survival, α-syn clearance, CTSD maturation, lysosomal activity, endoplasmic reticulum (ER) stress, oxidative stress, autophagic flux, and apoptosis were systematically evaluated. Rg1 improved locomotor performance and preserved dopaminergic neurons, promoted α-syn clearance, and enhanced CTSD maturation in lysosomes. These effects coincided with reduced ER and oxidative stress, normalized autophagic flux, and decreased apoptosis. Rg1 functions as a natural lysosomal enhancer, restoring lysosome-ER homeostasis and counteracting multiple pathogenic pathways in PD. The findings reveal a CTSD-dependent regulatory axis in α-syn homeostasis and highlight Rg1 as a promising multi-target therapeutic candidate for PD.
Dehydrocostus lactone (DHL), a sesquiterpene lactone isolated from Aucklandia lappa (Radix Aucklandiae), is a bioactive compound widely used in traditional medicine for its anti-inflammatory and antitumor properties. However, its precise molecular target and pharmacological mechanisms in hepatocellular carcinoma (HCC) remain largely undefined. This study aimed to elucidate the role of protein tyrosine kinase 7 (PTK7) in HCC and to investigate the antitumor efficacy and mechanism of DHL as a PTK7-targeting compound that inhibits the β-catenin signaling pathway. The single-cell transcriptomic data and spatial transcriptomic data, together with publicly available TCGA-LIHC and GEO datasets, were integrated to determine PTK7 expression patterns and pathway associations in HCC. Cell proliferation, apoptosis, mitochondrial membrane potential, and invasion assays were performed to evaluate DHL activity. Molecular docking, cellular thermal shift assay (CETSA), drug affinity responsive target stability (DARTS) assays, and co-immunoprecipitation assays were conducted to confirm that DHL directly inhibited interaction of PTK7 and β-catenin. The in vivo antitumor efficacy of DHL was assessed in Hep3B and Huh7 xenograft models. PTK7 was highly expressed in malignant HCC cells and positively correlated with β-catenin pathway activation. DHL treatment disrupted the PTK7-β-catenin interaction, inhibited proliferation, induced mitochondrial-dependent apoptosis, and reduced invasion by downregulating matrix metalloproteinase (MMP)-2 and MMP-9. DHL also decreased phosphorylation of protein kinase B (AKT), extracellular signal-regulated kinase (ERK), and β-catenin in a dose-dependent manner. CETSA and DARTS assays confirmed the direct inhibition of PTK7 by DHL. In vivo, DHL markedly suppressed tumor growth, achieving inhibition rates of ~34%-54% in Hep3B and 28%-58% in Huh7 xenograft models. DHL exerts significant antitumor effects in HCC by directly targeting PTK7 and inhibiting β-catenin signaling. These findings identify PTK7 as a potential therapeutic target and support the development of DHL as a promising natural agent for PTK7-directed HCC therapy.
Radioresistance remains a key barrier in breast cancer (BC) treatment. Baicalein, a flavonoid from Scutellaria baicalensis, has shown potential to enhance radiosensitivity. This study investigates its efficacy and mechanism in radioresistant BC models. Radioresistant BC cell models were established via fractionated irradiation and validated by clonogenic assays. Baicalein's radiosensitizing effects were assessed using clonogenic survival and DNA damage assays (γ-H2AX foci and neutral comet). KEGG and protein-protein interaction analyses identified Rac1 as a key mediator. Molecular docking and 100 ns molecular dynamics (MD) simulations evaluated the binding stability of the Rac1-baicalein complex, with binding free energy calculated via gmx_MMPBSA. Rac1 activity, immunoblotting, immunofluorescence, and comet assays demonstrated that baicalein inhibits Rac1 activity and delays DNA repair. In vivo xenograft studies (n = 5 mice/group) with immunohistochemistry validated Rac1 pathway inhibition. Baicalein enhanced radiosensitivity in BT549 (radiation enhancement ratio, ER = 2.20 ± 0.12) and MCF7-R cells (ER = 1.38 ± 0.10). Molecular docking revealed a strong binding affinity (-6.87 kcal/mol) between baicalein and Rac1, with MD simulations indicating a relatively stable interaction (binding free energy: -73.42 kJ/mol). Surface plasmon resonance (SPR) analysis revealed a direct interaction between baicalein and Rac1 protein with moderate affinity (KD = 2.91 × 10-6 M). High Rac1 expression correlated with poorer clinical outcomes in overall survival (OS), disease specific survival (DSS), and progress free interval (PFI). Baicalein delayed DNA repair after radiation by inhibiting Rac1 activity via the NHEJ pathway, an effect reversed by a constitutively active Rac1-Q61L plasmid. In vivo, baicalein inhibited Rac1, slowing tumor growth and enhancing radiosensitivity. Baicalein is a potential radiosensitizer, offering a novel strategy to overcome radioresistance in breast cancer therapy.
