Phylogenetic selection of Narcissus species for drug discovery

Plants are of immense importance in providing healthcare worldwide. With over 250 000 species of angiosperms alone, the potential for finding new medicinal plants and lead compounds for drug development is enormous. Some way of selecting plants for drug discovery programs is necessary. Phylogenies have great explanatory power and also enable a predictive perspective not offered by previous classifications of plants. Phylogenetic selection of target species is a new approach to drug discovery and the present study is the first attempt to correlate acetylcholine esterase (AChE) inhibitory activity and alkaloid distribution with a molecular phylogenetic hypothesis of Narcissus. The distribution of alkaloids with AChE inhibitory activity is significantly constrained by the phylogeny. Simultaneous evaluation of all available information of alkaloids and AChE inhibitory activity in a phylogenetic framework allowed us to discuss various strategies for selection of target species for further studies of AChE inhibitory activity.     

Protection of red blood cell acetylcholinesterase by oral huperzine A against ex vivo soman exposure: Next generation prophylaxis and sequestering of acetylcholinesterase over butyrylcholinesterase

As part of a phase Ib clinical trial to determine the tolerability and safety of the highly specific acetylcholinesterase (AChE) inhibitor huperzine A, twelve (12) healthy elderly individuals received an escalating dose regimen of huperzine A (100, 200, 300, and 400 μg doses, twice daily for a week at each dose), with three (3) individuals as controls receiving a placebo. Using the WRAIR whole blood cholinesterase assay, red blood cell AChE and plasma butyrylcholinesterase (BChE) were measured in unprocessed whole blood samples from the volunteers following each dose, and then for up to 48 h following the final and highest (400 μg) dose to monitor the profile of inhibition and recovery of AChE. Significant inhibition of AChE was observed, ranging from 30–40% after 100 μg to >50% at 400 μg, and peaking 1.5 h after the last dose. Gradual recovery of AChE activity then occurs, but even 48 h after the last dose red blood cell AChE was about 10% below control (pre-dose) values. Huperzine A levels in plasma peaked 1.5 h after the final 400 μg dose (5.47 ± 2.15 ng/mL). Plasma BChE was unaffected by huperzine A treatment (as expected).Aliquots of huperzine A-containing (from three individuals) and placebo blood samples were exposed ex vivo to the irreversible nerve agent soman (GD) for 10 min, followed by removal of unbound huperzine and soman from the blood by passing through a small C₁₈ reverse phase spin column. Eluted blood was diluted in buffer, and aliquots taken at various time intervals for AChE and BChE activity measurement to determine the time taken to achieve full return in activity of the free enzyme (dissociation from the active site of AChE by huperzine A), and thus the proportion of AChE that can be protected from soman exposure. Huperzine A-inhibited red blood cell (RBC) AChE activity was restored almost to the level that was initially inhibited by the drug. The increased doses of huperzine A used were well tolerated by these patients and in this ex vivo study sequestered more red blood cell AChE than has been previously demonstrated for pyridostigmine bromide (PB), indicating the potential improved prophylaxis against organophosphate (OP) poisoning.     

Potential therapeutic targets of huperzine A for Alzheimer's disease and vascular dementia

Huperzine A (HupA), a novel Lycopodium alkaloid isolated from Chinese folk medicine Huperzia serrata (Qian Ceng Ta), is a potent, selective and well-tolerated inhibitor of acetylcholinesterase (AChE). It has been proven to significantly improve the learning and memory impairment in Alzheimer's disease (AD) and vascular dementia (VaD) patients in China. Interestingly, our recent data indicate that HupA also possesses other protective functions. This paper will give an overview on the protective effects of HupA, which includes regulating beta-amyloid precursor protein (APP) metabolism, protecting against Abeta-mediated oxidative stress, apoptosis and mitochondrial dysfunction, as well as anti-inflammation. The multiple neuroprotective effects of HupA might yield additional beneficial effects in AD and VaD therapy.     

