Ca2+在水杨酸诱发的丹参培养细胞培养基碱化过程中的作用

利用质膜钙离子通道抑制剂LaCl3、异搏定(Verapamil,VP),钙离子载体A23187,内膜系统钙离子通道抑制剂2-APB和LiCl处理,研究水杨酸(SA)诱发的丹参培养细胞内Ca2+迸发在培养基碱化过程中的作用。结果显示:SA处理诱发丹参培养细胞培养基碱化,质膜钙离子通道抑制剂LaCl3和VP、内膜系统钙离子通道抑制剂2-APB和LiCl单独处理均可显著抑制SA处理诱发的培养基碱化过程,但质膜钙离子通道抑制剂对SA处理诱发的培养基碱化的抑制作用要显著强于内膜系统钙离子通道抑制剂;当两类钙离子通道抑制剂同时使用,培养基碱化过程被完全抑制,甚至培养基出现酸化趋势;钙离子载体A23187可以显著促进培养基碱化过程。以上结果说明,由水杨酸诱发的胞外Ca2+内流与胞内钙库Ca2+释放均参与了丹参培养基碱化的诱导过程,但胞外Ca2+内流的作用更重要。本研究揭示了SA诱发的Ca2+与丹参细胞培养基碱化之间的关系,为更深层次地阐明植物次生代谢调控机制提供理论基础。     

钙信使系统对机械刺激诱导的烟草悬浮培养细胞中H_2O_2爆发的调控

机械刺激可诱导烟草悬浮培养细胞H2O2的爆发,外源Ca2＋有强化作用,而Ca2＋螯合剂EGTA、质膜Ca2＋通道阻塞剂La3＋、胞内Ca2＋通道阻断剂钌红,以及钙调素拮抗剂氯丙嗪和三氟拉嗪则削弱机械刺激诱导的氧化爆发。这暗示以钙和钙调素为核心的钙信使系统对机械刺激诱发的烟草悬浮培养细胞中H2O2的爆发有调控作用。     

机械刺激诱导的烟草悬浮细胞一氧化氮产生途径及其与C矿和钙调素的关系

通过提高摇床转速对烟草细胞施加机械刺激（Ms）可诱导其胞内一氧化氮（No）的快速产生和一氧化氮合酶（Nos）活性的提高,这种MS诱导的NO产生可被N0清除剂cPTIO和NOS抑制剂L-NMMA显著抑制。此外,Ca2＋螯合剂EGTA、质膜Ca＋通道阻断剂La3＋、胞内Ca2＋通道拮抗剂钌红,以及钙调素抑制剂CPZ和TFP预处理均不同程度地抑制了机械刺激诱导的烟草细胞NO的产生,而机械刺激过程中钙调素活性显著上升并与NOS活性和NO含量的变化相一致。这些结果暗示着（类）Nos酶催化的精氨酸依赖途径可能是机械刺激诱发烟草细胞NO产生的主要途径,Ca2＋／CAM可能通过调节（类）NOS活性来调控No的产生。     

Ca~(2+)参与NO对蚕豆气孔运动的调控

观察了Ca2 + 、Ca2 + 的螯合剂和Ca2 + 通道抑制剂对NO调控的蚕豆气孔运动的影响。结果表明 ,NO的供体 1～ 10 0 μmol/LSNP (sodiumnitroprusside ,硝普纳 )可诱导气孔关闭 ;除去表皮条缓冲液中的Ca2 + 后 ,NO不再影响气孔的运动 ;Ca2 + 的螯合剂EGTA和BAPTA几乎可以完全抑制NO诱导的气孔关闭作用 ;胞内钙通道抑制剂钌红 (rutheniumred)和L型Ca2 + 通道阻断剂硝苯吡啶 (nifedipine)能够减弱SNP诱导气孔运动的关闭趋势 ;加入Ca2 + 通道抑制剂LaCl3 ,则外源NO失去其诱导气孔关闭的作用。说明在NO调控的气孔运动中 ,在NO信号途径的下游可能涉及来自胞内和胞外Ca2 + 的参与 ,并且胞外Ca2 + 更为重要。     

