氟中毒对家蚕幼虫血淋巴中γ-谷氨酰转肽酶活性的影响

试验测定了家蚕5龄幼虫血淋巴中γ-谷氨酰转肽酶(γ-GT)的活性和不同浓度NaF溶液添食处理对γ-GT活性的影响.结果表明,血淋巴中γ-GT的活性在5龄中期活性最强,龄初和龄末活性较弱.NaF溶液添食后γ-GT的活性明显增强,NaF溶液浓度越高,酶的活性越强.     

氟中毒对家蚕幼虫血淋巴中γ-谷氨酰转肽酶活性的影响

本试验测定了家蚕5龄幼虫血淋巴中γ-谷氨酰转肽酶(γ-GT)的活性和不同浓度NaF溶液添食处理对γ-GT活性的影响.结果表明,血淋巴中γ-GT的活性在5龄中期活性最强,龄初和龄末活性较弱.NaF溶液添食后γ-GT的活性明显增强,NaF溶液浓度越高,酶的活性越强.     

强痛宁对大鼠中枢脑区Ca~(2+)、Mg~(2+)-ATP酶活性影响

通过测定强痛宁作用下大鼠中枢脑区Ca~(2+),Mg~(2+)-ATP酶活性变化,探讨其麻醉镇痛作用与中枢脑区Ca~(2+),Mg~(2+)-ATP酶相关性。将24只纯种SD大鼠随机分成对照组、诱导组、麻醉组和催醒组,于各时间点冰上采集大鼠各脑区,采用比色法测定Ca~(2+),Mg~(2+)-ATP酶活性。结果表明:药物作用引起诱导期及麻醉期大鼠大脑皮质、小脑、海马、丘脑及脑干区Ca~(2+),Mg~(2+)-ATP酶活性降低,与对照组比较差异显著(P0.01或P0.05);催醒期各脑区Ca~(2+),Mg~(2+)-ATP酶活性恢复到麻前水平。结果提示,强痛宁作用下引起大鼠各脑区Ca~(2+),Mg~(2+)-ATP酶活性降低,阻碍了神经细胞对痛觉刺激信息传导,产生麻醉镇痛作用。     

NO 介导的 Ca2＋信号在干旱胁迫下紫花苜蓿种子萌发及抗氧化酶中的传导作用研究

本试验以紫花苜蓿种子为试验材料,添加外源一氧化氮供体硝普钠(SNP)、CaCl_2及抑制剂亚甲基蓝(methylene blue,MB)和LaCl_3,对种子进行浸种处理,以研究NO介导的Ca~(2+)信号在干旱胁迫下紫花苜蓿种子萌发及抗氧化酶中的传导作用。结果表明,15%PEG胁迫下紫花苜蓿种子萌发受到明显抑制,当外源添加NO或Ca~(2+)处理后萌发指标均有上升,外施0.1mmol/L SNP或10mmol/L CaCl_2都能有效缓解PEG对紫花苜蓿种子的胁迫伤害。干旱胁迫下NO+Ca~(2+)共处理时效果最为显著,萌发率较SNP处理提高了8.96%,较CaCl_2处理提高了19.67%。共处理时比SNP、CaCl_2处理时提高了种子淀粉酶活性、淀粉含量、可溶性糖、可溶性蛋白及脯氨酸含量,降低了MDA含量和超氧阴离子产生速率,显著提高了SOD,POD,CAT活性。其中淀粉酶活性、淀粉含量、可溶性蛋白、可溶性糖、脯氨酸含量以及POD活性的变化中,均表现出:NO和Ca~(2+)共处理下各指标变化要慢于单一处理。当添加外源NO的同时添加Ca~(2+)通道抑制剂La~(3+),NO的促进效果受到抑制,而添加外源Ca~(2+)的同时添加NO抑制剂亚甲基蓝,Ca~(2+)的促进效果受到抑制,表明NO经由Ca~(2+)信号通路调控干旱胁迫下紫花苜蓿的信号传导。     

