Development of cell surface saccharides on embryonic pancreatic cells

Using a battery of seven lectin-ferritin conjugates as probes for cell surface glycoconjugates, we have studied the pattern of plasmalemmal differentiation of cells in the embryonic rat pancreas from day 15 in utero to the early postpartum stage. Our results indicate that differentiation of plasmalemmal glycoconjugates on acinar, endocrine, and centroacinar cells is temporally correlated with development and is unique for each cell type, as indicated by lectin-ferritin binding. Specifically, (a) expression of adult cell surface saccharide phenotype can be detected on presumptive acinar cells as early as 15 d in utero, as indicated by soybean agglutinin binding, and precedes development of intracellular organelles characteristic of mature acinar cells; (b) maturation of the plasmalemma of acinar cells is reached after intracellular cytodifferentiation is completed, as indicated by appearance of Con A and fucoselectin binding sites only at day 19 of development; conversely, maturation of the endocrine cell plasmalemma is accompanied by "loss" (masking) of ricinus communis II agglutinin receptors; and (c) binding sites for fucose lectins and for soybean agglutinin are absent on endocrine and centroacinar cells at all stages examined. We conclude that acinar, centroacinar, and endocrine cells develop from a common progenitor cell(s) whose plasmalemmal carbohydrate composition resembles most closely that of the adult centroacinar cell. Finally, appearance of acinar lumina beginning at approximately 17 d in utero is accompanied by differenetiation of apical and basolateral plasmalemmal domains of epithelial cells, as indicated by enhanced binding of several lectin-ferritin conjugates to the apical plasmalemmal, a pattern that persists from this stage through adult life.     

Age-Specific Mortality During the 1918 Influenza Pandemic: Unravelling the Mystery of High Young Adult Mortality

The worldwide spread of a novel influenza A (H1N1) virus in 2009 showed that influenza remains a significant health threat, even for individuals in the prime of life. This paper focuses on the unusually high young adult mortality observed during the Spanish flu pandemic of 1918. Using historical records from Canada and the U.S., we report a peak of mortality at the exact age of 28 during the pandemic and argue that this increased mortality resulted from an early life exposure to influenza during the previous Russian flu pandemic of 1889–90. We posit that in specific instances, development of immunological memory to an influenza virus strain in early life may lead to a dysregulated immune response to antigenically novel strains encountered in later life, thereby increasing the risk of death. Exposure during critical periods of development could also create holes in the T cell repertoire and impair fetal maturation in general, thereby increasing mortality from infectious diseases later in life. Knowledge of the age-pattern of susceptibility to mortality from influenza could improve crisis management during future influenza pandemics.

“The war is over – and I must go” Egon Schiele, 1890–1918.

     

Energy,ecology and the distribution of microbial life

Mechanisms that govern the coexistence of multiple biological species have been studied intensively by ecologists since the turn of the nineteenth century. Microbial ecologists in the meantime have faced many fundamental challenges, such as the lack of an ecologically coherent species definition, lack of adequate methods for evaluating population sizes and community composition in nature, and enormous taxonomic and functional diversity. The accessibility of powerful, culture-independent molecular microbiology methods offers an opportunity to close the gap between microbial science and the main stream of ecological theory, with the promise of new insights and tools needed to meet the grand challenges humans face as planetary engineers and galactic explorers. We focus specifically on resources related to energy metabolism because of their direct links to elemental cycling in the Earth''s history, engineering applications and astrobiology. To what extent does the availability of energy resources structure microbial communities in nature? Our recent work on sulfur- and iron-oxidizing autotrophs suggests that apparently subtle variations in the concentration ratios of external electron donors and acceptors select for different microbial populations. We show that quantitative knowledge of microbial energy niches (population-specific patterns of energy resource use) can be used to predict variations in the abundance of specific taxa in microbial communities. Furthermore, we propose that resource ratio theory applied to micro-organisms will provide a useful framework for identifying how environmental communities are organized in space and time.     