湘西蛇足石杉中石杉碱甲、乙和丙含量的测定

采用超声萃取法提取湘西蛇足石杉总生物碱,用高效液相色谱法同时测定其石杉碱甲、乙和丙含量,并分析其在植株不同部位的分布。结果表明:湘西4个样地蛇足石杉中石杉碱甲、乙和丙含量基本一致,分别达到0.5‰、0.3‰和0.04‰;但植株不同部位三种石杉碱含量差异显著,其中石杉碱甲和乙的分布均为叶>茎>根,石杉碱丙则是根大于叶和茎。由此可知,湘西蛇足石杉具有资源优势,石杉碱在植株的分布具有明显的部位选择性;采用HPLC可同时检测石杉碱甲、乙和丙,方法简单快速、准确可靠。     

他克林和双他克林抗星形孢菌素致凋亡作用的比较

用星形孢菌素（STS）诱导NG108-15细胞和HeLa细胞凋亡,观察他克林和双他克林是否具有抗凋亡作用.相差显微镜和Hoechst 33342染色观察细胞及胞核形态;噻唑蓝(MTT)测定分析细胞损伤状况;DNA提取及琼脂糖凝胶电泳观察凋亡特征性梯带;蛋白质印迹分析Bcl-2和Bax的表达水平.结果表明,经0.1 mmol/L他克林预处理后,由STS诱导的NG108-15细胞凋亡受到明显抑制.双他克林预处理无保护作用.蛋白质印迹分析表明他克林可显著抑制Bax的表达,同时还可促进Bcl-2的表达.他克林和双他克林预处理对STS诱导的HeLa细胞损伤无明显的保护作用.结论为：a.他克林对STS诱导的神经细胞损伤有显著的保护作用,但这种保护作用与其AChE抑制作用似乎没有明显关联.b.他克林对STS损伤的保护作用有细胞选择性.     

A revision of the Verticillium fungicola species complex and its affinity with the genus Lecanicillium

Verticillium fungicola, the type species of Verticillium sect. Albo-erecta, and related taxa were studied using morphological and molecular techniques. Sequences of the ITS region and SSU rDNA suggest that V. fungicola is very close to members of the genus Lecanicillium and unrelated to other species that were originally accommodated in the same section. New combinations in Lecanicillium are proposed for Verticillium fungicola and its var. flavidum at species rank, L. f. var. aleophilum is retained as a variety. These taxa can be distinguished from each other by optimum and maximum temperatures for growth, in addition to ITS sequence differences. Morphologically, L. flavidum is also distinct by repeated branching of the conidiophores, whereas the two varieties of L. fungicola have a simple conidiophore axis.     

The determination of huperzine A in European Lycopodiaceae species by HPLC-UV-MS

A rapid qualitative HPLC-UV-MS method was developed for the detection of huperzine A in Lycopodiaceae species. HPLC coupled with mass spectrometry experiments allowed the identification of the alkaloid and enabled targeted analysis of complex matrices such as herbal extracts. Huperzine A was detected by single ion monitoring of the pseudomolecular ion [M + H]+ at m/z 243.2. The alkaloid was also detected on TLC in a bioautographic enzyme assay with acetylcholinesterase to show the activity of the compound. Four Lycopodiaceae species collected in Switzerland were tested by these methods and Huperzia selago (L.) Bernh. ex Schrank et Martius was the only species found to contain huperzine A.     

Variation amongEucalyptus species in resistance to eucalyptus longhorned borer in Southern California

Eucalyptus L'Héritier (Myrtaceae: Leptospermoideae) species are native to the Austro-Malaysian region, but have been widely planted in temperate and subtropical regions around the world. In most regions whereEucalyptus have been imported, the Eucalyptus Longhorned Borer (Phoracantha semipunctata F.) (Coleoptera: Cerambycidae) has been accidently introduced. Larvae of the beetle bore through the bark and mine along the cambium of stressed trees, usually killing their host. We report here the relative susceptibilities of 12Eucalyptus species in two mixed-species plantations in California, USA. These trees were stressed by water deficit resulting from a prolonged drought.Eucalyptus species that appeared resistant to the borer includedE. camaldulensis Dehnhardt,E. cladocalyx F. Muller,e. sideroxylon A. Cunn. ex Woolls, andE. trabutii (anE. camaldulensis hybrid). Species that were more susceptible to attack wereE. diversicolor F. Mueller,E. globulus LaBillardière,E. grandis Hill ex Maiden,E. nitens (Deane & Maiden),E. saligna Sm., andE. viminalis LaBillardière. Survival of trees was influenced by fine-scale moisture variation resulting from slope and irrigation effects. Resistance characteristics of theseEucalyptus species did not correlate with taxonomic relatedness or bark characteristics, but did correspond to drought tolerance traits in their native habitat.Eucalyptus species that were resistant to attack byP. semipunctata were those that are most tolerant of drought in Australia.     

