Polyamines induce rapid biosynthesis of nitric oxide (NO) in Arabidopsis thaliana seedlings

In this study, we examined the regulation by putrescine, spermidine and spermine of nitric oxide (NO) biosynthesis in Arabidopsis thaliana seedlings. Using a fluorimetric method employing the cell-impermeable NO-binding dye diaminorhodamine-4M (DAR-4M), we observed that the polyamines (PAs) spermidine and spermine greatly increased NO release in the seedlings, whereas arginine and putrescine had little or no effect. Spermine, the most active PA, stimulated NO release with no apparent lag phase. The response was quenched by addition of 2-aminoethyl-2-thiopseudourea (AET), an inhibitor of the animal nitric oxide synthase (NOS) and plant NO biosynthesis, and by 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-1-oxy-3-oxide (PTIO), an NO scavenger. By fluorescence microscopy, using the cell-permeable NO-binding dye diaminorhodamine-4M acetoxymethyl ester (DAR-4M AM), we observed that PAs induced NO biosynthesis in specific tissues in Arabidopsis seedlings. Spermine and spermidine increased NO biosynthesis in the elongation zone of the Arabidopsis root tip and in primary leaves, especially in the veins and trichomes, while in cotyledons little or no effect of PAs beyond the endogenous levels of NO-induced fluorescence was observed. We conclude that PAs induce NO biosynthesis in plants.     

Cations in component reactions of `malic'' enzyme catalysis

The ;malic' enzyme (EC 1.1.1.40) has been purified (300-fold) from wheat germ and its abilities to catalyse the decarboxylation and the hydrogenation of oxaloacetic acid and oxaloacetate esters was studied. The free 1-carboxyl group is essential for the interaction of oxaloacetates and substituted oxaloacetates with the enzyme via cations. The free 4-carboxyl group is required for the decarboxylation but is not indispensable for the hydrogenation. At high concentrations, cations inhibit the enzymic hydrogenation of oxaloacetic acid but not that of 4-ethyl oxaloacetate. A plausible inhibitory mechanism is proposed.     

The Effects of Rosuvastatin on the Serum Cortisol,Serum Lipid,and Serum Mevalonic Acid Levels in the Healthy Indian Male Population

In this open-label, balanced, randomized, placebo-controlled, parallel study, healthy male volunteers were randomly divided into two groups. Each group received either a single oral dose of rosuvastatin 20 mg or placebo. Estimations were done at predose on day 1 of dosing (baseline) and 24 h postdose after days 7 and 14. Serum cortisol and serum lipid levels were estimated using enzyme-linked immunosorbent assay kits and serum mevalonic acid (MVA) levels were measured using validated liquid chromatography–tandem mass spectrometry method. Rosuvastatin produced a statistically significant (P < 0.05) decrease in total cholesterol, low-density lipoprotein cholesterol, very low-density lipoprotein cholesterol, and triglycerides. However, the increase in high-density lipoprotein cholesterol and decrease in cortisol and MVA were not statistically significant when compared to the placebo-treated group. The study showed that rosuvastatin at a dose of 20 mg/day for a period of 14 days was very potent as cholesterol-lowering agent, without any significant change in serum cortisol level in the healthy Indian male population.     

A dysregulated endocannabinoid-eicosanoid network supports pathogenesis in a mouse model of Alzheimer's disease

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Highlights? Monoacylglycerol lipase (MAGL) is a major contributor to brain arachidonic acid pools ? MAGL mutation decreases neuroinflammation and amyloid β in Alzheimer's disease mouse ? MAGL blockade recapitulates brain prostaglandin and cytokine-lowering effects ? MAGL inhibitors may be a next-generation strategy for combating Alzheimer's disease     

Keratinocyte-derived Chemokine in Obesity: EXPRESSION, REGULATION, AND ROLE IN ADIPOSE MACROPHAGE INFILTRATION AND GLUCOSE HOMEOSTASIS*

