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ObjectiveTo explore the molecular mechanism of Sanhuang Xiexintang （SHXXT） in protecting stress gastric ulcer （SGU） in rats through network pharmacology， molecular docking， and animal experiments. MethodThe active ingredients and corresponding targets in SHXXT were collected and screened from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）， Traditional Chinese Medicine Information Database （TCMID）， Bioinformation Analysis Tool for Molecular Mechanism of Traditional Chinese Medicine （BATMAN-TCM）， and Swiss Target Prediction database. SGU-related targets were screened from the Online Mendelian Inheritance in Man （OMIM）， Therapeutic Target Database （TTD）， GeneCards database， and PharmGKB database. Herbal-ingredient-target （H-C-T） network was constructed by using Cytoscape 3.9.1 software. Protein-protein interaction （PPI） of drug and disease intersection targets was analyzed by using the Protein Interaction Platform （STRING） database. Gene ontology （GO） enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway analysis were conducted through the Database for Annotation Visualization and Integrated Discovery （DAVID）. The active ingredients and key targets were validated using AutodockVina 1.2.2 molecular docking software， and the experimental results were further validated through animal experiments. ResultThe 55 active ingredients were screened， and 255 potential target genes for SHXXT treatment of SGU were predicted. The PPI analysis showed that protein kinase B （Akt）， phosphatase and tensin homolog deleted on chromosome ten （PTEN）， tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β）， and cyclooxygenase-2 （COX-2） are the core targets of SHXXT for protecting SGU. GO and KEGG analyses showed that SHXXT may affect the development of SGU by regulating various biological processes such as the phosphoinositide 3-kinase （PI3K）/Akt signaling pathway and inflammatory processes. The molecular docking results showed that both the active ingredients and key targets had good binding ability. Animal experiments showed that compared with the blank group， the ulcer index （UI） of the model group was significantly increased （P<0.01）， and the serum levels of TNF-α and IL-1β significantly increased （P<0.01）. The phosphorylation level of PTEN in gastric mucosal tissue was significantly down-regulated （P<0.05）. The phosphorylation levels of PI3K， Akt， and nuclear factor kappa-B （NF-κB） were significantly up-regulated （P<0.05）. Compared with the model group， the UI of the treatment group was significantly reduced （P<0.01）， and the serum levels of TNF-α and IL-1β were significantly reduced （P<0.01）. The phosphorylation level of PTEN in gastric mucosal tissue was significantly up-regulated （P<0.01）， and the phosphorylation levels of PI3K， Akt， and NF-κB were significantly downregulated （P<0.01）. ConclusionThe application of network pharmacology prediction， molecular docking simulation， and animal experimental validation confirms that SHXXT regulates the PI3K/Akt/NF-κB signaling pathway to regulate the inflammatory response of rats and thus protects the gastric mucosa of SGU rats.
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Colorectal cancer （CRC） is one of the leading causes of cancer-related deaths worldwide， with increasing incidence and mortality rates. The disease often develops covertly and lacks specific symptoms in its early stages， leading to late-stage diagnoses in most patients. It has become a prominent research topic in the field of digestive system tumors. The exact mechanisms underlying CRC are not yet clear and involve factors such as genetics， gene mutations， inflammatory responses， and aberrant activation of tumor-related signaling pathways. Nuclear factor-kappa B （NF-κB） is a crucial transcription factor that participates in various biological processes， including inflammatory responses， immune responses， cell proliferation， and apoptosis. Research suggests that NF-κB， serving as a molecular link between inflammation and cancer， is highly expressed in CRC. It promotes the occurrence and development of CRC by regulating the activity of target genes such as cell pro-inflammatory factors， chemokines， angiogenic factors， metastasis factors， and anti-apoptotic proteins. Currently， common treatments for CRC include surgery， radiation therapy， and chemotherapy drugs like 5-fluorouracil. However， these treatments have limitations such as significant adverse reactions， high metastasis rates， and the development of drug resistance. Therefore， the search for effective， low-adverse-reaction drugs to replace or supplement current treatments is essential. In recent years， traditional Chinese medicine （TCM） has shown some effectiveness in preventing and treating CRC. TCM has been found to inhibit the growth of CRC cells by modulating the NF-κB signaling pathway， playing a positive role in the occurrence and progression of CRC. Based on the asthenia in origin and sthenia in superficiality and deficiency-excess in complexity in CRC， this article summarized and analyzed the mechanisms and effects of TCM interventions targeting the NF-κB signaling pathway in CRC， and reviewed advances of 10 Chinese medicinal compound formulas and 37 Chinese medicinal monomer components of different types， including flavonoids， phenols， alkaloids， glycosides， and terpenoids with the effects of dispelling pathogenic factors， reinforcing healthy qi， and removing toxins in the prevention and treatment of CRC by targeting the NF-κB pathway. It is found that Chinese medicine can inhibit CRC cell proliferation， invasion， metastasis， angiogenesis， and inflammation by modulating the NF-κB signaling pathway， induce cell apoptosis， restore drug and radiation sensitivity， and counteract CRC. This article is expected to provide insights and references for the in-depth exploration and treatment of CRC mechanisms.
