IFN-gamma-producing dendritic cells are important for priming of gut intraepithelial lymphocyte response against intracellular parasitic infection

The importance of intraepithelial lymphocytes (IEL) in immunoprotection against orally acquired pathogens is being increasingly recognized. Recent studies have demonstrated that Ag-specific IEL can be generated and can provide an important first line of defense against pathogens acquired via oral route. However, the mechanism involved in priming of IEL remains elusive. Our current study, using a microsporidial model of infection, demonstrates that priming of IEL is dependent on IFN-gamma-producing dendritic cells (DC) from mucosal sites. DC from mice lacking the IFN-gamma gene are unable to prime IEL, resulting in failure of these cells to proliferate and lyse pathogen-infected targets. Also, treatment of wild-type DC from Peyer's patches with Ab to IFN-gamma abrogates their ability to prime an IEL response against Encephalitozoon cuniculi in vitro. Moreover, when incubated with activated DC from IFN-gamma knockout mice, splenic CD8(+) T cells are not primed efficiently and exhibit reduced ability to home to the gut compartment. These data strongly suggest that IFN-gamma-producing DC from mucosal sites play an important role in the generation of an Ag-specific IEL response in the small intestine. To our knowledge, this report is the first demonstrating a role for IFN-gamma-producing DC from Peyer's patches in the development of Ag-specific IEL population and their trafficking to the gut epithelium.     

Mannose-binding lectin A-deficient mice have abrogated antigen-specific IgM responses and increased susceptibility to a nematode infection

To investigate the role of mannose-binding lectin-A (MBL-A) in protection against infectious disease, MBL-A(-/-)-deficient mice were generated. Using a well-characterized mouse model of human filarial nematode infection, nematode survival and protective immune responses were tested in vivo. Blood-borne Brugia malayi microfilariae survived for significantly longer time periods in MBL-A(-/-) than in wild-type (WT) mice. However, no differences in either splenic cytokine responses or induction of leukocytes in the blood were observed. A profound abrogation of Ag-specific IgM levels was measured in B. malayi-infected MBL-A(-/-) mice, and some IgG isotypes were higher than those observed in WT animals. To establish whether there was a defect in Ab responses per se in MBL-A(-/-) mice or the effect was specific to filarial infection, we immunized these mice with OVA or a carbohydrate-free protein. Significantly, Ag-specific IgM responses were defective to both of these Ags, and Ag-specific IgG responses were largely unaffected. Furthermore, in naive mice, total IgM levels did not differ between MBL-A(-/-) and WT mice. This article describes the first demonstration that MBL-A may function independently of MBL-C and suggests that MBL-A, like other C-type lectins and members of the complement cascade, is intimately involved in the priming of the humoral Ab response.     

Evolution and function of red pigmentation in land plants

BackgroundLand plants commonly produce red pigmentation as a response to environmental stressors, both abiotic and biotic. The type of pigment produced varies among different land plant lineages. In the majority of species they are flavonoids, a large branch of the phenylpropanoid pathway. Flavonoids that can confer red colours include 3-hydroxyanthocyanins, 3-deoxyanthocyanins, sphagnorubins and auronidins, which are the predominant red pigments in flowering plants, ferns, mosses and liverworts, respectively. However, some flowering plants have lost the capacity for anthocyanin biosynthesis and produce nitrogen-containing betalain pigments instead. Some terrestrial algal species also produce red pigmentation as an abiotic stress response, and these include both carotenoid and phenolic pigments.ScopeIn this review, we examine: which environmental triggers induce red pigmentation in non-reproductive tissues; theories on the functions of stress-induced pigmentation; the evolution of the biosynthetic pathways; and structure–function aspects of different pigment types. We also compare data on stress-induced pigmentation in land plants with those for terrestrial algae, and discuss possible explanations for the lack of red pigmentation in the hornwort lineage of land plants.ConclusionsThe evidence suggests that pigment biosynthetic pathways have evolved numerous times in land plants to provide compounds that have red colour to screen damaging photosynthetically active radiation but that also have secondary functions that provide specific benefits to the particular land plant lineage.     