The cover of living and dead corals from airborne remote sensing

Trends in coral cover are widely used to indicate the health of coral reefs but are costly to obtain from field survey over large areas. In situ studies of reflected spectra at the coral surface show that living and recently dead colonies can be distinguished. Here, we investigate whether such spectral differences can be detected using an airborne remote sensing instrument. The Compact Airborne Spectrographic Imager (Itres Research Ltd, Canada) was flown in two configurations: 10 spectral bands with 1-m² pixels and 6 spectral bands with 0.25-m² pixels. First, we show that an instrument with 10 spectral bands possesses adequate spectral resolution to distinguish living Porites, living Pocillopora spp., partially dead Porites, recently dead Porites (total colony mortality within 6 months), old dead (>6 months) Porites, Halimeda spp., and coralline red algae when there is no water column to confuse spectra. All substrata were distinguished using fourth-order spectral derivatives around 538 nm and 562 nm. Then, at a shallow site (Tivaru) at Rangiroa Atoll, Tuamotu Archipelago (French Polynesia), we show that live and dead coral can be distinguished from the air to a depth of at least 4 m using first- and fourth-order spectral derivatives between 562–580 nm. However, partially dead and recently dead Porites colonies could not be distinguished from an airborne platform. Spectral differences among substrata are then exploited to predict the cover of reef substrata in ten 25-m² plots at nearby Motu Nuhi (max depth 8 m). The actual cover in these plots was determined in situ using quadrats with a 0.01-m² grid. Considerable disparity occurred between field and image-based measures of substrate cover within individual 25-m² quadrats. At this small scale, disparity, measured as the absolute difference in cover between field and remote-sensing methods, reached 25% in some substrata but was always less than 10% for living coral (99% of which consisted of Porites spp.). At the scale of the reef (all ten 25-m² quadrats), however, disparities in percent cover between imagery and field data were less than 10% for all substrata and extremely low for some classes (e.g. <3% for living Porites, recently dead Porites and Halimeda). The least accurately estimated substrata were sand and coralline red algae, which were overestimated by absolute values 7.9% and 6.6%, respectively. The precision of sampling was similar for field and remote-sensing methods: field methods required 19 plots to detect a 10% difference in coral cover among three reefs with a statistical power of 95%. Remote-sensing methods required 21 plots. However, it took 1 h to acquire imagery over 92,500 m² of reef, which represents 3,700 plots of 25 m² each, compared with 3 days to survey 10 such plots underwater. There were no significant differences in accuracy between 1-m² and 0.25-m² image resolutions, suggesting that the advantage of using smaller pixels is offset by reduced spectral information and an increase in noise (noise was observed to be 1.6–1.8 times greater in 0.25-m² pixels). We show that airborne remote sensing can be used to monitor coral and algal cover over large areas, providing that water is shallow and clear, and that brown fleshy macroalgae are scarce, that depth is known independently (e.g. from sonar survey).     

Adenoviral gene transfer of eNOS: high-level expression in ex vivo expanded marrow stromal cells

Endothelial nitric oxide synthase (eNOS) is an attractive target for cardiovascular gene therapy. Marrow stromal cells (MSCs), also known as mesenchymal stem cells, hold great promise for use in adult stem cell-based cell and gene therapy. To determine the feasibility of adenoviral-mediated eNOS gene transfer into ex vivo expanded MSCs, rat MSCs (rMSCs) were isolated, expanded ex vivo, and transduced with Ad5RSVeNOS, an adenoviral vector containing the eNOS gene under the control of the Rous sarcoma virus promoter. The presence of eNOS protein in Ad5RSVeNOS-transduced rMSCs was confirmed by immunohistochemical and Western blot analysis. Transduction efficiency was dose dependent, and eNOS transgene expression in rMSCs persisted for =" BORDER="0">21 days in culture. The rMSCs retained multipotential differentiation capability after adenoviral-mediated eNOS gene transfer. Furthermore, intracavernosal injection of Ad5RSVeNOS-transduced rMSCs increased the expression of eNOS in the corpus cavernosum, and stem cells were identified within corporal sinusoids. These findings demonstrate that replication-deficient recombinant adenovirus can be used to engineer ex vivo expanded rMSCs and that high-level eNOS transgene expression can be achieved, pointing out the clinical potential of using this novel adult stem cell-based gene therapy method for the treatment of cardiovascular diseases. adenoviral vector; nitric oxide; gene expression; differentiation; gene therapy     

Vasodilator responses to adenosine and hyperemia are mediated by A(1) and A(2) receptors in the cat vascular bed

