Activated protein C mediates novel lung endothelial barrier enhancement: role of sphingosine 1-phosphate receptor transactivation

Increased endothelial cell (EC) permeability is central to the pathophysiology of inflammatory syndromes such as sepsis and acute lung injury (ALI). Activated protein C (APC), a serine protease critically involved in the regulation of coagulation and inflammatory processes, improves sepsis survival through an unknown mechanism. We hypothesized a direct effect of APC to both prevent increased EC permeability and to restore vascular integrity after edemagenic agonists. We measured changes in transendothelial electrical resistance (TER) and observed that APC produced concentration-dependent attenuation of TER reductions evoked by thrombin. We next explored known EC barrier-protective signaling pathways and observed dose-dependent APC-mediated increases in cortical myosin light chain (MLC) phosphorylation in concert with cortically distributed actin polymerization, findings highly suggestive of Rac GTPase involvement. We next determined that APC directly increases Rac1 activity, with inhibition of Rac1 activity significantly attenuating APC-mediated barrier protection to thrombin challenge. Finally, as these signaling events were similar to those evoked by the potent EC barrier-enhancing agonist, sphingosine 1-phosphate (S1P), we explored potential cross-talk between endothelial protein C receptor (EPCR) and S1P1, the receptors for APC and S1P, respectively. EPCR-blocking antibody (RCR-252) significantly attenuated both APC-mediated barrier protection and increased MLC phosphorylation. We next observed rapid, EPCR and PI 3-kinase-dependent, APC-mediated phosphorylation of S1P1 on threonine residues consistent with S1P1 receptor activation. Co-immunoprecipitation studies demonstrate an interaction between EPCR and S1P1 upon APC treatment. Targeted silencing of S1P1 expression using siRNA significantly reduced APC-mediated barrier protection against thrombin. These data suggest that novel EPCR ligation and S1P1 transactivation results in EC cytoskeletal rearrangement and barrier protection, components potentially critical to the improved survival of APC-treated patients with severe sepsis.     

Raiders of the lost bark: orangutan foraging strategies in a degraded landscape

Deforestation is rapidly transforming primary forests across the tropics into human-dominated landscapes. Consequently, conservationists need to understand how different taxa respond and adapt to these changes in order to develop appropriate management strategies. Our two year study seeks to determine how wild Sumatran orangutans (Pongo abelii) adapt to living in an isolated agroforest landscape by investigating the sex of crop-raiders related to population demographics, and their temporal variations in feeding behaviour and dietary composition. From focal animal sampling we found that nine identified females raided cultivated fruits more than the four males. Seasonal adaptations were shown through orangutan feeding habits that shifted from being predominantly fruit-based (56% of the total feeding time, then 22% on bark) to the fallback food of bark (44%, then 35% on fruits), when key cultivated resources such as jackfruit (Artocarpus integer), were unavailable. Cultivated fruits were mostly consumed in the afternoon and evening, when farmers had returned home. The finding that females take greater crop-raiding risks than males differs from previous human-primate conflict studies, probably because of the low risks associated (as farmers rarely retaliated) and low intraspecific competition between males. Thus, the behavioral ecology of orangutans living in this human-dominated landscape differs markedly from that in primary forest, where orangutans have a strictly wild food diet, even where primary rainforests directly borders farmland. The importance of wild food availability was clearly illustrated in this study with 21% of the total orangutan feeding time being allocated to feeding on cultivated fruits. As forests are increasingly converted to cultivation, humans and orangutans are predicted to come into conflict more frequently. This study reveals orangutan adaptations for coexisting with humans, e.g. changes in temporal foraging patterns, which should be used for guiding the development of specific human-wildlife conflict mitigation strategies to lessen future crop-raiding and conflicts.     