The influence of brain inflammation upon neuronal adenosine A2B receptors

Alzheimer's disease (AD) is associated with glial activation and increased levels of pro-inflammatory cytokines. Epidemiological results suggest that anti-inflammatory therapies can slow the onset of AD. Adenosine, acting at type-2 receptors, is an effective endogenous anti-inflammatory agent that can modulate inflammation both in the periphery and the brain. We investigated changes in the expression of adenosine type-2B (A2B) receptors and a related intracellular second messenger during chronic brain inflammation and following treatment with the non-steroidal anti-inflammatory drug flurbiprofen and its nitric oxide (NO)-donating derivative, HCT1026. Chronic infusion of lipopolysaccharide (LPS) into the 4th ventricle of young rats induced brain inflammation that was associated with microglial activation and reduced neuronal immunoreactivity for adenosine A2B receptors in the cortex. Daily administration of HCT1026, but not flurbiprofen, reduced microglial activation, prevented the down-regulation of A2B receptors and elevated tissue levels of cAMP. The results suggest that a therapy using an NO-releasing NSAID might significantly attenuate the processes that drive the pathology associated with AD and that this process may involve the activation of adenosine A2B receptors.     

Variation within species and inter-species comparison of seed dormancy and germination of four annual Lamium species

In an ecological context, knowledge of intra-species variation in dormancy and germination is necessary both for practical and theoretical reasons. We used four or five seed batches (replicates) of four closely related annuals co-occurring in arable fields in Sweden: Lamium amplexicaule, L. confertum, L. hybridum and L. purpureum. Seeds used for experiments stemmed from plants cultivated on two sites, each site harbouring one population of each species, thereby ensuring similar environmental history of seeds. Seeds were tested for germination when fresh and after three different pre-treatments (cold or warm stratification, or dry storage) for up to 24 weeks. Seeds were also sown outdoors. Despite substantial intra-species variation, there were clear differences between species. The general seed dormancy pattern, i.e. which environmental circumstances that affect dormancy, was similar for all species; dormancy reduction occurred during warm stratification or dry storage. Even though the response to warm stratification indicates a winter annual pattern, successful plants in Sweden were mostly spring emerged. Germination in autumn occurred, but plants survived winters poorly. Consequently, as cold stratification did not reduce dormancy, strong dormancy in combination with dormancy reduction during dry periods might explain spring germination. It is hypothesised that local adaptations occur through changes mainly in dormancy strength, i.e. how much effort is needed to reduce dormancy. Strong dormancy restricts the part of each seed batch that germinate during autumn, and thus reduces the risk of winter mortality, in Sweden.     

海三棱藨草及其近缘种基因组大小的测定

基因组大小是物种基因组的重要特征,通常用DNA C值来衡量,能够用于快速判断基因组倍性,并为分类学与进化生物学提供重要依据。海三棱藨草(Scirpus mariqueter)是长江口和杭州湾具有重要生态意义的标志性物种,被认为是扁秆藨草(S. planiculmis)和藨草(S. triqueter)的杂交种,因染色体小而难以准确确定倍性。近年来,部分研究者指出该物种的分类和命名存在疑点。该研究通过基因组Survey分析检测海三棱藨草样本CJ1的基因组特征,测序深度约为120 ×,并以绿豆(Vigna radiata)为参考标准,利用流式细胞术测定了海三棱藨草及其同域近缘种扁秆藨草和藨草以及海三棱藨草和扁秆藨草的杂交F1共13个样本的DNA C值和相对倍性。结果表明:(1)基因组Survey分析测得CJ1的基因组大小为244.12 Mbp,杂合率为0.68%,重复序列比例为42.38%,GC含量为37.25%。(2)流式细胞术测得来自不同区域的海三棱藨草各样本的基因组倍性相同,1C值在234.87 ～ 242.5 Mbp之间,其中CJ1的基因组大小与基因组Survey检测结果高度一致。(3)扁秆藨草的1C值在251.77 ～ 264.13 Mbp之间,藨草1C值为537.33 Mbp。根据上述基因组大小,认为海三棱藨草不可能是这两者的杂交种。该研究补充了海三棱藨草及其近缘种的基因组特征,为后续全基因组测序奠定基础,同时也否定了海三棱藨草起源于扁杆藨草和藨草杂交的假说。     

