Objective:To discuss the regulatory effect of physiological tensile stress on the differentiation of chondrocytes,and to clarify the associated signaling pathway mechanism.Methods:The ATDC5 chondrocytes were cultured in vitro and subjected to physiological tensile stress by four-point bending cell mechanical loading device.Initially,the cells were divided into control group and tensile stress group(2 000 μstrain/2 h group),and further divided into different stress magnitudes(1 000,2 000,and 3 000 μstrain)for 2 h,and 2 000 μstrain for different duration time(1,2,and 4 h)groups;the cells without tensile stress were used as control group.Real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of type Ⅱ collagen(Col-Ⅱ),type Ⅹ collagen(Col-Ⅹ),aggregated proteoglycom(Aggrecan),sex-determining region Y-box protein 9(SOX9),vascular endothelial growth factor(VEGF),proliferating cell nuclear antigen(PCNA),Nel-like molecule tyep 1(Nell-1),Runt-related transcription factor 2(Runx2),Indian hedgehog(Ihh),patched homolog 1(Ptch-1),GLI family zinc finger protein 1(Gli-1),and hedgehog interacting protein 1(Hhip-1)mRNA in the cells in various groups;Western blotting method was used to detect the expression levels of Nell-1,Runx2,and Ihh proteins in the cells in various groups.The ATDC5 cells were divided into control group,cyclopamine group,tensile stress group,and cyclopamine + tensile stress group.RT-qPCR method was used to detect the expression levels of Nell-1,Ihh,Ptch-1,Gli-1,and Hhip-1 mRNA in the cells in various groups;Western blotting method was used to detect the expression levels of Nell-1 and Ihh proteins in the cells in various groups.Results:Compared with control group,the expression levels of Col-Ⅱ,Col-Ⅹ,Aggrecan,SOX9,VEGF,and PCNA mRNA in the cells in 2 000 μstrain/2 h group were significantly increased(P<0.01);after treated with 2 000 μstrain tensile stress for different duration time(1,2,and 4 h)or different tensile stresses(1 000,2 000,and 3 000 μstrain)for 2 h,compared with control group,the expression levels of Runx2 mRNA in the cells in other groups were increased with the prolongation of time or the increasing of tensile stress(P<0.01),and the expression levels of Nell-1,Ihh,Ptch-1,Gli-1,and Hhip-1 mRNA were gradually increased(P<0.01),the expression levels reached the peaking at 2 000 μstrain/2 h,and then decreased but remained significantly higher than that in control group(P<0.01).The Western blotting results showed that the expression levels of Nell-1,Runx2,and Ihh proteins in the cells were consistent with the change trend of mRNA expression levels.After pre-treated with cyclopamine,compared with control group,the expression levels of Ihh,Ptch-1,Gli-1,and Hhip-1 mRNA in the cells in cyclopamine group were significantly decreased(P<0.01),and the expression levels of Ihh,Ptch-1,Gli-1,and Hhip-1 mRNA in the cells in tensile stress and cyclopamine+tensile stress groups were significantly increased(P<0.01);compared with cyclopamine group,the expression levels of Nell-1,Ihh,Ptch-1,Gli-1,and Hhip-1 mRNA in the cells in cyclopamine+tensile stress group were significantly increased(P<0.01);compared with tensile stress group,the expression levels of Ihh,Ptch-1,Gli-1,and Hhip-1 mRNA in the cells in cyclopamine + tensile stress group were significantly decreased(P<0.01).Compared with control group,the expression level of Ihh protein in the cells in cyclopamine group was significantly decreased(P<0.01),but there was no significant difference in expression level of Nell-1 protein in the cells between control group and cyclopamine group(P>0.05),while the expression levels of Nell-1 and Ihh proteins in the cells in tensile stress group and cyclopamine + tensile stress group were significantly increased(P<0.01);compared with cyclopamine group,the expression levels of Nell-1 and Ihh proteins in the cells in tensile stress group and cyclopamine + tensile stress group were significantly increased(P<0.01);compared with tensile stress group,in the expression levels of Nell-1 and Ihh proteins in the cells in cyclopamine + tensile stress group had no significant differences(P>0.05).Conclusion:After stimulated with physiological tensile stress,Nell-1 can activate the Ihh signaling pathway upstream,and regulate the differentiation of the ATDC5 chondrocytes.

