A cardinal role for cathepsin D in co-ordinating the host-mediated apoptosis of macrophages and killing of pneumococci

The bactericidal function of macrophages against pneumococci is enhanced by their apoptotic demise, which is controlled by the anti-apoptotic protein Mcl-1. Here, we show that lysosomal membrane permeabilization (LMP) and cytosolic translocation of activated cathepsin D occur prior to activation of a mitochondrial pathway of macrophage apoptosis. Pharmacological inhibition or knockout of cathepsin D during pneumococcal infection blocked macrophage apoptosis. As a result of cathepsin D activation, Mcl-1 interacted with its ubiquitin ligase Mule and expression declined. Inhibition of cathepsin D had no effect on early bacterial killing but inhibited the late phase of apoptosis-associated killing of pneumococci in vitro. Mice bearing a cathepsin D-/- hematopoietic system demonstrated reduced macrophage apoptosis in vivo, with decreased clearance of pneumococci and enhanced recruitment of neutrophils to control pulmonary infection. These findings establish an unexpected role for a cathepsin D-mediated lysosomal pathway of apoptosis in pulmonary host defense and underscore the importance of apoptosis-associated microbial killing to macrophage function

Hypertonic saline reduces inflammation and enhances the resolution of oleic acid induced acute lung injury

<p>Abstract</p> <p>Background</p> <p>Hypertonic saline (HTS) reduces the severity of lung injury in ischemia-reperfusion, endotoxin-induced and ventilation-induced lung injury. However, the potential for HTS to modulate the resolution of lung injury is not known. We investigated the potential for hypertonic saline to modulate the evolution and resolution of oleic acid induced lung injury.</p> <p>Methods</p> <p>Adult male Sprague Dawley rats were used in all experiments. <b><it>Series 1 </it></b>examined the potential for HTS to reduce the severity of evolving oleic acid (OA) induced acute lung injury. Following intravenous OA administration, animals were randomized to receive isotonic (Control, n = 12) or hypertonic saline (HTS, n = 12), and the extent of lung injury assessed after 6 hours. <b><it>Series 2 </it></b>examined the potential for HTS to enhance the resolution of oleic acid (OA) induced acute lung injury. Following intravenous OA administration, animals were randomized to receive isotonic (Control, n = 6) or hypertonic saline (HTS, n = 6), and the extent of lung injury assessed after 6 hours.</p> <p>Results</p> <p>In <b><it>Series I</it></b>, HTS significantly reduced bronchoalveolar lavage (BAL) neutrophil count compared to Control [61.5 ± 9.08 versus 102.6 ± 11.89 × 10³ cells.ml^-1]. However, there were no between group differences with regard to: A-a O2 gradient [11.9 ± 0.5 vs. 12.0 ± 0.5 KPa]; arterial PO2; static lung compliance, or histologic injury. In contrast, in <b><it>Series 2</it></b>, hypertonic saline significantly reduced histologic injury and reduced BAL neutrophil count [24.5 ± 5.9 versus 46.8 ± 4.4 × 10³ cells.ml^-1], and interleukin-6 levels [681.9 ± 190.4 versus 1365.7 ± 246.8 pg.ml^-1].</p> <p>Conclusion</p> <p>These findings demonstrate, for the first time, the potential for HTS to reduce pulmonary inflammation and enhance the resolution of oleic acid induced lung injury.</p

Ectopic Cdx2 Expression in Murine Esophagus Models an Intermediate Stage in the Emergence of Barrett's Esophagus