Cardiovascular disease (CVD) remains the leading cause of morbidity and mortality worldwide. While studies suggest that berry fruits offer beneficial effects in CVD prevention, the bioactive compounds within different berries vary, and the specific mechanisms behind their protective effects are not yet fully elucidated. Our study aimed to evaluate and compare the cardioprotective and anti-atherosclerotic effects of Morus alba (white mulberries, WMex) and Vaccinium myrtillus L. fruit extracts (blueberries, BBex), focusing on acute myocardial infarction (AMI), CVD risk factors, atherosclerosis development, and the underlying mechanisms. WMex and BBex were produced and their qualitative compositions were characterized. We evaluated the infarct size (IS) limiting potential of 8-weeks supplementation with both WMex and BBex at nutritional doses in mice with western diet (WD)-induced metabolic syndrome (MS). We further evaluated atherosclerosis development mitigation after 4-weeks supplementation with the extracts in both male and female WD-fed ApoE-/- mice as well as the putative cardioprotective and atheroprotective mechanisms. The total flavonoids in WMex and BBex were 49.50 and 98.92 mg rutin/g extract, respectively, while the total phenolic content was 50.74 and 110.00 mg gallic acid/g extract, respectively. Both extracts limited IS compared to Control by suppressing apoptosis. WMex upregulated endothelial nitric oxide synthase (eNOS) phosphorylation and pathway along with circulating nitrite levels. WMex suppressed the extent of atheromatic area in both male and female ApoE-/- mice, while BBex was effective in females without any of the extracts affecting estrogen levels. WMex downregulated NF-kB-ICAM-1 axis in the atherosclerotic area and reduced circulating triglyceride levels. None of the extracts caused signs of toxicity, supporting a safety profile. In conclusion, WMex exerts potent cardioprotective and anti-atherosclerotic properties in vivo and could be further exploited for CVD prevention.
Myocardial ischemia-reperfusion injury (MIRI) remains a major clinical challenge due to limited therapeutic options and the risk of complications such as hemorrhage. Dihydromyricetin (DMY), a flavonoid derived from Vine tea, has shown cardioprotective effects, but its mechanism of action in MIRI is not fully understood. This study aimed to investigate the therapeutic effects of DMY on MIRI and elucidate the underlying molecular mechanisms. A rat model of MIRI was established by left anterior descending coronary artery ligation. Rats received DMY or the positive control diltiazem (DIL) for 7 days post-injury. Cardiac damage was assessed by measuring cardiac troponin levels and histopathological analysis. The expression of chemokine-like factor 1 (CKLF1) and its downstream signaling pathways was examined using molecular and biochemical approaches. The interaction between DMY and CKLF1 was further validated using a CKLF1 agonist (C27) and CKLF1-knockout rats. CKLF1 expression was significantly upregulated in MIRI, correlating with inflammatory infiltration, tissue disorganization, and elevated cardiac troponin levels. Mechanistically, CKLF1 activation promoted phosphorylation of nuclear factor kappa-B (NF-κB) and subsequent assembly of the NLR family pyrin domain-containing 3 (NLRP3) inflammasome, leading to caspase-1-dependent pyroptosis. DMY treatment attenuated these effects by downregulating CKLF1 expression and disrupting its interaction with C-C chemokine receptor type 5 (CCR5) and NLRP3, thereby suppressing pyroptosis. Notably, activation of CKLF1 signaling by its agonist C27 reversed the protective effects of DMY. Moreover, while CKLF1 knockout modestly reduced pyroptosis-related protein expression, the anti-pyroptotic effect of DMY was abolished in knockout rats, indicating its dependence on CKLF1. These findings demonstrate that DMY alleviates MIRI by targeting the CKLF1/NF-κB/NLRP3 axis, thereby inhibiting pyroptosis and preserving cardiomyocyte integrity. The anti-pyroptotic effect of DMY is specifically dependent on CKLF1 expression. This study provides a novel mechanistic basis for developing targeted therapies against MIRI.
