Monitoring the caspase cascade in single apoptotic cells using a three-color fluorescent protein substrate

Fluorescence cross-correlation spectroscopy (FCCS) reveals information about the spatiotemporal coincidence of two spectrally well-defined fluorescent molecules in a small observation area at the level of single-molecule sensitivity. To simultaneously evaluate the activities of caspase-3 and caspase-9, we constructed a chimeral protein that consisted of tandemly fused enhanced cyan fluorescent protein (ECFP), monomeric red fluorescent protein (mCherry) and monomeric yellow fluorescent protein (Venus). In HeLa cell lysates, a combination of tumor necrosis factor-α (TNF-α)- and cycloheximide (CHX-)-induced apoptosis was monitored. In this, decreases of cross-correlation amplitudes were observed between ECFP and mCherry and between mCherry and Venus. Moreover, time-dependent monitoring of single cells revealed decreases in the cross-correlation amplitudes between ECFP and mCherry and between mCherry and Venus before morphologic changes were observed by laser scanning fluorescence microscopy (LSM). Thus, our method could predict the fate of the cell in the early apoptotic stage.     

HCVE2／NS1相对保守区多肽体外诱导慢丙肝患者PBMC增殖反应的研究

为了解ＨＣＶ感染后细胞免疫在其中的作用,对３１例慢丙肝及２０例正常献血员以ＭＴＴ法检测外周血单核细胞（ＰＢＭＣ）对ＨＣＶＥ２／ＮＳ１相对保守区多肽抗原及Ｃ２２、ＮＳ５的增殖反应。结果表明与正常人相比,慢性丙型肝炎患者ＰＢＭＣ对ＨＣＶＣ２２、Ｅ２／ＮＳ１抗原有明显的增殖反应（Ｐ＜００５）,而对ＮＳ５无明显增殖,其中以Ｃ２２抗原性最强。作者认为慢性丙型肝炎患者存在针对与ＨＣＶ相关抗原的细胞免疫,这种细胞免疫不能消除ＨＣＶ感染,而且与疾病的状态无关。     

Nitric oxide production is associated with response to brown planthopper infestation in rice

Nilaparvata lugens Stål, the brown planthopper (BPH), is one of the most destructive phloem-feeding insects of rice (Oryza sativa L.) throughout Asia. Here, we show that BPH feeding increases the level of endogenous nitric oxide (NO) in the leaf and sheath tissue of both resistant and susceptible rice cultivars. However, in the roots, the NO level increased in the resistant cultivar, but decreased in the susceptible one. A burst of NO production occurred in the sheath within 1 h of infestation with BPH. The production of NO in response to BPH feeding appears to be dependent primarily on the activity of nitric oxide synthase. The application of exogenous NO reduced plant water loss by its effect on both stomatal opening and root architecture. It also stimulated the expression of certain drought stress-related genes, reduced plant height and delayed leaf senescence. Over the short term, NO supplementation reduced the seedling mortality caused by BPH feeding. This suggests that NO signaling plays a role in the rice tolerance response to BPH feeding.     

Green fluorescent protein expression triggers proteome changes in breast cancer cells

Green fluorescent protein (GFP) is the most commonly used reporter of expression in cell biology despite evidence that it affects the cell physiology. The molecular mechanism of GFP-associated modifications has been largely unexplored. In this paper we investigated the proteome modifications following stable expression of GFP in breast cancer cells (MDA-MB-231). A combination of three different proteome analysis methods (2-DE, iTRAQ, label-free) was used to maximise proteome coverage. We found that GFP expression induces changes in expression of proteins that are associated with protein folding, cytoskeletal organisation and cellular immune response. In view of these findings, the use of GFP as a cell reporter should be carefully monitored.     

Effect of Long Term Exposure to High CO2 Concentrations on Photosynthetic Characteristics of Prunus Avium L. Plants

