Phospholipase D stimulation is required for sphingosine-1-phosphate activation of actin stress fibre assembly in human airway epithelial cells.

In human airway epithelial cells, sphingosine-1-phosphate (SPP) and lysophosphatidic acid (LPA) stimulated the production of phosphatidic acid (PA), which was inhibited by the primary alcohol butan-1-ol, but not by the inactive butan-2-ol, clearly indicating phospholipase D (PLD) involvement. Both SPP and LPA stimulated actin stress fibre formation, which was also butan-2-ol-insensitive and inhibited by butan-1-ol. SPP-induced PLD activation and cytoskeletal remodelling were insensitive to brefeldin A and toxin B from Clostridium difficile, which conversely blocked the effect of LPA, suggesting that the monomeric GTPases ADP ribosylation factor (ARF) and Rho are involved in LPA, but not in SPP responses. Pertussis toxin inhibited SPP- but not LPA-induced effects. PLD activation and stress fibre formation by both lysolipids were abolished by the tyrosine kinase inhibitor genistein. Addition of PA to cells caused a massive stress fibre assembly. In conclusion, PLD is one of the signalling components linking SPP-receptor activation to assembly of actin stress fibres.     

Diurnal Variation in Scent Marking Behavior in Captive Male and Female Common Marmosets, Callithrix jacchus

Chemical communication by scent-marking behavior in New World primates is used to prevent the access of potential competitors to a territory, to identify food resources and the reproductive condition of mates, among others. In common marmosets, primates of the Callitrichidae family, this behavior also occurs as olfactory identification of an individual or of the reproductive status of females. Despite this information, the diurnal variation and gender differences in the profile of this behavior remain to be investigated. The aims of this study were to establish the diurnal profile of the distribution of this behavior and the influence of the sex of markers. We used 18 adult common marmosets, Callithrix jacchus, 10 males and 8 females from 6 family groups (6 fathers and 4 sons; 4 mothers and 4 daughters). The frequency of scent-marking behavior was recorded for each animal over a period of 8 days, twice a week, for 4 weeks, starting when the animals left the nest box (approximately at 05:00 a.m.) until the end of the photophase, at about 05:00 p.m. A MANOVA test was performed to compare the frequency of scent-marking behavior at 2 hour intervals using pooled data for males and females. The results showed that significantly higher levels of scent-marking behavior occurred during the 03:00-05:00 p.m. interval compared to all other intervals. Lower values were recorded during the 11:00-13:00 interval and an effect of the sex factor was also found, with the values being higher for females than for males, although a significant difference was recorded only for the 07:00-09:00 interval. Minimal values for males were recorded during the 07:00-09:00 interval, whereas minimum values for females were recorded during the 11:00-13:00 interval. However, the highest values for both sexes continued to occur during the 15:00-17:00 interval. These results suggest that scent marking behavior in common marmosets has a preferential incidence at the end of the day and this might be occurring in association with feeding behavior. At this time these animals usually forage more to prepare for the night's fast. Since these animals can discriminate chemical clues as long as 24 hours after they have been left, the higher incidence of this behavior at this time probably will assure that the animals will localize feeding resources used on the preceding day. Significant elevation of scent marking behavior in females in relation to males was found only at 07:00-09:00 interval and seems to be associated with signalizing of reproductive status, preferential access to foraging or both.     

