Acute in vitro hypoxia and high-altitude (4,559 m) exposure decreases leukocyte oxygen consumption

Hypoxia impairs metabolic functions by decreasing activity and expression of ATP-consuming processes. To separate hypoxia from systemic effects, we tested whether hypoxia at high altitude affects basal and PMA-stimulated leukocyte metabolism and how this compares to acute (15 min) and 24 h of in vitro hypoxia. Leukocytes were prepared at low altitude and ～24 h after arrival at 4559 m. Mitochondrial oxygen consumption (JO?) was measured by respirometry, oxygen radicals by electron spin resonance spectroscopy, both at a Po? = 100 mmHg (JO?,???) and 20 mmHg (JO?,??). Acute hypoxia of leukocytes decreased JO? at low altitude. Exposure to high altitude decreased JO?,???, whereas JO?,?? was not affected. Acute hypoxia of low-altitude samples decreased the activity of complexes I, II, and III. At high altitude, activity of complexes I and III were decreased when measured in normoxia. Stimulation of leukocytes with PMA increased JO?,??? at low (twofold) and high altitude (five-fold). At both locations, PMA-stimulated JO? was decreased by acute hypoxia. Basal and PMA-stimulated reactive oxygen species (ROS) production were unchanged at high altitude. Separate in vitro experiments performed at low altitude show that ～75% of PMA-induced increase in JO? was due to increased extra-mitochondrial JO? (JO?(,res); in the presence of rotenone and antimycin A). JO?(,res) was doubled by PMA. Acute hypoxia decreased basal JO?(,res) by ～70% and PMA-stimulated JO?(,res) by about 50% in cells cultured in normoxia and hypoxia (1.5% O?; 24 h). Conversely, 24 h in vitro hypoxia decreased mitochondrial JO?,??? and JO?,??, extra-mitochondrial, basal, and PMA-stimulated JO? were not affected. These results show that 24 h of high altitude but not 24 h in vitro hypoxia decreased basal leukocyte metabolism, whereas PMA-induced JO? and ROS formation were not affected, indicating that prolonged high-altitude hypoxia impairs mitochondrial metabolism but does not impair respiratory burst. In contrast, acute hypoxia impairs respiratory burst at either altitude.     

Induction of humoral and CD8+ T cell responses are required for protection against lethal Ebola virus infection

Ebola virus (EBOV)-like particles (eVLP), composed of the EBOV glycoprotein and matrix viral protein (VP)40 with a lipid membrane, are a highly efficacious method of immunization against EBOV infection. The exact requirements for immunity against EBOV infection are poorly defined at this time. The goal of this work was to determine the requirements for EBOV immunity following eVLP vaccination. Vaccination of BALB/c or C57BL/6 mice with eVLPs in conjunction with QS-21 adjuvant resulted in mixed IgG subclass responses, a Th1-like memory cytokine response, and protection from lethal EBOV challenge. Further, this vaccination schedule led to the generation of both CD4(+) and CD8(+) IFN-gamma(+) T cells recognizing specific peptides within glycoprotein and VP40. The transfer of both serum and splenocytes, but not serum or splenocytes alone, from eVLP-vaccinated mice conferred protection against lethal EBOV infection in these studies. B cells were required for eVLP-mediated immunity to EBOV because B cell-deficient mice vaccinated with eVLPs were not protected from lethal EBOV challenge. We also found that CD8(+), but not CD4(+), T cells are absolutely required for eVLP-mediated protection against EBOV infection. Further, eVLP-induced protective mechanisms were perforin-independent, but IFN-gamma-dependent. Taken together, both EBOV-specific humoral and cytotoxic CD8(+) T cell responses are critical to mediate protection against filoviruses following eVLP vaccination.     