A novel amino-on CPG-support for the synthesis of 3'-aminoalkylated oligonucleotides

The synthesis of a novel amino-ON CPG support and its application in the synthesis of 3'-aminoalkylated oligonucleotides is reported. The release of oligonucleotides with free 3'-amino groups is accomplished by treatment with concentrated ammonia for 2 h at 55 degrees C.     

Ethanol blocks leukocyte recruitment and endothelial cell activation in vivo and in vitro

Immune system impairment and increased susceptibility to infection among alcohol abusers is a significant but not well-understood problem. We hypothesized that acute ethanol administration would inhibit leukocyte recruitment and endothelial cell activation during inflammation and infection. Using LPS and carrageenan air pouch models in mice, we found that physiological concentrations of ethanol (1-5 g/kg) significantly blocked leukocyte recruitment (50-90%). Because endothelial cell activation and immune cell-endothelial cell interactions are critical regulators of leukocyte recruitment, we analyzed the effect of acute ethanol exposure on endothelial cell activation in vivo using the localized Shwartzman reaction model. In this model, ethanol markedly suppressed leukocyte accumulation and endothelial cell adhesion molecule expression in a dose-dependent manner. Finally, we examined the direct effects of ethanol on endothelial cell activation and leukocyte-endothelial cell interactions in vitro. Ethanol, at concentrations within the range found in human blood after acute exposure and below the levels that induce cytotoxicity (0.1-0.5%), did not induce endothelial cell activation, but significantly inhibited TNF-mediated endothelial cell activation, as measured by adhesion molecule (E-selectin, ICAM-1, VCAM-1) expression and chemokine (IL-8, MCP-1, RANTES) production and leukocyte adhesion in vitro. Studies exploring the potential mechanism by which ethanol suppresses endothelial cell activation revealed that ethanol blocked NF-kappaB nuclear entry in an IkappaBalpha-dependent manner. These findings support the hypothesis that acute ethanol overexposure may increase the risk of infection and inhibit the host inflammatory response, in part, by blocking endothelial cell activation and subsequent immune cell-endothelial cell interactions required for efficient immune cell recruitment.     

An 18 kDa Acid Phosphatase from Chicken Heart Possesses Phosphotransferase Activity

A low molecular weight acid phosphatase was purified to homogeneity from chicken heart with a specific activity of 42 U/mg and a recovery of about 1%. Nearly 800 fold purification was achieved. The molecular weight was estimated to be 18 kDa by SDS-polyacrylamide gel electrophoresis. Para-nitrophenyl phosphate, phenyl phosphate and flavin mononucleotide were efficiently hydrolysed by the enzyme and found to be good substrates. Fluoride and tartrate had no inhibitory effect while phosphate, vanadate and molybdate strongly inhibited the enzyme. The acid phosphatase was stimulated in the presence of glycerol, ethylene glycol, methanol, ethanol and acetone, which reflected the phosphotransferase activity. When phosphate acceptors such as ethylene glycol concentrations were increased, the ratio of phosphate transfer to hydrolysis was also increased, demonstrating the presence of a transphosphorylation reaction where an acceptor can compete with water in the rate limiting step involving hydrolysis of a covalent phospho enzyme intermediate. Partition experiments carried out with two substrates, para-nitrophenyl phosphate and phenyl phosphate, revealed a constant product ratio of 1.7 for phosphotransfer to ethylene glycol versus hydrolysis, strongly supporting the existence of common covalent phospho enzyme intermediate. A constant ratio of K _cat/K _m, 4.3×10⁴, found at different ethylene glycol concentrations, also supported the idea that the rate limiting step was the hydrolysis of the phospho enzyme intermediate.     