家蚕氟化物中毒后血淋巴中脂质过氧化物和抗氧化物含量及抗氧化酶活性的变化

为了探究氟化物对家蚕的毒理作用,通过给家蚕5龄期幼虫添食NaF,检测氟化物中毒家蚕幼虫血淋巴中脂质过氧化物质丙二醛(MDA)、抗氧化物质谷胱甘肽(GSH)的含量变化以及谷胱甘肽过氧化物酶(GSH-Px)、谷胱甘肽硫转移酶(GST)等抗氧化物酶的活性变化。正常家蚕幼虫和氟化物中毒家蚕幼虫血淋巴中的MDA含量在5龄期均逐渐下降,但氟化物中毒家蚕的MDA平均浓度比正常家蚕升高了83.77%;GSH含量变化趋势为5龄第1天和第5天较高,第3天较低,但氟化物中毒家蚕5龄期的GSH浓度平均比正常家蚕下降了47.81%;氟化物中毒家蚕5龄期的GSH-Px和GST的平均活性分别比正常家蚕升高63.49%、39.22%,5龄第3天达到最高峰,5龄第1天和第5天较弱。初步分析认为家蚕幼虫血淋巴中MDA和GSH的含量变化以及GSH-Px和GST的活性变化,与家蚕氟化物中毒后体内氧自由基含量上升和脂质过氧化作用增强密切相关,可作为诊断家蚕氟化物中毒的生化指标。     

CaCl_2对干旱胁迫下苜蓿幼苗营养器官离子含量的影响

为了研究CaCl_2对干旱胁迫下苜蓿幼苗根、茎和叶离子含量的变化,试验采用水培方法测定了CaCl_2缓解聚乙二醇(PEG)胁迫下草原1号苜蓿幼苗不同营养器官中Na~+、K~+、Ca~(2+)、Mg~(2+)含量。结果表明:在20%PEG胁迫下,苜蓿幼苗根、茎、叶中Na~+含量增加,K~+、Ca~(2+)、Mg~(2+)含量减少。随着CaCl_2浓度的升高,Na+含量出现先降低后升高的变化趋势,K~+、Ca~(2+)(除根外)和Mg~(2+)含量均出现先增加后降低的变化趋势。根、茎、叶中Na+含量在CaCl_2浓度为10 mmol/L时最低,K~+、Ca~(2+)(除根外)和Mg~(2+)含量均在CaCl_2浓度为10 mmol/L最高。说明Ca~(2+)可改善苜蓿幼苗体内的离子平衡,缓解干旱胁迫造成的伤害,CaCl_2浓度为10 mmol/L时表现出对PEG渗透胁迫的较好缓解效应。     

家蚕氟中毒后中肠NADPH-细胞色素P450还原酶和NADPH-细胞色素C还原酶活性的变化

为了探究家蚕对NaF的代谢机制,以家蚕耐氟品种T6和氟化物敏感品种734为材料,从5龄起蚕开始分别添食50、100、200、400 mg/kg NaF溶液处理后的桑叶,检测蚕体中肠微粒体酶液中的黄素蛋白NADPH-细胞色素P450还原酶(CPR)和NADPH-细胞色素C还原酶(CR)的活性变化。氟物化敏感品种734的4个NaF处理组第3天的中肠CPR活性低于对照组,其余时间几乎都高于对照组,400 mg/kg NaF处理组在第4天的CPR活性最高,且各NaF处理组的CPR活性差异显著(P<0.05);耐氟品种T6 NaF处理组和对照组的中肠CPR活性整体上呈先升后降的趋势,几乎都在第2天达到最高值,各组之间的差异不显著(P>0.05)。氟化物敏感品种734的4个NaF处理组的中肠CR活性呈现先升后降的趋势,而对照组呈下降趋势;耐氟品种T6的50、100、400 mg/kg NaF处理组在第1～2天的中肠CR活性呈明显下降趋势,之后的变化相对较小,对照组的CR活性仅在第3～4天略高于NaF处理组,而其余时间NaF处理组的CR活性较高。2个家蚕品种添食不同浓度NaF后的中肠CR活性差异均不显著(P>0.05)。试验结果显示,耐氟品种T6在NaF作用下中肠的CPR和CR活性变化范围(分别为对照组的0.4～2.0倍和0.3～2.9倍)远小于氟化物敏感品种734这2种酶的活性变化范围(分别为对照组的0.6～9.3倍和0.4～4.6倍)。初步推测CPR和CR与家蚕对氟化物代谢具有一定的关联。     

