Inhibitory effect of active ingredients of Tripterygium wilfordii Hook.F.on human carboxylesterases

OBJECTIVE The inhibitory effect of active ingredients of Tripterygium wilfordii Hook.F.(TWHF)(celastrol,triptolide,triptonide,wilforlide A,wilforgine and wilforine)on human carboxylester⁃ase 1(CES1)and CES2 was detected to investigate the herb-drug interactions(HDIs)of TWHF.METHODS Human liver microsomes catalysed hydrolysis of 2-(2-benzoyl-3-methoxyphenyl)benzothi⁃azole(BMBT)and fluorescein diacetate(FD)were used as the probe reaction to phenotype the activity of CES1 and CES2,respectively.The residual activities of CES1 and CES2 were detected by ultrahigh performance liquid chromatography(UPLC)after intervention with celastrol,triptolide,triptonide,wilforlide A,wilforgine and wilforine(100μmol·L^(-1)).Kinetics analysis,involving half inhibitory concentra⁃tion(IC_(50)),inhibition type and kinetic parameter(Ki),and in vitro-in vivo extrapolation(IVIVE),was carried out to predict the HDIs between these compounds and CES-metabolizing drugs.Molecular docking was performed to analyze the ligand-enzyme interaction.RESULTS Out of the six main con⁃stituents of TWHF,only celastrol exhibited strong inhibition towards both CES1 and CES2,with the inhibitory rates of 97.45%(P<0.05)and 95.62%(P<0.05),respectively.The IC_(50)was 9.95 and 4.02 mol·L^(-1),respectively,and the types of inhibition were all non-competitive inhibition.Based on the kinetics analysis,the Ki values were calculated to be 5.10 and 10.55μmol·L^(-1)for the inhibition of celastrol on CES1 and CES2,respectively.IVIVE indicated that celastrol might disturb the metabolic hydrolysis of clinical drugs in vivo by inhibiting CES1.Molecular docking results showed that hydrogen bonds and hydrophobic contacts contributed to the interaction of celastrol and CESs.CONCLUSION The inhibitory effect of celastrol on CES1 and CES2 might cause HDIs with clinical drugs hydrolysed by CESs.

In vitro propagation of a medicinal plant: Tripterygium wilfordii Hook f.

In this study a reliable protocol was developed for the establishment of commercial in vitro cultures of Tripterygium wilfordii Hook f.. Juvenile shoots from one-year-old elite plants were used as the source of explants. New axillary shoots were obtained after 30 days of culture on a MS medium supplemented with BAP （2.0 mg.L^-1） and NAA （0.1 mg.L^-1）. The optimal multiplication medium was a modified MS medium supplemented with BAP （1.0 mg.L^-1） and NAA （0.1 mg.L^-1）. This yielded a multiplication rate of 2.4 for each subculture. Slightly more than 92% of shoots rooted when cultured on a modified MS medium containing IBA （0.2 mg.L^-1） and activated charcoal （0.5 mg.L^-1）. Activated charcoal promoted both a strong and a high rooting rate during the rooting phase. Plantlets were transferred to pots for a short acclimatization stage in a greenhouse where 95% of the plantlets survived. This highly reproducible procedure can be adopted for large-scale propagation of T. wilfordii.

Isolation and Purification of Triptolide from the Leaves of Tripterygium wilfordii Hook F

Triptolide is an important active component of Tripterygium wilfordii Hook F (TWHF) and possesses anti-inflammatory, immunosuppressive, male anti-fertility, and anticancer properties. A new method combining different techniques, including solid-liquid extraction, liquid-liquid partition, column chromatography and high-speed counter-current chromatography (HSCCC) but avoiding the use of chloroform, was developed for the isolation and purification of triptolide from the leaves of TWHF. 48 mg of triptolide at 96.5% purity was obtained from 1 kg of air-dried leaves of TWHF.

Effect of the Multiglycoside of Tripterygium Wilfordii Hook f. (Tii) on Cornea Allograft Rejection Model in Rabbit

Purpose:Toexamine the effect of Tii treatment of cornea graft survival in a rab-bit model.Methods:Tii was administrated orally after eccentrical corneal transplantation.Survival times were determined by biomicroscopy.Cytotoxic T lymphocytes(CTL)and delayed-type hypersensitivity(DTH)responses to donor alloantigens were assessed at ady 16after heterotopic corneal grafts.Results:Administration of Tii reduced the incidence and prologed the graft sur-vival time.Both CTLand DTH responses to donor alloantigens were severely ed-pressed in hosts treated with Tii.However,combination of Tii and cyclosporine further enhanced the immunosuppressive effects described above.Conclusions:Tii is a potent immunosuppressant with the ability to prolong allo-graft survival in the rabbit penetrating keratoplasty model and may have coordi-native effects with CsA through different mechanisms.Further studies are neces-sary to define any potentially coordinative role in the prevention of allograft rejec-tion in human keratoplasty.Eye Science 1995;11:168-172.