Paeoniflorin (PF), the primary bioactive component of Paeonia plants, exhibits neuroprotective and anti-inflammatory activities. However, its potential role in alcohol-induced depression (AID) through the microbiota-metabolite-brain axis remains unclear. This study aimed to investigate the antidepressant effect of PF and its underlying gut-brain communication mechanisms. A mouse model of chronic alcohol exposure was established and treated with PF. Behavioral tests (SPT, FST, TST, OFT) and histopathological examinations were conducted. Network pharmacology and molecular docking were used to predict targets, followed by experimental validation of microglial activation and the NF-κB/NLRP3 pathway. Multi-omics approaches including 16S rRNA sequencing and untargeted metabolomics were applied to analyze gut microbiota and fecal metabolites. PF significantly alleviated depressive-like behaviors, hippocampal damage, and oxidative stress, while restoring monoamine neurotransmitter levels. It suppressed microglial activation and the NF-κB/NLRP3 inflammasome cascade. Furthermore, PF reshaped gut microbiota composition (reducing Rikenellaceae and Prevotellaceae) and modulated metabolite profiles and upregulated the neuroprotective metabolite niacin, along with other identified metabolites. These findings demonstrate that PF alleviates Alcohol-Induced Depression (AID) by remodeling the microbiota-metabolite-brain axis and inhibiting neuroinflammation. The study highlights PF's therapeutic potential for alcohol-related mood disorders and underscores the gut-brain axis as a critical target for antidepressant therapy.
The global prevalence of inflammatory bowel disease (IBD) continues increasing, but effective targeted treatments remain elusive. Recently, mesenchymal stem cells (MSCs) have emerged as a promising candidate for IBD treatment, with energy metabolism playing a critical role. Berberine (BBR), a natural extract from Berberidaceae plants, has been shown to modulate various cell functions by reprogramming metabolism in immune-related diseases. This study aims to investigate the influence of BBR on the therapeutic effectiveness of MSCs in colitis. Using the colitis mice model, we compared the therapeutic effects of BBR treated MSCs (MSCBBR) and untreated MSCs by evaluating colon length, Disease Activity Index, and histopathological change. Various methods were employed to detect the differences of MSCBBR and MSCs, including seahorse analysis for energy metabolism, RNA-seq and qPCR for gene transcription, western blotting for protein expression, flow cytometry for phenotypic analysis, and cytokine production. Our results revealed that BBR treatment activated the AMPK signaling pathway, upregulated the expression of glucose transporter 1 (GLUT1), and promoted aerobic glycolysis in MSCs. Comparing to MSCs, MSCBBR exhibited the increased expression of tumor necrosis factor-stimulated gene-6 (TSG-6), enhanced immunomodulatory function and superior therapeutic efficacy. Inhibition of either glycolysis or TSG-6 expression impaired the therapeutic effect of MSCBBR, while overexpression of GLUT1 or TSG-6 obviously improved the therapeutic efficacy of MSCs. BBR treatment improved the therapeutic effect of MSCs in colitis by promoting aerobic glycolysis, providing a safe new approach for optimizing MSCs-based therapy in IBD through reprogramming energy metabolism.
This study investigated the efficacy of hydroxytyrosol (HT), an active ingredient renowned for its antioxidant and anti-inflammatory properties derived from olive leaves and oil, in mitigating skin photoaging and elucidating its underlying mechanisms. Utilizing a comprehensive methodology that included network pharmacology, molecular docking, molecular dynamics (MD) simulations, quantitative mass spectrometry-based proteomics, in vitro assays on keratinocytes, and in vivo experiments with a nude mouse model, we assessed the effects of HT against ultraviolet (UV) radiation-induced skin damage. Our results demonstrated that HT significantly reduces oxidative stress and inflammatory responses, inhibited matrix metalloproteinases (MMPs) activity, mitigates collagen and elastin degradation, enhanced moisture retention, and suppresses apoptosis in keratinocytes. Mechanistically, HT engaged critical signaling pathways such as phosphatidylinositol 3-kinase (PI3K)-protein kinase B (AKT), nuclear factor-kappa B (NF-κB), mitogen-activated protein kinase (MAPK), and Janus kinase (JAK)-signal transducer and activator of transcription (STAT), resulting in improved skin hydration, elasticity, collagen density, attenuation of the senescence-associated secretory phenotype (SASPs), and restoration of skin integrity in vivo. These findings provide a molecular basis for the anti-photoaging properties of HT, supporting its potential application as a novel phytotherapeutic agent for the management of UV-induced skin damage.
