青枯菌HPLC分析中样品制备方法的优化

本文研究了青枯菌细胞的制备方法对细胞生命活力和表面特性的影响。结果表明,在高效离子交换色谱分析（HPLC）中,采用5000×g离心10min收集菌体细胞、超纯水（＞16MΩ）悬浮和洗涤青枯菌、重复洗涤二次的制备方法,既可以避免培养基成分造成的干扰,又可以保持青枯菌细胞的生命活力和细胞表面的原有性质。     

The distribution of iron oxidation states in a constructed wetland as an indicator of its redox properties

Wastewater-treatment processes taking place inside constructed wetlands are closely connected with chemical properties of these systems. The aeration of a wetland via the roots of the vegetation (and a subsequent formation of redox-potential gradients) strongly influences the wastewater treatment efficiency, and thus it represents one of the most important characteristics of the wetland. The concentration ratios of individual iron oxidation states (Fe(II) and Fe(III)) were determined as the indicator of the redox properties of the constructed wetland reed bed during this study. Interstitial water from the wetland was sampled eleven times throughout the year 2005. The spectrophotometric method using 1,10-phenanthroline was properly optimized (limits of detection and quantification, sensitivity, linear dynamic range, repeatability, and accuracy values were assessed) and applied for iron determination. Most of iron, ca. 98%, is reduced to the Fe(II) form in raw wastewater and water from the inflow zone of the constructed wetland, however, at the outflow and in the vegetation bed both iron oxidation states are usually detected. The presence of Fe(III) in the reed bed (ca. 10-30% for some samples) demonstrates the aeration of the wetland by the vegetation (Phragmites australis) resulting in a re-oxidation of Fe(II).     

Biomechanical Plasticity Facilitates Invasion of Maritime Forests in the southern USA by Brazilian pepper (Schinus terebinthifolius)

Biomechanical plasticity and within-species growth form diversity are traits that can facilitate invasion by non-native plant species. We support this argument with evidence from the invasion of coastal habitats in northern Florida, USA, by Schinus terebinthifolius and describe some of the consequences of this invasion for overtopped saltmarsh plants. In crowded stands, Schinus grows more like a vine than a tree, with stem height : diameter ratios nearly twice than those observed in open-grown individuals but with no changes in wood density or the modulus of elasticity of stem material. When extracted from the surrounding vegetation, the formerly crowded Schinus stems buckle under their own weight. Schinus crowns also extend much further over adjacent saltmarsh than crowns of Juniperus virginiana, the only other tree species abundant in the study site. Along forest edges, the above-ground biomass of saltmarsh plants overtopped by Schinus crowns was reduced by more than an order of magnitude. The biomechanical plasticity of Schinus allows it to adapt its growth form to suit habitat conditions and can dominate the edges of salt marshes as a sprawling shrub and maritime forests as either a free-standing tree or a woody vine, depending on stand crowding     

Integrating the Study of Non-native Plant Invasions across Spatial Scales

Non-native (alien, exotic) plant invasions are affecting ecological processes and threatening biodiversity worldwide. Patterns of plant invasions, and the ecological processes which generate these patterns, vary across spatial scales. Thus, consideration of spatial scale may help to illuminate the mechanisms driving biological invasions, and offer insight into potential management strategies. We review the processes driving movement of non-native plants to new locations, and the patterns and processes at the new locations, as they are variously affected by spatial scale. Dispersal is greatly influenced by scale, with different mechanisms controlling global, regional and local dispersal. Patterns of invasion are rarely documented across multiple spatial scales, but research using multi-scale approaches has generated interesting new insights into the invasion process. The ecological effects of plant invasions are also scale-dependent, ranging from altered local community diversity and homogenization of the global flora, to modified biogeochemical cycles and disturbance regimes at regional or global scales. Therefore, the study and control of invasions would benefit from documenting invasion processes at multiple scales.     

