Prostaglandin synthesis and release by subpopulations of rat alveolar macrophages

Alveolar macrophages are the primary phagocytic cell of lung, but are also capable of a variety of other functions, which include initiating or modulating inflammatory and immune responses through the production of soluble mediators. One such group of mediators is the eicosanoids. Further, recent data indicate that alveolar macrophages are not functionally homogeneous, but are heterogeneous with several subpopulations that differ both morphologically and functionally. Considering the apparent importance of prostaglandin synthesis and release in inflammatory and immune responses, the current study was undertaken to determine whether alveolar macrophage subpopulations differ in their ability to synthesize and release prostaglandin (PG) E, PGI2, and thromboxane A2 after stimulation by calcium ionophore A23187, zymosan, or aggregated IgG. Alveolar macrophages were harvested by bronchoalveolar lavage and were separated into 18 density-defined fractions. Density-defined alveolar macrophages (DD-AM) showed marked heterogeneity in prostaglandin synthesis and release. Maximal PGE synthesis and release was seen as a single peak after calcium ionophore A23187 and zymosan stimulation. In contrast, two peaks in PGE synthesis were seen after aggregated IgG stimulation. PGI2 synthesis was seen as a single peak generated by different DD-AM after calcium ionophore A23187 and zymosan. In contrast, aggregated IgG stimulation of subpopulations exhibited uniform synthesis and release of PGI2. Thromboxane A2 synthesis and release was maximal from a broad range of various DD-AM after calcium ionophore A23187, zymosan, and aggregated IgG stimulation. The results demonstrate that DD-AM are heterogeneous in ability to synthesize and release prostaglandins which is dependent on the stimuli. Therefore, specific subpopulations of alveolar macrophages may be central to the control of the pulmonary inflammatory response through specific eicosanoid synthesis and release.     

Phylogenetic diversification of immunoglobulin genes and the antibody repertoire

Immunoglobulins are encoded by a large multigene system that undergoes somatic rearrangement and additional genetic change during the development of immunoglobulin-producing cells. Inducible antibody and antibody-like responses are found in all vertebrates. However, immunoglobulin possessing disulfide-bonded heavy and light chains and domain-type organization has been described only in representatives of the jawed vertebrates. High degrees of nucleotide and predicted amino acid sequence identity are evident when the segmental elements that constitute the immunoglobulin gene loci in phylogenetically divergent vertebrates are compared. However, the organization of gene loci and the manner in which the independent elements recombine (and diversify) vary markedly among different taxa. One striking pattern of gene organization is the "cluster type" that appears to be restricted to the chondrichthyes (cartilaginous fishes) and limits segmental rearrangement to closely linked elements. This type of gene organization is associated with both heavy- and light-chain gene loci. In some cases, the clusters are "joined" or "partially joined" in the germ line, in effect predetermining or partially predetermining, respectively, the encoded specificities (the assumption being that these are expressed) of the individual loci. By relating the sequences of transcribed gene products to their respective germ-line genes, it is evident that, in some cases, joined-type genes are expressed. This raises a question about the existence and/or nature of allelic exclusion in these species. The extensive variation in gene organization found throughout the vertebrate species may relate directly to the role of intersegmental (V<==>D<==>J) distances in the commitment of the individual antibody-producing cell to a particular genetic specificity. Thus, the evolution of this locus, perhaps more so than that of others, may reflect the interrelationships between genetic organization and function.      

Insulin-like growth factor type-I receptor internalization and recycling mediate the sustained phosphorylation of Akt

Previously we demonstrated that insulin-like growth factor-I mediates the sustained phosphorylation of Akt, which is essential for long term survival and protection of glial progenitors from glutamate toxicity. These prosurvival effects correlated with prolonged activation and stability of the insulin-like growth factor type-I receptor. In the present study, we investigated the mechanisms whereby insulin-like growth factor-I signaling, through the insulin-like growth factor type-I receptor, mediates the sustained phosphorylation of Akt. We showed that insulin-like growth factor-I stimulation induced loss of receptors from the cell surface but that surface receptors recovered over time. Blocking receptor internalization inhibited Akt phosphorylation, whereas inhibition of receptor trafficking blocked receptor recovery at the cell surface and the sustained phosphorylation of Akt. Moreover the insulin-like growth factor type-I receptor localized with the transferrin receptor and Rab11-positive endosomes in a ligand-dependent manner, further supporting the conclusion that this receptor follows a recycling pathway. Our results provide evidence that ligand stimulation leads to internalization of the insulin-like growth factor type-I receptor, which mediates Akt phosphorylation, and that receptor recycling sustains Akt phosphorylation in glial progenitors. Mathematical modeling of receptor trafficking further supports these results and predicts an additional kinetic state of the receptor consistent with sustained Akt phosphorylation.     

