雷公藤甲素通过PTEN表达调控Wnt/β-catenin通路抑制黑色素瘤的机制研究

目的:探讨雷公藤甲素的抗黑色素瘤作用是否与PTEN表达及Wnt/β-catenin通路相关。方法:实验分为5组:空白对照组、雷公藤甲素(20 nmol/L)组、β-catenin抑制剂(5μmol/L IWR-1-endo)组、雷公藤甲素+β-catenin抑制剂组、ATRA(100μmol/L)组。分别采用CCK8法、Annexin V-FITC/PI双染色法、实时定量PCR法和Western blot法检测各组A375细胞活力、凋亡率及PTEN、β-catenin、Bcl-2、Caspase-3、PCNA mRNA及蛋白的表达。结果:雷公藤甲素、β-catenin抑制剂和ARTA处理能抑制A375细胞的增殖并诱导其凋亡,在增强Caspase-3和PTEN表达的同时抑制β-catenin、Bcl-2、PCNA的表达。结论:雷公藤甲素能抑制黑色素瘤细胞A375的增殖并诱导其凋亡,该作用可能是通过增加PTEN的表达进而抑制Wnt/β-catenin通路的激活来实现。     

雷公腾内酯与多柔比星诱导耐药白血病细胞凋亡协同作用及其机制的探讨

目的：以耐多柔比星（adriamycin,ADM）的人急性髓系白血病（acute myeloid leukemia,AML）耐药细胞株HL60/ADM为研究对象,探讨IC20浓度雷公腾内酯（triptolide,TPL）能否提高ADM诱导耐药白血病细胞凋亡及其与Nrf2通路的关系。方法：流式细胞仪检测空白对照组、IC20浓度TPL单药组、ADM单药组和TPL联合ADM组处理HL-60/ADM后细胞凋亡率;实时荧光定量PCR检测各个处理组作用后,Nrf2及其下游基因醌氧化还原酶（quinone oxidoreductase,NQO1）、谷胱甘肽还原酶（glutathione reductase,GSR）及血红素加氧酶1（heme oxygenase 1,HO-1）的表达水平变化;蛋白质印迹法检测各处理组作用后Nrf2蛋白表达变化。结果：TPL单药组细胞凋亡率为（5.28±0.80）%,与空白对照组的（7.09±0.46）%比较差异无统计学意义,P=0.226;但该浓度TPL可使ADM组细胞凋亡率由（19.55±1.70）%提高到（72.62±4.83）%,是ADM单药组的3.71倍,P〈0.001。空白对照组、TPL单药组、ADM单药组及双药联合组Nrf2mRNA表达水平分别为1、0.742±0.052、0.619±0.042和0.241±0.010,NQO1分别为1、0.363±0.075、0.228±0.053和0.050±0.034;GSR分别为1、0.268±0.042、0.231±0.106和0.038±0.017;HO-1分别为1、0.495±0.023、0.282±0.099和0.048±0.036;各用药组Nrf2及其下游基因NQO1、GSR及HO-1的mRNA表达水平较对照组均出现显著下调,P〈0.001;其中双药联合组下调程度最大。蛋白质印迹法结果显示,用药组及联合组Nrf2表达水平较对照组均有不同程度下调,其中联合组下调最为明显。结论：IC20浓度雷公腾内酯可显著提高ADM诱导耐药白血病细胞凋亡,其分子机制与下调Nrf2通路有关。     

来氟米特与雷公藤多苷治疗IgA肾病对照研究

目的 观察来氟米特(LEF)、雷公藤多苷联合激素治疗IgA肾病的疗效和安全性.方法 收集符合条件的60例中等量蛋白尿IgA肾病患者随机分为2组,试验组LEF联合中等量激素治疗,对照组雷公藤多苷联合中等量激素治疗,观察治疗前、后1、3、6个月的相关临床指标变化,并进行评价.结果 试验组治疗后24 h尿蛋白定量显著减少(P<0.01),血清白蛋白显著升高(P<0.01),完全缓解率为46.4%,总有效率为85.7%.与对照组比较疗效差异无统计学意义(P>0.05).观察组起效快,不良反应轻微,患者耐受性好.结论 来氟米特联合激素可以作为治疗IgA肾病的选择之一,且安全、有效.     

Enhanced Anti-tumor Efficacy of Aspirin Combined with Triptolide in Cervical Cancer Cells

Background: The non-steroidal anti-inflammatory drug (NSAID) aspirin (acetylsalicylic acid) is an inhibitor ofcyclooxygenase enzymes. Recent studies have shown that aspirin could be used as an anti-tumor drug. Triptolide,the major compound extracted from the Chinese herb Tripteryglum wilfordii Hook.f, has now been shown that itcan inhibit tumor growth. The aim of this study was to analyze the anti-tumor efficiency of aspirin and triptolidein cervical cancer cells. Methods: Viability of cervical cancer cell lines was assessed by the MTT method atvarious concentrations of aspirin and triptolide. Siha and HeLa cell apoptotic analysis was performed by flowcytometry. Real time-PCR and Western Blotting were used to analyze the expression of Bcl-2/Bax, Cyclin D1 andp16. Results: Viability in the combination group was significantly decreased as compared with either drug usedalone. Expression change of Bcl-2/Bax, CyclinD1 and p16 appeared to play an important role in the synergistickilling effect on cervical cancer cell apoptosis. Conclusion: Aspirin and triptolide combination treatment mayhave synergistic anti-tumor effects on cervical cancer cells.     

