Objective To investigate the anti-HIV effects of ampelopsin and its interaction with HIV-1 coreceptor CXCR4. Methods Through anti-virus experiments in vitro, the inhibitory effect of ampelopsin on HIV-1 infection was ...Objective To investigate the anti-HIV effects of ampelopsin and its interaction with HIV-1 coreceptor CXCR4. Methods Through anti-virus experiments in vitro, the inhibitory effect of ampelopsin on HIV-1 infection was verified. Chemotaxis assay was performed to show the ability to induce PBMCs migration by ampelopsin, RANTES and SDF-1(. Fluorescence labelling monoclonal antibody was utilized to observe the interaction of ampelopsin and CXCR4. Mice immunosuppressant model was also established to detail the role ampelopsin played in regulating cellular immunological functions. Results Ampelopsin could protect sensitive cells against HIV-1 infection and dramatically reduce HIV-1 antigen P24 expression. HIV-1SF33 attaching to MT-4 cells was interfered by ampelopsin, and the EC50 was 0.175 mg/mL for cellular protection and 0.024 mg/mL for P24 inhibition. At co-cultivating phase, EC50 was 0.229 mg/mL and 0.197 mg/mL respectively. Furthermore, the EC50 was 0.179 mg/mL and 0.348 mg/mL in acute infection. Human PBMCs migration was induced after being challenged with ampelopsin or chemokines, and synergistic action was observed during co-treatment. Ampelopsin alone resulted in maximal chemotaxis at 1 mg/mL. HIV-1 co-receptor CXCR4 on the surface of PBMCs was decreased by internalization, which indicated the effect of ampelopsin on CXCR4. About 70% CXCR4 was reduced by ampelopsin at 1 mg/mL. Ampelopsin also augmented cellular immunological functions in immunosuppressive mice. Conclusion Ampelopsin displays a strong inhibitive role during HIV-1 absorption, incubation and acute infection. These results are coincident with its immune enhancement.展开更多
This study investigated the protective effects of ampelopsin on H2O2-induced oxidative injury in porcine kidney epithelial cell line, PK-15; cells were pretreated with medium containing 0, 15, 30, and 60 μg mL-1 ampe...This study investigated the protective effects of ampelopsin on H2O2-induced oxidative injury in porcine kidney epithelial cell line, PK-15; cells were pretreated with medium containing 0, 15, 30, and 60 μg mL-1 ampelopsin, respectively, for 1 h prior to the addition of 100 μmol L-1 H2O2. After 2 h, the cell viability, intracellular superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity, cellular malondialdehyde (MDA) content, and DNA strand breakage were determined. Results showed that ampelopsin pretreatment did not modify the SOD and GSH-Px inactivation but significantly decreased MDA production; except for the medium dose (30 μg mL-1), all the tested concentrations of ampelopsin did not significantly inhibit cell viability reduction and gave no influence on DNA damage. The results suggested that all the tested doses of ampelopsin pretreatment before H2O2 addition can protect PK-15 cell from H2O2-induced lipid peroxidation, which might not be mediated by the SOD and GSH-Px responsees. The appropriate dose of ampelopsin pretreatment prevents PK-15 cell from DNA damage and cell viability reduction.展开更多
Objective: To evaluate the cytotoxic effects of ampelopsin sodium(Amp-Na) and carboplatin(CBP) used alone or in combination on human non-small cell lung cancer(NSCLC) cells SPC-A1 in vitro and its related mechanism. M...Objective: To evaluate the cytotoxic effects of ampelopsin sodium(Amp-Na) and carboplatin(CBP) used alone or in combination on human non-small cell lung cancer(NSCLC) cells SPC-A1 in vitro and its related mechanism. Methods: Cytotoxic effects were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assays. The synergistic effects of the drugs were calculated with