A Study on the Interest Linkage Mechanism of the Taxus Industry Chain in Pengshui Miao and Tujia Autonomous County

At the Taxus base of Pengshui Miao and Tujia Autonomous County,Chongqing,the comprehensive development and utilization of Taxus was carried out with seedling cultivation,understory economy,product processing,ecotourism and other aspects as the main line.The benefit linkage mechanism of the operation of the Taxus industrial chain was explored and analyzed,so as to extend and supplement the Taxus industrial chain.The two models of"company+farmer"and"company+village collective+farmer"are relatively close benefit linkage models,which indirectly promote the increase of farmers income and smoothly guarantee the operation of the industrial chain,thereby playing a positive role during the development of the Taxus industry.This study provides reliable countermeasures and suggestions for the sustained and healthy development of the Taxus industry in Pengshui County.

Pharmacodynamics of frigid zone plant Taxus cuspidata S.et Z.against skin melanin deposition,oxidation,inflammation and allergy

Background:Taxus cuspidata S.et Z.is a precious species of frigid zone plant belonging to the Taxaceae family,which possesses anticancer,anti-inflammatory,hypoglycemic,and antibacterial pharmacological properties.While taxane extracted from Taxus chinensis has been reported to elicit antioxidant activities,whether Taxus cuspidata S.et Z.has skin-protective actions against injuries remained unknown.This study aims to explore the pharmacological effects of three Taxus extracts on skin melanin deposition,oxidation,inflammation,and allergy so as to provide new ideas for the prevention and treatment of various diseases related to skin damage.Methods:Skin melanin deposition was evaluated by measuring melanin content in the skin of guinea pigs by alkali lysis method.Antioxidant capacity was evaluated by measuring superoxide dismutase(SOD)concentration and glutathione(GSH)content in skin tissue homogenates of Kunming mice by SOD assay kit and micro reduced GSH assay kit.The quantitative real-time polymerase chain reaction(qRT-PCR)and western blotting were used to examine the levels of both SOD and recombinant glutathione peroxidase 4(GPX4).Skin inflammation was evaluated by xylene-induced ear swelling test and egg-white-induced paw swelling test in mice.In a mouse model of skin allergy induced by 4-aminopyridine(4-AP),allergy was determined by licking body counts and histamine concentrations in tissue homogenates using enzyme-linked immunosorbent assay(ELISA)kits.Two proinflammatory factors tumor necrosis factor(TNF)-αand interleukin(IL)-1βwere measured by qRT-PCR.Hematoxylin and eosin(HE)staining was conducted to assess the degree of skin lesion.Results:All three Taxus extracts including Taxus chinensis essential oil,Taxus chinensis extract and Taxus chinensis extract compound reduced the melanin deposits in the back skin relative to the non-treated control animals,of which Taxus chinensis essential oil produced the greatest effect.In contrast,the three Taxus extracts elevated SOD and GSH levels in the skin tissues,and the highest increase was seen with Taxus chinensis essential oil.Three Taxus extracts,especially Taxus chinensis essential oil,effectively reduce the rate of ear and paw swelling.All three Taxus extracts reduced the number of body licks,the levels of TNF-αand IL-1β,and the histamine content in tissue homogenates of mice and alleviated skin damage.Consistently,Taxus chinensis essential oil yielded the greatest magnitude of decreases.Conclusion:While all three Taxus extracts possessed the anti-skin melanin deposition,oxidation,and allergy properties,Taxus chinensis essential oil produced the superior effects.

立地条件差异对天然东北红豆杉(Taxus cuspidata)种间竞争的影响

立地因子空间梯度特征是影响种间竞争的重要因素。通过研究主要竞争树种及竞争强度随海拔、坡向、坡位和坡度等立地因子变化的规律,揭示不同立地条件对天然东北红豆杉种间竞争的影响,为濒危东北红豆杉种群的保护与恢复提供理论指导。研究结果表明:东北红豆杉的种间竞争压力主要来自紫椴、臭冷杉、色木槭、红松、青楷槭、枫桦、花楷槭等7个树种,主要竞争树种以小径级个体居多。立地因子差异对种间竞争强度和主要竞争树种的排序有明显影响。东北红豆杉在阴坡受到的种间竞争压力是阳坡的3.27倍,主要树种的竞争强度显著高于阳坡,竞争树种的排序发生更迭;随着坡位的提升,东北红豆杉受到的种间竞争压力明显增加,主要竞争种类和强度也发生变化;坡度增加,种间竞争强度增加,主要竞争树种排序发生变化;海拔对种间竞争的影响总体不大,但影响主要竞争树种的竞争强度和排序。