复合改性提高大豆分离蛋白功能性的研究

目的:利用中性蛋白酶和谷氨酰胺转胺酶(TG)对大豆分离蛋白(SPI)进行复合改性,探讨对其功能性的影响。方法:通过单因素和正交实验以及电泳分析,研究了中性蛋白酶和TG对SPI的影响趋势。结果:中性蛋白酶作用的最佳工艺条件为温度60℃、时间0.5h、pH7.0、酶用量4000U/g,SPI溶解性可达97.9%。结论:经中性蛋白酶作用后,再经过TG改性,所得的聚合物不仅有很大的相对分子质量,还可以改善SPI的溶解性,并且乳化性、发泡性均有提高。     

Decreased protein tyrosine phosphorylation and membrane fluidity in spermatozoa from infertile men with varicocele

Varicocele is a prevalent pathology among infertile men. The mechanisms linking this condition to infertility, however, are poorly understood. Our previous work showed a relationship between sperm functional quality and the ability of spermatozoa to respond to capacitating conditions with increased membrane fluidity and protein tyrosine phosphorylation. Given the reported association between varicocele, oxidative stress, and sperm dysfunction, we hypothesized that spermatozoa from infertile patients with varicocele might have a combined defect at the level of membrane fluidity and protein tyrosine phosphorylation. Semen samples from infertile patients with and without grade II/III left varicocele were evaluated for motion parameters (computer-assisted semen analysis [CASA]), hyperactivation (CASA), incidence and intensity of protein tyrosine phosphorylation (phosphotyrosine immunofluorescence and western blotting), and membrane fluidity (Laurdan fluorometry), before and after a capacitating incubation (6 hr at 37 degrees C in Ham's F10/BSA, 5% CO(2)). Spermatozoa from varicocele samples presented a decreased response to the capacitating challenge, showing significantly lower motility, hyperactivation, incidence and intensity of tyrosine phosphorylation, and membrane fluidity. The findings reported in this article indicate that the sperm dysfunction associated to infertile varicocele coexists with decreased sperm plasma membrane fluidity and tyrosine phosphorylation. These deficiencies represent potential new pathophysiological mechanisms underlying varicocele-related infertility.     

In vitro comparison of six different matrix systems for the cultivation of human chondrocytes

Summary In recent years, a great variety of different matrix systems for the cultivation of chondrocytes have been developed. Although some of these scaffolds show promising experimental results in vitro, the potential clinical value remains unclear. In this comparative study, we propagated human articular chondrocytes precultivated in monolayer culture on six different scaffolds (collagen gels, membranes and sponges) under standardized in vitro conditions. Mechanical properties of the matrix systems were not improved significantly by cultivation of human chondrocytes under the given in vitro conditions. The gel systems (CaReS, Ars Artho, Germany and Atelocollagen, Koken, Japan) showed a homogeneous cell distribution; chondrocytes propagated on Chondro-Gide (Geistlich Biomaterials, Switzerland) and Integra membranes (Integra, USA) were building multilayers. Only few cells penetrated the two Atelocollagen honeycomb sponges (Koken, Japan). During cultivation, chondrocytes propagated on all systems showed a partial morphological redifferentiation, which was best with regard to the gel systems. In general, only small amounts of collagen type-II protein could be detected in the pericellular region and chondrocytes failed to build a territorial matrix. During the first two weeks of cultivation, the two gel systems showed a significantly higher collagen type-II gene expression and a lower collagen type-I gene expression than the other investigated matrix systems. Although collagen gels seem to be superior when dealing with deep cartilage defects, membrane systems might rather be useful in improving conventional autologous chondrocyte transplantation or in combination with gel systems.     

Secretion of pro- and mature Rhizopus arrhizus lipases by Pichia pastoris and properties of the proteins