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ObjectiveTo investigate the mechanism by which Shenbai Jiedu prescription （SBJDF） inhibits the proliferation of colorectal cancer （CRC） HCT116 cells. MethodAfter 48 h treatment of HCT116 cells with SBJDF （0， 0.25， 0.5， 1， 2， 4 g·L-1）， the viability of HCT116 cells were determined by methyl thiazolyl tetrazolium （MTT） colorimetry. Following the classification of cells into blank control group and SBJDF （1， 2， 4 g·L-1） groups， the effect of SBJDF on HCT116 cell morphology was observed under an inverted microscope. The effects of SBJDF on the proliferation of HCT116 cells and mitochondrial membrane potential （Δψm） were detected by colony formation assay and JC-1 probe， respectively. The flow cytometry was then performed for determining cell cycle distribution and apoptosis. The effects of SBJDF on cell cycle-， apoptosis-， and nuclear factor kappa-B （NF-κB） signaling pathway-related proteins were determined by Western blot. ResultSBJDF effectively inhibited the vitality of HCT116 cells and changed their morphology in a concentration-dependent manner. Compared with the blank control group， SBJDF at 1， 2， 4 g·L-1 significantly reduced cell colony formation （P<0.05， P<0.01），and SBJDF at 2 and 4 g·L-1 arrested the HCT116 cell cycle at G0/G1 phase （P<0.05， P<0.01）. Compared with the blank control group， SBJDF at 1， 2， 4 g·L-1 remarkably down-regulated the protein expression of CyclinD1 （P<0.05， P<0.01）. SBJDF at 2 and 4 g·L-1 lowered the CyclinA2 and cyclin-dependent kinase 4 （CDK4） （P<0.05， P<0.01）. SBJDF at 4 g·L-1 reduced the cyclin-dependent kinase 1 （CDK1） （P<0.01）. Compared with the blank control group， SBJDF at 2 and 4 g·L-1 induced HCT116 cell apoptosis， down-regulated the protein expression of anti-apoptosis-related proteins Bcl-2 and Bcl-xl as well as the NF-κB signaling pathway-related proteins IκB kinase α （IKKα），inhibitor α of NF-κB （IκBα），and phospho-NF-κB p65 （p-p65） （P<0.05， P<0.01）， and diminished the mitochondrial membrane potential of HCT116 cells. ConclusionSBJDF inhibits the proliferation of HCT116 cells， which may be related to its inhibition of the activation of NF-κB signaling pathway and the induction of cell cycle arrest and apoptosis.
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ObjectiveTo observe the intervention of modified Sanrentang on the lipopolysaccharide-induced proliferation of rat glomerular mesangial cells， the phosphatidylinositol-3 kinase（PI3K）/protein kinase B（PKB/Akt）/nuclear factor kappa B（NF-κB） signaling pathway， and to investigate its mechanism in improving kidney inflammation in rats with immunoglobulin A nephropathy（IgAN）. MethodThe 18 rats were divided into 3 groups by serum pharmacology method： normal group， high-dose and low-dose （20.70，10.35 g·kg-1·d-1） groups with 6 rats in each group. Modified Sanrentang high- and low-dose groups were intragastric with the corresponding solution of modified Sanrentang， and normal group was intragastric with equal volume of distilled water. After 5 days of intragastric administration， blood samples were collected to prepare drug-containing serum. Rat mesangial （HBZY-1） were divided into five groups of normal group， LPS 10 mg·L-1 in the model group， benazepril（50 μmol·L-1）， modified Sanrentang high- and low-dose group. Preclude the use of methyl thiazolyl tetrazolium（MTT） method detect the proliferation activity of HBZY-1 cells， enzyme-linked immunosorbent assay（ELISA） was used to determine the content of each group type Ⅳ collagen（ColⅣ），Western blot and Real-time fluorescence quantitative polymerase chain reaction（Real-time PCR） were used to detect protein and mRNA expression levels of PI3K/Akt/NF-κB signaling pathway. ResultAs compared with the normal group， MTT assay showed that exposure to LPS significantly enhanced the proliferative activity， the ColⅣ was increased significantly of HBZY-1 cells（P<0.01）， p-Akt， p-p65 was increased significantly （P<0.01）. Compared with the model group， the proliferation and ColⅣ of rat chronic glomerulonephritis cells induced by LPS by inhibiting PI3K/Akt/NF-κB signaling pathway（P<0.01）， and the phosphorylation of Akt was significantly inhibited（P<0.01）， the expression levels of NF-κB p65 was reduced in modified Sanrentang high-dose group（P<0.01）. ConclusionModified Sanrentang could inhibit cell proliferation and the content of ColⅣ in rat mesangial cells induced by LPS， and its mechanism might be related to suppression of PI3K/Akt signaling pathway.