Objective:To discuss the effect of Jiegeng Yuanshen Tang(JGYST)on airway tissue inflammation and mucus secretion in the mice with allergic asthma,and to clarify the related mechanism.Methods:Forty male C57BL/J mice were randomly divided into control group,JGYST group,ovalbumin(OVA)group,and OVA + JGYST group.The mice in OVA group and OVA +JGYST group were sensitized with 50 μg OVA via intraperitoneal injection twice weekly,followed by 20 μg OVA nasal drops daily for 7 d to induce asthma;the mice in OVA +JGYST group were gavaged with 200 μL JGYST 1 h before each OVA challenge,and the administration lasted for 7 d;the mice in control group were given equivalent dose of PBS via intraperitoneal injection,nasal drops,and gavage;the mice in JGYST group were given the same dose of PBS for intraperitoneal and nasal administration and gavaged with the same dose of JGYST.The pathomorphology of lung tissue of the mice in various groups was observed by HE staining and periodic acid-Schiff(PAS)staining,and the inflammation and PAS scores were calculated;flow cytometry method was used to detect the numbers of eosinophils,neutrophils,helper T lymphocyte 1(Th1)cells,helper T lymphocyte 2(Th2)cells,and dendritic cells(DCs),as well as the percentage of mature DCs and level of reactive oxygen species(ROS)in lung tissue of the mice in various groups;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of interleukin-4(IL-4),interleukin-10(IL-10),and tumor necrosis factor-α(TNF-α)mRNA in lung tissue of the mice in various groups.Results:The HE and PAS staining results showed that the mice in control group had intact airway and alveolar structure,without infiltration of inflammatory cells or mucus secretion;compared with control group,there was a large number of infiltrating inflammatory cells in airway tissue of the mice in OVA group,and the inflammation and PAS scores were increased(P<0.01);compared with OVA group,the infiltration of inflammatory cells in airway tissue of the mice in JGYST group and OVA + JGYST group was decreased,and the inflammation and PAS scores were significantly decreased(P<0.01).The flow cytometry results showed that compared with control group,the numbers of eosinophils,Th2 cells,and DCs in lung tissue of the mice in OVA group were increased(P<0.05 or P<0.01),and the percentage of mature DCs and level of ROS were significantly increased(P<0.01);compared with OVA group,the numbers of eosinophils,Th2 cells,and DCs in lung tissue of the mice in JGYST group and OVA + JGYST group were decreased(P<0.01),and the percentage of mature DCs and level of ROS were significantly decreased(P<0.01).The RT-qPCR results showed that compared with control group,the expression levels of IL-4,IL-10,and TNF-α mRNA in lung tissue of the mice in OVA group were increased(P<0.01);compared with OVA group,the expression levels of IL-4 and TNF-α mRNA in lung tissue of the mice in JGYST group and OVA + JGYST group were decreased(P<0.01),while the expression level of IL-10 mRNA was increased(P<0.01).Conclusion:JGYST can alleviate the airway tissue inflammation and mucus secretion in the mice with allergic asthma,and its mechanism may be related to reducing the number of Th2 cells and DCs,decreasing the ROS level and expression level of proinflammatory cytokine,and increasing the expression level of anti-inflammatory cytokine.

Objective:To discuss the expression of programmed cell death-ligand 1(PD-L1)in the oral squamous cell carcinoma(OSCC)cells and its effect on biological behavior of the OSCC CAL27 cells,and to clarify the possible mechanism.Methods:Western blotting method was used to detect the expression levels of PD-L1 protein in the oral epithelial HOK cells and OSCC CAL27,TCA8113,and SCC15 cells;immunofluorescence staining method was used to detect the expression and localization of PD-L1 protein in the CAL27 cells.The CAL27 cells were divided into control group(transfected with si-NC)and si-PD-L1 group(transfected with si-PD-L1).Western blotting method was used to detect the interference efficiency of the cells in two groups;CCK-8 assay was used to detect the proliferative activities of the cells in two groups at different time points;plate clone formation assay was used to detect the numbers of clone formation of the cells in two groups;cell scratch healing assay was used to detect the scratch healing rates of the cells in two groups;Transwell chamber assay was used to detect the numbers of migration and invasion cells in two groups.Results:The expression level of PD-L1 protein in the OSCC cells was higher than that in the HOK cells(P<0.05 or P<0.01);PD-L1 expressed in the cytoplasm and nucleus of the CAL27 cells.The CCK-8 assay and plate clone formation assay results showed that compared with control group,the proliferative activities of the CAL27 cells in si-PD-L1 group at different time points were significantly decreased(P<0.05 or P<0.01),and the numbers of clone formation were significantly decreased(P<0.01).The cell scratch healing assay results showed that compared with control group,the scratch healing rates of the cells in si-PD-L1 group were significantly decreased(P<0.05 or P<0.01).The Transwell chamber assay results showed that compared with control group,the numbers of migration and invasion cells in si-PD-L1 group were significantly decreased(P<0.01).Conclusion:The expression of PD-L1 in the OSCC cells is higher than that in normal oral epithelial cells,and knocking down PD-L1 expression can inhibit the proliferation,clone formation,migration and invasion capabilities of the OSCC cells.