氟中毒家蚕幼虫血淋巴中谷胱甘肽过氧化物酶的活性

为了解家蚕氟中毒的机理 ,研究了氟中毒家蚕幼虫血淋巴中谷胱甘肽过氧化物酶 (GSH—Px)的活性。正常的家蚕 5龄幼虫血淋巴中GSH—Px活性在龄中期较强 ,龄初和龄末较弱。幼虫添食 10mmol/L的氟化钠溶液后 ,GSH—Px的活性显著增强     

新烟碱类杀虫剂吡虫啉对家蚕幼虫生长发育及食物利用的影响

吡虫啉(imidacloprid)是新烟碱类杀虫剂,在农林害虫防治中广泛使用。在已明确吡虫啉对家蚕的毒性基础上,采用质量法研究吡虫啉导致的家蚕慢性中毒对幼虫生长发育及食物利用的影响,并采用酶活力测定法分析吡虫啉对5龄幼虫中肠3种消化酶(淀粉酶、蔗糖酶、海藻糖酶)活性的影响。添食高浓度(10~(-1)和10~(-2)mg/L)吡虫啉药液对家蚕3龄和5龄幼虫的生长发育及食物利用有明显抑制作用,幼虫的体质量及其增加量、相对生长率、取食量、相对取食量、食物利用率、食物转化率均显著降低(P≤0.05),而近似消化率显著升高(P≤0.05);添食低浓度(10~(-3)、10~(-4)和10~(-5)mg/L)吡虫啉药液对家蚕3龄和5龄幼虫的生长发育及食物利用则有促进作用,幼虫的取食量、相对取食量、消化量、食物利用率和近似消化率均显著高于对照(P≤0.05),同时体质量及其增加量、相对生长率也升高。添食低浓度吡虫啉药液对5龄幼虫中肠内淀粉酶和海藻糖酶的活性起促进作用;而添食高浓度吡虫啉药液在幼虫5龄1~5 d时对中肠内淀粉酶和海藻糖酶的活性起抑制作用,随着幼虫的生长发育,酶活性逐渐恢复正常并高于对照。吡虫啉对5龄幼虫中肠的蔗糖酶活性具有抑制作用,且添食药液浓度越大,抑制作用越明显,至5龄后期又恢复到正常水平或高于对照。试验结果表明,家蚕添食不同浓度吡虫啉药液后的慢性中毒过程中,蚕体的生长发育、取食行为、对食物的利用和消化吸收等均发生了显著变化。     

家蚕谷胱甘肽-S-转移酶基因(BmGSTd1)的诱导表达定量分析

谷胱甘肽-S-转移酶(GST)在机体的解毒代谢和抗氧化中起重要作用。为探究家蚕谷胱甘肽-S-转移酶基因(BmGSTd1)在家蚕体内的解毒和抗性功能,采用实时荧光定量PCR方法,对BmGSTd1基因在正常饲养及添食NaF家蚕5龄幼虫不同组织中的转录水平进行检测,并采用家蚕Actin3、GAPDH、28S rRNA 3种内参照基因对检测结果进行归一化处理。BmGSTd1基因在家蚕5龄幼虫各组织的转录水平存在差异,且采用不同内参照基因结果不一致:分别以家蚕Actin3、GAPDH、28S rRNA基因为内参照,该基因转录水平最高的组织分别为中肠、丝腺、脂肪体。5龄第2天幼虫添食NaF对体内各组织中BmGSTd1基因的转录水平具有诱导作用,且在不同组织中诱导表达的情况不同,采用以上3种内参照基因进行归一化处理的结果数据表明,中肠和脂肪体组织中的诱导转录水平最高,可能与这2种组织是家蚕主要的解毒器官有关。研究认为,检测基因在不同组织中的转录水平,采用合适的内参照基因对保证检测结果的可靠性非常重要。     

