Enhanced endothelial nitric oxide-synthase activity in mice infected with Trypanosoma brucei

Infection of humans with Trypanosoma brucei causes sleeping sickness, which is invariably fatal if left untreated. The course of infection is characterised, among others, by multiple organ damage including cardiovascular dysfunctions such as hypotension and breakdown of the blood-brain barrier. The latter eventually leads to the parasite invasion into central nervous system and ultimately to the death of the patient. Nitric oxide (NO) synthesised from L-arginine via endothelial NO-synthase (eNOS) is involved in the control of vascular tone and permeability. The present study explores the effect of T. brucei infection on the endothelium-dependent in vitro vasomotor response of isolated mouse aortas. Aorta rings were suspended in organ chambers for isometric tension recording. The endothelium-dependent NO-mediated relaxation in response to acetylcholine (10(-9) to 10(-5) M) was markedly enhanced in the infected mice compared to controls (P<0.05), whereas the endothelium-independent vasodilation to an exogenous NO-donor, sodium nitroprusside, was comparable in both groups. Norepinephrine-stimulated contraction was also comparable in the absence or presence of the NO-synthase inhibitor N(omega)-Nitro-L-arginine methyl ester (L-NAME; 10(-4)M) in both groups. The enhanced endothelium-dependent relaxation in the infected mice correlated well with a 3.5-fold increase in eNOS protein level in these aortas as compared to those of control mice (P=0.05). Thus, T. brucei infection enhances eNOS protein expression in the endothelium, causing a pronounced vasodilation. Overproduction of NO in trypanosomiasis may be involved in the observed generalised hypotension and in an increased vascular permeability that facilitates T. brucei invasion into surrounding tissues and its penetration into the central nervous system in later phases of infection.     

‘兰箭3号’箭筈豌豆荚果发育动态及腹缝线结构研究

箭筈豌豆(Vicia sativa)是高海拔地区重要的一年生豆科牧草,但荚果成熟时的开裂现象会造成种子的严重损失。该研究以栽培品种‘兰箭3号’为对象,对其荚果在发育过程中的形态特征、水分含量、腹缝线表面结构及腹缝线横截面解剖结构的动态变化进行观察分析,以探讨箭筈豌豆荚果的裂荚机理,为生产中确定种子收获的适宜时间提供理论依据。结果显示:(1)‘兰箭3号’约在盛花后25~30d荚果变为浅棕色,此时荚果已完成生理成熟,且荚果的大小和干重均达到最大值,含水量降到最小值;盛花后25d荚果腹缝线出现裂缝,盛花后35d腹缝线完全裂开。(2)‘兰箭3号’于盛花后20d腹缝线处离层细胞开始解体;盛花后25d,内、中、外果皮的薄壁细胞均开始失水皱缩,其中内果皮的薄壁细胞部分已开始破裂,离层细胞及其下面的薄壁细胞完全解体,外部果瓣缘细胞内侧细胞壁破裂,但外侧异常加厚的细胞壁仍然保持完整并连接两个果瓣,使荚果不开裂;盛花后30~35d,内、中、外果皮的薄壁细胞完全失水,细胞壁皱缩在一起,同时外部果瓣缘细胞外侧细胞壁断裂成两部分,荚果的两个果瓣裂开。研究表明,盛花后25~30d荚果失去绿色变为浅棕色时是‘兰箭3号’的适宜收获时间,且离层和细胞失水产生的机械拉力是导致箭筈豌豆荚果开裂的主要原因,推测外部果瓣缘细胞外侧增厚融合的细胞壁很可能是‘兰箭3号’抵抗裂荚的关键结构。     

巨噬细胞产生NO.和O_2~-自由基的分子机理

建立了用顺磁共振（ＥＳＲ）和化学发光技术测定巨噬细胞产生ＮＯ和氧自由基的方法．捕捉到了巨噬细胞受佛波酯刺激产生的ＮＯ．和Ｏ－２自由基．测定了在不同浓度Ｌ－精氨酸存在时佛波酯刺激后巨噬细胞产生的ＮＯ自由基．研究了巨噬细胞产生的ＮＯ和氧自由基的分子机理．结果表明巨噬细胞不仅产生氧自由基而且产生ＮＯ自由基．ＮＡＤＰＨ氧化酶产生氧自由基的部位位于巨噬细胞膜的外侧．ＮＯ合成酶活化产生ＮＯ自由基比ＮＡＤＰＨ氧化酶活化产生氧自由基晚几分钟．     

Soil N and C trace gas fluxes and microbial soil N turnover in a sessile oak (Quercus petraea (Matt.) Liebl.) forest in Hungary