Objective:To discuss the inhibitory effect of chelerythrine(CHE)on the migration,invasion,and epithelial-mesenchymal transition(EMT)of the human ovarian cancer SKOV3 cells,and to clarify the associated mechanism.Methods:The SKOV3 cells were cultured in vitro and divided into control group and 2.5,5.0,10.0,20.0,and 40.0 μmol·L-1 CHE groups.Methylthiazolydiphenyl-tetrazolium(MTT)assay was used to detect the inhibitory rates of proliferation of the cells in various groups.The SKOV3 cells were cultured in vitro and divided into control group,transforming growth factor-β1(TGF-β1)group,TGF-β1+5 μmol·L-1 CHE group,and TGF-β1+10 μmol·L-1 CHE group.Cell scratch assay was used to detect the migration rates of the cells in various groups;Transwell chamber assay was used to detect the numbers of migration and invasion cells in various groups;Western blotting method was used to detect the expression levels of E-cadherin,N-cadherin,and Vimentin proteins in the cells in various groups;immunofluorescence staining method was used to detect the fluorescence intensities of E-cadherin and N-cadherin in the cells in various groups.Results:The MTT assay results showed that compared with control group,the inhibitory rates of proliferation of the cells in 5.0,10.0,20.0,and 40.0 μmol·L-1 CHE groups were significantly increased(P<0.05 or P<0.01).The cell scratch assay results showed that compared with control group,the migration rate of the cells in TGF-β1 group was increased(P<0.01);compared with TGF-β1 group,the migration rates of the cells in TGF-β1+5 μmol·L-1 CHE group and TGF-β1+10 μmol·L-1 CHE group were significantly decreased(P<0.01).The Transwell chamber assay results showed that compared with control group,the numbers of migration and invasion cells in TGF-β1 group were significantly increased(P<0.05);compared with TGF-β1 group,the numbers of migration and invasion cells in TGF-β1+5 μmo·l L-1 CHE group and TGF-β1+10 μmo·l L-1 CHE group were significantly decreased(P<0.01).The Western blotting results showed that compared with control group,the expression level of E-cadherin protein in the cells in TGF-β1 group was significantly decreased(P<0.01),while the expression levels of N-cadherin and Vimentin proteins were increased(P<0.05 or P<0.01);compared with TGF-β1 group,the expression levels of E-cadherin protein in the cells in TGF-β1+5 μmol·L-1 CHE group and TGF-β1+10 μmol·L-1 CHE group were significantly increased(P<0.01),and the expression levels of N-cadherin and Vimentin proteins were significantly decreased(P<0.01).The immunofluorescence staining results showed that compared with control group,the fluorescence intensity of E-cadherin in the cells in TGF-β1 group was decreased,and the fluorescence intensity of N-cadherin was increased;compared with TGF-β1 group,the fluorescence intensities of E-cadherin in the cells in TGF-β 1+5 μmol·L-1 CHE group and TGF-β1+10 μmol·L-1 CHE group were significantly increased,and the fluorescence intensities of N-cadherin were decreased.Conclusion:CHE can inhibit the proliferation,migration,invasion,and EMT of the human ovarian cancer SKOV3 cells.