Ovule Development, Formation of Pollination Drop and Pollination Process in Taxus chinensis (Taxaceae)

Pollen morphology and hydration, ovule development, formation of pollination drop and the pollination process of Taxus chinensis (Pilg.) Rehd were investigated in detail. The results showed that mature pollen was one_celled, irregular in shape, nonsaccate, and the surface was covered with orbicules. When pollen became hydrated, the intine swelled and the exine bursted. The exine usually remained on the surface of water or on the pollination drop all the time, while the rest portion of the pollen sank into the drop of water or the pollination drop. In the late August, the down_curved female buds could be observed in the field trees. The downturning of the female buds was an characteristic feature discriminating from the vegetative buds. At this stage ovular primordium was present in the female bud with a dome_shaped nucellar primordium encircled by a rudimentary integument. The integument grew fast and overgrew the nucellus. In the March of the following year, the ovule formed a micropylar canal about 1?000 μm in length, 200-300 μm in width. Except the micropyle, the other part of the ovule was enclosed by 20-25 scales. During the early stage of the ovule development, the nucellar surface clearly showed cell configuration; but a membrane_like structure covering the nucellar apex appeared in the late stage of the ovule development, and subsequently broke down with nucellar cell disintegration before pollen shedding. The disintegrated cell products might partake in the formation of the pollination drop. Before and during pollination, the nucellar cells contained abundant mitochondria, endospermic reticulum and dictysomes with vesicles; the plasma membrane was undulated and there was no accumulation of electron_dense substances in the pericytoplasm. Pollination drops usually occurred at night, and disappeared by noon in the field tree. 15 days before pollination, the in vitro ovules from the field trees already had the capability of producing pollination drops, which could last for 4-5 days indoors. However, after these drops received pollen, they did not withdraw noticeably compared with the unpollinated ones. The retraction of the pollination drop was presumed not to be a nonmetabolic outcome but rather a result of evaporation.

Productions of Taxol and Related Taxanes by Cell Suspension Cultures of Taxus yunnanensis

A high taxol yield cell line of Taxus yunnanensis Cheng et L. K. Fu keeps a high taxol_producing level after successive subcultures for more than eight years. In this study, eight taxanes were isolated from the suspension cell cultures of this cell line. Based on NMR and MS analyses, and comparison with literature data and standards, their structures were determined to be 2α,5α,10β_triacetoxy_14β_propionyloxy_4(20),11_taxadiene (1), 2α,5α,10β_triacetoxy_14β_(2′_methyl)_butyryloxy_4(20),11_taxadiene (2), 2α,5α,10β_14β_tetra_acetoxy_4 (20),11_taxadiene (3, taxuyunnanine C), 2α,5α,10β_triacetoxy_14β_(2′_methyl_3′_hydroxy)_butyryloxy_4(20),11_taxadiene (4, yunnanxane) and its 3′_epimer (5), baccatin Ⅳ (6), baccatin Ⅲ (7) and taxol (8), respectively. Among those compounds, 3, 5, 6 and 7 were reported to be isolated from the suspension cell cultures of T. yunnanensis for the first time. TLC and HPLC analyses indicated that the chemical constituents of the culture solution were similar to those of cultured cells. Moreover, the highest taxol content of this cell line reached 0.3% and the cell line could be applied for a large_scale culture.

FIREBIRD雷帕霉素洗脱支架与TAXUS紫杉醇洗脱支架成功置入后长期临床结果观察：单中心、大规模注册研究

目的:比较成功置入FIREBIRD雷帕霉素洗脱支架与TAXUS紫杉醇洗脱支架后长期临床观察结果。方法:本研究入选了2004-04至2006-10期间,在我院择期置入FIREBIRD雷帕霉素洗脱支架与TAXUS紫杉醇洗脱支架共计3 110例患者,并按置入支架的种类分为FIREBIRD组(2 274例)和TAXUS组(836例)。临床随访资料包括术后30天、1年和2年的死亡、心肌梗死、支架内血栓形成、靶病变血运重建、靶血管血运重建和主要心脏不良事件。本研究通过COX比例风险模型比较了倾向性评分匹配前后两组间的2年临床随访结果。结果:两组倾向性评分匹配前COX比例风险模型分析显示FIREBIRD组靶病变血运重建(风险比0.381,95%可信区间0.262～0.541)、靶血管血运重建(风险比0.512,95%可信区间0.383～0.681)以及主要心脏不良事件(风险比0.527,95%可信区间0.410～0.677)的风险低于TAXUS组。经过倾向性评分匹配后COX比例风险模型分析仍显示FIREBIRD组靶病变血运重建(风险比0.330,95%可信区间0.188～0.578)、靶血管血运重建(风险比0.412,95%可信区间0.263～0.643)以及主要心脏不良事件(风险比0.476,95%可信区间0.332～0.681)的风险低于TAXUS组。结论:与TAXUS紫杉醇洗脱支架相比,置入FIREBIRD雷帕霉素洗脱支架可降低靶病变血运重建、靶血管血运重建及主要心脏不良事件发生的风险。

