Adiponectin improves endothelial function in hyperlipidemic rats by reducing oxidative/nitrative stress and differential regulation of eNOS/iNOS activity

Plasma adiponectin level is significantly reduced in patients with metabolic syndrome, and vascular dysfunction is an important pathological event in these patients. However, whether adiponectin may protect endothelial cells and attenuate endothelial dysfunction caused by metabolic disorders remains largely unknown. Adult rats were fed with a regular or a high-fat diet for 14 wk. The aorta was isolated, and vascular segments were incubated with vehicle or the globular domain of adiponectin (gAd; 2 mug/ml) for 4 h. The effect of gAd on endothelial function, nitric oxide (NO) and superoxide production, nitrotyrosine formation, gp91(phox) expression, and endothelial nitric oxide synthase (eNOS)/inducible NOS (iNOS) activity/expression was determined. Severe endothelial dysfunction (maximal vasorelaxation in response to ACh: 70.3 +/- 3.3 vs. 95.2 +/- 2.5% in control, P < 0.01) was observed in hyperlipidemic aortic segments, and treatment with gAd significantly improved endothelial function (P < 0.01). Paradoxically, total NO production was significantly increased in hyperlipidemic vessels, and treatment with gAd slightly reduced, rather than increased, total NO production in these vessels. Treatment with gAd reduced (-78%, P < 0.01) superoxide production and peroxynitrite formation in hyperlipidemic vascular segments. Moreover, a moderate attenuation (-30%, P < 0.05) in gp91(phox) and iNOS overexpression in hyperlipidemic vessels was observed after gAd incubation. Treatment with gAd had no effect on eNOS expression but significantly increased eNOS phosphorylation (P < 0.01). Most noticeably, treatment with gAd significantly enhanced eNOS (+83%) but reduced iNOS (-70%, P < 0.01) activity in hyperlipidemic vessels. Collectively, these results demonstrated that adiponectin protects the endothelium against hyperlipidemic injury by multiple mechanisms, including promoting eNOS activity, inhibiting iNOS activity, preserving bioactive NO, and attenuating oxidative/nitrative stress.     

A short-form C-type lectin from amphioxus acts as a direct microbial killing protein via interaction with peptidoglycan and glucan

To investigate the evolution and immune function of C-type lectin in amphioxus, the primitive representative of the chordate phylum, we identified three C-type lectins consisting solely of a carbohydrate recognition domain and N-terminal signal peptide and found that they had distinct express patterns in special tissues and immune response to stimulations analyzed by quantitative real-time PCR. We characterized the biochemical and biological properties of AmphiCTL1, which was dramatically up-regulated in amphioxus challenged with Staphylococcus aureus, Saccharomyces cerevisiae, and zymosan. Immunohistochemistry demonstrated that the localization of AmphiCTL1 protein was exclusively detected in the inner folding tissues of the hepatic diverticulum. Recombinant AmphiCTL1 was characterized as a typical Ca2+-dependent carbohydrate-binding protein possessing hemagglutinating activity, preferentially bound to all examined four Gram-positive bacteria and two yeast strains, but had little binding activity toward four Gram-negative bacteria we tested. It aggregated S. aureus and S. cerevisiae in a Ca2+-dependent manner and specifically bound to insoluble peptidoglycan and glucan, but not to LPS, lipoteichoic acid, and mannan. Calcium increased the intensity of the interaction between AmphiCTL1 and those components, but was not essential. This lectin directly killed S. aureus and S. cerevisiae in a Ca2+-independent fashion, and its binding to microorganism cell wall polysaccharides such as peptidoglycan and glucan preceded microbial killing activity. These findings suggested that AmphiCTL1 acted as a direct microbial killing C-type lectin through binding microbial targets via interaction with peptidoglycan and glucan. Thus, AmphiCTL1 may be an evolutionarily primitive form of antimicrobial protein involved in lectin-mediated innate immunity.     

