Hepatic sinusoidal obstruction syndrome (HSOS) via exposure to pyrrolizidine alkaloids (PAs) is with high mortality and there is no effective treatment in clinics. Bear bile powder (BBP) is a famous traditional animal drug for curing a variety of hepatobiliary diseases such as cholestasis, inflammation, and fibrosis. Here, we aim to evaluate the protective effect of BBP against HSOS induced by senecionine, a highly hepatotoxic PA compound. Our results showed that BBP treatment protected mice from senecionine-induced HSOS dose-dependently, which was evident by improved liver histology including reduced infiltration of inflammatory cells and collagen positive cells, alleviated intrahepatic hemorrhage and hepatic sinusoidal endothelial cells, as well as decreased conventional serum liver function indicators. In addition, BBP treatment lowered matrix metalloproteinase 9 and pyrrole-protein adducts, two well-known markers positively associated with the severity of PA-induced HSOS. Further investigation showed that BBP treatment prevents the development of liver fibrosis by decreasing transforming growth factor beta and downstream fibrotic molecules. BBP treatment also alleviated senecionine-induced liver inflammation and lowered the pro-inflammatory cytokines, in which tauroursodeoxycholic acid played an important role. What's more, BBP treatment also decreased the accumulation of hydrophobic bile acids, such as cholic acid, taurocholic acid, glycocholic acid, as well. We concluded that BBP attenuates senecionine-induced HSOS in mice by repairing the bile acids homeostasis, preventing liver fibrosis, and alleviating liver inflammation. Our present study helps to pave the way to therapeutic approaches of the treatment of PA-induced liver injury in clinics.
Animals ; Bile ; Bile Acids and Salts ; Endothelial Cells/metabolism* ; Hepatic Veno-Occlusive Disease/pathology* ; Inflammation/pathology* ; Liver Cirrhosis/drug therapy* ; Mice ; Powders ; Pyrrolizidine Alkaloids/adverse effects* ; Ursidae

OBJECTIVE: To screen the active components from Fuzheng Huayu Recipe (FZHY) and redesign a new recipe composed of the active components, and validate the effect of active components formulation from FZHY against liver fibrosis. METHODS: Thirty-two components from FZHY were evaluated for their activities against liver fibrosis respectively, with 6 kinds of cell models in vitro, including oxidative stressed hepatocyte in L-02, hypoxia injured/proliferative hepatic sinusoidal endothelial cells in SK-HEP-1 and human hepatic sinusoidal endothelial cells (HHSEC), and activated hepatic stellate cell in LX-2. The comprehensive activity of each component against liver fibrosis was scored according to the role of original herbs in FZHY and cell functions in fibrogenesis. Totally 7 active components were selected and combined with equal proportion to form a novel active components formulation (ACF). The efficacy of ACF on liver fibrosis were evaluated on activation of LX-2 and proliferation of HHSEC in vitro and in liver fibrosis model mice induced by dimethylnitrosamine (DMN). Totally 72 mice were divided into 6 groups using a random number table, including normal, high-dose ACF control (20 µ mol/L × 7 components/kg body weight), model, low-, medium-, high-dose ACF groups (5, 10, 20 µ mol/L × 7 components/kg body weight, respectively). Hematoxylin eosin and Sirius red stainings were used to observe inflammation and fibrosis change of liver tissue; scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were utilized to observe the effect of ACF on ultrastructure of hepatic sinusoids. RESULTS: Fifteen components from FZHY showed higher scores for their activity on against liver fibrosis. Among them, 7 components including tanshinone II A, salvianolic acid B, cordycepin, amygdalin, quercetin, protopanaxatriol, and schizandrin B were recombined with equal proportions to form ACF. ACF at 1,2, 4 µ mol/L showed strong inhibitory effects on activation of LX-2 and proliferation of HHSEC in vitro (all P<0.01). Compared with the model group, ACF attenuated liver collagen deposition, improved sinusoidal capillarization in a dose-dependent manner (all P<0.05). CONCLUSION ACF exerts a satisfactory effect against experimental liver fibrosis and attenuates sinusoidal capillarization, which warrant a further research and development for herbal components formulation on liver fibrosis.
Animals ; Body Weight ; Drugs, Chinese Herbal/adverse effects* ; Endothelial Cells ; Liver ; Liver Cirrhosis/drug therapy* ; Mice

