SummaryAlthough there is a mounting body of evidence that eosinophils are recruited to sites of allergic inflammation by a number of ~-chemokines, particularly eotaxin and RANTES, the receptor that mediates these actions has not been identified. We have now cloned a G protein-coupled receptor, CC CKR3, from human eosinophils which, when stably expressed in AML14.3D10 cells bound eotaxin, MCP-3 and R_ANTES with Kas of 0.1, 2.7, and 3.1 nM, respectively. CC CKR3 also bound MCP-1 with lower affinity, but did not bind MIP-10~ or MIP-I[~. Eotaxin, RANTES, and to a lessor extent MCP-3, but not the other chemokines, activated CC CKR3 as determined by their ability to stimulate a CaZ+-flux. Competition binding studies on primary eosinophils gave binding at~inities for the different chemokines which were indistinguishable from those measured with CC CKR3. Since CC CKR3 is prominently expressed in eosinophils we conclude that CC CKR3 is the eosinophil eotaxin receptor. Eosinophils also express a much lower level of a second chemokine receptor, CC CKR1, which appears to be responsible for the effects of MIP-llx.

We reviewed prominent emerging infectious diseases of cetaceans, examined their potential to impact populations, re-assessed zoonotic risk and evaluated the role of environmental stressors. Cetacean morbilliviruses and papillomaviruses as well as Brucella spp. and Toxoplasma gondii are thought to interfere with population abundance by inducing high mortalities, lowering reproductive success or by synergistically increasing the virulence of other diseases. Severe cases of lobomycosis and lobomycosis-like disease (LLD) may contribute to the death of some dolphins. The zoonotic hazard of marine mammal brucellosis and toxoplasmosis may have been underestimated, attributable to frequent misdiagnoses and underreporting, particularly in developing countries and remote areas where carcass handling without protective gear and human consumption of fresh cetacean products are commonplace. Environmental factors seem to play a role in the emergence and pathogenicity of morbillivirus epidemics, lobomycosis/LLD, toxoplasmosis, poxvirus-associated tattoo skin disease and, in harbour porpoises, infectious diseases of multifactorial aetiology. Inshore and estuarine cetaceans incur higher risks than pelagic cetaceans due to habitats often severely altered by anthropogenic factors such as chemical and biological contamination, direct and indirect fisheries interactions, traumatic injuries from vessel collisions and climate change.

The guanine nucleotide-binding proteincoupled receptor superfamily binds a vast array of biological messengers including lipids, odorants, catecholamines, peptides, and proteins. While some small molecules bind to these receptors at a single interhelical site, we find that the binding domain on the receptor for the inflammatory protein C5a is more complex and consists of two distinct subsites. This more elaborate motif appears to be an evolutionary adaptation of the simpler paradigm to which a second interaction site has been added in the receptor N terminus. Surprisingly, occupation of only one of the subsites is required for receptor activation. The two-site motif is not unique to the C5a receptor but appears to be widely used by the superfamily to accommodate macromolecular ligands.The 74-aa glycoprotein C5a evokes a variety of responses in vivo and in vitro, implying that it is a principal mediator of inflammatory responses (1, 2). C5a is a potent chemotaxin and secretagogue for granulocytes and macrophages; it activates the respiratory burst in these cells and modulates their adhesive properties. The effects of C5a are amplified by its ability to stimulate the release of other mediators including histamine, prostaglandins, leukotrienes, interleukin (IL) 1, and IL-6 (1-3).All of the effects of C5a are initiated when it binds to its cell surface receptor, a member ofthe guanine nucleotide-binding protein (G protein)-coupled receptor superfamily (4, 5). The superfamily consists of over 100 members and binds a variety of ligands ranging in complexity from small molecules to moderately sized proteins. Despite this biologic diversity, a general model for the structure of these receptors has emerged: an extracellular N terminus, seven membranespanning helices connected by alternating intracellular and extracellular loops, and an intracellular C terminus (6, 7). The amino acid sequence of the C5a receptor is consistent with this model and like most members of the family has a short N terminus of about 30 residues in length (4, 5).Family members such as rhodopsin and the ,3adrenergic receptor bind their ligands at a single domain, which lies in the receptor's hydrophobic core, between the helices and below the upper plane of the cellular membrane (6, 8). However, it is unclear whether this binding motif is also used by other members of the superfamily, especially those that interact with more complex ligands like C5a, or whether the motif is altered to accommodate the larger agonists. The little information that exists comes largely from studies with the glycopeptide hormone receptors, a branch ofthe superfamily characterized by a greatly extended extracellular N terminus (9, 10). These receptors, in contrast to rhodopsin and the ,fadrenergic receptor, appear to bind ligands by means of this enlarged N terminus (11,12). We now report that the binding site of the C5a receptor is more complex and consists of two physically separable domains. The first domain is composed of the N terminus and possibly the exter...