Objective:To discuss the differential effects of apolipoprotein E(APOE)gene polymorphism in the neurotoxicity-reactive astrocytes,and to provide the theoretical basis for the study of the pathogenesis of Alzheimer's disease(AD).Methods:The primary cortical astrocytes from the APOE-knockout mice(APOE-/-)were isolated and cultured in vitro,and the purity of the cells was identified by immunofluorescence staining.The human APOE3 and APOE4 recombinant over-expression plasmids were constructed and separately transfected into the primary APOE-/-astrocytes,and the APOE-/-primary cells were regarded as control.Western blotting method was used to detect the expression levels of APOE and glial fibrillary acidic protein(GFAP)proteins in the cells;enzyme-linked immunosorbent assay(ELISA)method was used to detect the APOE level in the cellular culture supernatant.The inflammatory models were prepared with the primary astrocytes transfected with APOE3 and APOE4 and co-stimulated with interleukin-1α(IL-1α),tumor necrosis factor(TNF),and complement C1q.The cells were divided into APOE3+PBS group,APOE4+PBS group,APOE3+IL-1α+TNF+ C1q group,and APOE4+IL-1α+TNF+C1q group.Cell immunofluorescence staining method was used to observe the morphology of the cells in various groups;real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression levels of glypican 4(Gpc4),glypican 6(Gpc6),thrombospondin 1(Thbs1),thrombospondin 2(Thbs2),SPARC-like protein 1(Sparcl1)and glial cell line derived neurotrophic factor(GDNF),C3,and S100 calcium binding protein B(S100B)mRNA in the cells in various groups;microsphere phagocytosis assay was used to detect the phagocytic capacities of the cells in various groups;Western blotting was used to detect the protein expression levels of B-cell lymphoma 2(Bcl-2),and cysteinyl aspartate specific protease-3(Caspase-3)proteins in the cells in various groups.Results:Compared with APOE-/-group,the expression levels of APOE and GFAP proteins in the cells and the APOE level in the cellular culture supernatant in transfected APOE3 and transfected APOE4 groups were increased(P<0.01).The fluorescence microscope observation results showed that compared with APOE3+PBS and APOE4+PBS groups,the astrocytic processes in APOE3+IL-1α +TNF+Cq1 group and APOE4+IL-1α+TNF+Cq1 group became shorter and the cell bodies became larger;compared with APOE3+IL-1α +TNF+Cq1 group,the astrocytic processes in APOE4+IL-1α +TNF+Cq1 group were even shorter.Compared with APOE3+PBS and APOE4+PBS groups,the expression levels of Gpc4,Gpc6,Thbs1,Thbs2,and Sparcl1 mRNA in the cells in APOE3+IL-1α +TNF+Cq1 group and APOE4+IL-1α +TNF+Cq1 group were significantly decreased(P<0.01);compared with APOE3+IL-1α +TNF+Cq1 group,the expression levels of Gpc4,Gpc6,Thbs1,Thbs2,and Sparcl1 mRNA in the cells in APOE4+IL-1α +TNF+Cq1 group were significantly decreased(P<0.05 or P<0.01).Compared with APOE3+PBS and APOE4+PBS groups,the expression levels of GDNF mRNA in the cells in APOE3+IL-1α+TNF+Cq1 group and APOE4+ IL-1α +TNF+Cq1 group were decreased(P<0.01),and the expression levels of C3 and S100B mRNA were increased(P<0.01);compared with APOE3+IL-1α +TNF+Cq1 group,the expression level of GDNF mRNA in the cells in APOE4+IL-1α+TNF+Cq1 group was decreased(P<0.05),and the expression levels of C3 and S100B mRNA were increased(P<0.05).Compared with APOE3+ PBS group and APOE4+PBS group,the numbers of hagocytosis of microspheres in the cells in APOE3+ IL-1α +TNF+Cq1 group and APOE4+IL-1α +TNF+Cq1 group were significantly decreased;compared with APOE3+IL-1α+TNF+Cq1 group,the number of hagocytosis of microspheres in the cells in APOE4+IL-1α+TNF+Cq1 group was significantly decreased.Compared with APOE3+PBS group and APOE4+PBS group,the expression levels of Bcl-2 protein in the cells in APOE3+IL-1α+TNF+ Cq1 group and APOE4+IL-1α +TNF+Cq1 group were decreased(P<0.05 or P<0.01)and the expression levels of Caspase-3 protein were significantly increased(P<0.01);compared with APOE3+ IL-1α+TNF+Cq1 group,the expression level of Bcl-2 protein in the cells in APOE4+IL-1α+TNF+ Cq1 group was decreased(P<0.01),and the expression level of Caspase-3 protein was increased(P<0.05).Conclusion:The APOE4 genotype has a stronger ability to induce the inflammatory factors compared with APOE3;it can lead to a neurotoxicity-reactive astrocyte phenotype,increase the neurotoxicity,affect the astrocyte apoptosis,and aggravate the neuron damage.