Animals ; Apoptosis/drug effects* ; Autophagy/drug effects* ; Carnitine/pharmacology* ; Cells, Cultured ; Endothelial Cells ; Lipopolysaccharides ; Mice

This study aims to explore the effect of extract of Ginseng Radix et Rhizoma, Notoginseng Radix et Rhizoma, and Chuanxiong Rhizoma(hereinafter referred to as GNS) on the SIRT1-autophagy pathway of endothelial cell senescence induced by hydrogen peroxide(H_2O_2). To be specific, vascular endothelial cells were classified into the blank control group(control), model group(model), model + DMSO group(DMSO), resveratrol group(RESV), and GNS low-dose(GNS-L), medium-dose(GNS-M), and high-dose(GNS-H) groups. They were treated with H_2O_2 for senescence induction except the control. After intervention of cells in each group with corresponding drugs for 24 h, cell growth status was observed under an inverted microscope, and the formation of autophagosome under the transmission electron microscope. In addition, the changes of microtubule-associated protein 1 light chain 3β(LC3 B) were detected by immunofluorescence staining. The autophagy flux was tracked with the autophagy double-labeled adenovirus(mRFP-GFP-LC3) fusion protein. Dansylcadaverine(MDC) staining was employed to determine the autophagic vesicles, and Western blot the expression of sirtuin 1(SIRT1), ubiquitin-binding protein p62, and LC3Ⅱ. After H_2O_2 induction, cells demonstrated slow growth, decreased adhesion ability, raised number of SA-β-gal-stained blue ones, a certain number of autophagosomes with bilayer membrane and secondary lysosomes in the cytoplasm, and slight rise of autophagy flux level. Compared with the model group, GNS groups showed improved morphology, moderate adhesion ability, complete and smooth membrane, decreased SA-β-gal-stained blue cells, many autophagosomes, autophagic vesicles, and secondary lysosomes in the cytoplasm, increased autophagolysosomes, autophagy flux level, and fluorescence intensity of LC3 B and MDC, up-regulated expression of SIRT1 and LC3Ⅱ, and down-regulated expression of p62, suggesting the improvement of autophagy level. GNS can delay the senescence of vascular endothelial cells. After the intervention, the autophagy flux and related proteins SIRT1, LC3Ⅱand p62 changed significantly, and the autophagy level increased significantly. However, EX527 weakened the effect of Chinese medicine in delaying vascular senescence. GNS may delay the senescence of vascular endothelial cells through the SIRT1 autophagy pathway.
Autophagy ; Cells, Cultured ; Cellular Senescence ; Drugs, Chinese Herbal/pharmacology* ; Endothelial Cells/drug effects* ; Hydrogen Peroxide ; Panax/chemistry* ; Sirtuin 1/genetics*

Endothelial progenitor cells (EPCs) play an important role in diabetic vascular complications. A large number of studies have revealed that some clinical antihyperglycemics can improve the complications of diabetes by regulating the function of EPCs. Metformin can improve EPCs function in diabetic patients by regulating oxidative stress level or downstream signaling pathway of adenosine monophosphate activated protein kinase; Pioglitazone can delay the aging of EPCs by regulating telomerase activity; acarbose, sitagliptin and insulin can promote the proliferation, migration and adhesion of EPCs. In addition to lowering blood glucose, the effects of antihyperglycemics on EPCs may also be one of the mechanisms to improve the complications of diabetes. This article reviews the research progress on the regulation of EPC proliferation and function by antihyperglycemics.
Cell Movement/drug effects* ; Cells, Cultured ; Endothelial Progenitor Cells/drug effects* ; Humans ; Hypoglycemic Agents/pharmacology* ; Signal Transduction/drug effects*

