Development of a high-throughput screening assay for cytoprotective agents in rotenone-induced cell death

Parkinson’s disease (PD) is a neurodegenerative disease featured by selective loss of substantia nigra neurons. Rotenone administration in animals induces neurodegeneration accompanied by α-synuclein-positive Lewy body-like inclusions, recapturing typical histopathological features of PD. In an effort to screen for small-molecule agents to reverse rotenone-induced cytotoxicity, we developed and validated a sensitive and robust assay with neuroblastoma SK-N-SH cells. This assay was amenable to a high-throughput screening format with Z′ factor of 0.56. Robotic screening of a bioactive compound library led to the identification of carnosic acid that can effectively protect cells from rotenone treatment. Using a high-content image-based assay and Western blot analysis, we demonstrated that carnosic acid protects cells from rotenone stress by significant induction of HSP70 expression. Therefore, the assay reported here can be used to identify novel cytoprotective agents for clinical therapeutics of PD.     

A PORTABLE DIGITAL SOUND EMITTER FOR INTERACTIVE PLAYBACK OF ANIMAL VOCALISATIONS

ABSTRACT

A new portable digital sound emitter (DSE) for normal and interactive playback of sound signals in the field and in the laboratory is described together with two examples of applications of the DSE in interactive field experiments. The DSE may be loaded with a broad spectrum of digitised sounds, e.g. artificial signals or natural animal vocalisations, and it is controlled via the keyboard of a portable PC running a dedicated program. This program, which can be tailored to support a vast number of different demands, enables the experimenter to start and stop the analog output from the DSE at any time and to choose freely between the available sounds and playback modes. The use of the DSE for interactive playback experiments therefore is only limited by the ability of the operator to perceive the vocalisations of the participating animal and to operate the keyboard of the PC.     

Testing prediction accuracy in short-term ecological studies

Applied ecology is based on an assumption that a management action will result in a predicted outcome. Testing the prediction accuracy of ecological models is the most powerful way of evaluating the knowledge implicit in this cause-effect relationship, however, the prevalence of predictive modeling and prediction testing are spreading slowly in ecology. The challenge of prediction testing is particularly acute for small-scale studies, because withholding data for prediction testing (e.g., via k-fold cross validation) can reduce model precision. However, by necessity small-scale studies are common. We use one such study that explored small mammal abundance along an elevational gradient to test prediction accuracy of models with varying degrees of information content. For each of three small mammal species, we conducted 5000 iterations of the following process: (1) randomly selected 75 % of the data to develop generalized linear models of species abundance that used detailed site measurements as covariates, (2) used an information theoretic approach to compare the top model with detailed covariates to habitat type-only and null models constructed with the same data, (3) tested those models’ ability to predict the 25 % of the randomly withheld data, and (4) evaluated prediction accuracy with a quadratic loss function. Detailed models fit the model-evaluation data best but had greater expected prediction error when predicting out-of-sample data relative to the habitat type models. Relationships between species and detailed site variables may be evident only within the framework of explicitly hierarchical analyses. We show that even with a small but relatively typical dataset (n = 28 sampling locations across 125 km over two years), researchers can effectively compare models with different information content and measure models’ predictive power, thus evaluating their own ecological understanding and defining the limits of their inferences. Identifying the appropriate scope of inference through prediction testing is ecologically valuable and is attainable even with small datasets.     

Diversity,dynamics and reproduction in a community of small mammals in Upper Guinea,with emphasis on pygmy mice ecology

As part of a large survey on reservoirs of Lassa fever in Guinea, three villages were investigated in high endemic zone, close to Sierra Leone border. Biodiversity of the small mammal community is presented in this study through a standardized trapping in houses, cultivations and forest. Identification of the small mammals was based on morphology and by molecular technique for sibling species. Of the 1123 specimens collected in 2003–2005, we identified seventeen species (thirteen Muridae, four Soricidae), leading to high diversity (Shannon index = 1.6–1.8) and high equitability (evenness index = 0.7–0.8) in cultivations and forest. In houses conversely, the rodent community was dominated by Mastomys natalensis (95–98%), leading to low diversity and equitability. Dynamics and reproduction were investigated in two species of pygmy mice, Mus mattheyi and Mus minutoides, two species of Praomys, P. daltoni and P. rostratus, and in Mastomys erythroleucus. The pygmy mice were abundant in cultivations in early rainy season, and reproduced from rainy to dry season. Praomys daltoni was also found more abundant in cultivations and seemed to reproduce between rainy and dry season, whereas P. rostratus preferred forest and cultivations in late rainy season, and reproduced throughout the year. Finally, M. erythroleucus was more abundant in forest in dry season, and seemed to reproduce from late rainy to dry season. This species had a low occurrence (6.5%) in the Faranah’s zone, and probably lived at its southern limit in Guinea. The presence of other Murinae, such as M. natalensis, Praomys spp as possible competitors in the same habitats, is discussed. For the first time, this study relates population biology of pygmy mice with molecular identification.     