N6-(2-hydroxyethyl) adenosine (HEA), the main active component of Cordyceps species, has garnered attention for its multifaceted hypolipidemic and antihyperglycemic activities. However, the influence by which HEA affects the progression of nonalcoholic fatty liver disease (NAFLD) remains unclear. This study sought to investigate the efficacy and mechanisms of HEA in treating NAFLD. The mouse NAFLD models were induced by high-fat diet feeding or methionine-choline-deficient diet feeding. AML12 and HepG2 cells were used for the in vitro study. Lipidemic and glycemic parameters, untargeted lipidomic, cellular thermal shift assay, and so forth were used to explore the beneficial effects of HEA in NALFD. HEA effectively alleviated the progression of NAFLD by regulating glucolipid metabolism and insulin resistance both in vitro and in vivo. Lipidomic data suggested that HEA markedly reduced triglyceride levels by blocking hepatic de novo lipogenesis and shifting fatty acids into mitochondria for oxidation and into structural lipids. Moreover, stimulation of the supernatant of adipocytes with HEA was more effective than treatment with HEA only in terms of hepatic de novo lipogenesis in HepG2 cells. Mechanistically, HEA significantly reduced the population of JunB+ adipocytes, which exerts lower thermogenic capacity. Further studies verified that HEA interacted with ASP-12, ASP-13, and TYR-15 of JunB subunits through hydrogen bonding, leading to activated PGC-1α activity and governing thermogenic adipocyte heterogeneity and consequent biological responses. These findings highlight the capacity of HEA to alleviate NAFLD through the JunB pathway, paving a new way to treat NAFLD by influencing adipocyte functionality.
Psoralea corylifolia Linn. exhibits osteogenic effects; however, the mechanism by which its active component Bakuchiol (BAK) alleviates glucocorticoid-induced osteoporosis (GIOP) remains unclear. We aim to investigate BAK's therapeutic effects and potential mechanisms in GIOP. We treated GIOP mice with BAK. We evaluated BAK's therapeutic efficacy using micro-CT and histopathological staining. We performed transcriptomic analysis and identified significantly enriched GO pathways. We then used biochemical assays and Western blotting to examine the effects of BAK on osteogenic differentiation-related signaling factors in GIOP mice. We conducted in vitro experiments using osteoblasts. We employed RT-qPCR, Western blot, ALP & ARS staining, immunofluorescence and dual-luciferase reporter assays to assess BAK's influence on signaling pathways related to osteoblast differentiation. We used molecular docking, cellular thermal shift assay (CETSA), and drug affinity responsive target stability (DARTS) to confirm the direct binding interaction between BAK and FAT4. Finally, we silenced Fat4 to validate that BAK exerts its anti-GIOP effects by activating YAP1 through FAT4. BAK treatment significantly enhanced bone quality and strength in GIOP mice, and mitigated femoral pathological damage. BAK upregulated key osteoblast differentiation-related transcription factors and enhanced serum alkaline phosphatase activity. Western blot confirmed that BAK increased protein levels of YAP1 while reducing levels of FAT4 and DCHS1. In vitro, BAK similarly promoted osteoblast differentiation and activated YAP1 expression and Runx2 transcription. Molecular docking, CETSA and DARTS analyses demonstrated that BAK directly binds to FAT4. Silencing Fat4 abolished the pro-osteogenic effects of BAK. BAK likely exerts its therapeutic effects in GIOP by targeting FAT4, activating YAP1 and promoting osteoblast differentiation.