Influence of salinity and organic nutrient concentration on survival and growth of Vibrio cholerae in aquatic microcosms

Laboratory microcosms were employed to evaluate the influence of selected environmental parameters, organic nutrient concentration, and salinity on the growth and survival of a toxigenic strain of Vibrio cholerae LA4808. Over the range conditions tested, this strain of V. cholerae showed maximum response as determined by increased plate counts and direct microscopic counts in microcosms prepared with a chemically defined sea salts solution at a salinity of 25%, but with lower or higher salinity levels, the maximum population size declined. When added organic concentrations of less than 1,000 micrograms/liter were present, a marked salinity effect on the growth of V. cholerae was detected. However, at or above an organic nutrient concentration of 1,000 micrograms/liter, the need for an optimum salinity level was spared. From the results of this study, it is concluded that V. cholerae can grow under conditions of organic nutrient concentration and salinity typical of estuaries. Results obtained support the hypothesis that V. cholerae is an autochthonous member of the estuarine microbial community.     

Use of the API rapid NFT system for identifying nonfermentative and fermentative marine bacteria.

Thirty-five American Type Culture Collection type strains of marine bacteria were used to evaluate the Rapid NFT system (API Analab Products, Plainview, N.Y.) for use in identifying heterotrophic marine bacteria. The 21 biochemical and assimilation tests on the Rapid NFT test strips were treated according to the manufacturer's protocol, which included use of AUX medium (provided with the Rapid NFT system) for preparing assimilation tests, and by substituting phenol red broth base (BBL Microbiology Systems, Cockeysville, Md.) with and without an oil overlay for the AUX medium. A seven-digit numerical profile was obtained for each NFT test strip from each of the three procedures and matched to its corresponding number in the Rapid NFT identification codebook. Also, all biochemical and assimilation test results were analyzed with SASTAXAN and SAS/GRAPH programs (SAS Institute, Inc., Cary, N.C.); similarity matrices were computed for all 35 strains. For comparison purposes, bacterial strains were grouped at a similarity level of 70%. The results indicated a low efficacy of identification for all three procedures. In addition, similarity matrix analysis showed more cohesive grouping based on results of phenol red broth base-treated strains than for the AUX medium provided by the manufacturer. However, none of the three treatments provided exclusive grouping of type strains at the genus level. Thus, the reliability of the data obtained from the NFT system and modifications thereof should be evaluated carefully when environmental isolates are characterized.     

Is reproduction of the Australian house mouse (<Emphasis Type="Italic">Mus domesticus</Emphasis>) constrained by food? A large-scale field experiment

Food quantity and especially food quality are thought to be key factors driving reproductive changes in the house mouse, Mus domesticus, leading to outbreaks of house mouse populations in the Australian grain-growing region. Characteristic changes during an incipient mouse plague are an early start of breeding, a high proportion of females breeding at a young age and a prolonged breeding season. We conducted a large-scale food manipulation during an incipient mouse plague, which started with early breeding and relatively high spring numbers of mice. We measured background food availability in four farms throughout the study and conducted a food manipulation experiment from November to March in two of them. After harvest in December 100-200 kg/ha spilled grain remained in the stubble. This was depleted by March. In two treatment farms we added high-protein food pellets on a weekly basis between November and March and two farms served as controls. We measured changes in mouse numbers by capture-mark-recapture trappings and changes in reproduction by scoring embryos and recent placental scars at necropsy. Mouse numbers did not differ between treatments and controls. There were no differences in the litter size or the proportion of females breeding between treatments and controls. We observed the normal pattern of high litter size in spring and decreasing litter size towards the end of summer in treatments and controls. In all farms reproduction stopped in March. Mouse numbers were high but not at plague densities. Contrary to our prediction we did not observe food constraint affecting the reproduction of female mice. Our field experiment seems to rule out food quality as the driving factor for improved reproduction and formation of an outbreak of mice. We suggest that physiological mechanisms in mice might not enable them to take advantage of food with a high protein content in arid summers in southeastern Australian grain fields because of the lack of free-standing water.