Potential of the bean α‐amylase inhibitor αAI‐1 to inhibit α‐amylase activity in true bugs (Hemiptera)

True bugs (Hemiptera) are an important pest complex not controlled by Bt‐transgenic crops. An alternative source of resistance includes inhibitors of digestive enzymes, such as protease or amylase inhibitors. αAI‐1, an α‐amylase inhibitor from the common bean, inhibits gut‐associated α‐amylases of bruchid pests of grain legumes. Here we quantify the in vitro activity of α‐amylases of 12 hemipteran species from different taxonomic and functional groups and the in vitro inhibition of those α‐amylases by αAI‐1. α‐Amylase activity was detected in all species tested. However, susceptibility to αAI‐1 varied among the different groups. α‐Amylases of species in the Lygaeidae, Miridae and Nabidae were highly susceptible, whereas those in the Auchenorrhyncha (Cicadellidae, Membracidae) had a moderate susceptibility, and those in the Pentatomidae seemed to be tolerant to αAI‐1. The species with αAI‐1 susceptible α‐amylases represented families which include both important pest species but also predatory species. These findings suggest that αAI‐1‐expressing crops have potential to control true bugs in vivo.     

结核分枝杆菌感染小鼠的脾脏和肺脏组织荷菌量与病理变化

目的分析结核急性感染小鼠脾脏和肺脏的组织荷菌量以及病理的动态变化。方法以3×105CFU结核分枝杆菌标准株H37Rv尾静脉途径感染雌性C57BL/6J小鼠,测定感染后1 d、1周、2周、3周、4周、6周和8周肺脏及脾脏组织的荷菌量,脾脏和肺脏组织经HE染色分析病理变化。结果感染动物的脾脏荷菌量在前3周逐步提高,在4～8周脾脏荷菌量维持稳定,肺脏组织在2～8周组织荷菌量逐步升高。病理分析显示动物感染3周后肺脏和脾脏出现肉芽肿,在6～8周病理损伤加重。结论小鼠经尾静脉感染高剂量结核分枝杆菌在8周前表现为急性感染状态,适用于抗结核药物的体内药效学评价。     

小鼠结核分枝杆菌耐药模型的建立与评价

目的 建立小鼠结核分枝杆菌耐利福平株静脉感染小鼠模型.方法 30只BALB/c雌性小鼠经尾静脉感染结核分枝杆菌耐利福平株每只106 CFU,观察小鼠一般状况,并分别在感染后6周、10周、14周处死小鼠,进行脾、肺组织病理切片、抗酸染色、计脏器荷菌数.结果 感染后小鼠体重呈逐渐上升趋势,脏器病理变化随时间推移由急性炎症逐...     

Infection of H69AR cells with retroviral particles harboring interfering RNAi significantly reduced the multidrug resistance of these small cell lung cancer cells

Incubation of the drug-sensitive H69, a small cell lung cancer cell line, with increased concentrations of adriamycin yielded multidrug resistant (MDR) H69AR cells that over-express multidrug resistance-associated protein (MRP1). MRP1 co-transports its substrate with glutathione (GSH), leading to lower intracellular GSH. In this report we tested whether depleting intracellular GSH in MRP1-expressing cells could hyper-sensitize them to anticancer drugs or not. We have found that the GSH contents in MRP1-expressing cells are significantly lower than their corresponding control cells. The treatment with MRP1 substrate verapamil or the GSH synthetase inhibitor buthionine sulfoxi-mine significantly reduced the intracellular GSH contents in MRP1-expressing cells. Interestingly, depleting intracellular GSH contents can hyper-sensitize the MRP1-cDNA transfected BHK cells to daunomycin, but not the adriamycin-selected H69AR cells. Further analyses indicated that anti-apoptotic factor Bcl2 might be a factor responsible for the fact that depleting intracellular GSH could not hyper-sensitize H69AR cells to daunomycin. We hypothesized that knocking down the expression of Bcl2 could hyper-sensitize H69AR cells to daunomycin. Interestingly, infection of H69AR cells with retroviral particles harboring Bcl2 interfering RNAi not only reduced the expression of Bcl2, but also many factors that contribute to MDR, such as Bcl-xl, MRP1 and ABCC3, etc., leading to the MDR H69AR cells more sensitive to daunomycin than the parental H69 cell. Thus, although the mechanisms of the down-regulation of the genes contributing to MDR remain to be elucidated, retroviral particles harboring Bcl2 interfering RNAi could be used as an alternative way to sensitize the MDR cancer cells to anticancer drugs.     

2010年宜昌市铜绿假单胞菌的耐药性分析

了解宜昌市铜绿假单胞菌（Pseudomonas aeruginosa）临床分离株的耐药现状。宜昌市城区5所医院临床分离的铜绿假单胞菌菌株,用K-B法作药敏试验,并根据统计其耐药情况及耐药表型（模式）分析可能存在的耐药机制。临床分离的铜绿假单胞菌共1 575株,耐药率依次为阿米卡星7.1%、美罗培南17.2%、头孢吡肟20.4%、头孢哌酮/舒巴坦21.0%、哌拉西林/他唑巴坦22.5%、环丙沙星23.1%、庆大霉素23.4%、头孢他啶25.0%、亚胺培南25.2%、哌拉西林30.4%、氨曲南34.5%、复方新诺明59.0%、米诺环素75.6%。多重耐药（MDR）和泛耐药（PDR）株分别占41.5%和0.17%。对各种抗假单胞菌药物分别耐药的菌株仍有13%～25.7%对阿米卡星敏感,提示在严重铜绿假单胞菌感染患者的治疗中,β内酰胺类抗假单胞菌药加氨基糖苷类仍是一个很好的联合用药组合。细菌耐药性仍呈增长趋势,临床上感染多重耐药和泛耐药的铜绿假单胞菌的治疗仍很棘手,应合理使用抗生素,尽量延缓耐药菌株的出现。     

