Changes of Photosystem Ⅱ Electron Transport in the Chlorophyll-deficient Oilseed Rape Mutant Studied by Chlorophyll Fluorescence and Thermoluminescence

The photosystem Ⅱ (PSII) complex of photosynthetic membranes comprises a number of chlorophyll-binding proteins that are important to the electron flow. Here we report that the chlorophyll b-deficient mutant has de creased the amount of light-harvesting complexes with an increased amount of some core polypeptides of PSII,including CP43 and CP47. By means of chlorophyll fluorescence and thermoluminescence, we found that the ratio of Fv/Fm, qP and electron transport rate in the chlorophyll b-deficient mutant was higher compared to the wild type.In the chlorophyll b-deficient mutant, the decay of the primary electron acceptor quinones (QA-) reoxidation was decreased, measured by the fluorescence. Furthermore, the thermolumlnescence studies in the chlorophyll b deficient mutant showed that the B band (S2/S3QB-) decreased slightly and shifted up towards higher temperatures.In the presence of dichlorophenyl-dimethylurea, which is inhibited in the electron flow to the second electron acceptor quinines (QB) at the PSII acceptor side, the maximum of the Q band (S2QA-) was decreased slightly and shifted down to lower temperatures, compared to the wild type. Thus, the electron flow within PSll of the chlorophyll b-deficient mutant was down-regulated and characterized by faster oxidation of the primary electron acceptor quinine QA- via forward electron flow and slower reduction of the oxidation S states.     

Retention of Virulence in a Viable but Nonculturable Edwardsiella tarda Isolate

Edwardsiella tarda is pathogen of fish and other animals. The aim of this study was to investigate the viable but nonculturable (VBNC) state and virulence retention of this bacterium. Edwardsiella tarda CW7 was cultured in sterilized aged seawater at 4°C. Total cell counts remained constant throughout the 28-day period by acridine orange direct counting, while plate counts declined to undetectable levels (<0.1 CFU/ml) within 28 days by plate counting. The direct viable counts, on the other hand, declined to ca. 10⁹ CFU/ml active cells and remained fairly constant at this level by direct viable counting. These results indicated that a large population of cells existed in a viable but nonculturable state. VBNC E. tarda CW7 could resuscitate in experimental chick embryos and in the presence of nutrition with a temperature upshift. The resuscitative times were 6 days and 8 days, respectively. The morphological changes of VBNC, normal, and resuscitative E. tarda CW7 cells were studied with a scanning electron microscope. The results showed that when the cells entered into the VBNC state, they gradually changed in shape from short rods to coccoid and decreased in size, but the resuscitative cells did not show any obvious differences from the normal cells. The VBNC and the resuscitative E. tarda CW7 cells were intraperitoneally inoculated into turbot separately, and the fish inoculated with the resuscitative cells died within 7 days, which suggested that VBNC E. tarda CW7 might retain pathogenicity.     

缺氧诱导因子-1和缺氧诱导因子-2：结构、功能及调节

缺氧诱导因子-1（HIF-1）和缺氧诱导因子-2（HIF-2）是细胞应对缺氧时关键的转录因子,在生物体生理及病理过程中有重要的作用。HIF由一个α亚基和一个β亚基组成二聚体。在蛋白水平上,HIF的稳定性及转录活性受到多种机制的调控,除为人所熟知的O2/PHDs/pVHL降解途径及FIH-1羟基化作用外,分别针对HIF-1α和HIF-2α的特异性调控机制也相继被报道。从HIF-1α和HIF-2α的蛋白结构、稳定性调控、转录激活功能以及两者在细胞代谢、肿瘤发生中的作用等方面对两者的相似性和差异性进行综述。     

Hypoglycemic effects of exopolysaccharides produced by mycelial cultures of two different mushrooms Tremella fuciformis and Phellinus baumii in ob/ob mice