cDNA Cloning, Expression and Characterization of Taxadiene Synthase, a Diterpene Cyclase from Taxus chinensis

Taxadiene synthase, a diterpene cyclase, catalyzes the conversion of geranylgeranyl diphosphate (GGPP) to taxadiene, a key intermediate in Taxol biosynthesis in yew. A 2 151 bp cDNA fragment encoding taxadiene synthase of Taxus chinensis (Pilg.) Rehd. was cloned by homology-based PCR and cDNA library screening. The 5′-terminal 611 bp cDNA fragment of taxadiene synthase was isolated by PCR. The two fragments were ligated together and gave a 2*!712 bp cDNA fragment with a 2*!586 bp open reading frame (ORF), encoding 862 amino acid residues including a presumptive plastidial transit peptide. The taxadiene synthase of T. chinensis most closely resembles the one from T. brevifolia (97% identity). Heterologous overexpression of 2.5 kb cDNA fragment from T. chinensis was obtained using a fusion expression vector pET-32a and the Escherichia coli strain BL21trxB. The expressed proteins from E. coli BL21trxB were present as inclusion bodies. After the inclusion bodies were denatured, renatured and refolded, the recombinant enzyme was purified by a single step with a His-binding metal affinity column. The catalytic product of taxadiene synthase of T. chinensis was detected by capillary gas chromatography-mass spectrometry (GC-MS) and identified as taxa-4(5),11(12)-diene.

Cell Apoptosis Induced by Oligosaccharide in Suspension Cultures of Taxus chinensis

Taxol production of Taxus chinensis (Pilger) Rehd. var. mairei (Lemee et Lévl.) Cheng et L. K. Fu induced by oligosaccharide from Fusarium oxysporum f. vasinfectum (Atkinson) Snyder et Hansen was studied in suspension cultures, and it was found that oligosaccharide triggered cell apoptosis. Under transmission electron microscope the following morphological changes were observed: cell shrinkage, condensation of cytoplasm, nuclear fragmentation, and the increase of high electron density bodies in vacuole in great quantity. In oligosaccharide_treated cells, agarose gel electrophoresis revealed that DNA was digested into oligonucleosomal fragments that were times of 200 bp appearing as DNA ladders. Control cells were in normal physiological state, they were intact, abundant in organelle and with integral nucleus DNA, and the rate of taxol biosynthesis in these cells was very low. After the oligosaccharide to the culture system, the defense system of cells was elicited and the secondary metabolism was strengthened, i.e. phenolics were accumulated in the medium, the activity of polyphenol oxidase (PPO) was increased quickly and secondary wall of cells was thickened. The activity of L _phenylalanine ammonia lyase (PAL), the critical enzyme of the phenylpropanoid pathway, was increased promptly 1 h after elicitation. The rate of taxol production was improved sharply and the maximal taxol concentration at 72 h was six times that of control. Appearance of cell apoptosis was accompanied with the highest concentration of taxol in suspension cultures.

Isolation and Identification of a Taxol-producing Endophytic Fungus Identified from Taxus media

[Objective] The aim was to isolate and identify a taxol-producing endophytic fungus from Taxus media. [Method] 32 strains of endophytic fungi were identified form the inner bark of T. media,and their fermentation products were detected by high performance liquid chromatography (HPLC). [Result] Through the screening,a strain of taxol-producing endophytic fungi M57 was obtained,which could produce 45-50 μg/L of taxol,and M57 was defined as Rhizopus sp. through morphological observation and 18S rDNA sequence analysis. [Conclusion] The finding of Rhizopus sp. M57 provided a promising strain for producing taxol with taxol-producing fungi fermentation process.