The effect of two elevated carbon dioxide concentrations, 700 µmol(CO₂) mol^–1 (C700) and 1 400 µmol(CO₂) mol^–1 (C1400), on photosynthetic performances of 1-year-old Prunus avium L. plant was studied. Plants grown at C700 were characterised by increased net photosynthetic rate (P _N) as compared to those grown at C1400. Plant photosynthetic adjustment to C1400 resulted in 27 % higher P _N than in control at atmospheric CO₂ concentration (C _a) at the beginning of the experiment (3–4 weeks) with a consequent decline to the end of the experiment. Thus, 1 400 µmol(CO₂) mol^–1 had short-term stimulatory effect on plant P _N. Both chlorophyll (Chl) a and b concentrations dramatically decreased during exposure to C1400. Compensation irradiance was increased by 57 % in C700 and by 87 % in C1400. Photochemical efficiency () was affected by balloon environment, however, a clear stimulatory effect of C700 was detected. Opposite influence of both elevated CO₂ concentrations on P _Nmax was established: slight increase by C700 (2.7 % at C_a), but considerable decrease by C1400 (63 % at C_a). Exposure to C700 enhanced compensation irradiance by 42 %, while C1400 by only 21 %. Either C700 or C1400 did not reduce stomatal conductance (g _s). Leaf area per plant (LAR) was more stimulated by C700 than by C1400. High unit area leaf mass, specific leaf area, and dry matter accumulation in roots without affecting tissue density characterised plants grown in C1400. However, when considering the root : shoot ratio, these plants allocated less carbon to the roots than plants from other treatments.     

Similarities and differences between the responses induced in human phagocytes through activation of the medium chain fatty acid receptor GPR84 and the short chain fatty acid receptor FFA2R

GPR84 is a recently de-orphanized member of the G-protein coupled receptor (GPCR) family recognizing medium chain fatty acids, and has been suggested to play important roles in inflammation. Due to the lack of potent and selective GPR84 ligands, the basic knowledge related to GPR84 functions is very limited. In this study, we have characterized the GPR84 activation profile and regulation mechanism in human phagocytes, using two recently developed small molecules that specifically target GPR84 agonistically (ZQ16) and antagonistically (GLPG1205), respectively. Compared to our earlier characterization of the short chain fatty acid receptor FFA2R which is functionally expressed in neutrophils but not in monocytes, GPR84 is expressed in both cell types and in monocyte-derived macrophages. In neutrophils, the GPR84 agonist had an activation profile very similar to that of FFA2R. The GPR84-mediated superoxide release was low in naïve cells, but the response could be significantly primed by TNFα and by the actin cytoskeleton disrupting agent Latrunculin A. Similar to that of FFA2R, a desensitization mechanism bypassing the actin cytoskeleton was utilized by GPR84. All ZQ16-mediated cellular responses were sensitive to GLPG1205, confirming the GPR84-dependency. Finally, our data of in vivo transmigrated tissue neutrophils indicate that both GPR84 and FFA2R are involved in neutrophil recruitment processes in vivo.In summary, we show functional similarities but also some important differences between GPR84 and FFA2R in human phagocytes, thus providing some mechanistic insights into GPR84 regulation in blood neutrophils and cells recruited to an aseptic inflammatory site in vivo.     

黄瓜叶片衰老过程中抗坏血酸含量与生理指标关系的研究

研究了黄瓜叶片衰老过程中抗坏血酸(ASA)含量及相关生理指标的变化,进一步分析了ASA与各相关生理指标之间的关系。结果显示,随着叶片逐渐衰老,丙二醛(MDA)含量、相对电导率(REC)及过氧化物酶(POD)活性显著升高;光合色素、可溶性蛋白质含量、超氧化物歧化酶(SOD)与过氧化氢酶(CAT)活性及ASA含量显著降低;ASA与MDA含量、REC、POD活性之间呈显著负相关关系,ASA与CAT及SOD活性之间呈显著正相关关系。研究表明,ASA对减缓叶片衰老有重要作用。     

Effect of maturation duration on desiccation tolerance in hybrid larch (Larix×leptoeuropaea dengler) somatic embryos

Summary Cotyledonary somatic embryos ofLarix × leptoeuropaea that developed after various maturation times on media containing abscisic acid showed different frequencies of conversion into plants. Drying of these somatic embryos under high relative humidity (RH) before germination improved plantlet recovery and eliminated differences in the performance of somatic embryos matured for different times. However, dehydration of somatic embryos under 98% RH to a water content below that of zygotic embryos excised from mature seeds (0.97 and 1.36 g H₂O/g dry weight, respectively) showed a strong positive correlation between longer maturation time and desiccation tolerance. Drying somatic embryos at 4° C under 59% RH for 1 wk resulted in desiccation to a water content of 0.30 g H₂O/g dry weight, which was the closest to the hydration state of zygotic embryos in dried, stored seeds (0.20 g H₂O/g dry weight). Under this condition, only somatic embryos matured for 5 wk germinated and produced plantlets at a relatively high frequency (73 and 41%, respectively).