A Novel Diterpenoid,11-O-β-D-glucopyranosylneo-triptophenolide, from Tripterygium wilfordii Hook. f

A new diterpenoid was isolated from the ethanolic extract of the dried root bark of Tripterygium wilfordii Hook.f. It is the first example of abietane diterpenoid glycoside isolated from Tripterygium wilfordii Hook.f. Its structure was identified to be 11-O-b-D-glucopyranosyl- neotriptophenolide based on spectral methods.

雷公藤内酯醇(T10)通过抑制脊髓内胶质细胞的激活改善大鼠CFA诱导的炎性痛的机制研究

目的:观察腹腔注射雷公藤内酯醇(Triptolide,T10)对完全弗氏佐剂(complete Freund’s adjuvant,CFA)诱导的大鼠慢性炎性痛行为的改善作用并探讨其机制。方法:SD大鼠随机分为四组,即Saline+Veh组、Saline+T10组、CFA+Veh组和CFA+T10组。后两组大鼠于右侧足底注射CFA制备大鼠慢性炎性痛模型,前两组则在相同部位注射生理盐水。第二组和第四组大鼠分别于造模前1 h给予T10腹腔注射,随后每12 h给予腹腔注射药物1次,持续用药7 d;第一组和第三组则以相同方式给予100 ml/kg的生理盐水作为对照。采用辐射热法和机械刺激法连续观察给药后大鼠的痛行为;应用免疫组织化学染色和Western Blot方法观察连续给予T10 3d和7 d后大鼠腰膨大平面脊髓背角内小胶质细胞和星形胶质细胞的活化程度;应用实时定量PCR(RT-PCR)方法观察连续给药7 d后大鼠腰5背根神经节内炎性因子,如白细胞介素-6(interleukin-6,IL-6)、白细胞介素-1β(interleukin-1 beta,IL-1β)和肿瘤坏死因子-α(tumour necrosis factor-α,TNF-α)的表达变化。结果:(1)与CFA+Veh组相比,CFA+T10组大鼠机械缩足阈值及辐射热痛潜伏期显著下降(P<0.05),并持续到造模成功后7 d,表明腹腔注射T10可以明显改善CFA诱导的大鼠机械痛敏和辐射热痛敏。(2)免疫组织化学染色和Western Blot结果显示:造模后3 d和7 d后CFA+Veh组大鼠脊髓背角出现大量CFA诱导活化的小胶质细胞和星形胶质细胞;小胶质细胞标记物IBa-1和星形胶质细胞标记物GFAP的表达量分别在CFA造模后3 d和7 d显著增高(P<0.05),而给予T10 3 d和7 d后炎性痛大鼠腰膨大平面IBa-1和GFAP的活化程度均显著减轻(P<0.05)。(3)RT-PCR结果显示:与Saline+Veh组相比,CFA造模7 d后腰5背根神经节内炎性因子IL-1β、IL-6 mRNA和TNF-αmRNA的表达显著增高(P<0.05),腹腔注射T10 7 d后上述炎性因子的表达则显著降低(P<0.05)。结论:腹腔注射T10可以显著改善CFA诱导的大鼠慢性炎性痛,其机制可能与抑制脊髓背角小胶质细胞和星形胶质细胞的活化以及炎性因子IL-1β、IL-6和TNF-α的合成有关。

雷公藤多甙抑制EM患者腹腔巨噬细胞及相应细胞模型分泌TNF-α的实验研究

目的观察雷公藤多甙(TWP)对子宫内膜异位症(EM)患者腹腔巨噬细胞和小鼠腹腔活化巨噬细胞释放TNF-α的影响,以探讨TWP用于临床治疗EM的机制并建立细胞模型,为后续研究做准备。方法收集16例经腹腔镜检查确诊为EM患者腹腔液分离纯化巨噬细胞,并给予不同剂量的TWP进行共同孵育;同时分离纯化小鼠腹腔巨噬细胞,用脂多糖(LPS)激活以获得细胞模型,同样予TWP进行共同孵育。分别以间接MTT法和RT-PCR法在蛋白质水平和基因水平检测该药物影响激活状态巨噬细胞分泌TNF-α含量的变化。结果 EM患者腹腔液中TNF-α含量显著高于非EM妇女,在4～24 h后TWP(6.25～100μg/L)显著抑制了TNF-α的分泌。结论 TWP在蛋白质和基因水平均能有效抑制激活巨噬细胞分泌TNF-α。