Lymphocyte transendothelial migration toward smooth muscle cells in interleukin-1 β-stimulated co-cultures is related to fibronectin interactions with α4β1 and α5β1 integrins

We previously reported infiltration of immune-inflammatory cells in coronary arteries from cardiac allografts, associated with increased endothelial and smooth muscle cell fibronectin synthesis regulated by interleukin (IL)-1b?. We now investigate, using a porcine endothelial-smooth muscle cell co-culture system, whether IL-1b?-stimulated fibronectin production is functionally important in lymphocyte transendothelial migration. Lymphocytes were harvested from porcine peripheral blood and, in the unactivated state or following activation with phorbol myristic acetate (PMA) and IL-2, were characterized by fluorescence-activated cell sorter (FACS) analysis and added to a confluent endothelial monolayer on the upper chamber of a transwell system. Endothelial cells, as well as smooth muscle cells (in the bottom of the chamber), were stimulated with IL-1b?. Then transendothelial lymphocyte migration was determined in the presence of CS1 and RGD (fibronectin) peptides, blocking α₄b?₁ and α₅b?₁ integrin receptors on lymphocyte surfaces, respectively. A 55-70% inhibition of lymphocyte migration was observed when compared to control peptides. The combination of CS1 and RGD peptides did not significantly enhance the inhibitory effect of either peptide alone. A similar decrease in lymphocyte transendothelial migration toward smooth muscle cells was documented using a monoclonal antibody to cellular fibronectin. Furthermore, using smooth muscle cell conditioned medium; we reproduced the enhanced transendothelial lymphocyte migration as well as the inhibition with blocking peptides or fibronectin antibodies. Our data suggest that cytokine-mediated fibronectin synthesis in vascular cells recruits inflammatory cells through interactions of specific peptides with cell surface α₄b?₁ α₅b?₁ integrins. © 1995 Wiley-Liss, Inc.     

Observations on the invading gastropod <Emphasis Type="Italic">Rapana venosa</Emphasis> in Punta del Este,Maldonado Bay,Uruguay

We report on the first sightings of the invasive Rapa Whelk Rapana venosa in Maldonado Bay (Punta del Este Harbor and Gorriti Island) using in vivo, underwater observations and video surveys. The species was first detected in the Río de la Plata (Uruguay and Argentina) in 1999, and by 2004 it had extended its local distribution to Punta del Este at the eastern boundary of the estuary. Observations performed by SCUBA diving showed that R. venosa is preying on native mussels Mytilus edulis and Brachidontes spp., and that formerly abundant mussel beds are being seriously depleted due to a combination of human extraction, habitat deterioration and predation by the Rapa Whelk.     

Unidirectional potentiation of binding between two anti-FBP MAbs: Evaluation of involved mechanisms

The monoclonal antibody MOv19 directed to a folate binding protein shows temperature-dependent potentiation of binding of the noncompeting monoclonal antibody MOv18 to the relevant antigen, but the mechanism involved in this phinomenon had remained unclear. Use of chimeric versions of both monoclonal antibodies and the F(ab′)₂ and fan fragments of MOv19 revealed an increment in MOv18 binding in all combinations irrespective of the orgin of the Fc portin of the monoclonal antibody. The potentiating effect of bivalent MOv19 fragments on ¹²⁵l-MOv18 binding was similar to that of the entire monoclonal antibody and occurred at saturating concentrations of both reagents at which monovalent binding prevails. Similarly, the monovalent fragment also induced a significant increase in MOv18 bunding. Howener, the potentiation sccurred only at very high concentrations of antibody fragment. Homologous inhibition was drastically reduced using MOv19 Fab fragment, suggesting a low binding stability of the monovalent reagent. Immunoblotting analysis and binding in the presence of exogenous purified folate binding protein indicated a cross-linking between soluble and cell surface molecules mediated by the bivalent monoclonal antibodies. The extentof the increase in MOv18 binging at O°C with high amounts of exogenous folate binding protein was lower than that obtained at 37⁰C in the absence of added molecule. Release of ¹²⁵l-MOv18 from the cell surface was significantly higher in the absence of MOv19 than in its presence. Affinity constant values of ¹²⁵l-MOv18 binding evaluated in the presence of MOv19 or control monoclonal antibody MINT5 were comparable, whereas the number of binding sites per cell detected by ¹²⁵l-MOv18 was significantly higher in the presence of MOv19 than MINT5. Together, the data suggest that monoclonal antibody MOv19 induces a conformational change of the molecule it binds that increases the number of antigenic sites anvailable for MOv18 binding and, in turn, the binding stability of the latter, MOv19 bivalency also contributes to the MOv18 binding increment by cross-linking released and cell surface–anchored folate binding protein molecules. © Wiley-Liss, Inc.     