Analysis of the developmental expression of small VCP-interacting protein and its interaction with steroidogenic acute regulatory protein in Leydig cells

Small VCP-interacting protein (SVIP) is a 9-kDa protein that is composed of 76 amino acids, and it plays a role in the endoplasmic reticulum-associated protein degradation (ERAD) pathway. Recent studies have shown that SVIP is an androgen-responsive protein and its expression is regulated by androgens. Because no data are available regarding the cellular localization and expression of SVIP in the mouse testis, where androgens are highly expressed, immunohistochemistry and western blotting were performed. In the fetal testis, we found that moderate but consistent staining of SVIP is present in the cytoplasm of Leydig cells. In prepubertal and adult life, SVIP remains present in Leydig cells as well as in the cytoplasm of some peritubular and Sertoli cells. From postnatal day 15 onward, SVIP is strongly expressed in the cytoplasm of Leydig cells.Furthermore, TM3, MA-10 Leydig and Sertoli cell lines were also used to evaluate the expression of SVIP. To identify the interacting partners, such as steroidogenic acute regulatory (STAR) protein, colocalization studies were performed by fluorescence microscopy, showing that STAR colocalized with SVIP in the adult mouse testis. The expression changes of STAR were studied by using SVIP siRNAs in Leydig cell line cultures. Depletion of SVIP resulted in decreased expression of STAR. Additionally, the number and size of lipid droplets were significantly increased in SVIP-depleted Leydig cells. Taken together, our data identify SVIP as a marker of Leydig cell lineage and as a regulator of STAR protein expression and lipid droplet status in Leydig cells.     

Pseudo-natural products and natural product-inspired methods in chemical biology and drug discovery

Through evolution, nature has provided natural products (NPs) as a rich source of diverse bioactive material. Many drug discovery programs have used nature as an inspiration for the design of NP-like compound classes. These concepts are guided by the prevalidated biological relevance of NPs while going beyond the limitations of nature to produce chemical matter that could have unexpected or novel bioactivities. Herein, we discuss, compare, and highlight recent examples of NP-inspired methods with a focus on the pseudo-NP concept.     

14种中国典型极小种群野生植物繁育特性和人工繁殖研究进展

繁殖是植物种群更新与维持的重要环节。包括极小种群野生植物在内的受威胁物种, 其濒危原因是在长期演化过程中自身繁育力的衰退、生活力的下降等内在因素和人类的过度采挖和生境的破坏等外在因素共同作用的结果。对极小种群野生植物进行高效的人工繁殖, 能扩大种群数量并应用于迁地保护、自然回归和满足商品市场的需求, 有利于其种质资源的保护和可持续利用。为了保持物种的遗传多样性, 采用种子繁殖育苗是有效的方法, 扦插、嫁接和组织培养技术等无性繁殖方法则可用于对难以用种子繁殖的种类进行快速繁殖。本文对14种中国典型极小种群野生植物的繁殖特性和已有的人工繁殖方法进行了综述, 并简要介绍在其种苗繁殖研究方面取得的进展。其中利用播种繁殖成功的物种有12种, 共繁殖230,000株种苗; 利用扦插繁殖成功的物种有5种, 共繁殖33,100株种苗; 华盖木(Manglietiastrum sinicum)、河北梨(Pyrus hopeiensis)和黄梅秤锤树(Sinojackia huangmeiensis)采用嫁接繁殖出了2,415株种苗; 9个物种的组织培养技术获得成功, 共繁殖了24,850株种苗。这些种苗有些已应用于迁地保护和自然回归。上述研究结果为这14种极小种群野生植物的保护和利用提供了理论和技术基础, 也能为其他极小种群野生植物的保护和利用提供参考。     

