The stimulator of interferon genes(STING),an integral adaptor protein in the DNA-sensing pathway,plays a pivotal role in the innate immune response against infections.Additionally,it presents a valuable therapeutic target for infectious diseases and cancer.We observed that fangchinoline(Fan),a bis-benzylisoquinoline alkaloid(BBA),effectively impedes the replication of vesicular stomatitis virus(VSV),encephalomyocarditis virus(EMCV),influenza A virus(H1 N1),and herpes simplex virus-1(HSV-1)in vitro.Fan treatment significantly reduced the viral load,attenuated tissue inflammation,and improved survival in a viral sepsis mouse model.Mechanistically,Fan activates the antiviral response in a STING-dependent manner,leading to increased expression of interferon(1FN)and interferon-stimulated genes(ISGs)for potent antiviral effects in vivo and in vitro.Notably,Fan interacts with STING,preventing its degradation and thereby extending the activation of IFN-based antiviral responses.Collectively,our findings highlight the potential of Fan,which elicits antiviral immunity by suppressing STING degra-dation,as a promising candidate for antiviral therapy.

目的 探索防己诺林碱在细胞水平抗H1N1病毒的作用,阐明防己诺林碱调控细胞自噬抑制H1N1病毒复制的分子机制.方法 CCK-8法检测 0.312 5、0.625 0、1.250 0、2.500 0、5.000 0、10.000 0、20.000 0、40.000 0、60.000 0 μmol·L-1的防己诺林碱对MDCK细胞活力的影响;MDCK细胞设置对照组、模型组和防己诺林碱(2.5、5.0、10.0 μmol·L-1)组,除对照组外,以感染复数(MOI)为0.1的H1N1病毒感染MDCK细胞,加入防己诺林碱共同孵育12 h,同时设置防己诺林碱(10.0 μmol·L-1)与病毒共同孵育4、8 h组,通过实时荧光定量PCR(qRT-PCR,检测HIN1 mRNA表达)和Western blotting[H1N1病毒核蛋白(NP)]检测防己诺林碱对病毒复制的抑制作用;PI/Hoechst33342染色检测防己诺林碱(10.0 μmol·L-1)对MDCK细胞死亡的影响;qRT-PCR法检测防己诺林碱检测防己诺林碱预处理、病毒感染早期药物处理、病毒感染晚期药物处理以及吸附和进入阶段给药对病毒复制的影响;MDCK细胞转染EGFP-LC3或EGFP-mCherry-LC3双荧光质粒,观察防己诺林碱对EGFP-LC3的荧光强度、EGFP-mCherry-LC3共定位的影响,设置自噬诱导剂雷帕霉素(0.5 μmol·L-1)、自噬晚期抑制剂氯喹(10 μmol·L-1)、巴佛洛霉素A1(100 nmol·L-1)对照.转染过EGFP-LC3质粒的MDCK细胞感染H1N1病毒,免疫荧光检测防己诺林碱对LC3与胞内的病毒NP蛋白共定位的影响;体外培养A549细胞,以MOI为0.1的H1N1病毒感染,Western blotting检测防己诺林碱对LC3Ⅱ/LC3Ⅰ蛋白表达的影响.结果 防己诺林碱对MDCK细胞的半数抑制浓度(IC50)为40.19 μmol·L-1;与模型组比较,防己诺林碱以浓度相关性的方式抑制HIN1 mRNA表达,以浓度和时间相关性的方式抑制NP蛋白表达(P＜0.01、0.001);显著减少H1N1诱导的细胞死亡(P＜0.001);抑制H1N1病毒进入过程.与对照组比较,防己诺林碱处理诱导LC3荧光聚集,诱导EGFP-LC3和mCherry-LC3双荧光共定位显著增加(P＜0.001).与模型组比较,防己诺林碱处理明显减少NP的荧光数量(P＜0.001),同时造成LC3荧光积累并与胞内的病毒NP蛋白共定位;引起LC3Ⅱ积累(P＜0.01、0.001).结论 防己诺林碱同氯喹和巴佛洛霉素A1一致,阻断自噬途径,使病毒粒子在自噬体内滞留,破坏病毒生命周期.