Combined Effects of Curcumin and Triptolide on an Ovarian Cancer Cell Line

Background: As natural medicines in Asia, curcumin and triptolide extracted from different drug plants haveproven to possess anticancer potential and widely used for anti-cancer research. The present study attempted toclarify that curcumin and triptolide synergistically suppress ovarian cancer cell growth in vitro. Methods: To testsynergic effects, cell viability and apoptosis were analyzed after curcumin and triptolide combination treatmenton ovarian cancer cell lines. Synergistic effects on apoptosis induction were determined by lactate dehydrogenase(LDH) leakage assay, intracellular reactive oxygen species (ROS) assay, mitochondrial membrane potential(MMP) loss assay and flow cytometry analysis. Critical regulators of cell proliferation and apoptosis relatedwere analyzed by qRT-PCR and Western blotting. Results: We showed that the combination of curcumin andtriptolide could synergistically inhibit ovarian cancer cell growth, and induce apoptosis, which is accompanied byHSP27 and HSP70, indicating that HSP27 and HSP70 play the important role in the synergic effect. Conclusions:From the result present here, curcumin and triptolide combination with lower concentration have a synergisticanti-tumor effect on ovarian cancer and which will have a good potential in clinical applications.     

Triptolide inhibits proliferation and induces apoptosis of human melanoma A375 cells

Triptolide, a diterpenoid obtained from Tripteryglum wilfordii Hook.f, has attracted interest for its anti- tumor activities against human tumor cell lines in recent years. This report focuses on anti-proliferative and pro-apoptotic activities in human melanoma A375 cells assessed by CCK8 assay, Hoechst 33258 staining and flow cytometry. In addition, triptolide-induced arrest in the S phase was also observed. Caspase assays showed the apoptosis induced by triptolide was caspase-dependent and probably through intrinsic apoptotic pathways. Furthermore, expression of NF-κB (p65) and its downstream factors such as Bcl-2, Bcl-XL was down-regulated. Taken together, the data indicate that triptolide inhibits A375 cells proliferation and induces apoptosis by a caspase-dependent pathway and through a NF-κB-mediated mechanism.     

Simultaneous determination of triptolide and its prodrug MC002 in dog blood by LC–MS/MS and its application in pharmacokinetic studies

Ethnopharmacological relevance

Tripterygium wilfordii HOOK F (TWHF) is a traditional Chinese medicine used in the treatment of various autoimmune diseases and inflammatory disorders including rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), and skin diseases. Triptolide (TP) is one of the main active ingredients of this traditional Chinese medicine. MC002 is a novel semi-synthetic derivate of TP which is highly water soluble, acts as a prodrug and is converted to TP in vivo.

Aim of this study

A sensitive, rapid method for the simultaneous determination of TP and its chemo-unstable prodrug MC002 in dog blood was developed and validated using electrospray ionization (ESI) liquid chromatography–tandem mass spectrometry (LC–MS/MS). Using this method, a pharmacokinetic study of MC002 and TP following an intravenous drip infusion of 0.2 mg/kg MC002 in dogs was performed.

Materials and methods

Chemo-degradation of the prodrug in blood samples was inhibited by the addition of a small amount of sodium fluoride solution before using liquid–liquid extraction with ethyl acetate. The concentrations of MC002 and TP in dog blood were determined using the LC–MS/MS method.

Results

The quantitative method showed good precision and stability and is suitable for the assay of biological samples. The pharmacokinetic study showed that the elimination of MC002 was faster than that of TP, and the concentrations and AUC_0−t values of TP were higher than MC002. MC002 can rapidly convert to TP in vivo.

Conclusions

This validated method was successfully applied in a pharmacokinetic study of MC002 following an intravenous drip infusion in dogs. With the development of this new prodrug of TP as a promising anti-cancer drug, this method is suitable for its further analysis in clinical studies.     

雷公藤内酯醇对HaCaT细胞表皮生长因子受体磷酸化及其下游信号激酶MEK的影响

目的 探讨雷公藤内酯醇对表皮生长因子(EGF)信号刺激HaCaT细胞后EGF受体磷酸化以及下游MEK途径的影响.方法 在HaCaT细胞培养体系中,用外源性重组人EGF为刺激信号,用免疫印迹法检测总EGF受体及其磷酸化受体,以及总MEK及其磷酸化激酶的表达.结果 雷公藤内酯醇对rhEGF引起的细胞总EGF受体表达无明显影响,但抑制磷酸化EGF受体表达,抑制作用呈剂量依赖性,IC₅₀值为2.38×10^-10mol/L.对细胞总MEK1/2表达也不影响,但明显下调磷酸化MEK,抑制作用的IC₅₀值约为6.03×10^-11mol/L.结论 雷公藤内酯醇对EGF信号引起的EGF受体磷酸化及下游MEK激酶磷酸化均有抑制作用.     

卵巢癌中雷公藤内脂醇的研究进展

卵巢癌是严重威胁女性生殖健康的肿瘤之一,死亡率居各类妇科肿瘤的首位。多药耐药是治疗后复发、转移甚至死亡的主要原因。雷公藤内酯醇（triptolide,TP）具有很强的抗肿瘤活性,其在卵巢癌治疗方面的作用也引起了众多学者的重视,尤其是在多药耐药卵巢癌治疗中的应用。本文从抑制卵巢癌细胞增殖、诱导癌细胞凋亡两大方面综述雷公藤内酯醇在卵巢癌治疗中的研究进展。TP的毒性反应、半衰期短等缺点限制了其临床应用,纳米给药技术的出现为TP用于卵巢癌治疗提供了更广阔的应用前景。