水杨酸诱发的NO介导了丹参悬浮培养细胞中丹酚酸B的生物合成

水杨酸(SA)可诱导丹参悬浮培养细胞中一氧化氮(NO)产生、苯丙氨酸解氨酶(PAL)活化及丹酚酸B(Sal B)的生物合成。为了阐明NO对丹参悬浮培养细胞中Sal B生物合成的影响及作用机理,本实验利用NO供体硝普钠(SNP)、NO合成酶抑制剂L-NNA(Nω-nitro-L-arginine)、NO淬灭剂c PITO(carboxy-2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide)以及PAL抑制剂L-AOPP(L-2-aminooxygen-3-phenyl acrylic acid)分别处理丹参悬浮培养细胞,并对其胞内NO水平、PAL活性和Sal B积累量进行了检测。结果表明,硝普钠(SNP)处理显著促进了NO产生、PAL活性和Sal B的积累,而L-NNA和c PITO抑制上述过程,说明NO诱发PAL活性提高并参与了SA诱导的Sal B生物合成;L-AOPP显著抑制了PAL活性及Sal B积累,却对NO产生没有显著影响,揭示NO位于PAL的上游。这说明SA诱发的NO产生、PAL活化及Sal B合成之间存在因果关系,即NO通过激活PAL触发Sal B生物合成。     

Ca2+和钙调素对H2O2诱导的玉米幼苗耐热性的调控

外源H2O2预处理提高了玉米幼苗内源H2O2的含量和钙调素(CaM)活性,缓解了高温处理过程中CaM活性的下降,增加了玉米幼苗在高温胁迫下的存活率.H2O2诱导的玉米幼苗耐热性的形成可被外源Ca2 处理所加强,被Ca2 螯合剂EGTA、质膜Ca2 通道阻塞剂La3 、胞内Ca2 通道阻断剂RR(钌红),以及CaM抑制剂CPZ(氯丙嗪)和TFP(三氟拉嗪)所抑制,表明Ca2 和CaM参与了H2O2诱导的玉米幼苗耐热性形成的调控.     

水杨酸诱发的丹参悬浮培养细胞内H2O2迸发与其培养基碱化的关系

水杨酸(salicylic acid,SA)处理可诱导丹参悬浮培养细胞内H2O2产生及其培养基碱化。利用NADPH氧化酶抑制剂咪唑(imidazole,IMD)、H2O2淬灭剂二甲基硫脲(dimethylthiourea,DMTU)、质膜H+-ATPase抑制剂钒酸钠(Na3VO4)及激活剂壳梭孢菌素(fusicoccin,FC)处理丹参悬浮培养细胞,探讨SA诱导的H2O2迸发与培养基碱化之间的关系。结果表明,H2O2可促发培养基碱化,IMD和DMTU抑制SA诱发的培养基碱化,说明H2O2参与SA诱发的培养基碱化过程;SA抑制质膜H+-ATPase活性,Na3VO4引发培养基碱化并使H2O2迸发时间提前,FC处理逆转了SA诱导的培养基碱化及H2O2迸发,说明质膜H+-ATPase调控培养基pH值变化,培养基碱化促进了H2O2产生。因此,丹参悬浮培养细胞内H2O2水平与其培养基碱化程度之间相互关联、共同作用,协同响应SA的诱导。     

Ca2+-CaM对过氧化氢诱导玉米幼苗耐冷性的影响

H2O2预处理可提高玉米幼苗的耐冷性及其体内钙调素(CaM)活性。阻断胞内Ca2 库的动员(钌红处理)、降低细胞中Ca2 水平(EGTA处理)及抑制CaM活性(TFP和CPZ处理)均可完全消除H2O2诱导的玉米幼苗的耐冷性。阻止胞外Ca2 跨膜进入胞内(La3 处理)并不抑制、甚至还能轻微地提高H2O2诱导的耐冷性。高Ca2 (20mmol.L^-1)处理削弱H2O2诱导的耐冷性。这些结果表明,CaM及胞内Ca2 库在H2O2诱导的玉米幼苗耐冷性的形成过程中起重要作用,而质外体中高浓度Ca2 和跨膜进入胞内会削弱H2O2诱导的耐冷性。     

NO体外供体硝普钠诱导蚕豆气孔保卫细胞死亡机理研究

为分析NO在植物细胞死亡过程中的作用,以蚕豆表皮条和NO体外供体硝普钠(SNP)及NO信号途径抑制剂为材料,采用表皮条生物法,探讨SNP对蚕豆叶面保卫细胞的毒性机理.结果表明:(1)0.5～9 mmol· L-1的SNP可使蚕豆气孔保卫细胞活性降低,部分细胞死亡,且随着SNP浓度的增高细胞死亡率增高.(2)凋亡抑制剂Z-Asp-CH2-DCB或TLCK可显著降低SNP诱发的保卫细胞死亡率.(3)抗坏血酸(AsA)、过氧化氢酶(CAT)、Ca2+螯合剂EGTA或Ca2+通道抑制剂LaCl3与SNP共同作用时,细胞死亡率显著降低.(4)NO清除剂c-PTIO、MAPK激酶抑制剂PD98059和鸟苷酸环化酶抑制荆ODQ亦能有效阻止SNP诱发的细胞死亡.研究发现,较高浓度的SNP可诱导蚕豆保卫细胞程序性死亡,SNP诱发植物细胞死亡与胁迫组保卫细胞内NO、ROS和Ca2+水平升高有关,cGMP和MAPK参与了SNP诱发的细胞死亡.     