Atractylodes macrocephala Koidz. has long been used for invigorating Qi and traditionally prescribed for cancer treatment by Chinese medicine practitioners. Atractylenolide I (ATI) is a major active component in A. macrocephala and has inhibitory effects on various malignancies. Triple-negative breast cancer (TNBC)-related deaths are predominantly attributed to metastasis. However, the activity and mechanism of ATI on TNBC metastasis are unclear. Cell viability was detected by CCK-8 assay. Cell migration was determined by wound healing and transwell assays. The in vivo experiment was performed in 4 T1 spontaneous metastasis breast cancer nude mice. The mechanism was investigated by RNA-seq analysis, and further verified by bioinformatics, qPCR, western blot, and siRNA transfection analysis. ATI inhibited the growth of 4TI cells. Additionally, ATI repressed the wound healing, invasion, and migration activities of 4TI and MDA-MB-231 cells at a lower concentration. In vivo experiment revealed that while ATI (50 mg/kg) did not significantly inhibit tumor growth but markedly suppressed lung metastasis progression in a 4 T1 spontaneous metastasis mouse model. RNA-seq analysis of lung tissues showed that the extracellular matrix (ECM) signaling pathway was significantly suppressed by ATI. In addition, ATI inhibited the mRNA and protein levels of ECM-related members (SPP1, MMP9, and COL1A1) in vitro and in vivo. Bioinformatics analysis revealed that the overexpression of SPP1 correlated with the poor prognosis of breast cancer patients. Furthermore, ATI inhibited ECM activator pyrintegrin-induced wound healing. siRNA knockdown of SPP1 abolished ATI's inhibition on wound healing, cell migration, and expression of ECM-related genes/proteins in 4 T1 cells. Molecular docking showed that ATI interacted with Trp-106, Pro-44, and Leu-46 residues in SPP1 (-7.73 kcal/mol). Our findings suggest that ATI could suppress the lung metastasis of TNBC by inhibiting the SPP1-mediated ECM signaling pathway.
Diabetic nephropathy (DN) is a major complication of diabetes. Swietenine (Swi), a natural compound derived from the fruit of Swietenia macrophylla , has shown beneficial effects in alleviating diabetic complications. However, its underlying mechanism for treating DN remains unclear. This study aimed to elucidate the therapeutic effects and molecular mechanisms of Swi against DN through in vitro and in vivo experiments. A model of streptozotocin/high-fat diet (STZ/HFD)-induced Sprague-Dawley (SD) rats was employed to evaluate the effects of Swi on improving DN renal fibrosis and enhancing autophagy in vivo. Followed based on the results of proteomics, network pharmacology, and molecular docking, the potential mechanism of Swi in treating DN was predicted, and subsequently the cell model of high glucose induced human renal tubular epithelial cells (HK-2) was established in vitro, and the potential mechanism of Swi's therapeutic effect was further validated through Western blot analysis, immunofluorescence, immunohistochemistry, and flow cytometry. After Swi treatment, levels of urinary protein, uric acid, creatinine, and urea nitrogen were significantly reduced in DN rats. Pathological improvements included decreased thickness of the tubular basement membrane, reduced PAS-positive deposits, less glycogen accumulation, and alleviated fibrosis. The number of autolysosomes increased significantly, especially with high-dose Swi. Immunofluorescence of renal tissue showed enhanced LC3B fluorescence intensity, suggesting restored autophagy levels. Western blot analysis revealed upregulation of LC3B and Beclin-1 and downregulation of p62 after Swi treatment. Immunohistochemistry indicated reduced expression of fibronectin (Fn) and collagen I (COI-I), indicating decreased extracellular matrix accumulation and renal injury. In high glucose-induced human renal tubular epithelial cells (HK-2), similar trends were observed. Swi treatment upregulated p-AMPK and p-ULK1 expression and downregulated p-mTOR. Overall, Swi significantly improved renal function and pathological damage in DN rats by activating the AMPK/mTOR/ULK1 signaling pathway, enhancing autophagy, reducing ROS production, and ameliorating mitochondrial injury, thereby delaying renal fibrosis progression. In conclusion, Swi exerts protective effects against DN by promoting autophagy and mitigating oxidative stress, suggesting its potential as a therapeutic agent for DN.