Calcitonin gene-related peptide activates different signaling pathways in mesenteric lymphatics of guinea pigs

The effects of calcitonin gene-related peptide (CGRP) on constriction frequency, smooth muscle membrane potential (V(m)), and endothelial V(m) of guinea pig mesenteric lymphatics were examined in vitro. CGRP (1-100 nM) caused an endothelium-dependent decrease in the constriction frequency of perfused lymphatic vessels. The endothelium-dependent CGRP response was abolished by the CGRP-1 receptor antagonist CGRP-(8-37) (1 microM) and pertussis toxin (100 ng/ml). This action of CGRP was also blocked by the nitric oxide (NO) synthase inhibitor N(G)-nitro-L-arginine (L-NNA; 10 microM), an action that was reversed by the addition of L-arginine (100 microM). cGMP, adenylate cyclase, cAMP-dependent protein kinase (PKA), and ATP-sensitive K+ (K+(ATP)) channels were all implicated in the endothelium-dependent CGRP response because it was abolished by methylene blue (20 microM), 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (10 microM), dideoxyadenosine (10 microM), N-[2-(p-bromociannamylamino)-ethyl]-5-isoquinolinesulfonamide-dichloride (H89; 1 microM) and glibenclamide (10 microM). CGRP (100 nM), unlike acetylcholine, did not alter endothelial intracellular Ca2+ concentration or V(m). CGRP (100 nM) hyperpolarized the smooth muscle V(m), an effect inhibited by L-NNA, H89, or glibenclamide. CGRP (500 nM) also caused a decrease in constriction frequency. However, this was no longer blocked by CGRP-(8-37). CGRP (500 nM) also caused smooth muscle hyperpolarization, an action that was now not blocked by L-NNA (100 microM). It was most likely mediated by the activation of the cAMP/PKA pathway and the opening of K+(ATP) channels because it was abolished by H89 or glibenclamide. We conclude that CGRP, at low to moderate concentrations (i.e., 1-100 nM), decreases lymphatic constriction frequency primarily by the stimulation of CGRP-1 receptors coupled to pertussis toxin-sensitive G proteins and the release of NO from the endothelium or enhancement of the actions of endogenous NO. At high concentrations (i.e., 500 nM), CGRP also directly activates the smooth muscle independent of NO. Both mechanisms of activation ultimately cause the PKA-mediated opening of K+(ATP) channels and resultant hyperpolarization.     

Genetic Diversity and Population Structure of Ethiopian Chickpea (Cicer arietinum L.) Germplasm Accessions from Different Geographical Origins as Revealed by Microsatellite Markers

Genetic diversity and population structure were studied in 155 chickpea (Cicer arietinum L.) entries using 33 polymorphic microsatellite (SSR) markers. Molecular analysis of variance showed variations of 73% within and 27% among populations. Introduced genotypes exhibited highest polymorphism (70.27%) than the landraces (36?C57%). Collections from Shewa, Harerge, W. Gojam and S. Gonder regions also showed higher polymorphism (50?C57%) than the rest of the local accessions (36?C45%). Analyses of pairwise population Nei genetic distance and PhiPT coefficients, expected heterozygosity (He) and unbiased expected heterozygosity (UHe), Shannon??s information index (I) and percent polymorphism (% P) showed existence of high genetic variation between geographical regions. Accessions from adjoining geographical regions mostly showed more genetic similarities than those from origins far isolated apart. This could be associated with the ease and likelihood of inter-regional gene flow and seed movement particularly during times of drought. The 155 entries were grouped into five clusters following analysis of population structure. The first cluster (C₁) constituted accessions from Arsi; the second (C₂) from Gojam and Gonder; the third (C₃) from Harerge and E. and N. Shewa; and the fourth (C₄) from W. Shewa, Tigray, and Wello regions. The fifth cluster (C₅) was entirely consisted of improved genotypes. Improved genotypes of both Kabuli and Desi types distinctly fell into cluster five (C₅) regardless of their difference in seed types. The result has firmly established that introduction of genetic materials from exotic sources has broadened the genetic base of the national chickpea breeding program. Further implications of the findings as regards to chickpea germplasm management and its utilization in breeding program are also discussed.