Effects of triptolide on hippocampal microglial cells and astrocytes in the APP/PS1 double transgenic mouse model of Alzheimer＇s disease

The principal pathology of Alzheimer＇s disease includes neuronal extracellular deposition of amyloid-beta peptides and formation of senile pl aques, which in turn induce neuroinflammation in the brain. Triptolide, a natural extract from the vine-like herb Tripterygium wilfordii Hook F, has potent anti-inflammatory and immunosuppressive efficacy. Therefore, we determined if triptolide can inhibit activation and proliferation of microglial cells and astrocytes in the APP/PS1 double transgenic mouse model of Alzheimer＇s disease. We used 1 or 5 μg/kg/d triptolide to treat APP/PS1 double transgenic mice （aged 4-4.5 months） for 45 days. Unbiased stereology analysis found that triptolide dose-dependent- ly reduced the total number of microglial cells, and transformed microglial cells into the resting state. Further, triptolide （5 μg/kg/d） also reduced the total number of hippocampal astrocytes. Our in vivo test results indicate that triptolide suppresses activation and proliferation of microglial cells and astrocytes in the hippocampus of APP/PS 1 double transgenic mice with Alzheimer＇s disease.

Gene expression profile analyses of mice livers injured by Leigongteng

AIM： To analyze the gene expression profiles of mice livers injured by Leigongteng and explore the relationship between the differentially expressed genes and liver damage. METHODS： The experimental mice were randomly divided into a control group and a liver-injured group in which the mice were administrated 33 μγ, of triptolide/ kg per day for 30 d. Liver mRNAs were extracted from animals in both groups and were reverse-transcribed to cDNA with dUTP labeled by different fluorescence （Cy3, Cy5） as hybridization probes. The mixed probes were hybridized with oligonucleotide microarray chips. The fluorescent signal results were acquired by scanner and analyzed with software. RESULTS： Among the 35852 target genes, 29 genes were found to be significantly differentially expressed, with 20 genes up-regulated and 9 genes down-regulated. The reliability of the differentially expressed genes was validated by RT-PCR experiments of 5 randomly selected differentially expressed genes. CONCLUSION： Based on the biological functions of the differentially expressed genes, it is obvious that the occurrence and development of liver damage induced by Leigongteng in mice are highly associated with immune response, metabolism, apoptosis and the cell skeleton of liver cells. This might be important for elucidating the regulatory network of gene expression associated with liver damage and it may also be important for discovering the pathogenic mechanisms of liver damage induced by Leigongteng.

Im m unoregulatory Effects of Triptergium W ilfordii Hook at Antifertility Dose on Splenic NK Cells Activityin Male Mice

Effects of Tripterypium Wilfordii Hook f (TWH) on sperm atozoa in the epi- didym is and splenic NK cells activity in m ale m ice w ere observed using MTT assay and silver impregnation m ethods. The results show ed that the density, viability and m otility of the epididym alsperm atozoa in the experim entalgroupstreated w ith TWH w ere m ore significantly reduced than those in the controlgroup (P< 0.01). The head sw elling, head separation from tailin the groups treated w ith TWH w ere observed. The inhibition of splenicNK cellsactivity in m iceby TWH w asdose-dependent. Inhi- bition by TⅡand TWH athigh dose on the NK cells activity w as significant (P< 0.01 and P< 0.05), w hileinhibitory effectsof TWH atinterm ediateand low doseson the NK cells activity w ere notobserved (P> 0.05). Itw as concluded thatTWH at low er antifertility dose did not significantly inhibit the splenic NK cells activity. It m ightbe usefulforevaluating thetherapeuticeffectsof TWH in futureclinicalprac- tice.

Effects of GTW treatment on proteinuria and acute glomerular immune lesion in experimental mesangial proliferative glomerulonephritis