During two intensive field campaigns in summer and autumn 2004 nitrogen (N₂O, NO/NO₂) and carbon (CO₂, CH₄) trace gas exchange between soil and the atmosphere was measured in a sessile oak (Quercus petraea (Matt.) Liebl.) forest in Hungary. The climate can be described as continental temperate. Fluxes were measured with a fully automatic measuring system allowing for high temporal resolution. Mean N₂O emission rates were 1.5 μg N m⁻² h⁻¹ in summer and 3.4 μg N m⁻² h⁻¹ in autumn, respectively. Also mean NO emission rates were higher in autumn (8.4 μg N m⁻² h⁻¹) as compared to summer (6.0 μg N m⁻² h⁻¹). However, as NO₂ deposition rates continuously exceeded NO emission rates (−9.7 μg N m⁻² h⁻¹ in summer and −18.3 μg N m⁻² h⁻¹ in autumn), the forest soil always acted as a net NO _x sink. The mean value of CO₂ fluxes showed only little seasonal differences between summer (81.1 mg C m⁻² h⁻¹) and autumn (74.2 mg C m⁻² h⁻¹) measurements, likewise CH₄uptake (summer: −52.6 μg C m⁻² h⁻¹; autumn: −56.5 μg C m⁻² h⁻¹). In addition, the microbial soil processes net/gross N mineralization, net/gross nitrification and heterotrophic soil respiration as well as inorganic soil nitrogen concentrations and N₂O/CH₄ soil air concentrations in different soil depths were determined. The respiratory quotient (ΔCO_{2 resp} ΔO_{2 resp}⁻¹) for the uppermost mineral soil, which is needed for the calculation of gross nitrification via the Barometric Process Separation (BaPS) technique, was 0.8978 ± 0.008. The mean value of gross nitrification rates showed only little seasonal differences between summer (0.99 μg N kg⁻¹ SDW d⁻¹) and autumn measurements (0.89 μg N kg⁻¹ SDW d⁻¹). Gross rates of N mineralization were highest in the organic layer (20.1–137.9 μg N kg⁻¹ SDW d⁻¹) and significantly lower in the uppermost mineral layer (1.3–2.9 μg N kg⁻¹ SDW d⁻¹). Only for the organic layer seasonality in gross N mineralization rates could be demonstrated, with highest mean values in autumn, most likely caused by fresh litter decomposition. Gross mineralization rates of the organic layer were positively correlated with N₂O emissions and negatively correlated with CH₄ uptake, whereas soil CO₂ emissions were positively correlated with heterotrophic respiration in the uppermost mineral soil layer. The most important abiotic factor influencing C and N trace gas fluxes was soil moisture, while the influence of soil temperature on trace gas exchange rates was high only in autumn.     

BH(4) (tetrahydrobiopterin)-dependent activation, but not the expression, of inducible NOS (nitric oxide synthase)-2 in proinflammatory cytokine-stimulated, cultured normal human astrocytes is mediated by MEK-ERK kinases

Nitric oxide (NO) from astrocytes is one of the signalers used by the brain's extensive glial-neuronal-vascular network, but its excessive production by pro-inflammatory cytokine-stimulated glial cells can be cytodestructive. Here, we show how three pro-inflammatory cytokines (IL-1beta, TNF-alpha, and IFN-gamma) together stimulated the activation, but not the prior expression, of NOS-2 protein via a mechanism involving MEK-ERKs protein kinases in astrocytes from adult human cerebral temporal cortex. The cytokines triggered a transient burst of p38 MAPK activity and the production of NOS-2 mRNA which were followed by bursts of MEK-ERK activities, synthesis of the NOS-2 co-factor tetrahydrobiopterin (BH(4)), a build-up of NOS-2 protein and from it active NOS-2 enzyme. Selectively inhibiting MEK1/MEK2, but not the earlier burst of p38 MAPK activity, with a brief exposure to U0126 between 24 and 24.5 h after adding the cytokine triad affected neither NOS-2 expression nor NOS-2 protein accumulation but stopped BH(4) synthesis and the assembly of the NOS-2 protein into active NOS-2 enzyme. The complete blockage of active NOS-2 production by the brief exposure to U0126 was bypassed by simply adding BH(4) to the culture medium. Therefore, this cytokine triad triggered two completely separable, tandem operating mechanisms in normal human astrocytes, the first being NOS-2 gene expression and accumulation of NOS-2 protein and the second being the synthesis of the BH(4) factor needed to dimerize the NOS-2 protein into active, NO-making NOS-2 enzyme.     

