OBJECTIVES: To exple the mechanism of Qixiong Zuogui Granules (QXZG) for enhancing synaptic plasticity in aging rats. METHODS: Forty SD rats were randomized into control group, aging model group, donepezil treatment group, and QXZG treatment group (n=10). Except for the control rats, all the rats were subjected to daily intraperitoneal injection of D-galactose for 8 consecutive weeks to induce brain aging, and donepezil hydrochloride and QXZG suspension were administered by gavage during modeling. After the interventions, the rats were evaluated for general conditions, behavioral changes, oxidative stress indicators, hippocampal pathologies, and expressions of the brain-derived neurotrophic factor (BDNF)/tyrosine kinase receptor B (TrkB) pathway, p16, and synaptic plasticity-associated proteins. RESULTS: The rats in the model group exhibited obvious aging phenotypes such as yellowing of the teeth and hair, body weight loss, and impaired learning and memory abilities, with decreased serum SOD and GSH-Px activities and increased serum MDA level. The rat models also showed obvious pathological changes, reduced Nissl bodies, and elevated p16 protein expression in the hippocampal CA1 region, with significantly decreased expression levels of BDNF, TrkB, CREB and synaptic plasticity proteins SYN, GAP43, and PSD95. Treatment with QXZG alleviated the aging phenotypes in the rat models, improved their learning and memory abilities and pathological changes in the hippocampal CA1 region, reduced oxidative stress and p16 protein expression, and promoted the expressions of the BDNF/TrkB pathway proteins and synaptic plasticity proteins. CONCLUSIONS QXZG enhances synaptic plasticity and reduces oxidative stress in aging rats possibly by upregulating the BDNF/TrkB signaling pathway proteins, thereby delaying brain aging and improving learning and memory abilities of the rats.
Animals ; Brain-Derived Neurotrophic Factor/metabolism* ; Neuronal Plasticity/drug effects* ; Signal Transduction/drug effects* ; Rats, Sprague-Dawley ; Receptor, trkB/metabolism* ; Rats ; Aging ; Drugs, Chinese Herbal/pharmacology* ; Male ; Oxidative Stress ; Hippocampus/metabolism*

OBJECTIVES: To investigate the effect of cardiomyocytes-derived exosomes on lipopolysaccharide (LPS)-induced cardiomyocyte injury and its mechanism. METHODS: Exosomes isolated from rat cardiomyocytes with or without LPS treatment were co-cultured with rat lymphocytes. The lymphocytes with or without exosome treatment were co-cultured with LPS-induced rat cardiomyocytes for 48 h. Cardiomyocyte apoptosis was detected using flow cytometry, and the expressions of apoptosis marker proteins and the PI3K/AKT pathway proteins were detected using Western blotting. The effects of human recombinant IL-38 protein on apoptosis and protein expressions in LPS-induced cardiomyocytes were examined. RESULTS: Compared with normal cardiomyocyte-derived exosomes, the exosomes from LPS-induced cardiomyocytes significantly enhanced proliferation and increased mRNA and protein expression levels of IL-38 in rat lymphocytes. Bioinformatics analysis suggested that miR-1275 in the exosome played a key role in LPS-induced cardiomyocyte injury, and in dual luciferase reporter gene assay, miR-1275 mimics significantly increased luciferase activity of WT-IL-38. Co-culture with lymphocytes treated with exosomes from LPS-induced cardiomyocytes significantly inhibited apoptosis of LPS-induced cardiomyocytes. Treatment with recombinant IL-38 also effectively lowered apoptosis rate of LPS-induced cardiomyocytes, reduced cellular expression of Bax protein, and increased the protein expression levels of Bcl-2, p-PI3K and p-AKT. CONCLUSIONS miR-1275 in exosomes derived from LPS-induced cardiomyocytes mediates IL-38 up-regulation expression in lymphocytes to activate the PI3K/AKT pathway and inhibit LPS-induced cardiomyocyte apoptosis.
Apoptosis/drug effects* ; MicroRNAs/metabolism* ; Myocytes, Cardiac/metabolism* ; Animals ; Lipopolysaccharides ; Rats ; Exosomes/metabolism* ; Up-Regulation ; Interleukins/metabolism* ; Lymphocytes/cytology* ; Cells, Cultured ; Signal Transduction ; Coculture Techniques ; Phosphatidylinositol 3-Kinases/metabolism* ; Rats, Sprague-Dawley ; Humans ; Proto-Oncogene Proteins c-akt/metabolism*