丹参毛状根中水溶性酚酸类化合物的产生（简报）

由发根农杆菌（Agrobacteriumrhizogenes）ATCC15834诱导丹参叶片形成毛状根,经HPLC分析,证实其中含有原儿茶醛、丹酚酸A、丹酚酸B等酚酸类活性成分。其中丹酚酸A的含量为生药的2．17倍。     

酚酸类物质对三七幼苗的化感影响

该文研究了不同浓度的阿魏酸、对香豆酸、丁香酸、对羟基苯甲酸、香草酸5种酚酸类物质对三七幼苗生长和生理的影响。结果表明:处理后,三七幼苗的苗高、根长、可溶性蛋白质含量、根系活力、CAT以及POD活性均有所降低。其中,阿魏酸各处理组幼苗的苗高及POD活性均显著降低,50、100 mg·L~(-1)的对香豆酸以及100 mg·L~(-1)的香草酸处理组幼苗苗高也分别比对照显著降低16.19%、16.67%和29.29%;对香豆酸、丁香酸以及对羟基苯甲酸各处理组幼苗根长均显著低于对照;香草酸处理组幼苗的根系活力也显著低于对照,且幼苗的CAT活性在10、50、100 mg·L~(-1)丁香酸、对羟基苯甲酸以及香草酸处理下也达到了显著降低水平。此外,1 mg·L~(-1)阿魏酸以及100 mg·L~(-1)香草酸处理组幼苗的叶绿素含量也均显著降低;中高浓度的阿魏酸、对香豆酸、丁香酸、对羟基苯甲酸增加了三七幼苗的MDA含量,而香草酸在0.1、1、10、100 mg·L~(-1)浓度下显著降低幼苗的MDA含量;丁香酸、香草酸、对羟基苯甲酸以及中高浓度的对香豆酸增加了三七幼苗的SOD活性,且香草酸各处理组均达到了显著性水平。综上结果表明,5种酚酸类物质对三七幼苗均具有一定的化感抑制作用,但各酚酸物质的作用方式及强度并不完全一致,阿魏酸的化感影响较大,这为进一步研究三七的化感自毒作用提供了一定的理论参考。     

Effects of flaxseed,raw soybeans and calcium salts of fatty acids on apparent total tract digestibility,energy balance and milk fatty acid profile of transition cows

Oilseeds offer some protection to the access of ruminal microorganisms and may be an alternative to calcium salts of fatty acids (FA), which are not fully inert in the ruminal environment. This study aimed to evaluate the effects of different sources of FA supplementation on apparent total tract nutrient digestibility, milk yield and composition, and energy balance (EB) of cows during the transition period and early lactation. We compared diets rich in C18:2 and C18:3 FA. Multiparous Holstein cows were randomly assigned to receive one of the four diets: control (n=11); whole flaxseed (WF, n=10), 60 and 80 g/kg (diet dry matter (DM) basis) of WF during the prepartum and postpartum periods, respectively; whole raw soybeans (WS, n=10), 120 and 160 g/kg (diet DM basis) of WS during the prepartum and postpartum periods, respectively; and calcium salts of unsaturated fatty acids (CSFA, n=11), 24 and 32 g/kg (diet DM basis) of CSFA during the prepartum and postpartum periods, respectively. Dry cows fed WF had higher DM and net energy of lactation (NE_L) intake than those fed WS or CSFA. The FA supplementation did not alter DM and NDF apparent total tract digestibility, dry cows fed WF exhibited greater NDF total tract digestion than cows fed WS or CSFA. Feeding WS instead of CSFA did not alter NE_L intake and total tract digestion of nutrients, but increased milk fat yield and concentration. Calculated efficiency of milk yield was not altered by diets. FA supplementation increased EB during the postpartum period. Experimental diets increased long-chain FA (saturated and unsaturated FA) in milk. In addition, cows fed WS and CSFA had higher C18:1 trans-11 FA and C18:2 cis, and lower C18:3 FA in milk than those fed WF. Furthermore, cows fed CSFA had higher C18:1 trans-11 and cis-9, trans-11 FA than cows fed WS. Although supplemental C18:2 and C18:3 FA did not influence the milk yield of cows, they positively affected EB and increased unsaturated long-chain FA in milk fat.     