氟中毒对桑蚕碱性磷酸酶活性及蚕体重的影响

本文通过添食NaF的方法，探讨了氟中毒对天体重及中肠组织ALKP酶活性的影响，蚕品种之间的耐氟性存在显著差异，氟中毒后蚕体重显著降低，NaF对中肠组织ALKP酶有明显的抑制作用，氟中毒后蚕体重变化与中肠组织ALKP酶活性的变化相关性密切，具有同步性，1％Ca^2 对氟化物抑制作用有一定的缓解作用。     

Electrolyte composition of mink (Mustela vison) erythrocytes and active cation transporters of the cell membrane

Red blood cells from mink (Mustela vison) were characterized with respect to their electrolyte content and their cell membranes with respect to enzymatic activity for cation transport. The intra- and extracellular concentrations of Na+, K+, Cl-, Ca2+ and Mg2+ were determined in erythrocytes and plasma, respectively. Plasma and red cell water content was determined, and molal electrolyte concentrations were calculated. Red cells from male adult mink appeared to be of the low-K+, high-Na+ type as seen in other carnivorous species. The intracellular K+ concentration is slightly higher than the extracellular one and the plasma-to-cell chemical gradient for Na+ is weak, though even the molal concentrations may differ significantly. Consistent with the high intracellular Na+ and low K+ concentrations, a very low or no ouabain-sensitive Na+, K(+)-ATPase activity and no K(+)-activated pNPPase activity were found in the plasma membrane fraction from red cells. The Cl- and Mg2+ concentrations expressed per liter cell water were significantly higher in red cells than in plasma whereas the opposite was the case with Ca2+. The distribution of Cl- thus does not seem compatible with an inside-negative membrane potential in mink erythrocytes. In spite of a steep calcium gradient across the red cell membrane, neither a calmodulin-activated Ca(2+)-ATPase activity nor an ATP-activated Ca(2+)-pNPPase activity were detectable in the plasma membrane fraction. The origin of a supposed primary Ca2+ gradient for sustaining of osmotic balance thus seems uncertain.     

A metallo phosphatase activity present on the surface of Trypanosoma brucei procyclic forms

In this work, we describe how living cells of Trypanosoma brucei procyclic forms were able to hydrolyze extracellular p-nitrophenylphosphate (pNPP). These intact parasites, which had their viability determined by motility and the Trypan blue method, presented a low level of pNPP hydrolysis in the absence of any divalent metal (0.72+/-0.07 nmol pNP/mg min). Interestingly, in the presence of 5mM MgCl(2), ectophosphatase activity of 1.91+/-0.21 nmol pNP/mg min was observed. The ectophosphatase activity was also stimulated by MnCl(2), CoCl(2) and CuCl(2) but not by CaCl(2) and CdCl(2) and was inhibited by ZnCl(2). The addition of Mg(2+), Mn(2+), Co(2+) and Cu(2+) to extracellular medium increased the ectophosphatase activity in a dose-dependent manner. At 5mM pNPP, half-maximal stimulation of pNPP hydrolysis was obtained with 0.39+/-0.05 mM MgCl(2), 0.33+/-0.03 mM MnCl(2), 1.63+/-0.12 mM CoCl(2) and 2.04+/-0.33 mM CuCl(2). In the absence of any divalent metal (basal activity) the apparent K(m) for pNPP was 0.66+/-0.09 mM, while at saturating MgCl(2) concentrations the corresponding apparent K(m) for pNPP for Mg(2+)-stimulated phosphatase activity (difference between total minus basal phosphatase activity) was 0.27+/-0.03 mM. The Mg(2+)-stimulated pNPP hydrolysis was strongly inhibited by ZnCl(2) and vanadate, while the metal-independent basal phosphatase activity was less inhibited by these phosphotyrosyl phosphatase inhibitors.     