Optimization of RAPD Reaction System of Taxus cuspidate Sieb. et Zucc

[Objective] The research aimed to study the optimization conditions of RAPD reaction system of Taxus cuspidata.[Method] Using the leaves of Taxus cuspidata as materials,DNA was extracted by using modified CTAB method.The influences of template DNA concentration,primer concentration,dNTP concentration,Taq DNA polymerase amount and magnesium ion concentration on the amplification effects of RAPD of Taxus cuspidate were analyzed.[Result] Through analysis and comparison of various factors,the optimized reaction system was established as follows：in a total volume of 20 μl,containing 10 ng DNA,2.0 mmol/L Mg2＋,0.2 mmol/L dNTPs,15 pmol/μl random primer,1.0 U Taq polymerase,2 μl 10×buffer,and the supplementary ddH2O.[Conclusion] The research provided technical support for further discussion on the related studies with the sexual identification of Taxus cuspidate.

Chemical Constituents of One Species of Endophytic Fungus in Taxus chinensis

Three compounds were obtained from the mycelia of an endophytic fungus Gliocladium sp. (designated as strain F) of Taxus chinensis (Pilg.) Rehd. growing in Fujian Province, China. Their structures were determined on the basis of spectral analysis. (20S,22S)-4a-homo-22-hydroxy-4-oxaergosta-7,24(28)-dien-3-one was a novel compound. 4,8,12,16-tetramethyl-1,5,9,13-tetraoxacyclohexadecane-2,6,10,14-tetraone was firstly isolated from the genus ofGliocladium and 6,9-epoxyergosta-7,22-dien-3-ol was firstly obtained from the strain F.

Study on the Browning in Cell Suspension Culture of Taxus cuspidata

[Objective] This study aimed to investigate the browning of T. cuspidata cells in suspension culture and provide the guidance for the cell suspension culture of T. cuspidata. [Method] T. cuspidata callus was used as experimental materials, to explore the effect of different medium, N/P ratio, pH, shaking speed, illumination time and light intensity and other factors on browning of T. cuspidata cells in suspension culture. [Result] Non-browning callus was transferred to 2MB5 medium （pH 7.0） for illumination culture at 22℃ under light intensity of 1 500 lx with shaking speed of 90 r/min for 24 h. Results showed that the cell browning was significantly inhibited. [Conclusion] This study laid the foundation for cell suspension culture of T. cuspidata and had important significance to the large-scale industrial production of paclitaxel.

Cypher与Taxus支架治疗冠心病的冠状动脉造影随访结果

目的观察Cypher及Taxus药物洗脱支架治疗冠心病的疗效。方法应用冠状动脉造影随访162例接受Cypher及Taxus药物洗脱支架置入术的冠心病患者,其中Cypher组93例,Taxus组69例。结果Cypher组和Taxus组再狭窄率分别为10.8%和11.6%,靶病变再血管化率分别为5.4%和5.8%,两组比较差异均无统计学意义。支架内晚期管径丢失及支架节段晚期管径丢失在Cypher组低于Taxus组,两组间差异有统计学意义。结论药物洗脱支架可有效降低介入治疗术后再狭窄的发生率,但Cypher组支架内晚期管径丢失低于Taxus组。

A New Taxane Diterpenoid from the Seeds of Taxus yunnanensis

A new taxane diterpenoid with a rearranged 5/7/6-membered ring system was isolated from the seeds of the Chinese pew, Taxus yunnanensis. Its structure was established as 9 alpha-acetoxy-5 alpha-cinnamoy]-11(15 --> 1)-abeotaxa-4(20), 11-diene-10 beta, 13 alpha, 15-triol on the basis of spectroscopic analysis. Its relative stereochemistry is proposed from the results of a NOESY experment.

Determination of 10-Deacetyl baccatin Ⅲ and BaccatinⅢ in Taxus yunnanensis

报道了一种在Ｃ＿（１８）高效液相色谱柱上分离紫杉醇（Ｔａｘｏｌ）的前体化合物ＢａｃｃａｔｉｎⅢ和１０－去乙酰ＢａｃｃａｔｉｎⅢ的分析方法。所用流动相为甲醇－水（５２：４８），必要时可使用水－乙腈－四氢呋喃（６７：２５：８），流速１．０ｍｌ／ｍｉｎ，检测波长２２７ｎｍ。运用此方法对云南红豆杉不同部位（干皮、枝、叶、心木）的若干样品进行了测定，发现ＢａｃｃａｔｉｎⅢ和１０－去乙酰ＢａｃｃａｔｉｎⅢ在各部位间含量差异较大，即使同一部位也有很大差异。

Taxol Production in Suspension Cultures of Taxus chinensis as Affected by the Combination of Nutrient Feed with Dissolved Oxygen Control