OBJECTIVES: To investigate the mechanism of Qihuang Jianpi Zishen Granules (QJZ) for ameliorating renal damage in MRL/lpr mice. METHODS: With 6 female C57BL/6 mice as the normal control group, 30 female MRL/lpr mice were randomized into model group, QJZ treatment groups at low, moderate and high doses, and prednisone treatment group (n=6). After 8 weeks of treatment, the mice were examined for 24-h urine protein, creatinine and albumin levels, serum levels of IgG, complement 3 (C3), C4, anti-dsDNA, interferon γ (IFN‑γ) and interleukin 17 (IL-17). Kidney tissues were sampled for histopathological examination with HE staining and observation of glomerular ultrastructure changes using transmission electron microscopy (TEM). The expressions of MyD88/NF-κB pathway-related molecules in the kidney tissue were detected using RT-qPCR, Western blotting and immunohistochemistry. RESULTS: Compared with those in the model group, the mice treated with QJZ at the 3 doses and prednisone showed significant reductions in the renal injury biomarkers and serum IgG, anti-dsDNA, IFN‑γ and IL-17 levels and elevation of serum C3 and C4 levels. HE staining revealed lessened glomerular endothelial cell proliferation and mesangial thickening in all the treatment groups. TEM observation further demonstrated reduced electron-dense deposits and diminished inflammatory cell infiltration in the glomeruli in the intervention groups. QJZ at the 3 doses and prednisone treatment all significantly lowered renal expression levels of MyD88, NF-κB, p65 and p52 in the mouse models. CONCLUSIONS QJZ can improve renal damage in MRL/lpr mice possibly by inhibiting overactivation of the MyD88/NF-κB pathway.
Animals ; Drugs, Chinese Herbal/therapeutic use* ; Female ; Mice, Inbred C57BL ; Mice, Inbred MRL lpr ; Myeloid Differentiation Factor 88/metabolism* ; Mice ; NF-kappa B/metabolism* ; Signal Transduction/drug effects* ; Kidney/metabolism* ; Interleukin-17

OBJECTIVES: To investigate the therapeutic mechanism of Sangma Zhike Formula (SMZKF) for relieving cough sensitivity and airway inflammation in rats with postinfectious cough (PIC). METHODS: Male SD rat models were established by cigarette smoke exposure with intranasal LPS instillation and capsaicin aerosol inhalation. From day 19 following the start of PIC modeling, the rats received daily treatment with saline (model group), low-, medium-, and high-dose SMZKF, and compound methoxyphenamine (ASM) via gavage for 10 consecutive days (n=8). The assessments included behavioral changes, cough sensitivity (latency and frequency), lung histopathology, inflammatory cell counts and cytokine/mediator levels in the bronchoalveolar lavage fluid (BALF), oxidative stress markers in the lung tissue, and expressions of proteins related with cough hypersensitivity and pyroptosis. RESULTS: The rat models of PIC exhibited reduced mental alertness, accelerated respiration, and pronounced symptoms such as coughing, sneezing, and facial scratching with significantly shortened cough latency and increased 5-min cough frequency. Histopathological analysis revealed collapsed alveolar structures, thickened alveolar septa, and extensive inflammatory cell infiltration in the bronchi and peribronchial regions, accompanied by elevated bronchial and alveolar inflammation scores of the rat models. In the BALF, inflammatory cell counts and the levels of IL-1β, TNF-α, IL-6, COX-2, PGE-2, and TXA-2 were all markedly elevated, and the pulmonary oxidative stress markers (ROS and MDA) and myeloperoxidase (MPO) activity were also significantly increased. The pulmonary expressions of cough hypersensitivity-related proteins (TRPV1, SP, CGRP, and NK1R) and pyroptosis-associated markers (P-NF-κB, NLRP3, ACS, cleaved caspase-1, cleaved IL-1β, and GSDMD-N) were significantly upregulated in the model group. SMZKF interventions significantly ameliorated these pathological changes in the rat models, and high-dose SMZKF produced a similar therapeutic efficacy to that of ASM. CONCLUSIONS SMZKF alleviates cough sensitivity and airway inflammation in PIC rats possibly by inhibiting TRPV1-mediated SP/NK1R signaling and the NLRP3/caspase-1/GSDMD pyroptosis pathway.
Animals ; Cough/metabolism* ; Rats, Sprague-Dawley ; Pyroptosis/drug effects* ; Male ; TRPV Cation Channels/metabolism* ; Rats ; Drugs, Chinese Herbal/pharmacology* ; Inflammation ; Signal Transduction

