Proteome characterization of sea star coelomocytes--the innate immune effector cells of echinoderms

Sea star coelomic fluid is in contact with all internal organs, carrying signaling molecules and a large population of circulating cells, the coelomocytes. These cells, also known as echinoderm blood cells, are responsible for the innate immune responses and are also known to have an important role in the first stage of regeneration, i.e. wound closure, necessary to prevent disruption of the body fluid balance and to limit the invasion of pathogens. This study focuses on the proteome characterization of these multifunctional cells. The identification of 358 proteins was achieved using a combination of two techniques for protein separation (1-D SDS-PAGE followed by nanoLC and 2-D SDS-PAGE) and MALDI-TOF/TOF MS for protein identification. To our knowledge, the present report represents the first comprehensive list of sea star coelomocyte proteins, constituting an important database to validate many echinoderm-predicted proteins. Evidence for new pathways in these particular echinoderm cells are also described, and thus representing a valuable resource to stimulate future studies aiming to unravel the homology with vertebrate immune cells and particularly the origins of the immune system itself.     

Identification of differentially expressed proteins in blood plasma of control and cigarette smoke-exposed mice by 2-D DIGE/MS

Cigarette smoke exposure is known to induce obstructive lung disease and several cardiovascular disease states in humans and also in animal models. Smoking leads to oxidative stress and inflammation that are important in triggering pulmonary and cardiovascular disease. The objective of the current study was to quantify differences in expression levels of plasma proteins of cigarette smoke -exposed and control mice, at the time of disease onset, and identify these proteins for use as potential biomarkers of the onset of smoking-induced disease. We utilized 2-D DIGE/MS to characterize these proteomic changes. 2-D DIGE of plasma samples identified 11 differentially expressed proteins in cigarette smoke -exposed mice. From these 11 proteins, 9 were downregulated and 2 were upregulated. The proteins identified are involved in vascular function, coagulation, metabolism and immune function. Among these, the alterations in fibrinogen (2.2-fold decrease), α-1-antitrypsin (1.8-fold increase) and arginase (4.5-fold decrease) are of particular interest since these have been directly linked to cardiovascular and lung pathology. Differences in expression levels of these proteins were also confirmed by immunoblotting. Thus, we observe that chronic cigarette smoke exposure in mice leads to prominent changes in the protein expression profile of blood plasma and these changes in turn can potentially serve as markers predictive of the onset and progression of cardiovascular and pulmonary disease.     

Immunodepletion of albumin and immunoglobulin G from bovine plasma

Current MS-based proteomics has facilitated the identification of large numbers of proteins from complex mixtures. The bovine plasma proteome has the potential to provide a wealth of information concerning the biological state of an animal. However, during MS-based experiments, higher abundance proteins such as albumin and immunoglobulin G (IgG) can hinder the identification of potentially important proteins that are present in much lower abundance. While a variety of readily available technologies exist for the depletion of multiple high-abundance proteins from human, mouse and rat samples, there are few available for bovine. In this study, we report the depletion of >97% of albumin and >92% of IgG from bovine plasma.     

基于DUS测试的水稻标准品种形态性状多样性分析

对水稻DUS测试标准品种进行多样性评价,为DUS测试标准品种的选择和优化提供依据。以DUS测试指南中规定的49个水稻标准品种为材料,根据2008年在杭州田间试验对59个形态性状的调查数据,利用PopGen32软件计算了等位变异数、有效等位变异数和Shannon's多样性信息指数,对标准品种的形态性状多样性进行了评价;并根据其多样性参数采用NTSYS软件对标准品种进行了聚类分析。结果表明,59个形态性状在19份籼稻和30份粳稻品种中分别检测到201和262个等位变异,平均每个性状可以检测到3.4068（籼稻）和4.4407(粳稻)个等位变异,籼、粳稻类群中59个性状的平均Shannon's多样性指数分别为0.8318（0~1.7674）和0.9828（0~1.8547）。聚类分析结果表明,籼、粳稻群体内品种间的形态性状相似系数分别介于0.45~0.81和0.36~0.76,取相似系数0.70为阈值,可将19份籼稻和30份粳稻品种分别分为16和27类。多样性分析表明本研究的49个品种形态性状多样性丰富,具有标准品种性状描述的示例和校正作用;根据聚类分析结果,可将现有标准品种的数量减少至43个。本研究为水稻新品种DUS测试中标准品种的选择和优化提供了参考依据,同时也可供研制其他植物的DUS测试标准借鉴。     

动物离心机中大鼠行为状态无线视频采集装置的研制

目的：研制一种在正加速度作用下能实时观测大鼠行为状态的实验视频采集装置,为研究正加速度导致脑损伤的病理生理机制和防治措施提供新的方法。方法：依据正加速度对大鼠体位的要求和离心机特殊的环境,通过特制的大鼠座舱和无线视频采集单元来实现对动物离心机中正加速度作用下大鼠行为状态的实时观测;固定大鼠的座舱主要由胸部固定圈、身长调节圈、头部运动限制框构成;无线视频采集单元由2.4G无线摄像头、无线信号接收器、信号放大器、计算机以及视频采集软件组成。结果：座舱特殊的大鼠固定方式,既满足了正加速度对体位的要求,又不影响对大鼠行为状态的观测,2．4G无线视频采集技术保证视频信号在离心机高速旋转环境下的实时传输和观测,视频画面清晰流畅,并且能保存视频供回放分析。结论：动物离心机无线视频采集装置可用于正加速度作用下大鼠行为状态的实时观测与研究。     