coefficient of drug interaction(CDI). Cell cycle was determined by flow cytometry(FCM). The levels of p53, p21, cyclin E, cyclin D1, and phosphorylated cyclin-dependent kinase-2(p-CDK2) were evaluated by Western blot. Results: Amp-Na(6.25–200 μg/m L) and CBP(3.13–100 μg/m L) alone exhibited prominent cytotoxic activity in a concentration-dependent manner on SPC-A1 cells with 50% inhibitive concentration values of 57.07±14.46 and 34.97±6.30 μg/m L, respectively. Drug combinations were associated with significantly higher cytotoxic effects than each drug alone(P<0.05 or 0.01). The CDI analysis confirmed the synergy of Amp-Na and CBP on inhibiting cancer cell viability across a wide concentration range(CDI <1). FCM and Western blot showed that synergistic cytotoxic effects of Amp-Na and CBP were related to G1 arrested which mainly mediated by p21 through the inhibition of CDK2 activity independent of the p53 tumor suppressor pathway. Conclusions: AmpNa exhibits anticancer activities and enhances the antitumor activities of CBP through up-regulation of p21 and inhibition of CDK2 activity in human NSCLC cells SPC-A1. These results suggest that Amp-Na may be applied to enhance the anticancer action of CBP.展开更多
目的研究蛇葡萄素(ampelopsin,AMP)诱导人宫颈癌细胞SiHa发生自噬及其可能的作用机制。方法单丹磺酰胺(monodansylcadaverine,MDC)染色法观察AMP对SiHa和C-33A细胞自噬的影响;透射电子显微镜(transmission electron microscope,TEM)观察...目的研究蛇葡萄素(ampelopsin,AMP)诱导人宫颈癌细胞SiHa发生自噬及其可能的作用机制。方法单丹磺酰胺(monodansylcadaverine,MDC)染色法观察AMP对SiHa和C-33A细胞自噬的影响;透射电子显微镜(transmission electron microscope,TEM)观察经AMP干预的SiHa和C-33A细胞超微结构的变化;自噬双标腺病毒转染(mRFP-GFP-LC3)法检测SiHa细胞自噬流强度;Western blot法对SiHa细胞中自噬相关蛋白LC3Ⅱ/Ⅰ、P62、Beclin-1、Atg13及PI3K/AKT/mTOR信号通路蛋白的表达情况进行检测。结果MDC染色结果显示,随着AMP浓度的升高,代表自噬小体的亮绿色逐渐增多;TEM实验结果表明,与空白对照组相比,AMP组细胞线粒体嵴溶解消失,同时可见自噬小体和自噬溶酶体;mRFP-GFP-LC3实验结果提示,随着AMP浓度的升高,细胞内自噬小体和自噬溶酶体增多,提示自噬流增强;同时发现LC3Ⅱ/Ⅰ、Beclin-1、Atg13蛋白表达上调,P62、p-PI3K、p-AKT、p-mTOR蛋白表达下调,表明AMP可能通过抑制PI3K/AKT/mTOR信号通路诱导SiHa细胞发生自噬;加入3-MA后,与AMP组相比,3-MA和AMP联合组的p-PI3K、p-AKT、p-mTOR的表达水平上升,提示自噬被抑制后,PI3K/AKT/mTOR信号通路被逆转。结论AMP可诱导SiHa细胞发生自噬,其机制可能与抑制PI3K/AKT/mTOR信号通路的激活有关。展开更多
基金This work was supported by Science Foundation of Guangdong Province (No.97002).
文摘Objective To investigate the anti-HIV effects of ampelopsin and its interaction with HIV-1 coreceptor CXCR4. Methods Through anti-virus experiments in vitro, the inhibitory effect of ampelopsin on HIV-1 infection was verified. Chemotaxis assay was performed to show the ability to induce PBMCs migration by ampelopsin, RANTES and SDF-1(. Fluorescence labelling monoclonal antibody was utilized to observe the interaction of ampelopsin and CXCR4. Mice immunosuppressant model was also established to detail the role ampelopsin played in regulating cellular immunological functions. Results Ampelopsin could protect sensitive cells against HIV-1 infection and dramatically reduce HIV-1 antigen P24 expression. HIV-1SF33 attaching to MT-4 cells was interfered by ampelopsin, and the EC50 was 0.175 mg/mL for cellular protection and 0.024 mg/mL for P24 inhibition. At co-cultivating phase, EC50 was 0.229 mg/mL and 0.197 mg/mL respectively. Furthermore, the EC50 was 0.179 mg/mL and 0.348 mg/mL in acute infection. Human PBMCs migration was induced after being challenged with ampelopsin or chemokines, and synergistic action was observed during co-treatment. Ampelopsin alone resulted in maximal chemotaxis at 1 mg/mL. HIV-1 co-receptor CXCR4 on the surface of PBMCs was decreased by internalization, which indicated the effect of ampelopsin on CXCR4. About 70% CXCR4 was reduced by ampelopsin at 1 mg/mL. Ampelopsin also augmented cellular immunological functions in immunosuppressive mice. Conclusion Ampelopsin displays a strong inhibitive role during HIV-1 absorption, incubation and acute infection. These results are coincident with its immune enhancement.