Objective:To discuss the effect of ligustilide on the cardiac function and angiogenesis in the rats with heart failure,and to clarify its regulatory effect on protein kinase D1(PKD1)/hypoxia-inducible factor-1α(HIF-1α)/vascular endothelial growth factor(VEGF)pathway.Methods:The SD rats were randomly divided into sham operation group,model group,ligustilide group,PKD1/HIF-1α/VEGF signaling pathway inhibitor CID755673(CID)group,and ligustilide+CID group.The heart failure rat model was established by ligation of the left anterior descending coronary artery.The rats in ligustilide group were injected intravenously with 20 mg·kg-1 ligustilide,the rats in CID group were injected intraperitoneally with 50 mg·kg-1 CID,and the rats in ligustilide+CID group were injected intraperitoneally with 50 mg·kg-1 CID followed by intravenous injection of 20 mg·kg-1 ligustilide,once per day for 4 consecutive weeks.The cardiac function indexes of the rats in various groups were detected by echocardiography;the percentages of myocardial infarction areas of the rats in various groups were detected by 2,3,5-triphenyltetrazolium chloride(TTC)staining;the pathomorphology of myocardium tissue of the rats in various groups was observed by HE staining;the expression levels of PKD1,HIF-1α,CD31,and VEGF mRNA and proteins in ischemic area of myocardium tissue of the rats in various groups were detected by real-time fluorescence quantitative PCR(RT-qPCR)and Western blotting methods.Results:Compared with sham operation group,the rats in model group and CID group had altered myocardial cell morphology,increased intercellular gaps,disorganized arrangement,visible muscle fiber breaks and inflammatory cell infiltration;the rats in ligustilide group and ligustilide+CID group had relatively orderly myocardial fiber arrangement,fewer myocardial fiber breaks and decreased number of inflammatory cells.Compared with sham operation group,the left ventricular ejection fraction(LVEF)and left ventricular fractional shortening(LVFS)of the rats in model group were decreased(P<0.05),the left ventricular end-systolic diameter(LVESD)and left ventricular end-diastolic diameter(LVEDD)were increased(P<0.05),and the expression levels of PKD1,HIF-1α,CD31,and VEGF mRNA and proteins in myocardium tissue were decreased(P<0.05).Compared with model group,the LVEF and LVFS of the rats in ligustilide group were increased(P<0.05),the LVESD and LVEDD were decreased(P<0.05),the percentage of myocardium infarction area was decreased(P<0.05),and the expression levels of PKD1,HIF-1α,CD31,and VEGF mRNA and proteins in myocardium tissue were increased(P<0.05);compared with model group,the LVEF and LVFS of the rats in CID group were decreased(P<0.05),the LVESD and LVEDD were increased(P<0.05),the percentage of myocardium infarction area was increased(P<0.05),and the expression levels of PKD1,HIF-1α,CD31,and VEGF mRNA and proteins in myocardium tissue were decreased(P<0.05);compared with ligustilide group,the LVEF and LVFS of the rats in ligustilide+CID group were decreased(P<0.05),the LVESD and LVEDD were increased(P<0.05),the percentage of myocardium infarction area was increased(P<0.05),and the expression levels of PKD1,HIF-1α,CD31,and VEGF mRNA and proteins in myocardium tissue were decreased(P<0.05);compared with CID group,the LVEF and LVFS of the rats in ligustilide+CID group were increased(P<0.05),the LVESD and LVEDD were decreased(P<0.05),the percentage of myocardium infarction area was decreased(P<0.05),and the expression levels of PKD1,HIF-1α,CD31,and VEGF mRNA and proteins in myocardium tissue were increased(P<0.05).Conclusion:Ligustilide can promote the angiogenesis,reduce the myocardium infarction area,and improve the cardiac function in the rats with heart failure;it works through activation of the PKD1/HIF-1α/VEGF pathway.