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ObjectiveTo investigate the effect of Biejiajian Wan （BJJW） on transforming growth factor-β1 （TGF-β1）-induced epithelial-mesenchymal transition （EMT） of HepG2 cells， and explore its mechanism against EMT of hepatocellular carcinoma cells. MethodHepG2 cells were randomly divided into a blank group， a TGF-β1 model group （10 μg·L-1 TGF-β1）， a low-dose BJJW group （10 μg·L-1 TGF-β1+0.55 g·kg-1 BJJW）， a medium-dose BJJW group （10 μg·L-1 TGF-β1+1.1 g·kg-1 BJJW）， a high-dose BJJW group （10 μg·L-1 TGF-β1+2.2 g·kg-1 BJJW）， and a sorafenib group （10 μg·L-1 TGF-β1+0.03 g·kg-1 sorafenib）. The EMT model was induced by 10 μg·L-1 TGF-β1 in HepG2 cells. After treatment with corresponding medicated serum， cell counting kit -8 （CCK-8） assay was used to detect cell proliferation. Cell migration ability was detected by the Transwell assay and wound healing assay. The protein expression related to EMT and nuclear factor-kappa B （NF-κB） signaling pathway was detected by cell immunofluorescence assay and Western blot. ResultCompared with the blank group 4 days later， the TGF-β1 model group showed fusiform and loose cells with widened gap and antennae reaching out， decreased protein expression of E-cadherin （P<0.05）， and increased protein expression of N-cadherin and vimentin （P<0.05）， which indicated that the EMT model was properly induced in HepG2 cells by TGF-β1 stimulation for 4 days. After 48 hours of treatment with the corresponding medicated serum， each medication group showed inhibited proliferation of HepG2 cells that had undergone EMT， especially the low- and high-dose BJJW groups （P<0.01）， and the medium-dose BJJW group showed increased E-cadherin protein expression （P<0.05） and decreased p-p65， N-cadherin， and vimentin protein expression （P<0.05）， as compared with the TGF-β1 model group. As revealed by the transwell assay and wound healing assay， TGF-β1 enhanced the migration ability of HepG2 cells （P<0.05， P<0.01） compared with the results in the blank group， compared with the TGF-β1 model group， the medication groups showed inhibited migration ability of HepG2 cells （P<0.05， P<0.01）. Compared with the blank group， the TGF-β1 model group promoted the expression of p65 and Snail into the nucleus. Compared with the TGF-β1 model group， the medication groups inhibited the expression of p65 and Snail into the nucleus. ConclusionBJJW may inhibit the EMT， proliferation， and migration of HepG2 cells induced by TGF-β1 by suppressing the NF-κB signaling pathway to exert an anti-hepatocellular carcinoma effect.