The anti-diabetic activities of the exopolysaccharides (EPS) produced by submerged mycelial culture of two different mushrooms, Tremella fuciformis and Phellinus baumii, in ob/ob mice were investigated. All the animals were randomly divided into three groups with seven animals in each group: The control group received 0.9% NaCl solution; the diabetic groups were treated with EPS from T. fuciformis (Tf EPS) and P. baumii (Pb EPS) at the level of 200 mg/kg body weight using an oral zoned daily for 52 days. The plasma glucose levels in the EPS-fed mice were substantially reduced by about 52% (Tf EPS) and 32% (Pb EPS), respectively, as compared to control mice. The results of oral glucose tolerance test (OGTT) revealed that both EPS-fed groups significantly increased the glucose disposal after 52 days of EPS treatments. Furthermore, higher food efficiency ratios and reduced blood triglyceride levels were observed in the EPS-treated groups. Because peroxisome proliferator-activated receptor gamma (PPAR-γ) is indeed a key regulator of insulin action, we investigated the expression pattern of adipose tissue PPAR-γ messenger RNA (mRNA) and plasma levels of PPAR-γ. It was revealed that PPAR-γ was significantly activated in response to EPS treatments. The results suggested that both EPS exhibited considerable hypoglycemic effect and improved insulin sensitivity possibly through regulating PPAR-γ-mediated lipid metabolism. Our results indicated that two mushroom-derived EPS might be developed as potential oral hypoglycemic agents or functional foods for the management of non-insulin-dependent diabetes mellitus.     

Simultaneous determination of metformin and rosiglitazone in human plasma by liquid chromatography/tandem mass spectrometry with electrospray ionization: application to a pharmacokinetic study

A selective and sensitive high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (ESI-MS/MS) method for simultaneous determination of metformin and rosiglitazone in human plasma using phenformin as internal standard (IS) has been first developed and validated. Plasma samples were precipitated by acetonitrile and the analytes were separated on a prepacked Phenomenex Luna 5u CN 100A (150 mm x 2.0 mm I.D.) column using a mobile phase comprised of methanol:30 mM ammonium acetate pH 5.0 (80:20, v/v) delivered at 0.2 ml/min. Detection was performed on a Finnigan TSQ triple-quadrupole tandem mass spectrometer in positive ion selected reaction monitoring (SRM) mode using electrospray ionization. The ion transitions monitored were m/z 130.27-->71.11 for metformin, m/z 358.14-->135.07 for rosiglitazone and m/z 206.20-->105.19 for the IS. The standard curves were linear (r(2)>0.99) over the concentration range of 5-3000 ng/ml for metformin and 1.5-500 ng/ml for rosiglitazone with acceptable accuracy and precision, respectively. The within- and between-batch precisions were less than 15% of the relative standard deviation. The limit of detection (LOD) of both metformin and rosiglitazone was 1 ng/ml. The method described is precise and sensitive and has been successfully applied to the study of pharmacokinetics of compound metformin and rosiglitazone capsules in 12 healthy Chinese volunteers.     

Cardiovascular responses to intrathecal administration of endomorphins in anesthetized rats

Endomorphins (EMs), the endogenous, potent and selective mu-opioid receptor agonists, have been shown to decrease systemic arterial pressure (SAP) in rats after intravenous (i.v.) administration. In the present study, cardiovascular responses to intrathecal (i.t.) injection of EMs were investigated in urethane-anesthetized rats. It is noteworthy that EMs elicited decreases in SAP and heart rate (HR) in a dose-dependent manner; 10-300nmol/kg were injected intrathecally. Furthermore, these vasodepressor and bradycardic effects were significantly antagonized by naloxone (0.5mg/kg, i.t.). Interestingly, i.t. (5mg/kg) or i.v. (50mg/kg) administrations of N(omega)-nitro-l-arginine methylester (l-NAME) attenuated the vasodepressor and bradycardic effects. Moreover, pretreatment of the rats with muscarinic receptor antagonist atropine (2mg/kg, i.v.) and alpha-adrenoceptor antagonist phentolamine (1mg/kg, i.v.) significantly reduced the vasodepressor effects of EMs. Nevertheless, pretreatment with beta-adrenoceptor antagonist propranolol (2mg/kg, i.v.) could only block the bradycardia effects induced by EMs, but had no significant effects on the hypotension. In summary, all the results suggested that i.t. administration of EMs decreased SAP and HR which were possibly mediated by the activation of opioid receptors in the rat spinal cord. In addition, nitric oxide (NO) release in both the spinal cord and in peripheral tissues might regulate the cardiovascular activities of EMs, and the muscarinic receptor and adrenoceptor played an important role in the regulation of the cardiovascular responses to i.t. administration of EMs.     