Objective:To discuss the regulatory effect of berberine(BBR)on fatty acids in the human glioma T98G cells and its effect on the cell proliferation,migration,and invasion,and to clarify its potential mechanism.Methods:The T98G cells at logarithmic growth phase were divided into control group and different concentrations(25,50,and 100 mg·L-1)of BBR groups.Cell wound healing assay was used to detect the migration rates of the cells in various groups;Transwell chamber assay was used to detect the invasion rates of the cells in various groups.The T98G cells at logarithmic growth phase were divided into control group and 100 mg·L-1 BBR group,and Mass spectrometry was used to detect the fatty acid contents in the cells in two groups.The T98G cells at logarithmic growth phase were divided into control group and different concentrations(50,100,and 150 mg·L-1)of BBR groups;Western blotting method was used to detect the expression levels of phosphatidylinositol 3-kinase(PI3K),phosphorylated PI3K(p-PI3K),protein kinase B(AKT),phosphorylated AKT(p-AKT),sterol regulatory element-binding protein 1(SREBP-1),and fatty acid synthase(FASN)in the cells in various groups.The expression of FASN was suppressed by gene silencing technology,and the T98G cells at logarithmic growth phase were divided into control group,shFASN1 group,and shFASN2 group.Western blotting method was used to detect the expression levels of FASN protein in the cells in various groups;clone formation assay was used to detect the clone formation of the cells in various groups;cell wound healing assay was used to detect the migration rates of the cells in various groups.Results:Compared with control group,the migration rates and invasion rates of the cells in different concentrations of BBR groups were decreased in a concentration-dependent manner(P<0.01),and the fatty acid content in the cells in 100 mg·L-1 BBR group was significantly decreased(P<0.01).Compared with control group,the expression levels of p-PI3K,p-AKT,SREBP-1,and FASN proteins in the cells in 150 mg·L-1 BBR group were significantly decreased(P<0.05 or P<0.01),and the expression level of SREBP-1 protein in the cells in 100 and 150 mg·L-1 BBR groups were significantly decreased(P<0.01).After suppression of FASN expression,compared with control group,the expression levels of FASN protein in the cells in shFASN1 and shFASN2 groups were significantly decreased(P<0.01),and the expression level of FASN protein in the cells in shFASN2 group was lower than that in shFASN1 group(P<0.05);compared with control group,the numbers of clone formation and migration rates of the cells in shFASN1 and shFASN2 groups were significantly decreased(P<0.01),and the migration rate of the cells in shFASN2 group was significantly lower than that in shFASN1 group(P<0.05).Conclusion:BBR interferes with fatty acid synthesis in the glioma T98G cells by reducing the expression of the PI3K/AKT/SREBP-1/FASN pathway related proteins,and decrease their migration and invasion capabilities.

Objective:To investigate the efficacy of Balanophora involucrata Hook.f.in treatment of hyperuricemia(HUA)based on network pharmacology,molecular docking,and hyperuricemia models in vivo and in vitro,and to clarify the main targets of its active components and related signaling pathway mechanism.Methods:The potential targets of Balanophora involucrata Hook.f.in treatment of HUA were identified by Databases such as the Traditional Chinese Medicine Database in Taiwan,the Chinese Herbal Medicine Identification Database,Professional Chemical Database,TargetNet Database,SwissTargetPrediction Database,GeneCards,Therapeutic Target Database(TTD),DrugBank Database,DisGeNET Database,Online Mendelian Inheritance in Man(OMIM)Database,and Venny Database.STRING Database and Cytoscape software were used to construct the active component-predictive target network and protein-protein interaction(PPI)network for Balanophora involucrata Hook.f.;topological analysis was used to select the main active components and core targets;Gene Ontology(GO)functional and Kyoto Encyclopedia of Genes and Genomes(KEGG)signaling pathway enrichment analysis were performed by R software;AutoDock Vina software was used for molecular docking validation.The NRK-52E cells were divided into blank control group,blank administration group,model group,and different concentrations(2.0,10.0,and 50.0 μmol·L-1)of erythrodiol(EDT)groups.High-performance liquid chromatography culture(HPLC)was used to detect the uric acid(UA)levels in the cell culture supernatants in various groups.The male ICR mice were divided into blank control group,blank administration group,model group,and EDT group;the mice in the last two groups were used to prepare the HUA models;kits were used to detect the levels of UA,creatinine(Cr),and blood urea nitrogen(BUN)in serum of the mice in various groups;the bilateral kidney tissue of the mice was harvested and weighed;the kidney indexes of the mice in various groups were calculated;TUNEL staining was used to observe the apoptosis in kidney tissue of the mice in various groups;Western blotting method was used to detect the expression levels of protein kinase B(AKT),phosphorylated AKT(p-AKT),phosphoinositide 3-kinase(PI3K),phosphorylated PI3K(p-PI3K),B-cell lymphoma-2(Bcl-2),Bcl-2-associated X protein(Bax),and matrix metalloproteinase-9(MMP-9)proteins in kidney tissue of the mice in various groups.Results:Six active components of Balanophora involucrata Hook.f.were identified,involving 116 