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ObjectiveTo investigate the preventive and curative effect of Chaishao Liujunzi Tang （CSLJZT） on colonic mucosal injury induced by dextran sulfate sodium （DSS） in mice with ulcerative colitis （UC） and its mechanism. MethodFifty Balb/c male mice were randomly divided into normal group， model group， CSLJZT low-dose group， CSLJZT high-dose group， and sulfasalazine group. Except for the normal group， other groups were given 2.5% DSS freely for 7 d， and were given drug intervention after successful modeling for 7 d. Bodyweight， feces， and other general physiological statuses of mice were recorded every day， and disease activity index （DAI） scores were calculated.The colon length was measured， and stained by hematoxylin-eosin （HE） staining to observe the morphological changes of the colon.The enzyme-linked immunosorbent assay （ELISA） was used to determine the content of interleukin-1β （IL-1β）， myeloperoxidase （MPO）， and superoxide dismutase （SOD） in the serum. Western blot was used to determine the protein expression levels of Toll-like receptor 4 （TLR4）， myeloid differentiation factor 88 （MyD88）， nuclear factor kappa B （NF-κB）， inhibitor-kappa binding protein （IκB）， Caspase-1， and nucleotide-binding oligomerization domain-like receptor 3 （NLRP3） in the colon tissues. ResultAs compared with the normal group， mice in the model group had significantly decreased body weight （P<0.01）， severe diarrhea and hematochezia， and significantly increased DAI score （P<0.01）. As compared with the model group， the decreasing trend of body weight was significantly alleviated in the CSLJZT groups （P<0.01）， diarrhea and hematochezia were significantly improved， DAI score was significantly decreased （P<0.01）， and colon length increased （P<0.05）. HE staining showed that the pathological damage of colon tissue was significantly improved and the inflammatory cell infiltration was reduced in the CSLJZT groups as compared with the model group. As compared with the normal group， the serum levels of IL-1β and MPO were significantly higher （P<0.01） and SOD levels were significantly lower （P<0.01） of mice in the model group.Compared with the model group， the treated group reduced the serum IL-1β and MPO levels （P<0.01）， and raised the SOD level （P<0.01）. The results of Western blot showed that as compared with the normal group， the expression levels of TLR4， MyD88， NF-κB， Ccaspase-1， and NLRP3 proteins were significantly increased （P<0.01）， whereas the expression level of IκB protein was significantly decreased （P<0.01） in the colonic tissue of mice in the model group. As compared with the model group， the expression levels of TLR4， MyD88， NF-κB， Caspase-1， and NLRP3 proteins were decreased （P<0.01）， whereas the expression level of IκB protein was increased （P<0.01） in the colonic tissue of mice in the CSLJZT groups. ConclusionCSLJZT improves the inflammatory injury of the colon tissue in DSS-induced UC mice through TLR4/MyD88/NF-κB signaling pathway.
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ObjectiveTo investigate the preventive and curative effect of Chaishao Liujunzi Tang （CSLJZT） on colonic mucosal injury induced by dextran sulfate sodium （DSS） in mice with ulcerative colitis （UC） and its mechanism. MethodFifty Balb/c male mice were randomly divided into normal group， model group， CSLJZT low-dose group， CSLJZT high-dose group， and sulfasalazine group. Except for the normal group， other groups were given 2.5% DSS freely for 7 d， and were given drug intervention after successful modeling for 7 d. Bodyweight， feces， and other general physiological statuses of mice were recorded every day， and disease activity index （DAI） scores were calculated.The colon length was measured， and stained by hematoxylin-eosin （HE） staining to observe the morphological changes of the colon.The enzyme-linked immunosorbent assay （ELISA） was used to determine the content of interleukin-1β （IL-1β）， myeloperoxidase （MPO）， and superoxide dismutase （SOD） in the serum. Western blot was used to determine the protein expression levels of Toll-like receptor 4 （TLR4）， myeloid differentiation factor 88 （MyD88）， nuclear factor kappa B （NF-κB）， inhibitor-kappa binding protein （IκB）， Caspase-1， and nucleotide-binding oligomerization domain-like receptor 3 （NLRP3） in the colon tissues. ResultAs compared with the normal group， mice in the model group had significantly decreased body weight （P<0.01）， severe diarrhea and hematochezia， and significantly increased DAI score （P<0.01）. As compared with the model group， the decreasing trend of body weight was significantly alleviated in the CSLJZT groups （P<0.01）， diarrhea and hematochezia were significantly improved， DAI score was significantly decreased （P<0.01）， and colon length increased （P<0.05）. HE staining showed that the pathological damage of colon tissue was significantly improved and the inflammatory cell infiltration was reduced in the CSLJZT groups as compared with the model group. As compared with the normal group， the serum levels of IL-1β and MPO were significantly higher （P<0.01） and SOD levels were significantly lower （P<0.01） of mice in the model group.Compared with the model group， the treated group reduced the serum IL-1β and MPO levels （P<0.01）， and raised the SOD level （P<0.01）. The results of Western blot showed that as compared with the normal group， the expression levels of TLR4， MyD88， NF-κB， Ccaspase-1， and NLRP3 proteins were significantly increased （P<0.01）， whereas the expression level of IκB protein was significantly decreased （P<0.01） in the colonic tissue of mice in the model group. As compared with the model group， the expression levels of TLR4， MyD88， NF-κB， Caspase-1， and NLRP3 proteins were decreased （P<0.01）， whereas the expression level of IκB protein was increased （P<0.01） in the colonic tissue of mice in the CSLJZT groups. ConclusionCSLJZT improves the inflammatory injury of the colon tissue in DSS-induced UC mice through TLR4/MyD88/NF-κB signaling pathway.