Influence of the combination of nutrient feed with dissolved oxygen control on taxol production in suspension cultures of Taxus chinensis (Pilg.) Rehd. was investigated in bioreactors. Addition of feeding medium with 20 g/L sucrose on day 16 enhanced both the biomass and taxol production in 5-L bioreactors. Further investigation of the fed-batch cultures in a 20-L bioreactor showed that cultivation under a low dissolved oxygen tension (DOT) of 20% for the entire culture resulted in the highest taxol content of 0.98 mg/g DW, while the taxol production was lower than that with 40% and 60% DOT. Moreover, taxol accumulation was remarkably improved by the cultivation of cells initially under DOT of 60% for 20 d followed by changing the DOT to 20% for another 12 d. By combining the use of these two strategies, a maximum taxol content of 18.7 mg/L was obtained in a 20-L bioreactor.

Identification of Taxus cuspidata Sieb.et Zucc endophytic fungi─ new recorded-genus-species of China and the metabolite

A total of 94 isolates of endophytic fungi were isolated from the bark of 200-yr.-old Taxus cuspidata Sieb. et Zucc. in the primeval forest of the Changbai Mountain Natural Reserve, and 19 species of endophytic fungi were identified. Among the taxa 2 new species and 1 new varied species as well as the 6 known species of China had been described in a previous article published on Journal of Forestry Research, Vol. 14 (4): 290-294 (2003). This paper made a detail description on the morpho-logical characteristics of the 9 new recorded-genus-species and 1 new record species with illustrations, including Staphy-lotrichum coccosporum Meyer et Nicot, Botryodema lateritium Papendoorf & Upadhyay, Oedocephalum glomerulosum (Rul-liard) Sacc., Trichosporonoides oedocephalis Haskins & Spencer, Beniowskia sphaeroidea (Kal Chbrenner et Caeke) Mason, Thermomyces verrucosus Pugh, Blakeman & Morgan-jones, Gilmaniella humicola Brron , Arthrinium sphaerosperma (Per.:Fr.) Ellis, Ambrosiella brunnea (Verrall) Batra, and Cytosporina notha Diedz (1 new record species). And the habitats, hosts, distribu-tions and fruiting periods were recorded for the new recorded-genus-species. The determination result of thin layer chroma-tography (TLC) showed that the liquid fermentation metabolites of the strains could not react positively with taxoids developer - the vanillic aldehyde and dilute bismuth potassium iodide.

Cypher支架并不优于Taxus支架

一项临床试验表明，J&J公司的Cypher西罗莫司(sirolimus)药物涂膜冠状动脉支架(Ⅰ)在预防支架再狭窄方面并不优于Boston Scientific公司的Taxus紫杉醇(paclitaxel)药物涂膜支架(Ⅱ)。

Population structure and distribution pattern of Taxus cuspidata in Muling region of Heilongjiang Province, China

An investigation was conducted on distribution pattern, site condition and population structure of yew Taxus cuspidata Sieb. et Zucc. in Muling Forest Bureau of Heilongjiang Province, China in April, 2005. Results showed that yew is mainly distributed under the main storey of natural mixed forest of conifer and broadleaf, the soil moisture content of the yew site is high （40%-60%）, the pH value of soil is relatively lower （4.7-5.5）, and that the population structure of wild yew is not rational, belonging to the degeneration population, which is one of the reasons leading to the population decline. Although the site conditions of Muling area are suitable for the growth of wild yew, the population of wild yew shows a decline tendency, due to the fact that the middle-sized adult yew trees have been cut, young yews are often grazed by wildlife, and that the trunks of adult yew tend to be hollow.

Optimization of Cell Line Building for Taxus chinensis var. mairei

[Objective] The paper aims to optimize the abduction and domestication conditions of Taxus chinensis var.Mairei callus.[Method] We compared the efficiencies of callus between different explants and investigated the multiplication conditions of callus and suspended cell culture conditions with the buds,young stems and young leaves from T.chinensis var as the explants.[Results] The effect was the best with the bud as the explants; the best way for sterilizing the explants of T.chinensis var mairei was：streptomycin detergent for 2 h ＋ suds for 3 h ＋ 75% alcohol disinfection for 30 s ＋ 10% sodium hypochlorite solution for 25 min ＋ 1‰ mercury chloride for 10 min; the optimum formula of callus subculture was：B5 ＋ 4.0 mg/L NAA ＋ 0.5 mg/L 2,4-D ＋ 0.2 mg/L GA ＋ 0.5 mg/L 6-BA ＋ 2 g/L AC.[Conclusion] This research built the high efficient regeneration system of T.chinensis var.