香花油茶——山茶属短柱茶组一新种

描述了山茶属短柱茶组新种Camellia osmantha Ye CX,Ma JL et Ye H.新种为灌木,花白色,微有香气.其苞被不分化为苞片和萼片,多数,10～12枚,花瓣6～8枚,均易脱落;雄蕊长短不一,大部分分离;花柱短,基部连生,先端分离,与短柱茶组特征一致.新种花有香气与窄叶短柱茶C.fluviantlis Hand.-Mzt.接近,但前者叶常为倒卵形,具尾尖,后者叶狭披针形,渐尖.     

Metabolic Processes in Alzheimer''s Disease: Adenine Nucleotide Content and Production of 14CO2 from [U-14C]Glucose In Vitro in Human Neocortex

Samples of neocortex removed at diagnostic craniotomy from patients with Alzheimer's disease and incubated in vitro showed an increased production of 14CO2 from [U-14C]glucose compared with neurosurgical controls. This was a feature of incubations in the presence of both 5 mM K+ (142% control) and 31 mM K+ (126%). Specific labelling of the amino acid pool was unaltered, suggesting that the apparent increase of CO2 production was not merely a reflection of changes in dilution of the radiolabel from glucose. The content of adenine nucleotides was significantly less than control values in the tissue from patients with Alzheimer's disease after in vitro incubations but the adenylate energy charge was unchanged, indicating that normal energy metabolism was not grossly impaired in these preparations.     

Regional brain metabolism with cytochrome c oxidase histochemistry in a PS1/A246E mouse model of autosomal dominant Alzheimer's disease: Correlations with behavior and oxidative stress

Mitochondrial dysfunction and brain metabolic alteration are early neurofunctional aspects in Alzheimer's disease (AD). Regional brain metabolism was analyzed by cytochrome c oxidase (COX) histochemistry in PS1-A246E mouse mutants, a model of autosomal dominant AD overexpressing beta-amyloid (Aβ) peptide without amyloidosis or cell degeneration. Immunohistochemical samples were analyzed on adjacent sections for regional Aβ1-42 levels, as well as DNA oxidative damage with 8-hydroxy-2-deoxyguanosine (8-OHdG). COX activity increased in the basal forebrain nuclear complex, specific parts of the amygdala and hippocampus, as well as in striatum and connected regions. On the contrary, a hypometabolism was observed in midline thalamic, interpeduncular, and pedonculopontine nuclei. The integration of these regions in circuitries subserving emotions, arousal, and cognitive functions may explain why neurochemical alterations in specific brain regions were linearly correlated with psychomotor slowing and disinhibition previously reported in the mutant. As the PS1-A246E model appears to mimick prodromal AD, the results support the existence of mitochondrial abnormalities prior to AD-related cognitive deficits. However, since affected PS1-A246E brain regions were not primarily those altered in AD-associated histopathological features and did not systematically display either Aβ overexpression or higher 8-OHdG immunolabelling, the hypermetabolism observed seems to comprise a compensatory reaction to early mitochondrial abnormalities; furthermore, neuronal synaptic function should be considered as particularly relevant in COX activity changes.     

Variation of individual electromorphs in Microtus pennsylvanicus and Peromyscus leucopus

Blood samples were taken from individual meadow voles (Microtus pennsylvanicus) and white-footed mice (Peromyscus leucopus) every three weeks. Two serum proteins, albumin and transferrin, were examined using acrylamide electrophoresis and densitometry. In individuals of both species, electrophoretic mobility and banding pattern changed. Such changes have been documented in other proteins, and may reflect changes in the internal or external environment of the individuals. Individual changes in serum proteins may seriously affect their interpretation, when used as genetic markers in population studies.     