Crystallization and preliminary crystallographic studies of the precursor and mature forms of a neutral lipase from the fungus Rhizopus delemar

A neutral lipase from the filamentous fungus Rhizopus delemar has been crystallized in both its proenzyme and mature forms. Although the latter crystallizes readily and produces a variety of crystal forms, only one was found to be suitable for X-ray studies. It is monoclinic (C2, a = 92.8 Å, b = 128.9 Å, c = 78.3 Å, β = 135.8) with two molecules in the asymmetric unit related by a noncrystallographic diad. The prolipase crystals are orthorhombic (P2₁2₁2₁, with a = 79.8 Å, b = 115.2 Å, c = 73.0 Å) and also contain a pair of molecules in the asymmetric unit. Initial results of molecular replacement calculations using the refined coordinates of the related lipase from Rhizomucor miehei identified the correct orientations and positions of the protein molecules in the unit cells of crystals of both proenzyme and the mature form. © 1994 John Wiley & Sons, Inc.     

1H, 13C, and 15N backbone, side-chain, and heme chemical shift assignments for oxidized and reduced forms of the monoheme c-type cytochrome ApcA isolated from the acidophilic metal-reducing bacterium Acidiphilium cryptum

We report the ¹H, ¹³C, and ¹⁵N chemical shift assignments of both oxidized and reduced forms of an abundant periplasmic c-type cytochrome, designated ApcA, isolated from the acidophilic gram-negative facultatively anaerobic metal-reducing alphaproteobacterium Acidiphilium cryptum. These resonance assignments prove that ApcA is a monoheme cytochrome c ₂ and the product of the Acry_2099 gene. An absence of resonance peaks in the NMR spectra for the 21N-terminal residues suggests that a predicted N-terminal signal sequence is cleaved. We also describe the preparation and purification of the protein in labeled form from laboratory cultures of A. cryptum growing on ¹³C- and ¹⁵N- labeled substrates.     

Coastal wetland restoration through the lens of Odum's theory of ecosystem development

Advancing ecological restoration assessments requires a more detailed consideration of species interactions and ecosystem processes. Most restoration projects rely on a few metrics not always directly linked with ecological theory. Here, we used Odum's theory of ecosystem development to assess and compare the ecosystem structure and services of created marshes (4–6 years old) with preexisting, reference marshes in a brackish water region of the Mississippi River Delta. We built ecosystem models for created and reference marshes that integrated large datasets of stomach contents, stable isotopes, and taxa abundances. Despite strong resemblance in community structure, created marshes were at an earlier succession stage compared to the reference marshes, having lower biomass (including exploited species), higher biomass turnover and production, less dependence on detritus, lower material cycling, and less energy flowing through specialist pathways. Although preserving preexisting marshes should be a priority, created marshes may still be an important tool for the restoration of coastal areas and their ecosystem services. In addition, our results show that comparisons of species biodiversity alone may fail to capture essential differences in ecosystem processes between habitats, which reinforces the importance of ecosystem modeling approaches to assess restoration projects.     

DO SUTURE ZONES EXIST?

Remington (1968) argued that 13 suture zones exist in North America. Remington defined a suture zone as, "a band of geographic overlap between major biotic assemblages, including some pairs of species or semispecies which hybridize in the zone" (p. 322). Although initially controversial, the idea that suture zones exist has picked up momentum over the past decade, due largely to the phylogeographic work of Hewitt, Avise, and their colleagues. Nevertheless, the reality of suture zones has not yet been subjected to rigorous analysis using statistical and geographic information system (GIS) approaches. To test for the existence of Remington's suture zones, we first identified 117 terrestrial hybrid zones in Canada and the United States through a literature search for the key words "cline," "contact zone," "hybrid zone," and "hybridization" in articles published between 1970 and 2002. The 117 hybrid zones were mapped using a GIS approach and compared with a digitized version of Remington's original suture zone map. Overall, there does appear to be an association between hybrid zones and suture zones, but this association is largely attributable to clustering of hybrid zones in only two of the 13 suture zones recognized by Remington. The results suggest that evolutionary biologists should retain some skepticism toward Remington's suture zones.