Human anti-CXCR4 antibodies undergo VH replacement, exhibit functional V-region sulfation, and define CXCR4 antigenic heterogeneity

The chemokine receptor CXCR4 and its ligand stromal-derived factor-1 (SDF-1/CXCL12) are essential for many biological processes and various pathological conditions. However, the relationship between CXCR4 antigenic structure and SDF-1-mediated biological responses is poorly understood. In this report, a panel of human anti-CXCR4 Abs were isolated and used to explore CXCR4 antigenic heterogeneity and function. Multiple fixed CXCR4 antigenic isoforms were detected on the surface of hemopoietic cells. Epitope mapping studies demonstrated the complex nature of the surface-exposed CXCR4 epitopes. Ab-mediated inhibition of chemotaxis correlated strongly with binding affinity, epitope recognition, as well as the level of CXCR4 isoform expression. In addition, detailed genetic analyses of these Abs showed evidence of V(H) replacement. Importantly, structural and biochemical studies demonstrated tyrosine sulfation in novel regions of the V genes that contributed bidirectionally to the binding activity of the Abs. These data provide the first evidence that functional tyrosine sulfation occurs in self-reactive Abs and suggest a potential new mechanism that may contribute to the pathogenesis of Ab-mediated autoimmune disease. These Abs also provide valuable tools to explore the selective in vivo targeting of CXCR4 isoforms that may be preferentially expressed in certain disease states and involved in steady-state CXCR4-SDF-1 homeostasis.     

S100B protein is released from rat neonatal neurons, astrocytes, and microglia by in vitro trauma and anti-S100 increases trauma-induced delayed neuronal injury and negates the protective effect of exogenous S100B on neurons

S100B protein is found in brain, has been used as a marker for brain injury and is neurotrophic. Using a well-characterized in vitro model of brain cell trauma, we have previously shown that strain injury causes S100B release from neonatal rat neuronal plus glial cultures and that exogenous S100B reduces delayed post-traumatic neuronal damage even when given at 6 or 24 h post-trauma. The purpose of the current studies was to measure post-traumatic S100B release by specific brain cell types and to examine the effect of an antibody to S100 on post-traumatic delayed (48 h) neuronal injury and the protective effect of exogenous S100B. Neonatal rat cortical cells grown on a deformable elastic membrane were subjected to a strain (stretch) injury produced by a 50 ms displacement of the membrane. S100B was measured with an ELISA kit. Trauma released S100B from pure cultures of astrocytes, microglia, and neurons. Anti-S100 reduced released S100B to below detectable levels, increased delayed neuronal injury in traumatized cells and negated the protective effect of exogenous S100B on injured cells. Heat denatured anti-S100 did not exacerbate injury. These studies provide further evidence for a protective role for S100B following neuronal trauma.     

两株滇产广义美味牛肝菌的分离培养及其分子鉴定

采用组织分离法从广义美味牛肝菌子实体分离获得2株稳定的菌株,初步研究了两菌株的分离和培养条件。用ITS序列分析,对分离菌株进行了分子鉴定。基于ITS序列构建的系统树表明两株菌属于美味牛肝菌复合群,并与夏生牛肝菌Boletus aestivalis（Paul.）Fr.有较近的亲缘关系。     

可溶性纤维连接蛋白可活化RhoA并引起癌细胞骨架相关改变

细胞外基质中的纤维连接蛋白可以使细胞表面的整合素受体聚集起来,引起RhoA介导的信号通路的活化,从而导致细胞骨架的重组和细胞迁移的调节。然而,大部分纤维连接蛋白以可溶形式存在于血浆中,这些可溶性纤维连接蛋白是否有相似的效应仍有待于进一步的研究。实验发现,向细胞培养液中加入可溶性纤维连接蛋白,可使胃癌细胞系SGC-7901中的RhoA由GDP结合的非活性形式转变为GTP结合的活性形式,与其底物结合的量增加,而α5β1整合素的抗体可以阻断这一活化过程;可溶性纤维连接蛋白可诱导细胞聚合体形式的肌动蛋白(F-肌动蛋白)的形成。在人前列腺癌细胞系PC-3中,可溶性纤维连接蛋白可引起细胞从多角形向圆形的形态改变,α5β1整合素的抗体可阻断这一改变。以上结果显示可溶性纤维连接蛋白能与α5β1整合素结合并诱导RhoA介导的信号转导。     