极小种群野生植物坡垒的生境特征及其对幼苗多度的影响

极小种群野生植物坡垒(Hopea hainanensis)曾经是热带低地雨林的优势种, 但由于商业采伐和刀耕火种等严重人类干扰及自然更新困难, 致使其种群数量急剧下降到最小可存活的界限, 急需开展种群的拯救恢复工作。而对于坡垒生境适宜性及更新限制的了解, 是进行种群保护及恢复的基础。本文在对野生坡垒种群及其生境因子调查测定的基础上, 分析了生物与非生物生境特征及其对坡垒种群更新幼苗多度的影响。结果表明: 坡垒种群从幼苗至幼树阶段存在着严重的增补限制。坡度小、土壤含水量和有效磷含量高、母株胸径和冠幅较大、伴生种胸高断面积中等的环境是坡垒幼苗较为适宜的生境, 且坡垒幼苗多度与坡度及土壤pH值显著负相关, 与土壤含水量及土壤全磷含量显著正相关。这些研究结果为极小种群野生植物坡垒的就地保护与种群复壮提供了科学依据。     

微生境对黄梅秤锤树野生种群叶片功能性状的影响

植物功能性状可以反映植物应对环境变化的适应策略。本文以黄梅秤锤树(Sinojackia huangmeiensis)当前唯一野生种群为对象, 比较了3种微生境(湖边、种群中心、耕地边)中该物种的叶片功能性状均值、种内变异和叶片生态化学计量特征的差异, 分析了黄梅秤锤树对湖岸带微生境变化的响应及其适应策略。结果表明: (1) 3种微生境中土壤C、N、P含量没有显著性差异(P > 0.05), 但土壤C∶N和C∶P具有显著性差异(P < 0.05), 土壤类型和养分条件有所不同。(2)黄梅秤锤树叶片功能性状的比较用单因素方差分析和贝叶斯方差分析得出的结果一致, 均为叶长、叶面积和比叶面积在中心区域显著高于湖边(P < 0.05), 而耕地边与湖边和中心区域均没有显著差异(P > 0.05); 叶N含量在湖边显著高于中心区域和耕地边(P < 0.05), 而中心区域和耕地边间没有显著差异(P > 0.05); 叶宽、叶长/叶宽、叶干物质含量、叶C和叶P含量在3种微生境间都没有显著性差异(P > 0.05)。(3)黄梅秤锤树叶片的N∶P在湖边显著高于中心区域和耕地边(P < 0.05), C∶N在湖边显著小于中心区域和耕地边(P < 0.05), N∶P和C∶N在中心区域和耕地边没有显著性差异(P > 0.05), C∶P在3种微生境间都没有显著性差异(P > 0.05)。(4)黄梅秤锤树叶片功能性状的总体变异程度在0.02-0.28之间, 其中叶片C和N含量在湖边和中心区域的种内变异程度显著较低, 表明3种生境中湖边和中心区域黄梅秤锤树种群的稳定性相对较差。(5)湖边黄梅秤锤树主要通过增加叶N含量促进生长; 中心区域黄梅秤锤树主要通过增加叶面积和比叶面积以及提高叶N的利用效率来提高光捕获能力促进生长; 耕地边黄梅秤锤树的叶N含量和叶面积、比叶面积都处于中等水平, 通过性状之间的共同作用使植株生长达到最佳水平。以上结果表明, 由于微地形、水位波动和土壤环境条件的差异, 黄梅秤锤树对3种生境中的适应策略有所不同, 并且不是通过单一性状调整来适应环境的变化, 而是通过多种性状之间的权衡达到更好的适应效果。     

结核分枝杆菌小RNA及其作用的研究进展

细菌小RNA是一类长度在50～500个核苷酸之间的不具有编码蛋白质功能,但具有转录后调控作用的RNA,在细菌中参与调控细菌多种生理和病理活动,如调节细菌代谢和毒力作用等过程。近年来,在结核分枝杆菌已经鉴定出近200种小RNA,并证明这些小RNA参与结核分枝杆菌的生理和病理过程。本文对结核分枝杆菌小RNA在细菌生长繁殖、毒力因子调控、细菌耐药和巨噬细胞内应激环境的适应等方面的作用进行综述。