目的 比较防己黄芪汤中活性成分在大鼠体内的药动学行为,并研究防己黄芪汤及其主要入血成分粉防己碱抑制刀豆球蛋白A(concanavalin A,ConA)诱导T淋巴细胞增殖的作用机制.方法 采用超高效液相色谱-串联质谱法(UPLC-MS/MS)检测大鼠ig不同剂量防己黄芪汤及粉防己碱后,入血成分在不同时间点的血药浓度,并分析其药动学行为.采用MTT法和3H-胸腺嘧啶核苷(3H-TdR)掺入法检测防己黄芪汤对T淋巴细胞毒性及增殖的抑制作用;采用ELISA法测定上清液中白细胞介素-17(interleukin-17,IL-17)、IL-2、γ-干扰素(γ-interferon,IFN-γ)水平;采用流式细胞仪检测调节性T细胞(regulatory T cell,Treg)数量.结果 防己黄芪汤在大鼠体内主要入血成分为防己诺林碱、木兰花碱、毛蕊异黄酮葡萄糖苷、甘草苷、黄芪甲苷、甘草次酸和粉防己碱.除防己诺林碱外,其他6个成分在体内的吸收具有剂量相关性.粉防己碱、防己诺林碱和甘草次酸在体内吸收速率慢,消除速率慢,驻留时间长[平均滞留时间(MRT)＞10 h、清除率(CL)＜8 L/(h·kg)];木兰花碱、毛蕊异黄酮葡萄糖苷和甘草苷在体内具有吸收和消除速度快、体内驻留时间短的特点[达峰时间(tmax)＜1 h、MRT＜5.2 h、CL＞50U(h·kg)].与单一成分给药相比,防己黄芪汤中的粉防己碱的吸收程度明显增加,而消除速率明显降低.防己黄芪汤可以抑制T淋巴细胞的增殖,还可降低细胞上清液中促炎因子IL-2、IL-17和IFN-γ水平(P＜0.05、0.01),增加Treg的分化数量(P＜0.05、0.01),防己黄芪汤主要入血成分之一粉防己碱可降低细胞上清液中IL-17水平并增加Treg分化数量(P＜0.05、0.01).结论 君药防己和黄芪的特征成分是防己黄芪汤的主要入血成分;君药防己含有的粉防己碱与防己黄芪汤中其他成分存在协同作用.防己黄芪汤可抑制T淋巴细胞的增殖,其作用机制可能是通过调节Treg的增殖分化过程,使激活的免疫反应受到影响,而防己黄芪汤中的粉防己碱可能起到重要作用.

目的 基于线粒体凋亡通路探究防己诺林碱(fangchinoline,FAN)对前列腺癌PC3细胞生物学行为的影响.方法 将前列腺癌PC3 细胞按照FAN处理浓度分为 4 组,分别为FAN 0 μmol/L组、FAN 5 μmol/L组、FAN 10 μmol/L组和FAN 20 μmol/L组,各组依次采用FAN 0 μmol/L(DMSO替代)、5 μmol/L、10 μmol/L和 20 μmol/L进行处理,孵育 48h后进行实验.CCK-8法检测PC3 细胞增殖,平板克隆法检测PC3 细胞克隆形成能力,流式细胞仪检测PC3 细胞凋亡,Transwell实验检测PC3 细胞侵袭,划痕实验检测PC3 细胞迁移,Western blot检测PC3 细胞自噬相关蛋白和线粒体凋亡通路蛋白.结果 与FAN 0 μmol/L组相比,FAN 5 μmol/L组、FAN 10 μmol/L组和FAN 20 μmol/L组的PC3 细胞增殖能力、克隆形成能力、侵袭和迁移能力明显降低(P＜0.001),凋亡率明显升高(P＜0.001);与FAN 5 μmol/L组和FAN 10 μmol/L组相比,FAN 20 μmol/L组的PC3 细胞增殖能力、克隆形成能力、侵袭和迁移能力明显降低(P＜0.001);与FAN 0 μmol/L组相比,FAN 5 μmol/L组、FAN 10 μmol/L组和FAN 20 μmol/L组的PC3细胞中P62和Bcl-2 蛋白表达明显降低(P＜0.001),LC3B、Bax和Caspase-3 蛋白表达明显升高(P＜0.001).结论 FAN可能通过线粒体凋亡通路抑制细胞增殖、侵袭和迁移,促进其自噬和凋亡,并呈现剂量依赖性.

目的 探究防己诺林碱对增生性瘢痕的抑制作用.方法 提取原代瘢痕组织成纤维细胞,分别用DMSO(对照组),10、20、40μmol/L的防己诺林碱处理成纤维细胞24 h后,通过MTT实验检测对其增殖能力的影响;通过流式细胞术及Hochest33258染色检测对其增殖、凋亡能力的影响;通过western blot和real time PCR检测防己诺林碱对瘢痕组织成纤维细胞中Cyclin D1和Bcl-2表达水平的影响.结果 10、20、40μmol/L的防己诺林碱作用瘢痕组织成纤维细胞24 h后,细胞的抑制率分别为(21.32±4.87)％、(27.13±1.71)％和(29.86±5.59)％,与对照组相比差异均具有统计学意义;防己诺林碱还可抑制细胞周期进程,促进其凋亡;Western blot和real time PCR检测结果显示,防己诺林碱可抑制Cyclin D1和Bcl-2在瘢痕组织成纤维细胞中的表达.结论 防己诺林碱可抑制瘢痕组织成纤维细胞的生长.