ABSTRACT:We succinctly review and document new cases of diseases of the skin and the skeletal system and external traumata in cetaceans from Ecuador, Colombia, Peru, Chile, Argentina, Uruguay, Brazil, and Venezuela. The survey revealed 590 cases diagnosed with a significant pathology, injury or malformation on a total of 7635 specimens of 12 odontocete species examined or observed in . Tattoo skin disease (TSD), lobomycosis-like disease (LLD) and cutaneous diseases of unknown aetiology seem to be emerging in several populations. TSD was confirmed in eight species from the SE Pacific and SW Atlantic. LLD affected only inshore Tursiops truncatus but was found in four tropical countries, namely Colombia, Ecuador, Peru and Brazil. Lobomycosis was confirmed by histology in one male from the Tramandaí estuary, southern Brazil. All LLD-affected specimens were encountered in the vicinity of major ports and cities and a possible association with chemical or organic water pollution is suspected. Whitish velvety cutaneous marks associated with scars occurred in inshore T. truncatus, Sotalia guianensis and Pseudorca crassidens. Large, rounded lesions were seen in a Cephalorhynchus eutropia calf and a C. commersonii. Cutaneous wounds and scars as well as body traumata possibly related to net entanglements and boat collisions were observed in 73 delphinids and Phocoena spinipinnis. Traumatic injuries resulted in the partial or complete amputation and other disfiguring scars of appendages in 17 cases. Fractures of the skull, ribs and vertebrae thought to be caused by fisheries-related interactions or boat collisions were seen in single individuals of Delphinus capensis, Lagenorhynchus obscurus, T. truncatus, S. guianensis and Ziphius cavirostris. Prevalence of osteopathology in small cetaceans from Peru, Brazil and Venezuela ranged widely, from 5.4% to 69.1%. In four species from Peru, lytic cranial lesions were the most frequently observed disease (5.4%-42.9%), followed by hyperostosis and ankylosing spondylitis in offshore (31%, n=42) and inshore (15.4%, n=26) T. truncatus. Fractures and other bone traumata were present in 47.2% of 53 axial skeletons of S. guianensis from the northern Rio de Janeiro state (Brazil) in 1987-1998. A high prevalence (48.4%, n=31) of, apparently congenital, malformations of cervical vertebrae, observed in a 2001-2006 sample, may be explained by a hypothetical genetic bottleneck in this population. Malformations with deficient ossification would clearly increase susceptibility for fractures. This study demonstrates the utility of a continent-wide analysis to discern epizootiological trends more readily than any local study could provide. Secondly, it underscores the need for focussed research on the effects of human activities on the spread of diseases in cetaceans, particularly in near-shore populations that utilize highly degraded coastal habitats. RESUMEN:En este trabajo revisamos y documentamos brevemente nuevos casos de enfermedades de piel y de esqueleto, y traumas externos en cetáceos de Ec...