氟中毒对家蚕幼虫中肠组织糖原分解代谢调节的影响

用酶偶合系统测定了氟中毒对家蚕幼虫中肠组织糖原磷酸化酶活性的影响，其结果人氟中毒使中肠糖原磷酸化酶ａ型和ｂ型的活性均明显下降，特别是ｂ型的活性下降幅度较大。由此时以认为，氟中毒抑制了家蚕幼虫中肠组织内糖的分解代谢，使细胞的能量供应减少。     

添食氟化物对家蚕中肠组织细胞色素P450和b5含量的影响

为了探讨家蚕对NaF的代谢机制,以家蚕耐氟品种T6和敏感品种734为材料,5龄起蚕分别添食用200、400mg/LNaF溶液浸泡后的桑叶,检测蚕体内氧化酶系重要成员细胞色素P450和b5的含量变化。添食NaF后的168h内,耐氟品种T6中肠组织的细胞色素P450含量比添食清水对照组高3～7倍,细胞色素b5含量的增长与细胞色素P450含量的增长具有较高的相关性(R2=0.8220);添食不同浓度NaF溶液的差异不显著。添食NaF后的24～48h,敏感品种734中肠组织的细胞色素P450含量迅速增加,为添食清水对照组的1～2倍,随后逐渐下降,细胞色素b5含量的增长与细胞色素P450含量的增长无较好的线性关系(R2=0.4727);添食不同浓度NaF溶液的差异不显著。在氟化物作用下,耐氟品种T6中肠组织的细胞色素P450、b5含量显著增加,对氟化物敏感品种734中肠组织的细胞色素P450、b5含量变化相对较小,推测细胞色素P450和b5与家蚕对氟化物的代谢具有一定关联。     

影响家蚕中肠碱性磷酸酶活性的若干因素

研究了影响家蚕中肠碱性磷酸酶活性的若干因素,在供试的蚕品种间,酶的活性差异显著,且与茧质成绩呈正相关;NPV与CPV对中肠碱性磷酸酶活性的影响不同,即CPV感染明显降低了此酶的活性,而NPV则无其影响;苏云金杆菌感染直接破坏中肠组织,引起酶活性的陡降;各种化学物质中以Mg2+对中肠碱性磷酸酶的激活作用最明显,体外条件下维生素C,Ca2-,柠檬酸,Cl-对酶有抑制作用。     

Characterization of an ecto-ATPase of Tritrichomonas foetus.

In this work, we describe the ability of living Tritrichomonas foetus to hydrolyze extracellular ATP. The addition of MgCl(2) to the assay medium increased the ecto-ATPase activity in a dose-dependent manner. At 5mM ATP, half maximal stimulation of ATP hydrolysis was obtained with 0.46mM MgCl(2). The ecto-ATPase activity was also stimulated by MnCl(2) and CaCl(2), but not by SrCl(2). The Mg(2+)-dependent ATPase presents two apparent K(m) values for Mg-ATP(2-) (K(m1)=0.03 mM and K(m2)=2.01 mM). ATP was the best substrate for this enzyme, although other nucleotides such as ITP, CTP, UTP also produced high reaction rates. GTP produced a low reaction rate and ADP was not a substrate for this enzyme. The Mg(2+)-dependent ecto-ATPase activity was insensitive to inhibitors of other ATPase and phosphatase activities, such as oligomycin, sodium azide, bafilomycin A(1), ouabain, furosemide, vanadate, molybdate, sodium fluoride and levamizole. The acid phosphatase inhibitors (vanadate and molybdate) inhibited about 60-70% of the Mg(2+)-independent ecto-ATPase activity, suggesting that the ATP hydrolysis measured in the absence of any metal divalent could, at least in part, also be catalyzed by an ecto-phosphatase present in this cell. In order to confirm the observed Mg(2+)-dependent activity as an ecto-ATPase, we used an impermeant inhibitor, 4,4'-diisothiocyanostylbene-2',2'-disulfonic acid (DIDS) as well as suramin, an antagonist of P(2) purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg(2+)-dependent ATPase activity in a dose-dependent manner. This ecto-ATPase was stimulated by more than 90% by 50mM D-galactose. Since previous results showed that D-galactose exposed on the surface of host cells is involved with T. foetus adhesion, the Mg(2+)-dependent ecto-ATPase may be involved with cellular adhesion and possible pathogenicity.