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Objective::To observe the effect of berberine and 6-shogaol, main components of Coptiae Rhizoma and Zingiberis Rhizoma, on the inflammatory signaling pathway of Toll-like receptors 4 (TLR4)/nuclear factor kappa B (NF-κB) in colonic epithelial cells of mice with ulcerative colitis. Method::Fifty Kunming mice were randomly divided into normal group, model group, berberine group (100 mg·kg-1), 6-shogaol group (100 mg·kg-1), and 6-shogaol combined with berberine group (200 mg·kg-1), with 10 mice in each group. A mouse model of ulcerative colitis was established through oral administration with 2% dextroan sulfate for two weeks. Each group was given corresponding drugs by gavage, while normal group and model group were given equal amount of normal saline. Serum and colon tissue samples were taken 20 days after administration. Enzyme-linked immunosorbent method was used to detect serum interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) expressions, and real-time fluorescence quantitative polymerase chain reaction(Real-time PCR) and Western blot method were used to detect TLR4, NF-κB p65 mRNA and protein expressions in colon epithelial tissue. Result::Compared with the normal group, relative expressions of TLR4 and NF-κB p65 mRNA and protein were increased in the model group (P<0.01), and the contents of serum IL-1β and TNF-α were increased (P<0.01). Compared with the model group, relative expressions of TLR4 and NF-κB p65 mRNA and protein were significantly decreased in 6-shogaol group, berberine group and 6-shogaol combined with berberine group (P<0.01), and the contents of serum IL-1β and TNF-α were significantly decreased (P<0.01). Among the three groups, 6-shogaol combined with berberine group had the strongest effect (P<0.01). Conclusion::Both 6-shogaol and berberine can inhibit colonic inflammation, reduce inflammatory damage and treat ulcerative colitis. The combined application of 6-shogaol and berberine has a significant synergism effect. The mechanism is related to the excessive activation of TLR4/NF-κB pathway and the regulation of non-controllable intestinal inflammation.
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Objective::To established the model of chronic alcoholic liver injury in rats by long-term(8 weeks) alcoholic gavage, to study the effects of Tibetan medicine Lagotis brachystachys extracts on Toll-like receptor(TLR)2/myeloid differentiation factor 88(MyD88)/nuclear factor kappa B (NF-κB)and NOD like receptor protein 3(NALP3) signaling pathways and study preliminary the mechanism of action of chronic alcoholic liver injury. Method::Sixty male Sprague-Dawley rats were randomly divided into normal group, model group, bifendate positive drug group (0.1 g·kg-1) and L. brachystachys low, medium and high-dose groups (0.5, 1, 2 g·kg-1), the corresponding drugs were given at 10 mL·kg-1 in each morning, and the 56 degree Liquor was administered by the afternoon gradient alcoholic gavage method.After 8 weeks, the levels of serum aspartate transaminase (AST), serum alanineaminotransfease(ALT), serum total cholesterol(TC), triglyceride(TG), interleukin-1β(IL-1β), and the liver levels of L-glutathione(GSH)were measured. The expression of TLR2, MyD88, NF-κB and NALP3 protein in liver were detected by Western blot.Hematoxylin-eosin (HE) staining was used to observe the pathological changes of liver tissue. Result::Compared with normal group, the serum levels of AST, ALT, TC, TG and IL-1β in model group were significantly increased (P<0.05, P<0.01). Compared with model group, the serum AST, ALT, TC, TG and IL-1β levels were decreased in the various doses of L. brachystachys, and the high dose group was particularly effective (P<0.05, P<0.01). Compared with normal group, the GSH level in the liver homogenate of model group decreased significantly, and the difference was not statistically significant. The levels of TLR2, MyD88, NF-κB and NALP3 in the liver tissue of model group were significantly increased (P<0.05, P<0.01). The GSH levels in the liver and the protein expression of TLR2, MyD88, NF-κB and NALP3 were decreased in L. brachystachys group (P<0.05, P<0.01). The liver pathological section showed that L. brachystachys can improve the pathological changes of rat liver tissue. Conclusion::L. brachystachys can protect liver from alcohol-induced chronic liver injury in rats. The mechanism was related to TLR2/MyD88/NF-κB and NALP3 signaling pathway.