The chemokine receptors CCR5 and CXCR4 act synergistically with CD4 in an ordered multistep mechanism to allow the binding and entry of human immunodeficiency virus type 1 (HIV-1). The efficiency of such a coordinated mechanism depends on the spatial distribution of the participating molecules on the cell surface. Immunoelectron microscopy was performed to address the subcellular localization of the chemokine receptors and CD4 at high resolution. Cells were fixed, cryoprocessed, and frozen; 80-nm cryosections were double labeled with combinations of CCR5, CXCR4, and CD4 antibodies and then stained with immunogold. Surprisingly, CCR5, CXCR4, and CD4 were found predominantly on microvilli and appeared to form homogeneous microclusters in all cell types examined, including macrophages and T cells. Further, while mixed microclusters were not observed, homogeneous microclusters of CD4 and the chemokine receptors were frequently separated by distances less than the diameter of an HIV-1 virion. Such distributions are likely to facilitate cooperative interactions with HIV-1 during virus adsorption to and penetration of human leukocytes and have significant implications for development of therapeutically useful inhibitors of the entry process. Although the mechanism underlying clustering is not understood, clusters were observed in small trans-Golgi vesicles, implying that they were organized shortly after synthesis and well before insertion into the cellular membrane. Chemokine receptors normally act as sensors, detecting concentration gradients of their ligands and thus providing directional information for cellular migration during both normal homeostasis and inflammatory responses. Localization of these sensors on the microvilli should enable more precise monitoring of their environment, improving efficiency of the chemotactic process. Moreover, since selectins, some integrins, and actin are also located on or in the microvillus, this organelle has many of the major elements required for chemotaxis.Human immunodeficiency virus (HIV) therapies have been highly successful in slowing disease progression, increasing health and well-being, and prolonging life. However, viral resistance is now becoming common, and since most existing drugs target only two viral proteins, reverse transcriptase and protease, cross-resistance is a significant problem. One solution to the issue of resistance is development of new complementary therapies based on novel mechanisms of action. The discovery that the chemokine receptors CCR5 and CXCR4, in addition to CD4, are required for viral entry not only furthered understanding of the fusion and infection process but provided two new targets for therapeutic intervention (3,12,14,17,18,22,44).The entry mechanism as currently understood is an ordered process in which the viral envelope protein, gp120, following interaction with CD4, undergoes a conformational change allowing binding to the appropriate chemokine receptor, CCR5 for macrophagetropic or R5 strains, and CXCR4 for T-celltropic or X4 s...

IP10 and MIG are two members of the CXC branch of the chemokine superfamily whose expression is dramatically up-regulated by interferon (IFN)-gamma. The proteins act largely on natural killer (NK)-cells and activated T-cells and have been implicated in mediating some of the effects of IFN-gamma and lipopolysaccharides (LPSs), as well as T-cell-dependent anti-tumor responses. Recently both chemokines have been shown to be functional agonists of the same G-protein-coupled receptor, CXCR3. We now report the pharmacological characterization of CXCR3 and find that, when heterologously expressed, CXCR3 binds IP10 and MIG with Ki values of 0.14 and 4.9 nM, respectively. The receptor has very modest affinity for SDF-1alpha and little or no affinity for other CXC-chemokines. The properties of the endogenous receptor expressed on activated T-cells are similar. Surprisingly, several CC-chemokines, particularly eotaxin and MCP-4, also compete with moderate affinity for the binding of IP10 to CXCR3. Eotaxin does not activate CXCR3 but, in CXCR3-transfected cells, can block IP10-mediated receptor activation. Eotaxin, therefore, may be a natural CXCR3 antagonist.

Human immunodeficiency virus type 1 (HIV-1) fuses with cells after sequential interactions between its envelope glycoproteins, CD4 and a coreceptor, usually CC chemokine receptor 5 (CCR5) or CXC receptor 4 (CXCR4). CMPD 167 is a CCR5-specific small molecule with potent antiviral activity in vitro. We show that CMPD 167 caused a rapid and substantial (4–200-fold) decrease in plasma viremia in six rhesus macaques chronically infected with simian immunodeficiency virus (SIV) strains SIVmac251 or SIVB670, but not in an animal infected with the X4 simian–human immunodeficiency virus (SHIV), SHIV-89.6P. In three of the SIV-infected animals, viremia reduction was sustained. In one, there was a rapid, but partial, rebound and in another, there was a rapid and complete rebound. There was a substantial delay (>21 d) between the end of therapy and the onset of full viremia rebound in two animals. We also evaluated whether vaginal administration of gel-formulated CMPD 167 could prevent vaginal transmission of the R5 virus, SHIV-162P4. Complete protection occurred in only 2 of 11 animals, but early viral replication was significantly less in the 11 CMPD 167-recipients than in 9 controls receiving carrier gel. These findings support the development of small molecule CCR5 inhibitors as antiviral therapies, and possibly as components of a topical microbicide to prevent HIV-1 sexual transmission.

The marine form of the South American dolphin Sotalia fluviatilis has an extensive and possibly continuous distribution, from the Florianópolis region, Brazil (27°35′S, 48°34′W), north to Panama (~9°22′N, 79°54′W). The high number of records from 25–20°S is due to the presence of many observers in those latitudes. The freshwater form of this species inhabits the Amazon and Orinoco drainages. It is commonly seen in the Amazon, and has been found as far inland as southern Peru. The southern limit of the range of the marine form of Sotalia is associated with the confluence zone of the Brazil and Falkland currents, suggesting that low sea-surface temperature is a limiting factor, whereas in fresh water the distribution of Amazonian Sotalia seems more closely related to the movements and concentrated occurrence of prey.