Obese adipose tissue (AT) is associated with chronic inflammation, and we hypothesized that the keratinocyte-derived chemokine (KC), the mouse ortholog of human interleukin-8, plays a role in obesity-mediated AT inflammation and the subsequent manifestation of insulin resistance. KC expression is increased in the AT and plasma of genetically (ob/ob) and high fat diet-induced obese mouse models, and this increase may be mediated by the elevated leptin and tumor necrosis factor-α levels associated with obesity. Obesity-induced KC expression occurs primarily in stromal vascular cells and not in adipocytes, and it is high in preadipocytes and decreases during adipogenesis. Although KC has no effect on adipogenesis, it induces adipocyte expression of inflammatory factors and the insulin resistance mediator, suppressor of cytokine signaling 3. Using chimeric mice deficient in the KC receptor CXCR2 in their bone marrow, we show that the lack of CXCR2 in hematopoietic cells is sufficient to protect from adipose and skeletal muscle macrophage recruitment and development of insulin resistance in diet-induced obese mice. These studies suggest that KC and its receptor CXCR2 are potential targets for the development of new therapeutic approaches for treatment of obesity-related insulin resistance, type II diabetes, and related cardiovascular diseases.Obesity is characterized by systemic low grade inflammation that appears to contribute to the genesis of insulin resistance (IR),³ type 2 diabetes, and increased risk for cardiovascular diseases (reviewed in Ref. 1). Furthermore, adipose tissue (AT) produces a variety of inflammatory factors, and its excessive development in obesity is associated with accumulation of AT macrophages (ATMs) (1), whose recruitment and proinflammatory activation are required for the development of IR in obese mice (reviewed in Ref. 2). An important question concerning ATMs is/are the trigger(s) driving the recruitment of these cells in obesity.Efforts at identifying factors that attract and recruit ATMs have mostly focused on the CC chemokine MCP-1 (monocyte chemoattractant protein-1) and its receptor CCR2. These studies have led to contradicting results with several publications showing that MCP-1 and CCR2 are important for ATM recruitment and the subsequent development of IR (3–5), whereas others show no involvement of this chemokine and its receptor in these processes (6–8). Furthermore, the studies that claim a role for MCP-1 and CCR2 in ATM recruitment and IR show that deficiency of the ligand or the receptor did not result in normalization of ATM content, indicating that other factors also participate in ATM recruitment. These findings suggest that the precise role of the MCP-1/CCR2 axis in ATM recruitment and IR is unclear, and that other chemokines and their receptors could also play a role in these processes. One such chemokine is interleukin 8 (IL-8), the prototypical CXC chemokine known to recruit and activate monocytes and to attract polymorphonuclear leukocytes to sites of inflammation (9). IL-8 is elevated in plasma of obese subjects (10, 11) and correlates with adiposity and insulin sensitivity, suggesting an involvement of this chemokine in obesity-related health complications (12–14). Additionally, IL-8 is implicated in the pathogenesis of atherosclerosis and cardiovascular disease, two obesity-associated disorders (15). Finally, IL-8 is an angiogenic factor, and angiogenesis is a hallmark of AT expansion in obesity (16). Although these findings suggest an important role for IL-8 in AT biology and pathology, little is known regarding the mechanism of regulation of IL-8 in obesity and its role in AT biology and pathology. This is probably due, in part, to the absence of suitable animal models because mice and rats do not have a clear-cut IL-8 ortholog (17).Although rodent keratinocyte-derived chemokine (KC) shows the highest homology with human growth-related oncogene (GRO-α), it appears to be the closest equivalent to IL-8, as judged by its pattern of expression and putative function (18). Monocytes express the KC receptor (CXCR2), and KC triggers monocyte arrest on early atherosclerotic endothelium, one of the first steps in the invasion of tissues by macrophages (19). Interaction of monocyte CXCR2 with its ligand KC leads to up-regulation of α₄β₁ integrin affinity and firm adhesion to the endothelium (19). Furthermore, both KC and its receptor play a central role in macrophage infiltration and accumulation in atherosclerotic lesions in mice (20, 21). However, no information is currently available regarding the role of KC in macrophage recruitment in obese AT or its role in AT biology and pathology.In this study, we show that KC expression is elevated in AT and plasma of genetically (ob/ob) and diet-induced obese (DIO) mice, probably as the result of increased leptin and tumor necrosis factor α (TNF-α) levels associated with obesity. We also show that obesity-induced KC is mostly derived from nonadipocyte sources in AT and that KC does not affect adipocyte differentiation but does increase pro-inflammatory cytokine expression in adipocytes. Finally, we show in a DIO model in chimeric mice lacking CXCR2 on their macrophages that the KC receptor plays an important role in macrophage accumulation in adipose and skeletal muscle tissue and subsequent development of IR.     

KSGal6ST Is Essential for the 6-Sulfation of Galactose within Keratan Sulfate in Early Postnatal Brain

Keratan sulfate (KS) comprises repeating disaccharides of galactose (Gal) and N-acetylglucosamine (GlcNAc). Residues of Gal and GlcNAc in KS are potentially modified with sulfate at their C-6 positions. The 5D4 monoclonal antibody recognizes KS structures containing Gal and GlcNAc, both 6-sulfated, and has been used most extensively to evaluate KS expression in mammalian brains. We previously showed that GlcNAc6ST1 is an enzyme responsible for the synthesis of the 5D4 epitope in developing brain and in the adult brain, where it is induced after injury. It has been unclear which sulfotransferase is responsible for Gal-6-sulfation within the 5D4 KS epitope in developing brains. We produced mice deficient in KSGal6ST, a Gal-6-sulfotransferase. Western blotting and immunoprecipitation revealed that all 5D4-immunoreactivity to proteins, including phosphacan, were abolished in KSGal6ST-deficient postnatal brains. Likewise, the 5D4 epitope, expressed primarily in the cortical marginal zone and subplate and dorsal thalamus, was eliminated in KSGal6ST-deficient mice. Disaccharide analysis showed the loss of Gal-6-sulfate in KS of the KSGal6ST-deficient brains. Transfection studies revealed that GlcNAc6ST1 and KSGal6ST cooperated in the expression of the 5D4 KS epitope in HeLa cells. These results indicate that KSGal6ST is essential for C-6 sulfation of Gal within KS in early postnatal brains.     

Dragon (repulsive guidance molecule b) inhibits IL-6 expression in macrophages

Despite the strong interest in the NK cell-mediated immunity toward malignant cells and viruses, there is a relative lack of data on the interplay between NK cells and filamentous fungi, especially Aspergillus fumigatus, which is the major cause of invasive aspergillosis. By studying the in vitro interaction between human NK cells and A. fumigatus, we found only germinated morphologies to be highly immunogenic, able to induce a Th1-like response, and capable of upregulating cytokines such as IFN-γ and TNF-α. Moreover, priming NK cells with human rIL-2 and stimulating NK cells by direct NK cell-pathogen contact were essential to induce damage against A. fumigatus. However, the most interesting finding was that NK cells did not mediate anti-Aspergillus cytotoxicity through degranulation of their cytotoxic proteins (perforin, granzymes, granulysine), but via an alternative mechanism involving soluble factor(s). To our knowledge, our study is the first to demonstrate that IFN-γ, released by NK cells, directly damages A. fumigatus, attributing new properties to both human NK cells and IFN-γ and suggesting them as possible therapeutic tools against IA.