Conservation of peculiar structural properties by the hemoglobins of anguilloid eels (Teleostei)

The three species investigated, the European eel, Anguilla anguilla (L.), the Mediterranean moray, Muraena helena L., and the conger eel, Conger coner (L.), represent three different superfamilies of the suborder Anguilloidei (infradivision Elopomorpha). Their hemoglobin systems show peculiar structural properties, which distinuish them from all other teleost species studied. They present acidic and basic components differing greatly in their isoelectric points; the basic components have the highest isoelectric points detected in teleost hemoglobins. While there is one major basic component, multiplicity is present in the acidic components in Muraena and Conger. The polyeptides of the acidic comonents show the same electrophoretic mobility in 8 M urea. In the three species, the electroploretic mobility with urea-SDS of the hemoglobin polypeptides shows a shorter β polypeptide in the basic components. These features had never been investigated among Elopomorpha, and are likely to be phylogenetically relevant.     

The early intracellular signaling pathway for the insulin/insulin-like growth factor receptor family in the mammalian central nervous system

Several studies support the idea that the polypeptides belonging to the family of insulin and insulin-like growth factors (IGFs) play an important role in brain development and continue to be produced in discrete areas of the adult brain. In numerous neuronal populations within the olfactory bulb, the cerebral and cerebellar cortex, the hippocampus, some diencephalic and brainstem nuclei, the spinal cord and the retina, specific insulin and IGF receptors, as well as crucial components of the intracellular receptor signaling pathway have been demonstrated. Thus, mature neurons are endowed with the cellular machinery to respond to insulin and IGF stimulation. Studies in vitro and in vivo, using normal and transgenic animals, have led to the hypothesis that, in the adult brain, IGF-I not only acts as a trophic factor, but also as a neuromodulator of some higher brain functions, such as long-term potentiation and depression. Furthermore, a trophic effect on certain neuronal populations becomes clearly evident in the ischemic brain or neurodegenerative disorders. Thus, the analysis of the early intracellular signaling pathway for the insulin/IGF receptor family in the brain is providing us with new intriguing findings on the way the mammalian brain is sculpted and operates.     

Lectin-Like Oxidized LDL Receptor-1 Is an Enhancer of Tumor Angiogenesis in Human Prostate Cancer Cells

Altered expression and function of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) has been associated with several diseases such as endothelial dysfunction, atherosclerosis and obesity. In these pathologies, oxLDL/LOX-1 activates signaling pathways that promote cell proliferation, cell motility and angiogenesis. Recent studies have indicated that olr1 mRNA is over-expressed in stage III and IV of human prostatic adenocarcinomas. However, the function of LOX-1 in prostate cancer angiogenesis remains to be determined. Our aim was to analyze the contribution of oxLDL and LOX-1 to tumor angiogenesis using C4-2 prostate cancer cells. We analyzed the expression of pro-angiogenic molecules and angiogenesis on prostate cancer tumor xenografts, using prostate cancer cell models with overexpression or knockdown of LOX-1 receptor. Our results demonstrate that the activation of LOX-1 using oxLDL increases cell proliferation, and the expression of the pro-angiogenic molecules VEGF, MMP-2, and MMP-9 in a dose-dependent manner. Noticeably, these effects were prevented in the C4-2 prostate cancer model when LOX-1 expression was knocked down. The angiogenic effect of LOX-1 activated with oxLDL was further demonstrated using the aortic ring assay and the xenograft model of tumor growth on chorioallantoic membrane of chicken embryos. Consequently, we propose that LOX-1 activation by oxLDL is an important event that enhances tumor angiogenesis in human prostate cancer cells.