OBJECTIVES: To verify whether hesperetin (Hes) alleviates doxorubicin (DOX)-induced cardiotoxicity by reducing inflammation via regulating the AMPK/NLRP3 pathway. METHODS: C57/bl6 mice and H9c2 cells treated with DOX to mimic cardiotoxicity were randomly divided into Sham (or control) group, DOX group, DOX+Hes group, DOX+Hes+compound C (CC, an AMPK inhibitor) group. Cardiac function and myocardial pathologies of the mice were evaluated, and the changes in H9c2 cell morphology and viability were assessed. Lactate dehydrogenase (LDH) activity in mouse myocardial tissues and H9c2 cells was measured using ELISA, and H9c2 cell apoptosis was detected with TUNEL staining. In both H9c2 cells and the myocardial tissues of the mice, cellular expression levels of TNF-α, IL-6 and IL-1β mRNAs and cleaved caspase-3, Bcl2, Bax, IL-1β, IL-18, p-AMPK, AMPK, p-mTOR, mTOR, NLRP3, ASC and caspase-1 proteins were detected using RT-PCR and Western blotting. RESULTS: DOX treatment caused cell swelling, decreased cell viability and increased LDH activity in H9c2 cells, resulting also in significantly increased cell apoptosis and cleaved caspase-3 expression and decreased Bcl2/Bax ratio. The DOX-treated mice showed obvious myocardial fiber swelling and inflammatory infiltration, decreased cardiac function and significantly increased myocardial LDH activity. In H9c2 cells, DOX treatment significantly increased the mRNA expressions of TNF-α, IL-6 and IL-1β and protein expressions of IL-1β and IL-18, lowered the expressions of p-AMPK and p-mTOR, and increased the expressions of NLRP3, ASC and caspase-1. Hes treatment obviously reduced these toxic effects of DOX in H9c2 cells, but its protective effects were blocked by application of compound C. CONCLUSIONS Hes reduces DOX-induced cardiotoxicity by inhibiting inflammation via regulating the AMPK/NLRP3 pathway.
Animals ; Doxorubicin/toxicity* ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Mice, Inbred C57BL ; Mice ; Signal Transduction/drug effects* ; Cardiotoxicity ; AMP-Activated Protein Kinases/metabolism* ; Apoptosis/drug effects* ; Cell Line ; Myocytes, Cardiac/drug effects* ; Rats

OBJECTIVES: To investigate the synergistic mechanism of the traditional Chinese medicine Rosa laevigata Michx. (RLM) for treatment of pulmonary arterial hypertension (PAH). METHODS: Network pharmacological analysis was carried out to screen the active ingredients of RLM and PAH disease targets and construct the "component-target-disease" interaction network, followed by gene enrichment analysis and molecular docking studies. In the cell experiments, primary cultures of rat pulmonary arterial smooth muscle cells were exposed to hypoxia for 24 h and treated with solvent or 100, 200 and 300 mg/mL RLM, and the changes in cell proliferation were detected using Western blotting for PCNA and immunofluorescence staining. In the animal experiment, male SD rats were randomized into 5 control group, monocrotaline (MCT) solvent group, and MCT with RLM (100, 200 and 300 mg/mL) treatment groups. HE staining and immunofluorescence staining were used to observe histopathological changes in the pulmonary blood vessels of the rats. RESULTS: Seven core active ingredients (including β-sitosterol and kaempferol) in RLM and 39 key disease targets were identified, and molecular docking showed that SRC was a high-affinity target. KEGG enrichment analysis showed that the differential genes were significantly enriched in calcium signaling and PI3K-AKT pathways. In rat pulmonary arterial smooth muscle cells, hypoxic exposure significantly up-regulated cellular expression of PCNA and phosphorylation levels of Src and AKT1, which were obviously lowered by RLM treatment. In RLM-treated rat models, the mean pulmonary artery pressure and right ventricular hypertrophy index (Fulton index) were significantly reduced, the tricuspid annular plane systolic excursion (TAPSE) was improved, and pulmonary vascular wall thickening and fibrosis were obviously ameliorated. CONCLUSIONS RLM inhibits pulmonary arterial smooth muscle cell proliferation in rat models of hypertension possibly by regulating the Src-AKT1 axis, suggesting the potential of RLM as a new natural drug for treatment of pulmonary hypertension.
Animals ; Cell Proliferation/drug effects* ; Proto-Oncogene Proteins c-akt/metabolism* ; Rats, Sprague-Dawley ; Pulmonary Artery/cytology* ; Male ; Rats ; Myocytes, Smooth Muscle/cytology* ; Hypertension, Pulmonary/pathology* ; Drugs, Chinese Herbal/pharmacology* ; Signal Transduction/drug effects* ; Muscle, Smooth, Vascular/cytology* ; src-Family Kinases/metabolism* ; Cells, Cultured