Identification of glutamine chain lengths of endogenous Folypoly-γ-glytamates in rat tissues

A simplified procedure for the determination of the glutamate chain lengths of endogenous tissue folate is described.Natural pteroylpoly-γ-glutamates in tissue extracts, irrespective of their one-carbon moiety, were converted by a reductive cleavage procedure to a homologous series of p-aminobenzoylpoly-γ-glutamates, differing only in glutamate chain length. Desalting and concentration of the extracts were achieved by absorbing the derivatives on active charcoal followed by their elution with ethanol: ammonia. Aminobenzoylpolyglutamates were separated by DEAE- cellulose chromatography and quantitated by a colorimetric procedure for primary aromatic amines.The major endogenous folates in rat liver and kidney were pteroylpentaglutamate derivatives with smaller amounts of pteroyltetra- and hexaglutamate.     

氨基酸L-和D-异构体对离体小麦胚植株生长的影响

5种氨基酸L-和D-异构体对离体小麦胚植株生长的影响结果表明,在1~7 mmol.L-1浓度范围内,脯氨酸异构体均不抑制植株生长;缬氨酸和蛋氨酸的L-型严重抑制生长,D-型不抑制或轻微抑制;丙氨酸则相反,L-型无抑制作用,D-型是严重抑制类型;丝氨酸的L-型轻微抑制生长,D-型严重抑制生长。生长结果也表明,DL-异构体的抑制作用介于D-型和L-型之间。     

氨基酸分析仪测定饲料添加剂——叶酸的方法

用６ｍｏｌ／Ｌ盐酸于１１０℃条件下水解饲料添加剂——叶酸,使之游离出谷氨酸,用氢氧化钠中和调节ｐＨ到２,氨基酸分析仪测定谷氨酸含量,经与标准叶酸水解样品比较,计算出叶酸的纯度。该方法重现性好,变异系数ＣＶ＝０．０８％,平均回收率为９８．３４％,浓度与峰面积呈线性相关,相关系数ｒ＝０．９９８７,可随氨基酸分析同时进行,不需改变任何分析条件。     

Separation and simultaneous determination of nalidixic acid, hydroxynalidixic acid and carboxynalidixic acid in serum and urine by micellar electrokinetic capillary chromatography

Separation in capillary electrophoresis is governed by various factors, including buffer type, buffer concentration, pH, temperature, voltage and micelles. Through proper adjustment of these parameters, nalidixic acid and its two major metabolites, 7-hydroxynalidixic and 7-carboxynalidixic, could be separated by micellar electrokinetic capillary chromatography using an electrophoretic electrolyte consisting of 50 mM borate buffer (pH 9) containing 25 mM sodium dodecyl sulphate and 10% acetonitrile. A linear relationship between concentration and peak area for each compound was obtained in the concentration range 0.15–100 μg ml⁻¹, with a correlation coefficient greater than 0.999 and detection limits in the 0.2–0.7 ng ml⁻¹ range. Intra- and inter-day precision values of about 0.8–1.2% RSD (n=11) and 1.3–2.0% RSD (n=30), respectively, were obtained. The method has been applied to the analysis of nalidixic acid and its two major metabolites in serum and urine with limits of sensitivity lower than 0.8 ng ml⁻¹.     

Characterization of Transgenic Rice Plants Expressing an Arabidopsis FAD7

Fatty acid -3 desaturase (FAD) is the key enzyme catalyzing the formation of trienoic fatty acids. We utilized an Arabidopsis FAD7 gene and the seven independent transgenic rice plants harbouring 1 to 3 copies of this gene were generated. The expression of FAD7 mRNA was different among independent transgenic lines regardless of the copy number. The total linolenic acid (18:3) contents reduced by about 7 – 32 % in transgenic rice plants but the linoleic acid (18:2) content increased accordingly. With or without wounding treatments, the jasmonate content was higher in transgenic lines than in wild-type rice plant. The transgenic lines overproducing jasmonate also showed increased expression of PR1b mRNA and allene oxide synthase inresponse to wounding.     

Achievements and challenges of sialic acid research

Sialic acids are one of the most important molecules of life, since they occupy the terminal position on macromolecules and cell membranes and are involved in many biological and pathological phenomena. The structures of sialic acids, comprising a family of over 40 neuraminic acid derivatives, have been elucidated. However, many aspects of the regulation of their metabolism at the enzyme and gene levels, as well as of their functions remain mysterious. Sialic acids play a dual role, not only are they indispensable for the protection to and adaptation of life, but are also utilised by life-threatening infectious microorganisms. In this article the present state of knowledge in sialobiology, with an emphasis on my personal experience in this research area, is outlined including a discussion of necessary future work in this fascinating field of cell biology.