OBJECTIVE: To elucidate the correlation between CKLF-like marvel transmembrane domain containing member (CMTM5) gene and the risk of in-stent restenosis (ISR) with coronary artery disease (CAD) patients and to detect the effects and mechanisms of CMTM5-stimulated genes on human vascular endothelial cells (ECs) proliferation and migration. METHODS: A total of 124 hospitalized patients in Shijitan Hospital were enrolled in this study. All the CAD patients were detected with platelet reactivity and grouped into two groups according to platelet reactivity; ISR was conformed by coronary angiography; RT-PCR method was used to detect CMTM5 gene expression; The CMTM5 over expression, reduction and control EC lines were established; Cell count, MTT, Brdu and flow cytometry methods were used to detect the proliferation of ECs, scratch and transwell experiments to test the migration of ECs, Western blot was used to detect signal path expressions. RESULTS: CMTM5 gene expression in HAPR (High on aspirin platelet reactivity) group was 1.72 times compared with No-HAPR group, which was significantly higher than No-HAPR group. HAPR group ISR rate was 25.8% (8 cases), the incidence of No-HAPR ISR group was 9.7% (9 cases), and the results showed that in HAPR group, the incidence of ISR was significantly higher than that in No-HAPR group (P=0.04, OR=0.04, 95%CI=1.16－7.52), which showed that CMTM5 gene was significantly correlated with the risk of ISR. In HAPR group ISR rate was 25.8% (8 cases), the incidence of ISR in No-HAPR group was 9.7% (9 cases), and the results showed that the risk of ISR in HAPR group was significantly higher than that in No-HAPR group. All the results showed that CMTM5 was significantly correlated with the risk of ISR in CAD patients (P < 0.05). CMTM5 overexpression inhibited the proliferation and migration ability of ECs (P < 0.05), PI3K/Akt signaling pathways were involved in the role of regulation on ECs. CONCLUSION Our results revealed that CMTM5 gene was closely related with ISR, CMTM5 overexpression may repress ECs proliferation and migration through regulating PI3K-Akt signaling.
Chemokines ; Coronary Artery Disease/surgery* ; Coronary Restenosis ; Drug-Eluting Stents/adverse effects* ; Endothelial Cells ; Humans ; MARVEL Domain-Containing Proteins ; Phosphatidylinositol 3-Kinases ; Tumor Suppressor Proteins

Ten fractions(A-J) were prepared by separation of Longxue Tongluo Capsules(LTC) by using silica gel column chromatography and orthogonal experimental design,showing similar chemical profiles with different abundances of peaks.These ten samples were assessed with UHPLC-QE OrbitrapHRMS for 97 common peaks.For the pharmacological activity experiment,three kinds of in vitro cell models including lipopolysaccharide(LPS)-induced BV-2 microglial cells NO release model,oxygen-glucose deprivation/reoxygenation(OGD/R)-treated HUVEC vascular endothelial cells injury model,and OGD/R-treated PC-12 nerve cells injury model were employed to evaluated the bioactivity of each fraction.Based on the contribution of each identified component,grey relation analysis and partial least squares(PLS) analysis were performed to establish component-activity relationship of LTC,identify the potential active components.After that,validation of the potential active components in LTC was carried out by using the same models.The results indicated that 4 phenolic compounds including 7,4'-dihydroxyhomoisoflavanone,loureirin C,4,4'-dihydroxy-2,6-dimethoxydihydrochalcone,and homoisosocotrin-4'-ol,might be the active components for anti-neuroinflammation effect;five phenolic compounds such as 3,5,7,4'-tetrahydroxyhomoisoflavanone,loureirin D,7,4'-dihydroxyhomoisoflavane,and 5,7-dihydroxy-4'-methoxy-8-methyflavane,might have positive effects on the vascular endothelial injury;three phenolic compounds including 5,7,4'-trihydroxyflavanone,7,4'-dihydroxy-5-methoxyhomoisoflavane,and loureirin D,might be the active components in LTC against neuronal injury.
Brain Ischemia ; drug therapy ; Capsules ; Cell Line ; Drugs, Chinese Herbal ; pharmacology ; Glucose ; Human Umbilical Vein Endothelial Cells ; drug effects ; Humans ; Microglia ; drug effects ; Oxygen