OBJECTIVES: To investigate the mechanism by which Guijianyu ameliorates podocyte injury in a mouse model of diabetic kidney disease (DKD) induced by advanced glycation endproducts (AGEs). METHODS: Sixty db/db mouse models of DKD were randomized equally into 5 groups for treatment with saline, Guijianyu extract at 3 doses or irbesartan for 12 weeks, and the changes in renal pathology and structure were observed using transmission electron microscopy, and the expressions of related genes and key proteins were detected using RT-qPCR and immunohistochemistry. In cultured MPC-5 cells incubated with 50 mg/L AGEs-BSA for 24 h, the effect of different concentrations of Guijianyu extract on cell viability was examined with CCK-8 assay; Western blotting was performed to detect the protein expressions of RAGE, VEGFA, TNF-α, NF-κB(p65), IL-6 and caspase-3, and the mRNA expressions of RAGE, NF-κB (p65), VEGFA and IL-6 were detected with RT-qPCR. RESULTS: In mouse models of DKD, treatment with high-dose Guijianyu extract significantly reduced renal expressions of RAGE, VEGFA, NF-κB(p65), and IL-6 proteins and the mRNA expressions of RAGE, NF-κB, and IL-6. In MPC-5 cells, exposure to AGEs significantly reduced cell viability and increased the protein expressions of RAGE, NF‑κB (p65), VEGFA, TNF-α, IL-6 and caspase-3 (P<0.05) and mRNA expressions of RAGE, NF-κB (p65), VEGFA, and IL-6. Treatment with Guijianyu extract obviously improved cell viability and reduced the expressions of RAGE, NF-κB(p65), VEGFA, TNF-α, IL-6, and caspase-3. Furthermore, Guijianyu extract effectively reversed RAGE agonist-induced elevation of protein expressions of RAGE, VEGFA, TNF-α, IL-6, and caspase-3 and mRNA expressions of RAGE, NF-κB (p65), IL-6, and VEGFA in MPC-5 cells. CONCLUSIONS Guijianyu extract ameliorates AGEs-induced mouse renal podocyte injury in DKD by inhibiting the activation of AGEs-RAGE signaling pathway and reducing the expressions of pro-inflammatory cytokines and vascular endothelial growth factors.
Animals ; Glycation End Products, Advanced ; Drugs, Chinese Herbal/pharmacology* ; Mice ; Signal Transduction/drug effects* ; Podocytes/pathology* ; Diabetic Nephropathies/drug therapy* ; Receptor for Advanced Glycation End Products ; Vascular Endothelial Growth Factor A/metabolism* ; Interleukin-6/metabolism* ; Male

OBJECTIVES: To explore the mechanism of cinnamic acid (CA) for improving doxorubicin-induced myocardial injury (DIC) in mice. METHODS: Network pharmacology analysis was used to obtain the key targets of CA and DIC. Male C57BL/6J mice were randomized into Sham, DOX, CA (25, 50 and 100 mg/kg)+DOX, and CA+Ferrostatin-1+DOX groups, and their myocardial function and pathology were examined by echocardiography and HE staining. Serum levels of CK-MB, LDH, MDA, IL-6, TNF‑α and myocardial ROS level were detected, and the expression levels of TLR4 and ferroptosis pathway proteins in myocardial tissue were detected by Western blotting. Cultured murine cardiomyocytes (HL-1 cells) with or without transfection with a small interfering RNA targeting TLR4 (si-TLR4) were treated with DOX or Erastin, and the cellular ROS content was measured by DCFH-DA staining; the expression level of GPX4 was detected using immunofluorescence staining. RESULTS: Network pharmacology analysis suggested that CA may improve DIC through TLR4 signaling. DOX treatment caused obvious myocardial injury in mice, which showed significantly increased serum levels of CK-MB, LDH, MDA, IL-6, TNF-α and myocardial ROS level with decreased myocardial levels of SLC7A11 and GPX4 proteins and increased levels of TLR4 and PTGS2 proteins. All these changes in the mouse models were significantly alleviated by treatment with CA, and the mice receiving CA or ferrostatin-1 treatment exhibited increased myocardial expressions of SLC7A11 and GPX4 proteins and lowered expressions of TLR4 and PTGS2 proteins. In cultured HL-1 cells, treatment with DOX and Erastin both obviously increased intracellular ROS level and decreased cellular GPX4 expression level, and these changes were strongly attenuated by TLR4 interference. CONCLUSIONS CA, as a potent herbal monomer, can effectively alleviate DIC in mice by inhibiting TLR4-mediated ferroptosis.
Animals ; Ferroptosis/drug effects* ; Toll-Like Receptor 4/metabolism* ; Myocytes, Cardiac/metabolism* ; Mice, Inbred C57BL ; Mice ; Male ; Doxorubicin/adverse effects* ; Cinnamates/pharmacology* ; Signal Transduction ; Reactive Oxygen Species/metabolism*