目的 建立中药材防己超高效液相色谱指纹图谱,结合化学计量学方法评价其质量.方法 采用UPLC法建立中药材防己指纹图谱,以全谱图色谱峰峰面积及共有峰峰面积为变量,利用 SIMCA-P(11.5)等软件进行相似度分析、主成分分析和聚类分析,并测定粉防己碱和防己诺林碱的含量.结果 防己药材指纹图谱共生成8个共有峰,指认出2个色谱峰,相似度介于0.706～0.997;主成分 分析、聚类分析和相似度分析结果之间互相验证,很好地体现了不同样品之间的内在质量差异;防己药材中粉防己碱和防己诺林碱含量在4.8747～20.2266、4.2407～9.5440mg/g之间.结论 超高效 液相指纹图谱结合化学计量学方法可适用于不同防己药材之间的内在质量评价.

Objective:With the development of the society,the number of people who catch the nephrotic syndrome (NS) is going up roughly.As we all know,traditional Chinese medicine (TCM),especially Fangji Huangqi Decoction (FHD),has a long history with good curative effects on NS.However,the mechanism of FHD treating NS has not been clearly elucidated.Methods:In this study,TCMSP platform was employed to screen active compounds of each herb of Fangji Huangqi Decoction combined with literatures.Furthermore,PharmMapper and SEA were used to predict and screen the active targets of FHD,and the HOME-NCBI-GENE,GeneCards and OMIM database were used to screen the active targets of NS.The GO and KEGG pathways involved in the targets were analyzed by ClueGO.Finally,contribution index was used to screen the active ingredients of FHD in the treatment of NS.Results:After drug-likeness (DL) and bioavailability (OB) filtering,43 compounds were selected from FHD,interacting with 85 NS-related targets.Systematic analysis of the constructed networks revealed that it was mainly involved in PI3K-Akt signaling pathway,MAPK signaling pathway,T cell receptor signaling pathway and TNF signaling pathway.The contribution index of every active ingredient also indicated five compounds,including astragaloside Ⅳ,quercetin,glycyrrhizic acid,glycyrrhizin and fangchinoline.Conclusions:These results have successfully predicted the pharmacodynamic material basis and the mechanism efficiency of FHD in the treatment of NS.

Tetrandrine (TET) and fangchinoline (FAN) are dominant bisbenzylisoquinoline (BBIQ) alkaloids from the roots of Stephania tetrandra of the family Menispermaceae.BBIQ alkaloids comprise two benzylisoquino-line units linked by oxygen bridges.The molecular structures of TET and FAN are exactly the same,except that TET has a methoxy (-OCH3) group,while FAN has a hydroxyl (-OH) group at C7.In this overview,the current knowledge on the chemistry,pharmacology and anticancer properties of TET and FAN have been updated.The focus is on colon and breast cancer cells,because they are most susceptible to TET and FAN,respectively.Against colon cancer cells,TET inhibits cell proliferation and tumor growth by inducing apoptosis and G1 cell cycle arrest,and suppresses adhesion,migration and invasion of cells.Against breast cancer cells,FAN inhibits cell proliferation by inducing apoptosis,G1-phase cell cycle arrest and inhibits cell migration.The processes involve various molecular mechanisms and signaling pathways.Some insights on the ability of TET and FAN to reverse multi-drug resistance in cancer cells and suggestions for future research are provided.

运用硅胶、反相硅胶、凝胶柱色谱,以及半制备高效液相色谱技术,从粉防己70％乙醇水提取物中分离得到15个生物碱类化合物,通过质谱、核磁共振波谱,以及单晶衍射数据,分别鉴定为tetrandrasideA (1)、(Z)-N-formyl-nomuciferin (2)、(E)-N-formyl-nornuciferin (3)、salutaridine (4)、salutaridine N-oxide (5)、(E)-3-(4-hydroxy-3-methoxy-phenyl)-N-[2-(4-hydroxy-3-methoxyphenyl)ethyl]-2 propenamide (6)、dauriporphine (7)、 sinomenine (8)、 liriodenine (9)、α-magnoflorine (10)、(1S)-4'-β-glucosylcoclaurine (11)、tetrandrine (12)、fangchinoline (13)、tetrandrine 2'-β-oxide (14)和tetrandrine 2'-α-oxide (15).其中,化合物1为新的生物碱苷类化合物,化合物2～11为首次从粉防己中分离得到.这些化合物对肺癌耐药细胞株H1299均显示了很好的细胞毒活性,化合物9的活性最佳,其IC50为5.38 μmol·L-1.