Barrett's esophagus (BE) is an intestinal metaplasia that occurs in the setting of chronic acid and bile reflux and is associated with a risk for adenocarcinoma. Expression of intestine-specific transcription factors in the esophagus likely contributes to metaplasia development. Our objective was to explore the effects of an intestine-specific transcription factor when expressed in the mouse esophageal epithelium. Transgenic mice were derived in which the transcription factor Cdx2 is expressed in squamous epithelium using the murine Keratin-14 gene promoter. Effects of the transgene upon cell proliferation and differentiation, gene expression, and barrier integrity were explored. K14-Cdx2 mice express the Cdx2 transgene in esophageal squamous tissues. Cdx2 expression was associated with reduced basal epithelial cell proliferation and altered cell morphology. Ultrastructurally two changes were noted. Cdx2 expression was associated with dilated space between the basal cells and diminished cell-cell adhesion caused by reduced Desmocollin-3 mRNA and protein expression. This compromised epithelial barrier function, as the measured trans-epithelial electrical resistance (TEER) of the K14-Cdx2 epithelium was significantly reduced compared to controls (1189 Ohm*cm2 ±343.5 to 508 Ohm*cm2±92.48, p = 0.0532). Secondly, basal cells with features of a transitional cell type, intermediate between keratinocytes and columnar Barrett's epithelial cells, were observed. These cells had reduced keratin bundles and increased endoplasmic reticulum levels, suggesting the adoption of secretory-cell features. Moreover, at the ultrastructural level they resembled “Distinctive” cells associated with multilayered epithelium. Treatment of the K14-Cdx2 mice with 5′-Azacytidine elicited expression of BE-associated genes including Cdx1, Krt18, and Slc26a3/Dra, suggesting the phenotype could be advanced under certain conditions. We conclude that ectopic Cdx2 expression in keratinocytes alters cell proliferation, barrier function, and differentiation. These altered cells represent a transitional cell type between normal squamous and columnar BE cells. The K14-Cdx2 mice represent a useful model to study progression from squamous epithelium to BE

Spatio-temporal Models of Lymphangiogenesis in Wound Healing

Several studies suggest that one possible cause of impaired wound healing is failed or insufficient lymphangiogenesis, that is the formation of new lymphatic capillaries. Although many mathematical models have been developed to describe the formation of blood capillaries (angiogenesis), very few have been proposed for the regeneration of the lymphatic network. Lymphangiogenesis is a markedly different process from angiogenesis, occurring at different times and in response to different chemical stimuli. Two main hypotheses have been proposed: 1) lymphatic capillaries sprout from existing interrupted ones at the edge of the wound in analogy to the blood angiogenesis case; 2) lymphatic endothelial cells first pool in the wound region following the lymph flow and then, once sufficiently populated, start to form a network. Here we present two PDE models describing lymphangiogenesis according to these two different hypotheses. Further, we include the effect of advection due to interstitial flow and lymph flow coming from open capillaries. The variables represent different cell densities and growth factor concentrations, and where possible the parameters are estimated from biological data. The models are then solved numerically and the results are compared with the available biological literature.Comment: 29 pages, 9 Figures, 6 Tables (39 figure files in total

Saturation sampling for spatial variation in multiple air pollutants across an inversion-prone metropolitan area of complex terrain

Background: Characterizing intra-urban variation in air quality is important for epidemiological investigation of health outcomes and disparities. To date, however, few studies have been designed to capture spatial variation during select hours of the day, or to examine the roles of meteorology and complex terrain in shaping intra-urban exposure gradients. Methods: We designed a spatial saturation monitoring study to target local air pollution sources, and to understand the role of topography and temperature inversions on fine-scale pollution variation by systematically allocating sampling locations across gradients in emissions sources (vehicle traffic, industrial facilities) and topography (elevation) in the Pittsburgh area. Street-level integrated samples of fine particulate matter (PM2.5), black carbon (BC), nitrogen dioxide (NO2), sulfur dioxide (SO2), and ozone (O3) were collected during morning rush and probable inversion hours (6-11 AM), during summer and winter. We hypothesized that pollution concentrations would be: 1) higher under inversion conditions, 2) exacerbated in lower-elevation areas, and 3) vary by season. Results: During July-August 2011 and January-March 2012, we observed wide spatial and seasonal variability in pollution concentrations, exceeding the range measured at regulatory monitors. We identified elevated concentrations of multiple pollutants at lower-elevation sites, and a positive association between inversion frequency and NO2 concentration. We examined temporal adjustment methods for deriving seasonal concentration estimates, and found that the appropriate reference temporal trend differs between pollutants. Conclusions: Our time-stratified spatial saturation approach found some evidence for modification of inversion-concentration relationships by topography, and provided useful insights for refining and interpreting GIS-based pollution source indicators for Land Use Regression modeling