To examine the effect of multi-glycoside of Tripterygium wilfordii Hook. f. （GTW） on proteinuria and acute glomerular immune lesion in experimental mesangial proliferative glomerulonephritis （MsPGN） induced by anti-Thyl. 1 monoelonal antibody （mAb） 1-22-3. The reversible model of MsPGN with anti-Thyl. 1 mAb 1-22-3 was established. After 7 days of oral treatment with GTW （ 100 mg/kg per day） and vehicle （distilled water, 5 ml/kg per day）, its effects on proteinuria, renal functions, mesangial morphological change, glomerular macrophage accumulation, and mRNA expressions of cytokines （PDGF-BB and MCP-1 ） were evaluated by light microscope （LM）, immunofluorescence （IF）, and reverse transcription polymerase chain reaction （RT-PCR）. It was found that GTW ameliorated proteinuria （on day 3 and day 7）, mesangial proliferation （total cell number, matrix expansion, a-smooth muscle actin expression, and collagen type Ⅰ expression） and macrophage accumulation （ED3^＋ ） in experimental MsPGN. In addition, GTW significantly suppressed the increased mRNA expressions for MCP-1 （67.6% to eontrol group, P 〈 0.01） together with the tendency to reduce the expression of PDGF （24.44% to control group） on day 7. It is concluded that GTW can not only decrease proteinuria, but also ameliorate acute mesangial alterations and glomerular activated macrophage accumulation probably by reduction of cytokines. These data indicate that GTW is an effective agent for early MsPGN.

雷公藤药材总生物碱含量及影响因素的研究

目的：考察雷公藤药材总生物碱的含量状况及其变化规律。方法：采用有机溶剂提取和氧化铝柱色谱分离进行样品前处理,紫外分光光度法在268 nm波长下测定吸收度。结果与结论：一般韧皮部含量是木质部含量的2~5倍,二者含量变化呈现一致性。从不同种和类型看,昆明山海棠含量较高,雷公藤F次之,东北雷公藤最低;但每一类型内,不同居群间差别较小。含量较高的居群均来自雷公藤F和昆明山海棠,它们是江西遂川、安徽黄山、浙江景宁和云南昆明。居群内个体间的差别在不同居群中程度有所不同,浙江景宁、福建泰宁、云南昆明等居群内多样性丰富,有利于优良种质的选择。在同一植株内,药材粗细对总生物碱的含量影响不大。木质部总生物碱的含量呈现随季节变化的规律,3月含量最高,然后降低,12月降为最低。韧皮部的总生物碱含量变化规律不明显。

HPLC法测定雷公藤及其片剂中雷公藤红素

目的建立雷公藤及其片剂中雷公藤红素的HPLC测定方法.方法 HPLC外标法.Agilent Zorbax SC-C18柱为色谱柱,甲醇-1%醋酸溶液(87:13)为流动相,检测波长为425 nm,体积流量为1.0 mL/min.结果雷公藤红素线性范围为40.96～204.8μg/mL(r=0.999 6),药材的平均回收率为98.37%,RSD为1.01%(n=9),制剂的平均回收率为98.59%,RSD为1.18%(n=9).结论本方法操作简便、准确,可作为雷公藤类药物中雷公藤红素的定量分析方法.

雷公藤与昆明山海棠采样调查报告

目的:通过采样调查,获取雷公藤与昆明山海棠野生资源现状、种植、商品流通等方面的资料。方法:在全国13个样地采集样品,比较雷公藤和昆明山海棠的形态特征与生长习性的差异;调查雷公藤与昆明山海棠的野生资源现状、种植信息、民间混淆种及不同市售药材的商品特征。结果:(1)雷公藤和昆明山海棠主要采集于林下或林地边缘,光照和水分对雷公藤属植物的分布有一定的影响;(2)雷公藤与昆明山海棠的野生资源量正在萎缩,两种植物种植历史较短,种植技术水平较低;(3)雷公藤的采收加工尚缺乏规范,市售药材将雷公藤属植物的根、根茎、茎均作商品药材销售,饮片规格多样。结论:雷公藤与昆明山海棠的野生资源正在萎缩,人工种植、采收加工及市售药材的管理等方面的研究尚需规范深入的开展。

雷公滕及铅、镉对小鼠睾丸间质细胞一氧化氮合酶活性的影响

目的 探讨雷公藤及铅、镉等重金属元素对睾丸间质细胞一氧化氮合酶 (NOS)活性的影响。 方法 用 NADPH- d黄递酶组织化学方法 ,观察给予氯化镉、醋酸铅、雷公藤 1、2、3、4周后 ,小鼠睾丸间质 NOS活性的变化。 结果 正常组间质细胞呈 NOS强阳性反应 ,而曲精小管生精细胞均呈 NOS阴性反应。给予雷公藤 2周内NOS反应无变化 ,第 3和第 4周后 ,NOS阳性的睾丸间质细胞数量无明显变化 ,但 NOS活性稍降低。给予醋酸铅和氯化镉 1周后 ,NOS阳性的睾丸间质细胞数量明显减少 ,并随给药时间延长而加重 ,NOS活性逐渐降低 ,给药 4周后 NOS阳性的睾丸间质细胞几乎不着色。 结论 雷公藤虽然具有明显抗生育作用 ,但对 NOS阳性的睾丸间质细胞数量及 NOS活性无明显影响 ;而铅、镉等对睾丸有毒理作用的重金属元素 ,不但可使 NOS阳性的睾丸间质细胞减少 ,NOS活性也明显降低。