PURPOSE: Wound represents a major health challenge as they consume a large amount of healthcare resources to improve patient's quality of life. Many scientific studies have been conducted in search of ideal biomaterials with wound-healing activity for clinical use and collagen has been proven to be a suitable candidate biomaterial. This study intended to investigate the wound healing activity of collagen peptides derived from jellyfish following oral administration. METHODS: In this study, collagen was extracted from the jellyfish--Rhopilema esculentum using 1% pepsin. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and fourier transform infrared (FTIR) were used to identify and determine the molecular weight of the jellyfish collagen. Collagenase II, papain and alkaline proteinase were used to breakdown jellyfish collagen into collagen peptides. Wound scratch assay (in vitro) was done to determine migration potential of human umbilical vein endothelial cells (HUVEC) covering the artificial wound created on the cell monolayer following treatment with collagen peptides. In vivo studies were conducted to determine the effects of collagen peptides on wound healing by examining wound contraction, re-epithelialization, tissue regeneration and collagen deposition on the wounded skin of mice. Confidence level (p < 0.05) was considered significant using GraphPad Prism software. RESULTS: The yield of collagen was 4.31%. The SDS-PAGE and FTIR showed that extracted collagen from jellyfish was type I. Enzymatic hydrolysis of this collagen using collagenase II produced collagen peptides (CP) and hydrolysis with alkaline proteinase/papain resulted into collagen peptides (CP). Tricine SDS-PAGE revealed that collagen peptides consisted of protein fragments with molecular weight <25 kDa. Wound scratch assay showed that there were significant effects on the scratch closure on cells treated with collagen peptides at a concentration of 6.25 μg/mL for 48 h as compared to the vehicle treated cells. Overall treatment with collagen peptide on mice with full thickness excised wounds had a positive result in wound contraction as compared with the control. Histological assessment of peptides treated mice models showed remarkable sign of re-epithelialization, tissue regeneration and increased collagen deposition. Immunohistochemistry of the skin sections showed a significant increase in β-fibroblast growth factor (β-FGF) and the transforming growth factor-β (TGF-β) expression on collagen peptides treated group. CONCLUSION Collagen peptides derived from the jellyfish-Rhopilema esculentum can accelerate the wound healing process thus could be a therapeutic potential product that may be beneficial in wound clinics in the future.
Administration, Oral ; Animals ; Collagen ; administration & dosage ; isolation & purification ; metabolism ; pharmacology ; Fibroblast Growth Factors ; metabolism ; Human Umbilical Vein Endothelial Cells ; Humans ; Male ; Regeneration ; Scyphozoa ; chemistry ; Skin ; metabolism ; Skin Physiological Phenomena ; Stimulation, Chemical ; Transforming Growth Factor beta1 ; metabolism ; Wound Healing ; drug effects