The gene ontology categorizer

The Gene Ontology Categorizer, developed jointly by the Los Alamos National Laboratory and Procter & Gamble Corp., provides a capability for the categorization task in the Gene Ontology (GO): given a list of genes of interest, what are the best nodes of the GO to summarize or categorize that list? The motivating question is from a drug discovery process, where after some gene expression analysis experiment, we wish to understand the overall effect of some cell treatment or condition by identifying 'where' in the GO the differentially expressed genes fall: 'clustered' together in one place? in two places? uniformly spread throughout the GO? 'high', or 'low'? In order to address this need, we view bio-ontologies more as combinatorially structured databases than facilities for logical inference, and draw on the discrete mathematics of finite partially ordered sets (posets) to develop data representation and algorithms appropriate for the GO. In doing so, we have laid the foundations for a general set of methods to address not just the categorization task, but also other tasks (e.g. distances in ontologies and ontology merger and exchange) in both the GO and other bio-ontologies (such as the Enzyme Commission database or the MEdical Subject Headings) cast as hierarchically structured taxonomic knowledge systems.     

What can we learn from noncoding regions of similarity between genomes?

Background  

In addition to known protein-coding genes, large amounts of apparently non-coding sequence are conserved between the human and mouse genomes. It seems reasonable to assume that these conserved regions are more likely to contain functional elements than less-conserved portions of the genome.     

An interlaboratory comparison of physiological and genetic properties of four Saccharomyces cerevisiae strains

To select a Saccharomyces cerevisiae reference strain amenable to experimental techniques used in (molecular) genetic, physiological and biochemical engineering research, a variety of properties were studied in four diploid, prototrophic laboratory strains. The following parameters were investigated: 1) maximum specific growth rate in shake-flask cultures; 2) biomass yields on glucose during growth on defined media in batch cultures and steady-state chemostat cultures under controlled conditions with respect to pH and dissolved oxygen concentration; 3) the critical specific growth rate above which aerobic fermentation becomes apparent in glucose-limited accelerostat cultures; 4) sporulation and mating efficiency; and 5) transformation efficiency via the lithium-acetate, bicine, and electroporation methods. On the basis of physiological as well as genetic properties, strains from the CEN.PK family were selected as a platform for cell-factory research on the stoichiometry and kinetics of growth and product formation.     

Adaptation of acidogenic sludge to increasing glycerol concentrations for biohydrogen production

Hydrogen is a promising alternative as an energetic carrier and its production by dark fermentation from wastewater has been recently proposed, with special attention to crude glycerol as potential substrate. In this study, two different feeding strategies were evaluated for replacing the glucose substrate by glycerol substrate: a one-step strategy (glucose was replaced abruptly by glycerol) and a step-by-step strategy (progressive decrease of glucose concentration and increase of glycerol concentration from 0 to 5 g L^?1), in a continuous stirred tank reactor (12 h of hydraulic retention time (HRT), pH 5.5, 35 °C). While the one-step strategy led to biomass washout and unsuccessful H₂ production, the step-by-step strategy was efficient for biomass adaptation, reaching acceptable hydrogen yields (0.4?±?0.1 mol_H2?mol^?1 _{glycerol consumed}) around 33 % of the theoretical yield independently of the glycerol concentration. Microbial community structure was investigated by single-strand conformation polymorphism (SSCP) and denaturing gradient gel electrophoresis (DGGE) fingerprinting techniques, targeting either the total community (16S ribosomal RNA (rRNA) gene) or the functional Clostridium population involved in H₂ production (hydA gene), as well as by 454 pyrosequencing of the total community. Multivariate analysis of fingerprinting and pyrosequencing results revealed the influence of the feeding strategy on the bacterial community structure and suggested the progressive structural adaptation of the community to increasing glycerol concentrations, through the emergence and selection of specific species, highly correlated to environmental parameters. Particularly, this work highlighted an interesting shift of dominant community members (putatively responsible of hydrogen production in the continuous stirred tank reactor (CSTR)) according to the gradient of glycerol proportion in the feed, from the family Veillonellaceae to the genera Prevotella and Clostridium sp., putatively responsible of hydrogen production in the CSTR.