Hemodynamic responses to adenosine, the A(1) receptor agonists N(6)-cyclopentyladenosine (CPA) and adenosine amine congener (ADAC), and the A(2) receptor agonist 5'-(N-cyclopropyl)-carboxamido-adenosine (CPCA) were investigated in the hindquarter vascular bed of the cat under constant-flow conditions. Injections of adenosine, CPA, ADAC, CPCA, ATP, and adenosine 5'-O-(3-thiotriphosphate) (ATPgamma S) into the perfusion circuit induced dose-related decreases in perfusion pressure. Vasodilator responses to the A(1) agonists were reduced by the A(1) receptor antagonists KW-3902 and CGS-15943, whereas responses to CPCA were reduced by the A(2) antagonist KF-17837. Vasodilator responses to adenosine were reduced by KW-3902, CGS-15943, and by KF-17837, suggesting a role for both A(1) and A(2) receptors. Vasodilator responses to ATP and the nonhydrolyzable ATP analog ATP gamma S were not attenuated by CGS-15943 or KF-17837. After treatment with the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine methyl ester, the cyclooxygenase inhibitor sodium meclofenamate, or the ATP-dependent K(+) (K) channel antagonists U-37883A or glibenclamide, responses to adenosine and ATP were not altered. Responses to adenosine, CPA, and CPCA were increased in duration by rolipram, a type 4 cAMP phosphodiesterase inhibitor, but were not altered by zaprinast, a type 5 cGMP phosphodiesterase inhibitor. When blood flow was interrupted for a 30-s period, the magnitude and duration of the reactive vasodilator response were reduced by A(1) and A(2) receptor antagonists. These data suggest that vasodilator responses to adenosine and the A(1) and A(2) agonists studied are not dependent on the release of cyclooxygenase products, nitric oxide, or the opening of K channels in the regional vascular bed of the cat. The present data suggest a role for cAMP in mediating responses to adenosine and suggest that vasodilator responses to adenosine and to reactive hyperemia are mediated in part by A(1) and A(2) receptors in the hindquarter vascular bed of the cat.     

Overexpression of arginase in the aged mouse penis impairs erectile function and decreases eNOS activity: influence of in vivo gene therapy of anti-arginase

Since both increased nitric oxide (NO) synthase (NOS) abundance and diminished NO signaling have been reported in the aging penis, the role of NO in the adaptations of aging remains controversial. Here we tested the hypothesis that arginase, an enzyme that competes with NOS for the substrate l-arginine, contributes to erectile dysfunction with advanced age in the B6/129 mouse strain. Arginase protein abundance, mRNA expression, and enzyme activity were elevated in aged compared with young penile endothelial cells. In addition, endothelial NOS (NOS3) protein abundance was greater in aged versus young penile endothelial cells, whereas NOS activity and cGMP levels were reduced. Calcium-dependent l-arginine-to-l-citrulline conversion and cGMP formation increased significantly in aged mouse penes in the presence of the arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH). However, there was no effect on l-arginine-to-l-citrulline conversion or cGMP accumulation in the endothelium from young mouse penes. To assess the functional role of arginase in the inhibition of NOS pathway responsiveness in the penis, we evaluated the effects of ABH and an adeno-associated virus encoding an antisense sequence to arginase I (AAVanti-arginase) on erectile function in vivo. ABH and AAVanti-arginase enhanced endothelium-dependent erectile responses in the aged mice without altering endothelium-independent responses. Paralleling our in vitro observations, ABH or AAVanti-arginase did not affect vascular responses in the young mice. Inhibition of the arginase pathway improves endothelial function in the aging penile circulation, suggesting that the arginase pathway may be exploited to improve erectile dysfunction associated with aging.     

Gene transfer to pre-hematopoietic and committed hematopoietic precursors in the early mouse Yolk Sac: a comparative study between <Emphasis Type="Italic">in situ</Emphasis> electroporation and retroviral transduction

Background  

Hematopoietic development in vertebrate embryos results from the sequential contribution of two pools of precursors independently generated. While intra-embryonic precursors harbour the features of hematopoietic stem cells (HSC), precursors formed earlier in the yolk sac (YS) display limited differentiation and self-renewal potentials. The mechanisms leading to the generation of the precursors in both sites are still largely unknown, as are the molecular basis underlying their different potential. A possible approach to assess the role of candidate genes is to transfer or modulate their expression/activity in both sites. We thus designed and compared transduction protocols to target either native extra-embryonic precursors, or hematopoietic precursors.