Consistent manufacturing and quality control of a highly complex recombinant polyclonal antibody product for human therapeutic use

The beneficial effect of antibody therapy in human disease has become well established mainly for the treatment of cancer and immunological disorders. The inherent monospecificity of mAbs present limitations to mAb therapy which have become apparent notably in addressing complex entities like infectious agents or heterogenic endogenous targets. For such indications mixtures of antibodies comprising a combination of specificities would convey more potent biological effect which could translate into therapeutic efficacy. Recombinant polyclonal antibodies (rpAb) consisting of a defined number of well-characterized mAbs constitute a new class of target specific antibody therapy. We have developed a cost-efficient cell banking and single-batch manufacturing concept for the production of such products and demonstrate that a complex pAb composition, rozrolimupab, comprising 25 individual antibodies can be manufactured in a highly consistent manner in a scaled-up manufacturing process. We present a strategy for the release and characterization of antibody mixtures which constitute a complete series of chemistry, manufacturing, and control (CMC) analytical methods to address identity, purity, quantity, potency, and general characteristics. Finally we document selected quality attributes of rozrolimupab based on a battery of assays at the genetic-, protein-, and functional level and demonstrate that the manufactured rozrolimupab batches are highly pure and very uniform in their composition.     

The modular structure of haemagglutinin/adhesin regions in gingipains of Porphyromonas gingivalis

High-molecular-weight arginine- and lysine-specific (Kgp) gingipains are essential virulence factors expressed by the oral pathogen Porphyromonas gingivalis. Haemagglutinin/adhesin (HA) regions of these proteases have been implicated in targeting catalytic domains to biological substrates and in other adhesive functions. We now report the crystal structure of the K3 adhesin domain/module of Kgp, which folds into the distinct β-jelly roll sandwich topology previously observed for K2. A conserved structural feature of K3, previously observed in the Kgp K2 module, is the half-way point anchoring of the surface exposed loops via an arginine residue found in otherwise highly variable sequences. Small-angle X-ray scattering data for the recombinant construct K1K2K3 confirmed a structure comprising a tandem repeat of three homologous modules, K1, K2 and K3 while also indicating an unusual 'y'-shape arrangement of the modules connected by variable linker sequences. Only the K2 and K3 modules and a K1K2 construct were observed to be potently haemolytic. K2, K3 and the K1K2 construct showed preferential recognition of haem-albumin over albumin whereas only low affinity binding was detected for K1 and the K1K2K3 construct. The data indicate replication of some biological functions over the three adhesin domains of Kgp while other functions are restricted.     

Diversity and Biogeography of Rhizobia Isolated from Root Nodules of <Emphasis Type="Italic">Glycine max</Emphasis> Grown in Hebei Province,China

A total of 215 rhizobial strains were isolated and analyzed with 16S rRNA gene, 16S–23S intergenic spacer, housekeeping genes atpD, recA, and glnII, and symbiotic genes nifH and nodC to understand the genetic diversity of soybean rhizobia in Hebei province, China. All the strains except one were symbiotic bacteria classified into nine genospecies in the genera of Bradyrhizobium and Sinorhizobium. Surveys on the distribution of these rhizobia in different regions showed that Bradyrhizobium japonicum and Bradyrhizobium elkanii strains were found only in neutral to slightly alkaline soils whereas Bradyrhizobium yuanmingense, Bradyrhizobium liaoningense-related strains and strains of five Sinorhizobium genospecies were found in alkaline–saline soils. Correspondence and canonical correspondence analyses on the relationship of rhizobial distribution and their soil characteristics reveal that high soil pH, electrical conductivity, and potassium content favor distribution of the B. yuanmingense and the five Sinorhizobium species but inhibit B. japonicum and B. elkanii. High contents of available phosphorus and organic matters benefit Sinorhizobium fredii and B. liaoningense-related strains and inhibit the others groups mentioned above. The symbiotic gene (nifH and nodC) lineages among B. elkanii, B. japonicum, B. yuanmingense, and Sinorhizobium spp. were observed in the strains, signifying that vertical gene transfer was the main mechanism to maintain these genes in the soybean rhizobia. However, lateral transfer of symbiotic genes commonly in Sinorhizobium spp. and rarely in Bradyrhizobium spp. was also detected. These results showed the genetic diversity, the biogeography, and the soil determinant factors of soybean rhizobia in Hebei province of China.