ABSTRACT
Free fatty acids (FFA) have generally been proposed to regulate pancreatic insulin release by an intracellular mechanism involving inhibition of CPT-1. The recently de-orphanized G-protein coupled receptor, FFA(1)R/GPR40, has been shown to be essential for fatty-acid-stimulated insulin release in MIN6 mouse insulinoma cells. The CPT-1 inhibitor, 2-bromo palmitate (2BrP), was investigated for its ability to interact with mouse FFA(1)R/GPR40. It was found to inhibit phosphatidyl inositol hydrolysis induced by linoleic acid (LA) (100 muM in all experiments) in HEK293 cells transfected with FFA(1)R/GPR40 and in the MIN6 subclone, MIN6c4. 2BrP also inhibited LA-stimulated insulin release from mouse pancreatic islets. Mouse islets were subjected to antisense intervention by treatment with a FFA(1)R/GPR40-specific morpholino oligonucleotide for 48 h. Antisense treatment of islets suppressed LA-stimulated insulin release by 50% and by almost 100% when islets were pretreated with LA for 30 min before applying the antisense. Antisense treatment had no effect on tolbutamide-stimulated insulin release. Confocal microscopy using an FFA(1)R/GPR40-specific antibody revealed receptor expression largely localized to the plasma membrane of insulin-producing cells. Pretreating the islets with LA for 30 min followed by antisense oligonucleotide treatment for 48 h reduced the FFA(1)R/GPR40 immunoreactivity to background levels. The results demonstrate that FFA(1)R/GPR40 is inhibited by the CPT-1 inhibitor, 2BrP, and confirm that FFA(1)R/GPR40 is indeed necessary, at least in part, for fatty-acid-stimulated insulin release.
Subject(s)
Fatty Acids/metabolism , Insulin/metabolism , Receptors, G-Protein-Coupled/metabolism , Animals , Cell Line , Female , Humans , Hypoglycemic Agents/metabolism , Insulin Secretion , Islets of Langerhans/cytology , Islets of Langerhans/metabolism , Linoleic Acid/metabolism , Mice , Oligonucleotides, Antisense/metabolism , Palmitates/metabolism , Phosphatidylinositols/metabolism , Receptors, G-Protein-Coupled/geneticsABSTRACT
In developing countries, planning in the forestry sector has been seen as an appropriate instrument to prepare and implement government policies and programs. Despite its potential and recent advancements in, for example, remote sensing and infrastructure, tropical forest land-use planning is often formal and non-integrated with agriculture. It rarely involves all legitimate stakeholders and neglects taking into account actual land-use. The socio-economic and environmental consequences of these shortcomings emphasise the need for alternative ways of approaching planning. This article summarises the idea, structure and current status of the Area Production Model (APM), originally developed in the 1980s, which is now gaining interest as a land-use planning tool in Africa and Asia. It describes the development over time of production and consumption in agriculture and forestry within a defined geographical area operating under different assumptions on management, land use and socio-economic and macro-economic changes. From a narrow perspective, the APM is a fairly simple computerised tool for generating scenarios. In a broad sense, it is a concept comprising the whole planning process including organisation, inventory, data analysis, consensus building and strategy. A case-study in Laos, based on verified data for a historical period of 45 years, and a review of different APM applications in the world over the last 15 years are presented as a base for conclusions about its potential and shortcomings. In some cases where the APM concept was used in training courses and planning exercises involving stakeholders, it generated a strong interest in collecting and analysing relevant information. It provides the means of addressing a number of shortcomings in current planning.
Subject(s)
Conservation of Natural Resources , Developing Countries , Models, Theoretical , Trees , Economics , Forecasting , Geography , Laos , Policy Making , Public Policy , Social ConditionsABSTRACT
The leukotriene B(4) receptor (BLTR) is a seven-transmembrane chemoattractant receptor that is important in pro-inflammatory responses. We have produced the first widely applicable monoclonal antibodies against the human BLTR and confirmed the antibody specificity using flow cytometric analysis of three different cell lines stably expressing the recombinant receptor. The antibodies did not cross-react with the recently cloned second LTB(4) receptor, BLTR2, or the Cys LT1 and Cys LT2 receptors. Functional analysis in combination with two-color flow cytometry showed that the BLTR antibodies bind to cells that are activated by LTB(4). The antibodies were shown to recognize BLTR in cell ELISA and immunocytochemistry. Endogenous expression of BLTR in CD15-positive blood leukocytes and in differentiated HL-60 cells was also demonstrated with the antibodies.
Subject(s)
Antibodies, Monoclonal , Receptors, Leukotriene B4/analysis , Receptors, Leukotriene B4/immunology , 3T3 Cells , Animals , Antibody Specificity , Flow Cytometry , Genes, Reporter , HeLa Cells , Humans , Immunohistochemistry , Mice , Receptors, Leukotriene B4/genetics , Recombinant Proteins/analysis , Recombinant Proteins/immunology , TransfectionABSTRACT
The genomic region containing the genes encoding the first leukotriene B(4) receptor, BLTR, as well as the recently cloned second leukotriene B(4)-activated receptor, BLTR2, was mapped by (a) sequence analysis of a human bacterial artificial chromosome (BAC) library containing a 15-kb segment corresponding to chromosome 14q11. 2-12 where the BLTR/BLTR2 genes were previously shown to be located, together with (b) sequence analysis of 83 expressed sequence tags (ESTs) from this region. The BLTR gene includes four different 5' untranslated regions (UTRs) and a mutual acceptor site for the exon containing the intronless open reading frame. The BLTR2 gene is intronless and overlapped by a 5' UTR splice version of BLTR and, on the reverse strand, of the apoptosis-related CIDE-B gene. This indicates a complex posttranscriptional gene regulation. Further adding to the complexity of the region is evidence of a fourth putative and novel gene, most homologous to the rat adenylyl cyclase IV gene.
Subject(s)
Chromosomes, Human, Pair 14/genetics , Physical Chromosome Mapping , Receptors, Leukotriene B4/genetics , 3' Untranslated Regions/genetics , 5' Untranslated Regions/genetics , Alternative Splicing/genetics , Apoptosis Regulatory Proteins , Chromosomes, Bacterial/genetics , DNA, Complementary/analysis , DNA, Complementary/genetics , Expressed Sequence Tags , Genomic Library , Humans , Proteins/genetics , Restriction Mapping , Sequence Analysis, DNAABSTRACT
By homology screening using BLAST searches of expressed sequence tags (ESTs), we have found a previously unidentified cDNA encoding a putative seven-transmembrane receptor with highest similarity to the leukotriene B(4) receptor, BLTR. Analysis of calcium flow in transfected cells, along with sequence analysis, revealed that the EST encoded a functionally inactive protein, lacking the segment corresponding to the C-terminal part of the putative receptor protein. The missing segment was obtained by PCR amplification of a human leukocyte cDNA library and ligated to the truncated EST cDNA. The novel cDNA encodes a full-length receptor with 39% identity to the previously cloned BLTR. Studies of intracellular calcium flow of transfected HeLa cells exposed to various leukotrienes showed that also the novel BLTR-like receptor can be activated by leukotriene B(4), and it is therefore tentatively named BLTR2.