OBJECTIVES: To investigate the effects of Naoluo Xintong Decoction (NLXTD) on proliferation of rat brain microvascular endothelial cells (BMECs) after oxygen-glucose deprivation/reoxygenation (OGD/R) injury and role of the HIF-1α/VEGF pathway in mediating its effect. METHODS: Using a BMEC model of OGD/R, we tested the effects of 10% NLXTD-medicated rat serum, alone or in combination with 2ME2 or 10% NAKL, on cell proliferation, migration, tube-forming ability and permeability using CCK-8 assay, Transwell chamber assay, tube formation assay and permeability assay. Cellular expressions of VEGF and Notch were detected using ELISA and laser confocal immunofluorescence analysis, and the expressions of HIF-1α, VEGFR2, Notch1, ERK and P-ERK1/2 proteins were detected with Western blotting. RESULTS: OGD/R injury significantly decreased viability of BMECs. NLXTD treatment of the cells with OGD/R could significantly promoted cell proliferation, migration and tube formation ability, but these effects were strongly attenuated by application of 2ME2. NLXTD treatment also significantly increased the percentages of VEGF- and Notch-positive cells in the cell models and obviously enhanced the expression levels of HIF-1α, VEGFR2, Notch1 and P-ERK1/2. CONCLUSIONS NLXTD promotes proliferation, migration, and tube formation of rat BMECs after OGD/R injury possibly by activating the HIF-1α/VEGF signaling pathway.
Animals ; Hypoxia-Inducible Factor 1, alpha Subunit/metabolism* ; Drugs, Chinese Herbal/pharmacology* ; Vascular Endothelial Growth Factor A/metabolism* ; Endothelial Cells/metabolism* ; Rats ; Cell Proliferation/drug effects* ; Signal Transduction/drug effects* ; Glucose ; Brain/blood supply* ; Cells, Cultured ; Rats, Sprague-Dawley ; Vascular Endothelial Growth Factor Receptor-2/metabolism* ; Oxygen/metabolism* ; Cell Hypoxia

OBJECTIVES: To explore the therapeutic mechanism of Flos Sophorae (FS) for treatment of psoriasis. METHODS: The active ingredients, targets and psoriasis-related disease targets of FS were obtained from TCMSP, GeneCards, OMIM, DisGeNET and String databases, and Cytoscape 3.8.0 software was used to construct the "FS -active ingredient-key target-signaling pathway-psoriasis" network. GO and KEGG enrichment analyses of the key targets were conducted, and molecular docking was performed using Discovery Studio 2019. In a BALB/c mouse model of imiquimod-induced psoriasis, the effects of vaseline, FS at high, medium and low doses (3.00, 1.50 and 0.75 g/kg, respectively) and a positive drug, given 1 week before and during modeling, were evaluated on body weight changes, spleen coefficient, psoriasis area and severity index (PASI) score and skin pathological changes. Phosphorylation levels of PI3K and AKT proteins were detected using immunohistochemistry and Western blotting. RESULTS: A total of 10 active components and 110 key targets were screened. GO and KEGG pathway enrichment analysis suggested that FS improved psoriasis primarily through the PI3K/AKT, TNF, and IL-17 signaling pathways. Molecular docking showed that both quercetin and kaempferol could spontaneously bind to AKT1, TNF and other sites. In the mouse model of psoriasis, treatment with low-dose FS significantly improved epidermal thickening, increased body weight, lowered PASI score, and reduced phosphorylation levels of PI3K and AKT proteins. CONCLUSIONS The therapeutic mechanism of FS for psoriasis involves multiple components, targets, and pathways that mediate the inhibition of the phosphorylation levels of PI3K and AKT proteins to suppress the activation of the PI3K/AKT signaling pathway.
Animals ; Psoriasis/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Mice ; Signal Transduction/drug effects* ; Mice, Inbred BALB C ; Phosphatidylinositol 3-Kinases/metabolism* ; Molecular Docking Simulation ; Disease Models, Animal ; Drugs, Chinese Herbal/therapeutic use* ; Imiquimod ; Phosphorylation