Rhodococcus opacus B4: a promising bacterium for production of biofuels and biobased chemicals

Bacterial lipids have relevant applications in the production of renewable fuels and biobased oleochemicals. The genus Rhodococcus is one of the most relevant lipid producers due to its capability to accumulate those compounds, mainly triacylglycerols (TAG), when cultivated on different defined substrates, namely sugars, organic acids and hydrocarbons but also on complex carbon sources present in industrial wastes. In this work, the production of storage lipids by Rhodococcus opacus B4 using glucose, acetate and hexadecane is reported for the first time and its productivity compared with Rhodococcus opacus PD630, the best TAG producer bacterium reported. Both strains accumulated mainly TAG from all carbon sources, being influenced by the carbon source itself and by the duration of the accumulation period. R. opacus B4 produced 0.09 and 0.14 g L1 at 24 and 72 h, with hexadecane as carbon source, which was 2 and 3.3 fold higher than the volumetric production obtained by R. opacus PD630. Both strains presented similar fatty acids (FA) profiles in intact cells while in TAG produced fraction, R. opacus B4 revealed a higher variability in fatty acid composition than R. opacus PD630, when both strains were cultivated on hexadecane. The obtained results open new perspectives for the use of R. opacus B4 to produce TAG, in particular using oily (alkane-contaminated) waste and wastewater as cheap raw-materials. Combining TAG production with hydrocarbons degradation is a promising strategy to achieve environmental remediation while producing added value compounds.This work was financially supported by the Portuguese Science Foundation (FCT) and European Social Fund (ESF, POPH-QREN) through the Grant given to A.R. Castro (SFRH/BD/64500/2009), the FCT Strategic Project of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684) and project RECI/BBB-EBI/0179/2012 (FCOMP-01-0124-FEDER-027462)

Inclusive education for Internally Displaced Children in Kenya::Children perceptions of their learning and development needs in post-conflict schooling

Abstract The Kenyan society has been characterised by tribal-political-instigated violence since the declaration of multiparty democracy in 1991. The 2007/8 post-election violence (PEV) particularly saw the scattering of families where some children lost months or years of schooling; others were permanently excluded from education, while the participation and achievement of those arriving in school were characterised by complex needs and experiences. This paper aims to analyse literature and report on findings from creative activities with 16 conflict-affected children (9–12 years) regarding their experiences and understandings of inclusive education during their post-conflict school-life. I conducted an intrinsic case study with aspects of ethnography in a post-conflict community primary school in Kenya whose majority (71%) pupil population was attributed to internal displacement following the 2007/8 PEV. Children perceived inclusive education in regard to their own learning and development needs as involving access and acceptance in the new school community, social-emotional development, ‘peer-keeping’ and community-consciousness. Whilst valuing their teachers’ pursuit for inclusion-sensitive practices, children’s understanding of their own circumstances resulted in group-made strategies like ‘peer-keeping’ and peer-constructed rules of interaction, helping them negotiate systemic constrains and distortion of values. This study underscores the fundamental role of children in social integration in post-conflict circumstances

The Formation and Evolution of the First Massive Black Holes

The first massive astrophysical black holes likely formed at high redshifts (z>10) at the centers of low mass (~10^6 Msun) dark matter concentrations. These black holes grow by mergers and gas accretion, evolve into the population of bright quasars observed at lower redshifts, and eventually leave the supermassive black hole remnants that are ubiquitous at the centers of galaxies in the nearby universe. The astrophysical processes responsible for the formation of the earliest seed black holes are poorly understood. The purpose of this review is threefold: (1) to describe theoretical expectations for the formation and growth of the earliest black holes within the general paradigm of hierarchical cold dark matter cosmologies, (2) to summarize several relevant recent observations that have implications for the formation of the earliest black holes, and (3) to look into the future and assess the power of forthcoming observations to probe the physics of the first active galactic nuclei.Comment: 39 pages, review for "Supermassive Black Holes in the Distant Universe", Ed. A. J. Barger, Kluwer Academic Publisher