Mouse Complete Stasis Model of Inferior Vena Cava Thrombosis

Venous thromboembolism (VTE) includes both deep vein thrombosis (DVT) and pulmonary embolism (PE). In the United States (U.S.), the high morbidity and mortality rates make VTE a serious health concern ^1-2. After heart disease and stroke, VTE is the third most common vascular disease ³. In the U.S. alone, there is an estimated 900,000 people affected each year, with 300,000 deaths occurring annually ³. A reliable in vivo animal model to study the mechanisms of this disease is necessary.The advantages of using the mouse complete stasis model of inferior vena cava thrombosis are several. The mouse model allows for the administration of very small volumes of limited availability test agents, reducing costs dramatically. Most promising is the potential for mice with gene knockouts that allow specific inflammatory and coagulation factor functions to be delineated. Current molecular assays allow for the quantitation of vein wall, thrombus, whole blood, and plasma for assays. However, a major concern involving this model is the operative size constraints and the friability of the vessels. Also, due to the small IVC sample weight (mean 0.005 grams) it is necessary to increase animal numbers for accurate statistical analysis for tissue, thrombus, and blood assays such as real-time polymerase chain reaction (RT-PCR), western blot, enzyme-linked immunosorbent (ELISA), zymography, vein wall and thrombus cellular analysis, and whole blood and plasma assays ^4-8.The major disadvantage with the stasis model is that the lack of blood flow inhibits the maximal effect of administered systemic therapeutic agents on the thrombus and vein wall.     

Effects of stressors in adolescence on learning and memory in rodent models

This article is part of a Special Issue "Puberty and Adolescence".     

Insulin-like Growth Factor I (IGF-I)-induced Chronic Gliosis and Retinal Stress Lead to Neurodegeneration in a Mouse Model of Retinopathy

Insulin-like growth factor I (IGF-I) exerts multiple effects on different retinal cell types in both physiological and pathological conditions. Despite the growth factor''s extensively described neuroprotective actions, transgenic mice with increased intraocular levels of IGF-I showed progressive impairment of electroretinographic amplitudes up to complete loss of response, with loss of photoreceptors and bipolar, ganglion, and amacrine neurons. Neurodegeneration was preceded by the overexpression of genes related to retinal stress, acute-phase response, and gliosis, suggesting that IGF-I altered normal retinal homeostasis. Indeed, gliosis and microgliosis were present from an early age in transgenic mice, before other alterations occurred, and were accompanied by signs of oxidative stress and impaired glutamate recycling. Older mice also showed overproduction of pro-inflammatory cytokines. Our results suggest that, when chronically increased, intraocular IGF-I is responsible for the induction of deleterious cellular processes that can lead to neurodegeneration, and they highlight the importance that this growth factor may have in the pathogenesis of conditions such as ischemic or diabetic retinopathy.     

Validation of shear wave elastography in skeletal muscle

Skeletal muscle is a very dynamic tissue, thus accurate quantification of skeletal muscle stiffness throughout its functional range is crucial to improve the physical functioning and independence following pathology. Shear wave elastography (SWE) is an ultrasound-based technique that characterizes tissue mechanical properties based on the propagation of remotely induced shear waves. The objective of this study is to validate SWE throughout the functional range of motion of skeletal muscle for three ultrasound transducer orientations. We hypothesized that combining traditional materials testing (MTS) techniques with SWE measurements will show increased stiffness measures with increasing tensile load, and will correlate well with each other for trials in which the transducer is parallel to underlying muscle fibers. To evaluate this hypothesis, we monitored the deformation throughout tensile loading of four porcine brachialis whole-muscle tissue specimens, while simultaneously making SWE measurements of the same specimen. We used regression to examine the correlation between Young′s modulus from MTS and shear modulus from SWE for each of the transducer orientations. We applied a generalized linear model to account for repeated testing. Model parameters were estimated via generalized estimating equations. The regression coefficient was 0.1944, with a 95% confidence interval of (0.1463–0.2425) for parallel transducer trials. Shear waves did not propagate well for both the 45° and perpendicular transducer orientations. Both parallel SWE and MTS showed increased stiffness with increasing tensile load. This study provides the necessary first step for additional studies that can evaluate the distribution of stiffness throughout muscle.     

Coagulant thrombin-like enzyme (barnettobin) from Bothrops barnetti venom: Molecular sequence analysis of its cDNA and biochemical properties

The thrombin-like enzyme from Bothrops barnetti named barnettobin was purified. We report some biochemical features of barnettobin including the complete amino acid sequence that was deduced from the cDNA. Snake venom serine proteases affect several steps of human hemostasis ranging from the blood coagulation cascade to platelet function. Barnettobin is a monomeric glycoprotein of 52 kDa as shown by reducing SDS-PAGE, and contains approx. 52% carbohydrate by mass which could be removed by N-glycosidase. The complete amino acid sequence was deduced from the cDNA sequence. Its sequence contains a single chain of 233 amino acid including three N-glycosylation sites. The sequence exhibits significant homology with those of mammalian serine proteases e.g. thrombin and with homologous TLEs. Its specific coagulant activity was 251.7 NIH thrombin units/mg, releasing fibrinopeptide A from human fibrinogen and showed defibrinogenating effect in mouse. Both coagulant and amidolytic activities were inhibited by PMSF. N-deglycosylation impaired its temperature and pH stability. Its cDNA sequence with 750 bp encodes a protein of 233 residues. Indications that carbohydrate moieties may play a role in the interaction with substrates are presented. Barnettobin is a new defibrinogenating agent which may provide an opportunity for the development of new types of anti-thrombotic drugs.