基金supported by the National Basic Re-search Program of China (2004CB117507)supportprovided by the Key Technologies R&D Program of China during the 11th Five-Year Plan period(2006BAD14B02-8)
文摘This study investigated the protective effects of ampelopsin on H2O2-induced oxidative injury in porcine kidney epithelial cell line, PK-15; cells were pretreated with medium containing 0, 15, 30, and 60 μg mL-1 ampelopsin, respectively, for 1 h prior to the addition of 100 μmol L-1 H2O2. After 2 h, the cell viability, intracellular superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity, cellular malondialdehyde (MDA) content, and DNA strand breakage were determined. Results showed that ampelopsin pretreatment did not modify the SOD and GSH-Px inactivation but significantly decreased MDA production; except for the medium dose (30 μg mL-1), all the tested concentrations of ampelopsin did not significantly inhibit cell viability reduction and gave no influence on DNA damage. The results suggested that all the tested doses of ampelopsin pretreatment before H2O2 addition can protect PK-15 cell from H2O2-induced lipid peroxidation, which might not be mediated by the SOD and GSH-Px responsees. The appropriate dose of ampelopsin pretreatment prevents PK-15 cell from DNA damage and cell viability reduction.
基金Supported by the Natural Science Foundation of Gansu Province,China(No.0710RJZA044)
文摘Objective: To evaluate the cytotoxic effects of ampelopsin sodium(Amp-Na) and carboplatin(CBP) used alone or in combination on human non-small cell lung cancer(NSCLC) cells SPC-A1 in vitro and its related mechanism. Methods: Cytotoxic effects were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assays. The synergistic effects of the drugs were calculated with coefficient of drug interaction(CDI). Cell cycle was determined by flow cytometry(FCM). The levels of p53, p21, cyclin E, cyclin D1, and phosphorylated cyclin-dependent kinase-2(p-CDK2) were evaluated by Western blot. Results: Amp-Na(6.25–200 μg/m L) and CBP(3.13–100 μg/m L) alone exhibited prominent cytotoxic activity in a concentration-dependent manner on SPC-A1 cells with 50% inhibitive concentration values of 57.07±14.46 and 34.97±6.30 μg/m L, respectively. Drug combinations were associated with significantly higher cytotoxic effects than each drug alone(P<0.05 or 0.01). The CDI analysis confirmed the synergy of Amp-Na and CBP on inhibiting cancer cell viability across a wide concentration range(CDI <1). FCM and Western blot showed that synergistic cytotoxic effects of Amp-Na and CBP were related to G1 arrested which mainly mediated by p21 through the inhibition of CDK2 activity independent of the p53 tumor suppressor pathway. Conclusions: AmpNa exhibits anticancer activities and enhances the antitumor activities of CBP through up-regulation of p21 and inhibition of CDK2 activity in human NSCLC cells SPC-A1. These results suggest that Amp-Na may be applied to enhance the anticancer action of CBP.
文摘目的研究蛇葡萄素(ampelopsin,AMP)诱导人宫颈癌细胞SiHa发生自噬及其可能的作用机制。方法单丹磺酰胺(monodansylcadaverine,MDC)染色法观察AMP对SiHa和C-33A细胞自噬的影响;透射电子显微镜(transmission electron microscope,TEM)观察经AMP干预的SiHa和C-33A细胞超微结构的变化;自噬双标腺病毒转染(mRFP-GFP-LC3)法检测SiHa细胞自噬流强度;Western blot法对SiHa细胞中自噬相关蛋白LC3Ⅱ/Ⅰ、P62、Beclin-1、Atg13及PI3K/AKT/mTOR信号通路蛋白的表达情况进行检测。结果MDC染色结果显示,随着AMP浓度的升高,代表自噬小体的亮绿色逐渐增多;TEM实验结果表明,与空白对照组相比,AMP组细胞线粒体嵴溶解消失,同时可见自噬小体和自噬溶酶体;mRFP-GFP-LC3实验结果提示,随着AMP浓度的升高,细胞内自噬小体和自噬溶酶体增多,提示自噬流增强;同时发现LC3Ⅱ/Ⅰ、Beclin-1、Atg13蛋白表达上调,P62、p-PI3K、p-AKT、p-mTOR蛋白表达下调,表明AMP可能通过抑制PI3K/AKT/mTOR信号通路诱导SiHa细胞发生自噬;加入3-MA后,与AMP组相比,3-MA和AMP联合组的p-PI3K、p-AKT、p-mTOR的表达水平上升,提示自噬被抑制后,PI3K/AKT/mTOR信号通路被逆转。结论AMP可诱导SiHa细胞发生自噬,其机制可能与抑制PI3K/AKT/mTOR信号通路的激活有关。