The lipases of Rhizopus spp. share a high 1,3-regiospecificity toward triacylglycerols, which makes them important enzymes in lipid modification. In the present study, the extracellularly active production of recombinant Rhizopus arrhizus lipase was carried out with genes encoding the mature region (mRAL) and the mRAL having the prosequence (ProRAL) in Pichia pastoris. Two transformed P. pastoris clones containing the multicopy of mRAL and ProRAL genes were separately selected for the production of recombinant enzymes. In a fed-batch cultivation, where methanol feeding was controlled by an on-line methanol analyzer, the supernatant contained 91 mg/L recombinant pro-form lipase (rProRAL) and 80 mg/L recombinant mature lipase (rRAL) after 92 h of cultivation. rProRAL and rRAL were purified by ultrafiltration, SP-Sepharose Rast Flow chromatography, and Butyl-Sepharose Fast Flow chromatography. Molecular weights of rProRAL and rRAL are 32 kDa and 29 kDa, respectively. The amino-terminal analysis showed that the 32-kDa protein was mRAL attached with 28 amino acids of the carboxy-terminal part of the prosequence (rPro28RAL). The specific lipase activities of mRAL attached with 28 amino acids of the carboxy-terminal part of the prosequence (rPro28RAL) and rRAL were 1543 U/mg and 2437 U/mg. The rPro28RAL was more stable than rRAL at pH 4.0–7.0, whereas rRAL was more stable at pH 7.0–10.0. The rPro28RAL had the highest lipase activity toward tributyrin (C₄), whereas rRAL had the highest lipase activity toward tricaprylin (C₈).     

Inactivation of calcium channels in vascular smooth muscle myocytes

Many of the structural domains involved in Ca²⁺ channel (CACN) inactivation are also involved in determining their sensitivity to antagonist inhibition. We hypothesize that differences in inactivation properties and their structural determinants may suggest candidate domains as targets for the development of novel, selective antagonists. The characteristics of Ca²⁺ current (I_Ca) inactivation, steady-state inactivation (SSIN), and recovery from inactivation were studied in freshly dispersed smooth muscle cells from rabbit portal vein (RPV) using whole-cell, voltage-clamp methods. The time course of inactivation could be represented by two time constants. Increasing I_Ca by increasing [Ca²⁺]_o or with more negative holding potentials decreased both time constants. With Sr²⁺, Ba²⁺, or Na⁺ as the charge carrier, I_Ca inactivation was also represented by two time constants, both of which were larger than those found with Ca²⁺. With Ca²⁺, Sr²⁺, or Ba²⁺ as the charge carrier, both time constants had minimum values near the voltage associated with maximum current. When Na⁺ (140 mM) was the charge carrier, voltages for I_max (−20 mV) or τ_min (o mV) did not correspond. SSIN of I_Ca had a half-maximum voltage of −32±4 mV for Ca²⁺, −43 mV±5 mV for Sr²⁺, −41±5 mV for Ba²⁺, and −68±6 mV for Na⁺. The slope factor for SSIN per e-fold voltage change was 6.5±0.2 mV for Ca²⁺, 6.8±0.3 for Sr²⁺, and 6.6±0.2 for Ba²⁺, representing four equivalent charges. When Na⁺ or Li⁺ was the charge carrier, the slope factor was 13.5±0.7 mV, representing two equivalent charges. For I_Ca in rat left ventricular (rLV) myocytes, there was no difference in the slope factor of SSIN for Ca²⁺ and Na⁺. The rate of recovery of I_Ca from inactivation varied inversely with recovery voltage and was independent of the charge carrier. These results suggest that inactivation of I_Ca in PV myocytes possess an intrinsic voltage dependence that is modified by Ca²⁺. For RPV but not rLV I_Ca, the charge of the permeating ion confers the voltage-dependency of SSIN.     

Biophysical studies on the differentiation of human CD14+ monocytes into dendritic cells

Dendritic cells (DCs), which are the most efficient antigen-presenting cells (APCs) currently known, can be derived from CD14⁺ monocytes (DC predecessor cells) in vitro. Immature DCs actively take up antigens and pathogens, generate major histocompatability complex-peptide complexes, and migrate from the sites of antigen acquisition to secondary lymphoid organs to become mature dendritic cells that interact with and stimulate T-lymphocytes. During this process, the cells must undergo deformation to translocate through several barriers, including the basement membrane and interstitial connective tissue in the blood vessel wall. To further understand the mechanisms of the activation of immunological responses and the migration from peripheral tissue to secondary lymphoid organs, we have applied biophysical and microrheological methods to study the development processes of DCs in vitro. The results showed that membrane fluidity, osmotic fragility, membrane viscoelastic properties, infrared spectroscopy, and cytoskeleton organization of DCs exhibit significant differences in different developmental stages. These authors contributed equally to this work.