滇产与日产松茸的IGS1-RFLP比较分析

应用Cfr13I限制性内切酶对采自于云南省16个市县的127个松茸子实体进行了IGS1-RFLP比较分析,发现126个松茸子实体属于A类型,来自大理白族自治州剑川县的1例(TF89)为C类型。通过对A类型的IGS1序列分析发现产自云南的松茸有一个CTTT的简单重复,滇产松茸IGS1序列差异不明显。滇产松茸的IGS1-RFLP与日产松茸的主要类型十分相似,两地松茸可能是同源的。     

p90 RSK-1 associates with and inhibits neuronal nitric oxide synthase

Evidence is presented that RSK1 (ribosomal S6 kinase 1), a downstream target of MAPK (mitogen-activated protein kinase), directly phosphorylates nNOS (neuronal nitric oxide synthase) on Ser847 in response to mitogens. The phosphorylation thus increases greatly following EGF (epidermal growth factor) treatment of rat pituitary tumour GH3 cells and is reduced by exposure to the MEK (MAPK/extracellular-signal-regulated kinase kinase) inhibitor PD98059. Furthermore, it is significantly enhanced by expression of wild-type RSK1 and antagonized by kinase-inactive RSK1 or specific reduction of endogenous RSK1. EGF treatment of HEK-293 (human embryonic kidney) cells, expressing RSK1 and nNOS, led to inhibition of NOS enzyme activity, associated with an increase in phosphorylation of nNOS at Ser847, as is also the case in an in vitro assay. In addition, these phenomena were significantly blocked by treatment with the RSK inhibitor Ro31-8220. Cells expressing mutant nNOS (S847A) proved resistant to phosphorylation and decrease of NOS activity. Within minutes of adding EGF to transfected cells, RSK1 associated with nNOS and subsequently dissociated following more prolonged agonist stimulation. EGF-induced formation of the nNOS-RSK1 complex was significantly decreased by PD98059 treatment. Treatment with EGF further revealed phosphorylation of nNOS on Ser847 in rat hippocampal neurons and cerebellar granule cells. This EGF-induced phosphorylation was partially blocked by PD98059 and Ro31-8220. Together, these data provide substantial evidence that RSK1 associates with and phosphorylates nNOS on Ser847 following mitogen stimulation and suggest a novel role for RSK1 in the regulation of nitric oxide function in brain.     

Normal cellular prion protein is a ligand of selectins: binding requires Le(X) but is inhibited by sLe(X)

The normal PrP(C) (cellular prion protein) contains sLe(X) [sialyl-Le(X) (Lewis X)] and Le(X). sLe(X) is a ligand of selectins. To examine whether PrP(C) is a ligand of selectins, we generated three human PrP(C)-Ig fusion proteins: one with Le(X), one with sLe(X), and the other with neither Le(X) nor sLe(X). Only Le(X)-PrP(C)-Ig binds E-, L- and P-selectins. Binding is Ca(2+)-dependent and occurs with nanomolar affinity. Removal of sialic acid on sLe(X)-PrP(C)-Ig enables the fusion protein to bind all selectins. These findings were confirmed with brain-derived PrP(C). The selectins precipitated PrP(C) in human brain in a Ca(2+)-dependent manner. Treatment of brain homogenates with neuraminidase increased the amounts of PrP(C) precipitated. Therefore the presence of sialic acid prevents the binding of PrP(C) in human brain to selectins. Hence, human brain PrP(C) interacts with selectins in a manner that is distinct from interactions in peripheral tissues. Alternations in these interactions may have pathological consequences.     

Genetic relationships among Chinese pigs and other pig populations from Hunan Province, China

In this study, protein-level polymorphisms of transferrin, pre-albumin, hemopexin, ceruloplasmin and amylase were investigated in Hunan native pigs and Large Yorkshire pigs collected from Hunan (a province of China), allowing calculations of allele frequencies, average heterozygosities, inbreeding coefficients and genetic distances. The genetic relationship between Southeast Asian native pigs and American pigs was more distant than those among Southeast Asian native pig breeds. The genetic relationship between Southeast Asian native pig breeds and Hampshire pigs was the most distant.