intersecting targets and 14 core targets.The enrichment analysis yielded 1 828 GO terms and 145 signaling pathways.The molecular docking results showed that EDT had good binding activity with MMP-9.The high uric acid cell experiment results showed that compared with blank control group,the UA level in the cells in model group was significantly increased(P<0.01);compared with model group,the UA levels in the cells in 2.0,10.0,and 50.0 μmol·L-1 EDT groups were significantly decreased(P<0.01).Compared with blank control group,the levels of UA,Cr,and BUN in serum of the mice in model group were increased(P<0.01),and the kidney indexes were significantly increased(P<0.01);compared with model group,the levels of UA,Cr,and BUN in serum of the mice in EDT group were decreased(P<0.05 or P<0.01),and the kidney index was significantly decreased(P<0.05 or P<0.01).Compared with blank control group,the number of apoptotic cells in kidney tissue of the mice in model group was increased;compared with model group,the number of the apoptotic cells in kidney tissue of the mice in EDT group was significantly decreased.Compared with blank control group,the ratios of p-AKT/AKT and p-PI3K/PI3K and expression level of Bcl-2 protein in kidney tissue of the mice in model group were significantly decreased(P<0.05 or P<0.01),while the expression levels of Bax and MMP-9 proteins were significantly increased(P<0.01);compared with model group,the ratios of p-AKT/AKT and p-PI3K/PI3K and expression level of Bcl-2 protein in kidney tissue of the mice in EDT group were significantly increased(P<0.05 or P<0.01),and the expression levels of Bax and MMP-9 proteins were significantly decreased(P<0.01).Conclusion:The active component of Balanophora involucrata Hook.f.,EDT,has a UA-decreasing effect and may inhibit the apoptosis and alleviate the kidney injury by activating the PI3K/AKT signaling pathway.

Objective:To discuss the repairment effect of intra-articular injection of adipose derived stem cells(ADSCs)on articular cartilage destruction in the temporomandibular joint osteoarthritis(TMJOA)model rabbits,and to clarify the possible mechanism.Methods:Twenty-seven rabbits were randomly divided into control group,model group,and ADSCs group.The ADSCs of the rabbits were extracted and cultured.The rabbit TMJOA model was prepared by monosodium-iodoacetate(MIA)injection technique.The temporomandibular joint cavity of the TMJOA model rabbits in ADSCs group was given two continuous intra-articular injections of 1.0×106 mL-1 ADSCs,while the rabbits in control and model group were given sequivalent volume of saline into the temporomandibular joint cavity.After 8 weeks,Micro-CT scan was performed on the temporomandibular joints of the rabbits in various groups;the bone volume fraction(BV/TV),bone surface area/bone volume(BS/BV),trabecular thickness(Tb.Th),trabecular separation(Tb.Sp),and trabecular number(Tb.N)of condyles tissue of the rabbits in various groups were analyzed;HE staining was used to observe the pathomorphology of condyles tissue of the rabbits in various groups;immunohistochemistry was used to detect the localization and expression levels of SRY-related high mobility group box gene 9(SOX9),matrix metalloproteinase-13(MMP-13),and vascular endothelial growth factor(VEGF)proteins in condyles tissue of the rabbits in various groups;Western blotting method was used to detect the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in various groups.Results:The micro-CT scan results showed that compared with control group,the BV/TV,Tb.Th,and Tb.N of condyles tissue of the rabbits in model group were significantly decreased(P<0.05),while the BS/BV and Tb.Sp were significantly increased(P<0.05);compared with model group,the BV/TV,Tb.Th,and Tb.N in condyles tissue of the rabbits in ADSCs group were significantly increased(P<0.05),and the BS/BV and Tb.Sp were significantly decreased(P<0.05).The HE staining results showed that the condylar cartilage surface of the rabbits in control group was smooth with clear layers and intact structure;compared with control group,the surface of condyles tissue of the rabbits in model group was irregular with thickened hypertrophic layer and areas of cell depletion and clustering;compared with model group,the pathological damage of condyles tissue of the rabbits in ADSCs group was significantly decreased.The immunohistochemical staining results showed that compared with control group and ADSCs group,the number of brown granule in condyles tissue of the rabbits in model group was increased,mainly concentrated in the hypertrophic layer,especially in the bone cartilage junction site and the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in model group were significantly increased(P<0.05);compared with model group,the number of brown granule in condyles tissue of the rabbits in ADSCs group was significantly decreased,and the expression levels of SOX9,MMP-13,and VEGF proteins were significantly decreased(P<0.05).The Western blotting results showed that compared with control group,the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in model group were significantly increased(P<0.05);compared with model group,the expression levels of SOX9,MMP-13,and VEGF proteins in condyles tissue of the rabbits in ADSCs group were significantly decreased(P<0.05).Conclusion:Intra-articular injection of ADSCs can effectively repair the cartilage destruction in TMJOA,alleviate the cartilage injury,and mitigate the progression of osteoarthritis.