The effect of triptolide( TP) on VEGFA,SDF-1,CXCR4 pathway were investigated in vitro to explore the mechanism in improving platelet activation in patients with ankylosing spondylitis( AS). Peripheral blood mononuclear cells( PBMC) were used for the experiment and divided into 4 groups: normal group( NC),model group( MC),triptolide group( TP),and AMD3100 group. The optimal concentration of TP was measured by the MTT method. The expressions of TNF-α,IL-1β,IL-4,IL-10,VEGFA and VEGFR were detected by ELISA. The expressions of SDF-1,CXCR4 and VEGFA were detected by real-time quantitative PCR( RT-qPCR).The expressions of SDF-1,CXCR4,VEGFA and VEGFR were detected by Western blot. The expression levels of CD62 p,CD40 L and PDGFA were detected by immunofluorescence. MTT results showed that medium-dose TP had the strongest inhibitory effect on cells at24 h. The results of ELISA and PCR showed that TP inhibited mRNA expressions of IL-1β,TNF-α,VEGFA,VEGFR and SDF-1,CXCR4 and VEGFA. The results of Western blot indicated that TP inhibited SDF-1,CXCR4 and VEGFA,VEGFR protein expressions; immunofluorescence results indicate that TP can inhibit the expressions of CD62 p,CD40 L,PDGFA. TP may regulate platelet activation by down-regulating SDF-1,CXCR4,VEGFA and VEGFR mRNA expressions,thereby down-regulating IL-1β and TNF-αexpressions,and up-regulating the expressions of IL-4 and IL-10 cytokines.
Cells, Cultured ; Chemokine CXCL12 ; metabolism ; Cytokines ; metabolism ; Diterpenes ; pharmacology ; Epoxy Compounds ; pharmacology ; Heterocyclic Compounds ; pharmacology ; Humans ; Leukocytes, Mononuclear ; drug effects ; Phenanthrenes ; pharmacology ; Platelet Activation ; Receptors, CXCR4 ; metabolism ; Spondylitis, Ankylosing ; Vascular Endothelial Growth Factor A ; metabolism

To observe the effect of Tripterygium Glycosides Tablets on angiogenesis of rats with type Ⅱ collagen-induced arthritis( CIA) and on the tube formation of human umbilical vein endothelial cells( HUVEC) in vitro. The HUVEC were induced by 20 μg·L-1 vascular endothelial growth factor( VEGF) in vitro,and were treated with 0. 1,1,10 mg·L-1 Tripterygium Glycosides Tablets continuously for 7 hours. The numbers of branches of tube formation were measured. SD rats were immunized to establish CIA. CIA rats were treated with 9,18,36 mg·kg-1·d-1 Tripterygium Glycosides Tablets for 42 days. Histopathological examination( HE) was performed to observe the vascular morphology and vascular density in the synovial membrane of the inflamed joints. Immunohistochemistry and immunofluorescence were performed to observe the expression of platelets-endothelial cell adhesion molecule( CD31) and αsmooth muscle actin( αSMA) in synovial membrane. Immunohistochemistry and Western blot were performed to observe the expression of hypoxia-inducible factors 1α( HIF1α) and angiotensin 1( Ang1) in the synovial tissue. The results showed that the numbers of branches of tube formation of HUVEC induced by VEGF were improved,and declined significantly after treated by Tripterygium Glycosides Tablets. Compared with the normal group,the vascular density,CD31 positive expression,CD31 +/αSMA-immature and total vascular positive expression in the synovial membrane of the model group were significantly increased,and so as HIF1α and Ang1 in the synovium. Tripterygium Glycosides Tablets reduced the synovial vascular density and inhibited the positive expression of CD31,CD31+/αSMA-immature blood vessels and total vascular,but has no effect on CD31+/αSMA+mature blood vessels. Tripterygium Glycosides Tablets also inhibited the expression of HIF1α and Ang1 in synovial membrane of inflammatory joints. Our results demonstrated that Tripterygium Glycosides Tablets could inhibit the angiogenesis of synovial tissue in CIA rats and the tube formation of HUVEC,which is related to the down-regulation of HIF1α/Ang1 signal axis.
Angiogenesis Inhibitors ; pharmacology ; Angiotensin I ; metabolism ; Animals ; Arthritis, Experimental ; chemically induced ; drug therapy ; Drugs, Chinese Herbal ; pharmacology ; Glycosides ; pharmacology ; Human Umbilical Vein Endothelial Cells ; drug effects ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Synovial Membrane ; drug effects ; Tablets ; Tripterygium ; chemistry ; Vascular Endothelial Growth Factor A