Pseudohypericin and hyperforin in two Turkish Hypericum species: Variation among plant parts and phenological stages

The genus Hypericum has received considerable interest from scientists, as it contains the variety of structurally diverse natural products which possess a wide array of biological properties, mainly hypericins and hyperforin. In the present study, variations of pseudohypericin and hyperforin were investigated in two Turkish species of Hypericum, namely Hypericum perfoliatum and Hypericum origanifolium. Wild growing plants were harvested at vegetative, floral budding, flowering, fresh fruiting and mature fruiting stages, and dissected into stem, leaf and reproductive tissues and assayed for chemical contents by high performance liquid chromatography method. Content of pseudohypericin and hyperforin in samples of the whole plant increased during the course of ontogenesis in both species. The highest levels of the chemicals were reached at full flowering (2.62 mg/g dry weight (DW) pseudohypericin and 1.84 mg/g DW hyperforin for H. perfoliatum; 0.93 mg/g DW pseudohypericin and 1.63 mg/g DW hyperforin for H. origanifolium). Among different reproductive parts, full opened flowers produced the highest amount of pseudohypericin (1.18 mg/g DW) and hyperforin (4.36 mg/g DW) in H. origanifolium. Similarly, the highest pseudohypericin accumulation was observed in full opened flowers in H. perfoliatum (7.41 mg/g DW) while floral buds of this species produced the highest amount of hyperforin (7.80 mg/g DW). These data can be useful when elucidating the medicinal properties of the species and the chemosystematic significance of hyperforin and pseudohypericin in the relationships among species of Hypericum.     

Evaluation of Beauveria bassiana and B. brongniartii strains and four wild-type fungal species against adults of Bactrocera oleae and Ceratitis capitata

The virulence of two isolates of the hyphomycete fungi, Beauveria bassianaand B. brongniartii, and additional fungal species isolated from diseased Bactrocera oleae pupae and Sesamia nonagrioideslarvae were assessed against adults of the olive fruit fly B. oleae and the Mediterranean fruit fly Ceratitis capitata (Diptera: Tephritidae). Contact and oral bioassays revealed that moderate to high mortality rates for the olive fruit fly occurred when the adults were exposed to conidia of Mucor hiemalis, Penicillium aurantiogriseum, P. chrysogenum and B. bassianaisolates. A strain of M. hiemalis isolated from S. nonagrioides larvae was the most toxic resulting in 85.2% mortality to the olive fruit fly adults. B. brongniartiiand B. bassiana were the most pathogenic to the C. capitataadults causing 97.4 and 85.6% mortality. Metabolites collected from the M. hiemalis and P. chrysogenum isolates were toxic to adults of both species.     

Phylogeny and biogeography of Chinese and Australasian Polystichum ferns as inferred from chloroplast trnL-F and rps4-trnS sequence data

Polystichum, one of the largest genera of ferns, occurs worldwide with the greatest diversity in southwest China and adjacent regions. Although there have been studies of Chinese Polystichum on its traditional classification, geographic distributions, and even a few on its molecular systematics, its relationships to other species outside China remain little known. Here, we investigated the phylogeny and biogeography of the Polystichum species from China and Australasia. The evolutionary relationships among 42 Polystichum species found in China (29 taxa) and Australasia (13 taxa) were inferred from phylogenetic analyses of two chloroplast DNA sequence data sets: rps4-trnS and trnL-F intergenic spacers. The divergence time between Chinese and Australasian Polystichum was estimated. The results indicated that the Australasian species comprise a monophyletic group that is nested within the Chinese diversity, and that the New Zealand species are likewise a monophyletic group nested within the Australasian species. The divergence time estimates suggested that Chinese Polystichum migrated into Australasia from around 40 Ma ago, and from there to New Zealand from about 14 Ma. The diversification of the New Zealand Polystichum species began about 10 Ma. These results indicated that Polystichum probably originated in eastern Asia and migrated into Australasia: first into Australia and then into New Zealand.