Objective:To discuss the effect of human umbilical cord mesenchymal stem cells culture supernatant(hUCMSCs-Sup)on the proliferation,apoptosis,and endometrium receptivity of the human endometrial stromal cells(hEndoSCs)treated with mifepristone(Ms),and to clarify the possible mechanism.Methods:The hEndoSCs were cultured in vitro and divided into control group and 40,60,80,and 100 μmol·L-1 Ms groups.The survival rates of the cells in various groups were detected by MTT assay.The hEndoSCs were divided into control group,40 μmol·L-1 Ms group,and 60 μmol·L-1 Ms group.The apoptotic rates of the cells in various groups were detected by flow cytometry;the expression levels of apoptosis-related protein B-cell lymphoma-2(Bcl-2)and Bcl-2-associated X protein(Bax)proteins in the cells in various groups were detected by Western blotting method,and the ratio of Bcl-2/Bax was calculated.After treated with hUCMSCs-Sup,the hEndoSCs were divided into control group,Ms group,Ms+hUCMSCs-Sup group,and Ms+hUCMSCs-Sup+3-methyladenine(3-MA)group.The survival rates of the cells in various groups were detected by MTT assay;the apoptotic rates of the cells in various groups were detected by flow cytometry;the expression levels of microtubule-associated protein 1 light chain 3B-Ⅱ(LC3B-Ⅱ)and microtubule-associated protein 1 light chain 3B-I(LC3B-Ⅰ)proteins in the cells in various groups were detected by Western blotting method,and the ratio of LC3B-Ⅱ/LC3B-Ⅰwas calculated;the expression levels of endometrium receptivity marker molecules mRNA in the cells in various groups were detected by real-time fluorescence quantitative PCR(RT-qPCR)method.Results:Compared with control group,the survival rates of the cells in 40,60,80,and 100 μmol·L-1 Ms groups were significantly decreased(P<0.05)in a time-dependent and dose-dependent manner.Compared with control group,the apoptotic rates of the cells in 40 and 60 μmol·L-1 Ms groups were significantly increased(P<0.05),and the ratios of Bcl-2/Bax were significantly decreased(P<0.05).After treated with hUCMSCs-Sup,compared with control group,the survival rate of the cells and ratio of LC3B-Ⅱ/LC3B-Ⅰ in the cells in Ms group were significantly decreased(P<0.05),the apoptotic rate was significantly increased(P<0.05),and the expression levels of homeobox A10(HOXA10),leukemia inhibitory factor(LIF),and integrin subunit beta 3(ITGB3)mRNA in the cells were significantly decreased(P<0.05);compared with Ms group,the survival rate of the cells and ratio of LC3B-Ⅱ/LC3B-Ⅰin the cells in Ms+hUCMSCs-Sup group were significantly increased(P<0.05),the apoptotic rate was significantly decreased(P<0.05),and the expression levels of HOXA10,LIF,and ITGB3 mRNA in the cells were significantly increased(P<0.05);compared with Ms+hUCMSCs-Sup group,the survival rate of the cells and ratio of LC3B-Ⅱ/LC3B-Ⅰ in the cells in Ms+hUCMSCs-Sup+3-MA group were significantly decreased(P<0.05).Conclusion:hUCMSCs-Sup can increase the survival rate and decrease the apoptotic rate of the hEndoSCs after treated with Ms,and increase the endometrium receptivity,and its mechanism may be associated with the activation of autophagy of the hEndoSCs by hUCMSCs-Sup.