雷公藤有效成分制备的研究进展

简要介绍了近年来有关中药雷公藤有效成分的制备方法及研究进展 。

雷公藤诱导细胞凋亡的研究进展

雷公藤 Tripterygium wilfordii提取物在临床上常用于治疗类风湿关节炎等多种疾病 ,其疗效已被广泛认可。近 5年来国内外学者从其提取物诱导细胞凋亡的角度 ,对其药理机制进行了细胞分子水平的探讨 。

系膜增殖性肾小球肾炎的新疗法——雷公藤多甙新治疗方案的疗效观察

目的：应用双倍剂量雷公藤多甙（ＴⅡ）继以间歇用药维持治疗方案治疗不同临床病理类型的原发性系膜增殖性肾小球肾炎，前瞻性观察其疗效及副反应，确定此一新疗法的临床价值。方法：７９例无氮质血症的原发性系膜增殖性肾小球肾炎患者（ＩｇＡＮ４７例，ＩｇＭＮ１５例，ＭｓＰＧＮ１７例）在连续服用ＴⅡ２ｍｇ／（ｋｇ·ｄ）４周后改为间歇疗法，每日２ｍｇ／ｋｇ，服２周停２周，交替进行，平均随访１９．３周，观察蛋白尿、血尿变化及副反应。结果：４７例尿检异常型ＩｇＡＮ患者中，１９例达临床缓解（４０．５％），５例改善（１０．６％），２３例无效（４８．９％）。其中表现为单纯蛋白尿者疗效最佳，其次为单纯血尿者，蛋白尿伴血尿者疗效最差。１７例ＭｓＰＧＮ患者中，１１例达临床缓解（６４．７％），６例无效（３５．３％）。同样以单纯蛋白尿者疗效最佳，其次为蛋白尿伴血尿者，单纯血尿疗效最差。１５例ＩｇＭＮ患者中，８例达临床缓解（５３．３％），３例改善（２０％），４例无效（２６．７％）。１９例以往曾用足量激素治疗无效的患者中有１２例（６３．２％）经双倍剂量ＴⅡ治疗后获缓解。临床缓解的３８例患者中，缓解时间在４周以内者２６例（６８．４％），６周内者３２例（?

双倍剂量雷公藤多甙治疗原发性肾病综合征的近期疗效

目的：首次应用双倍剂量雷公藤多甙（ＴⅡ）治疗原发性肾病综合征，前瞻性观察其疗效。方法：１８例单纯肾病综合征患者（８例ＩｇＡＮ，４例ＩｇＭＮ，６例ＭｓＰＧＮ）均服用ＴⅡ２ｍｇ／（ｋｇ．ｄ）４周，观察蛋白尿变化及副反应。结果：１５例肾病综合征获得完全缓解，总缓解率达８３３％，其中ＩｇＡＮ（７例），ＩｇＭＮ（３例），ＭｓＰＧＮ（５例）的缓解率分别为８７５％，８３３％，７５０％。１５例中１２例在服药２周内获得缓解，部分对皮质激素不敏感者双倍剂量ＴⅡ仍然有效，无效仅２例。尿蛋白平板电泳分析标明对ＴⅡ敏感者尿蛋白呈高度选择性，而对ＴⅡ不敏感者含有较多大分子及小分子量蛋白质。所有患者均能耐受双倍剂量ＴⅡ，消化道反应及对肝细胞的损伤作用轻微，无一例出现严重白细胞减少。分别有２例患者在ＴⅡ迅速减量及继发上呼吸道感染后肾病综合征复发，但重新采用大剂量ＴⅡ或延长治疗时间仍有效。结论：双倍剂量ＴⅡ对临床表现为单纯肾病综合征，组织学病变为系膜增生的多种肾小球疾病具有良好的疗效。

雷公藤多甙抑制小鼠腹腔巨噬细胞产生一氧化氮

雷公藤多甙（ｐｏｌｙｇｌｙｃｏｓｉｄｅｏｆＴｒｉｐｔｅｒｙｇｉｕｍｗｉｌｆｏｒｄｉＨｏｏｋ．ｆ．，ＴＷＰ）临床广泛用于治疗类风湿性关节炎等多种自身免疫性疾病，既可抑制细胞免疫又可抑制体液免疫［１］．但其在免疫抑制过程中对一氧化氮合酶（ＮＯＳ）的影响却...