Structure-activity relationships of pyrrole based S-nitrosoglutathione reductase inhibitors: pyrrole regioisomers and propionic acid replacement

S-Nitrosoglutathione reductase (GSNOR) is a member of the alcohol dehydrogenase family (ADH) that regulates the levels of S-nitrosothiols (SNOs) through catabolism of S-nitrosoglutathione (GSNO). GSNO and SNOs are implicated in the pathogenesis of many diseases including those in respiratory, cardiovascular, and gastrointestinal systems. The pyrrole based N6022 was recently identified as a potent, selective, reversible, and efficacious GSNOR inhibitor which is currently undergoing clinical development. We describe here the synthesis and structure-activity relationships (SAR) of novel pyrrole based analogues of N6022 focusing on scaffold modification and propionic acid replacement. We identified equally potent and novel GSNOR inhibitors having pyrrole regioisomers as scaffolds using a structure based approach.     

Sphingomyelinase causes endothelium-dependent vasorelaxation through endothelial nitric oxide production without cytosolic Ca(2+) elevation

Neutral sphingomyelinase (N-SMase) elevated nitric oxide (NO) production without affecting intracellular Ca(2+) concentration ([Ca(2+)](i)) in endothelial cells in situ on aortic valves, and induced prominent endothelium-dependent relaxation of coronary arteries, which was blocked by N(omega)-monomethyl-L-arginine, a NO synthase (NOS) inhibitor. N-SMase induced translocation of endothelial NOS (eNOS) from plasma membrane caveolae to intracellular region, eNOS phosphorylation on serine 1179, and an increase of ceramide level in endothelial cells. Membrane-permeable ceramide (C(8)-ceramide) mimicked the responses to N-SMase. We propose the involvement of N-SMase and ceramide in Ca(2+)-independent eNOS activation and NO production in endothelial cells in situ, linking to endothelium-dependent vasorelaxation.     

Influence of partial defoliation of green pepper on the senescence,growth, and nitrate reductase of the remaining leaf

Summary Removal of all but one leaf from pepper plants prevented the senescence of the remaining leaf and caused increases of approximately 140, 200, and 200%, respectively in leaf area, weight, and nitrate reductase activity. Development of the fruit (fresh and dry weight increases) was only approximately 65% of that of fruit on control plants.     

Kiwifruit extracts inhibit cytokine production by lipopolysaccharide-activated macrophages, and intestinal epithelial cells isolated from IL10 gene deficient mice

Inflammatory bowel disease (IBD) is a chronic, inflammatory disorder of the gastrointestinal tract involving an inappropriate immune response to commensal microorganisms in a genetically susceptible host. This study examined the effects of aqueous and ethyl acetate extracts of gold kiwifruit (Actinidia chinensis) or green kiwifruit (Actinidia deliciosa) using in vitro models of IBD. These models comprised primary macrophages and intestinal epithelial cells isolated from C57BL/5J and interleukin-10 gene deficient (Il10^−/−) mice and RAW 264.7, a murine macrophage-like cell line. All four kiwifruit extracts reduced the activation of these models after lipopolysaccharide stimulation, decreasing nitric oxide and cytokine secretion by both Il10^−/− and wild-type cells. The ethyl acetate extracts exhibited the highest anti-inflammatory activity, with almost complete suppression of lipopolysaccharide-stimulated macrophage activation. These results suggest that kiwifruit extracts have significant anti-inflammatory activity relevant to IBD. We suggest that the Il10^−/− mouse is a suitable model for further study of these compounds.     

链脲佐菌素致胰岛NO自由基损伤模型的建立和应用

以链脲佐菌素Streptozotocin（简称STZ）为糖尿病的诱因,以NO自由基含量为响应指标,建立了体外小鼠胰岛水平糖尿病药物筛选模型。当STZ作用浓度在0～50mmol/L内变化时,培养液中被检测到的NO大部分是来源于STZ溶于水后释放出的,而很小一部分是由胰岛培养物自身释放的,后者稳定在30～35mmol/L之间。另一方面,NO含量与胰岛素分泌量的剂量关系表明NO的增加伴随着胰岛素分泌量的下降,这标志着NO对胰岛功能的氧化性损伤,从而验证了NO作为该模型响应参量的可靠性。最终确定STZ致胰岛NO自由基损伤模型中STZ的作用浓度为5.0mmol/L,此时NO含量和胰岛素分泌量分别为STZ未加入前的10.81倍和0.43倍。最后应用该模型,快捷地考察了不同铬含量的魔芋葡甘露寡糖铬络合物（简称KOSCr）清除NO自由基的能力。