Dysregulated proliferation and differentiation of bone marrow mesenchymal stem cells (BMSCs) represent a key pathophysiological mechanism in osteoporosis. Recent studies have demonstrated a significant association between ferroptosis and the advancement of osteoporosis, suggesting that targeting ferroptosis could offer novel therapeutic approaches for osteoporosis treatment. Curcumin, a natural antioxidant, has shown therapeutic potential in bone-related disorders; however, its precise mechanisms for modulating BMSC function-particularly via ferroptosis-related pathways-remain poorly characterized. This study investigated whether curcumin alleviates iron overload-induced BMSC dysfunction by targeting ferroptosis, specifically elucidating its molecular mechanisms in promoting osteogenic differentiation and mitigating cellular senescence. Iron-overloaded BMSC in vitro models and in vivo murine systems were established to model osteoporosis-related microenvironments. Curcumin was administered to assess its effects on cellular and systemic outcomes, including bone microstructure, mechanical property, differentiation capacity, senescence markers, iron metabolism, and redox homeostasis by using micro-CT, RNA-seq, RT-qPCR, western blot, immunohistochemical, immunofluorescence, and transmission electron microscope (TEM). Furthermore, Nrf2 siRNA and the Nrf2 inhibitor ML385 were utilized to interrogate curcumin's mechanism of action in iron-overloaded BMSCs. In vivo, curcumin treatment significantly attenuated iron overload-induced bone microstructural damage, mechanical property, and elevated Nrf2 and GPX4 expression in BMSCs. In vitro, curcumin mitigated iron overload-induced ferroptosis in BMSCs by upregulating Nrf2 expression, thereby increasing GPX4 levels. This mechanism consequently delayed cellular senescence and promoted osteogenic differentiation. Our findings establish the Nrf2/GPX4 axis as a critical therapeutic target of curcumin for ameliorating iron overload-induced osteoporosis. This mechanistic insight provides a foundation for developing novel therapeutics against age-related and postmenopausal osteoporosis.
Mulberry leaf, as a traditional Chinese medicinal plant, has been utilized in the treatment of various diseases, including diabetes, cardiovascular diseases, inflammatory disorders, and liver diseases. However, the mechanisms underlying its therapeutic effects on non-alcoholic fatty liver disease (NAFLD) remain unclear. Therefore, this study aims to investigate the potential mechanisms of mulberry leaf extract (MLE) in the treatment of NAFLD. The chemical composition of MLE was analyzed using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). The NAFLD mice model was induced by a high-fat, high-fructose, and high-cholesterol (HFFC) diet, followed by intervention with MLE. The results indicated that the administration of MLE notably reduced the obesity (p < 0.05), oxidative stress (p < 0.05), inflammation (p < 0.05), and ECM deposition (p < 0.05) induced by the HFFC diet, restored the parameters of liver function, and attenuated the pathological changes. Utilizing a combination of integrated liver non-targeted metabolomics, network pharmacology, and transcriptomic approaches, we deciphered the molecular mechanisms by which MLE exerted its therapeutic effects in the treatment of NAFLD. In detail, our findings revealed that MLE suppressed the TGFβ1/Smad3 and NF-κB signaling pathways to ameliorate fibrosis and inflammation. This study provided novel insights into the correlation between MLE and NAFLD progression, offering a scientific foundation for the prospective use of MLE in the treatment of NAFLD.
Idiopathic pulmonary fibrosis is a chronic, progressive disease in older adults with unclear pathogenesis and a lack of effective drugs. Columbianadin, a natural coumarin analog isolated from Angelicae pubescentis Radix, has a wide range of pharmacological effects; however, its effects on pulmonary fibrosis are unknown. This study investigates the anti-pulmonary fibrosis effects of columbianadin and their underlying mechanisms of action. An in vivo model of mouse lung fibrosis was established, and mice were randomly assigned to different doses of columbianadin. The effects of 5'-adenosine monophosphate-activated protein kinase (AMPK) on the anti-pulmonary fibrosis and anti-cellular senescence effects of columbianadin was observed by combining AMPK inhibitor and columbianadin. Cellular senescence was induced in vitro by hydrogen peroxide and treated with different concentrations of columbianadin, and we observed the effect of AMPK on the anti-cellular senescence effect of columbianadin by specifically silencing the AMPK gene. Columbianadin reduced the expression levels of collagen type I alpha 1 (col1-a1), alpha-smooth muscle actin (a-SMA), p21, and p16 in lung tissues of mice with pulmonary fibrosis, and these effects were inhibited by AMPK inhibitors. Similarly, Columbianadin reduced the expression levels of p21 and p16 in senescent cells. In addition, we found that columbianadin promoted Sirt1 and Sirt3 expression as well as AMPK phosphorylation, whereas the anti-cellular senescence effect of columbianadin and the effect of promoting the expression of Sirt1 and Sirt3 were suppressed by specific silencing of the AMPK gene. Columbianadin exerts its anti-pulmonary fibrosis effect by inhibiting cellular senescence via the AMPK-Sirt1/3 pathway. The present study provided new insight into a novel treatment of pulmonary fibrosis.