Regulation of Ergothioneine Biosynthesis and Its Effect on Mycobacterium tuberculosis Growth and Infectivity

Ergothioneine (EGT) is synthesized in mycobacteria, but limited knowledge exists regarding its synthesis, physiological role, and regulation. We have identified Rv3701c from Mycobacterium tuberculosis to encode for EgtD, a required histidine methyltransferase that catalyzes first biosynthesis step in EGT biosynthesis. EgtD was found to be phosphorylated by the serine/threonine protein kinase PknD. PknD phosphorylates EgtD both in vitro and in a cell-based system on Thr²¹³. The phosphomimetic (T213E) but not the phosphoablative (T213A) mutant of EgtD failed to restore EGT synthesis in a ΔegtD mutant. The findings together with observed elevated levels of EGT in a pknD transposon mutant during in vitro growth suggests that EgtD phosphorylation by PknD negatively regulates EGT biosynthesis. We further showed that EGT is required in a nutrient-starved model of persistence and is needed for long term infection of murine macrophages.     

蛋白酶抗性人睫状神经营养因子突变体基因的构建及其在巴斯德毕赤酵母中的表达

AX15是一种比天然睫状神经营养因子具有更高的生物学活性、更好的稳定性和可溶性的hCNTF突变体。在巴斯德毕赤酵母中表达时AX15易发生降解。氨基酸序列分析表明降解位于由12和13位氨基酸残基组成的双碱性氨基酸之后。根据KEX2蛋白酶的底物专一性把双碱性氨基酸从RR变为RK,构建了KEX2抗性的AX15突变体AX15（R13K）。AX15（R13K）的稳定性得到了显著的提高,在诱导100 h后也未发生降解。利用超滤浓缩和凝胶过滤得到了纯度>90%的AX15（R13K）。TF-1细胞存活实验表明AX15（R13K）具有与AX15相同的生物学活性。蛋白酶抗性人睫状神经营养因子突变体可能具有更好的体内稳定性,在临床应用上具有潜在的优势。     

玉蜀黍赤霉(Gibberella zeae)对多菌灵的室内抗药突变体的诱导及其抗药性遗传分析

通过紫外线照射和药剂驯化的方法均获得了玉蜀黍赤霉野生敏感菌株对多菌灵(carbendazim,MBC)的室内抗药突变体。这些抗药突变体一部分表现低抗(low resistance．LR),即能在临界致死剂量1．4μg／mL多菌灵浓度下生长,但不能在10μg／mL浓度以上生长,且对二氯苯胺甲酸甲酯(N-3,5-dichlorophenyl carbamate,MDPC)不表现负交互抗药性;另一部分表现高抗(high resistance．HR)．即能在100μg／mL多菌灵浓度下生长,并与田间高抗菌株一样,对MDPC表现负交互抗药性;没有获得类似田间的中抗(moderate resistance,MR)菌株,即能在10μg／mL多菌灵浓度下快速生长,在100μg／mL浓度以上被完全抑制的突变体。通过药剂驯化的方法还获得了田间中抗(MR)菌株的高抗(HR)突变体,但这些突变体与MR一样对MDPC仍然不表现负交互抗药性。抗药性遗传研究表明。在所研究的抗药突变体中,抗药性在自交和无性繁殖后代中能稳定遗传;室内抗药突变体和田间抗药菌株对多菌灵的抗药性由同一个主效基因控制,但它们发生突变的位点不同或者同一碱基位点发生了不同的突变;对MDPC的敏感性也是由单个基因控制的,该基因与控制多菌灵抗性的基因是等位基因,当该基因发生对MDPC的敏感性增加的突变时会使病菌对多菌灵产生高水平抗性。     

寡核苷酸芯片研究结核分枝杆菌临床株和无毒株诱导的巨噬细胞U937凋亡相关基因的差异表达

结核病仍然是人类健康的主要威胁,结核分枝杆菌诱导的巨噬细胞凋亡是宿主防御反应之一,研究凋亡相关基因的差异表达有助于认识结核分枝杆菌致病机理和发现新的药物靶标。利用包括19200个基因或基因片段的DNA芯片研究巨噬细胞株U937对临床和实验室菌株感染的差异表达,Northem blotting和RT—PCR验证了芯片研究结果。Mtb H37Rv感染下调bcl—2,vitaminD受体、干扰素调控因子3、细胞色素氧化酶C表达,幅度分别为2-,3-,3-,2．5-倍,临床菌株感染上调SOD2、SOD3、丝氨酸蛋白酶、toll—like受体2、signal transducer and activator(STAT1)、hypoxia—inducible factor22等表达,幅度分别为2．9-,2．5-,2．5-,22-,2．4-,5．9-倍。结果提示,临床菌株感染更多促进凋亡,限制宿主的杀灭机理。该研究为进一步研究导致这些差异表达的结核分枝杆菌成分提供了基础。