Lysosomal accumulation and recycling of lipopolysaccharide to the cell surface of murine macrophages, an in vitro and in vivo study.

In this study, we detailed in a time-dependent manner the trafficking, the recycling, and the structural fate of Brucella abortus LPS in murine peritoneal macrophages by immunofluorescence, ELISA, and biochemical analyses. The intracellular pathway of B. abortus LPS, a nonclassical endotoxin, was investigated both in vivo after LPS injection in the peritoneal cavity of mice and in vitro after LPS incubation with macrophages. We also followed LPS trafficking after infection of macrophages with B. abortus strain 19. After binding to the cell surface and internalization, Brucella LPS is routed from early endosomes to lysosomes with unusual slow kinetics. It accumulates there for at least 24 h. Later, LPS leaves lysosomes and reaches the macrophage cell surface. This recycling pathway is also observed for LPS released by Brucella S19 following in vitro infection. Indeed, by 72 h postinfection, bacteria are degraded by macrophages and LPS is located inside lysosomes dispersed at the cell periphery. From 72 h onward, LPS is gradually detected at the plasma membrane. In each case, the LPS present at the cell surface is found in large clusters with the O-chain facing the extracellular medium. Both the antigenicity and heterogenicity of the O-chain moiety are preserved during the intracellular trafficking. We demonstrate that LPS is not cleared by macrophages either in vitro or in vivo after 3 mo, exposing its immunogenic moiety toward the extracellular medium.     

Biogeographical patterns and Rapoport's rule in southeastern Pacific benthic polychaetes of the Chilean coast

Recently three biogeographical units were identified along the Chilean coast (the Magellanic Province, an Intermediate Area, and the Peruvian Province), however few studies have focused on the factors and dynamic processes that formed these spatial units (e.g. Rapoport's rule and its causal mechanisms). In this study we used benthic polychaetes of the Chilean coast to evaluate patterns of latitudinal distribution and species richness, and the existence of the three main biogeographical provinces described for the Chilean coast. Additionally, we evaluated the latitudinal Rapoport effects and geometric constraint as a null hypothesis explaining the species richness distribution.
We found that benthic polychaete diversity increased towards southern latitudes. The cluster and ordination (non-metric MultiDimensional Scaling, nMDS) analyses of the distribution data, presented only two statistically significant (bootstrapping techniques) biogeographic provinces along the Chilean coast, with a break occurring between 41° and 42°S. While, our results did not support a latitudinal Rapoport effect, they did support the view that latitudinal Rapoport effects are a local phenomenon, occurring only for the Northeastern Pacific marine taxa. The relationship between latitudinal range extent and mean latitude indicated the existence of two hard boundaries at either extreme of the Chilean coast, limiting the geographical ranges of the species. However, geometric constraints tested using a Monte Carlo simulation approach showed a weak level of mid-domain effect on species richness. Finally, we propose that geometric constraint together with the geomorphology and historical characteristics of the Chilean coast explain the biogeographical patterns of benthic polychaete taxa in Chile.     

Resprouting in the Mediterranean‐type shrub Erica australis afffected by soil resource availability

Abstract. Soil resource availability may affect plant regeneration by resprouting in disturbed environments directly, by affecting plant growth rates, or indirectly by determining allocation to storage in the resprouting organs. Allocation to storage may be higher in stressful, low resource‐supply soils, but under such conditions plant growth rates may be lower. These factors could act in opposite directions leading to poorly known effects on resprouting. This paper analyses the role played by soil resources in the production and growth of resprouts after removal of above‐ground plant tissues in the Mediterranean shrub Erica australis. At 13 sites, differing in substrate, we cut the base of the stems of six plants of E. australis and allowed them to resprout and grow for two years. Soils were chemically analysed and plant water potential measured during the summer at all sites to characterize soil resource availability. We used stepwise regression analysis to determine the relationships between the resprouting response [mean site values of the number of resprouts (RN), maximum length (RML) and biomass (RB)] and soil nutrient content and plant water potential at each site. During the first two years of resprouting there were statistically significant differences among sites in the variables characterizing the resprouting response. RML was always different among sites and had little relationship with lignotuber area. RN was less different among sites and was always positively correlated with lignotuber area. RB was different among sites after the two years of growth. During the first months of resprouting, RN and RML were highly and positively related to the water status of the plant during summer. At later dates soil fertility variables came into play, explaining significant amounts of variance of the resprouting variables. Soil extractable cations content was the main variable accounting for RML and RB. Our results indicate that resprout growth of E. australis is positively affected by high water availability at the beginning of the resprouting response and negatively so by high soil extractable cation content at later periods. Some of these factors had previously shown to be related, with an opposite sign, to the development of a relatively larger lignotuber. Indeed, RML and RB measured in the second year of resprouting were significantly and negatively correlated with some indices of biomass allocation to the lignotuber at each site. This indicates that sites favouring allocation to the resprouting organ may not favour resprout growth.     

Leiurus quinquestriatus venom inhibits different kinds of Ca2+-dependent K+ channels

A minor protein component of Leiurus quinquestriatus venom has been reported to inhibit selectively the apamin-insensitive Ca2+-dependent K+ channels of mammalian skeletal muscle (Miller, C., Moczydlowski, E., Latorre, R. and Phillips, M. (1985) Nature 313, 316-318). We report the effect of the venom on both the apamin-insensitive channels of the human erythrocyte, the Ehrlich cell and the rat thymocyte and the apamin-sensitive channel of the guinea pig hepatocyte. The venom inhibited Ca2+-dependent K+ transport in all the cases with a Ki value within the range of 1 to 10 micrograms/ml, similar to that reported previously in muscle. Valinomycin-induced K+ transport was also antagonized by the venom but its sensitivity was about 1/10 as much as that of the Ca2+-dependent K+ channel.     

Effects of microcin B17 on microcin B17-immune cells

When microcin B17-immune cells are treated with microcin B17 they show many of the physiological effects displayed by microcin B17-sensitive cells treated in the same way. DNA replication stops immediately and several SOS functions are subsequently induced. In sensitive cells these effects are irreversible and lead to cell death, whereas in immune cells they are reversible and there is no loss of viability. This is an unusual mechanism of immunity because it does not prevent the primary action of the microcin. The implications of this mechanism concerning the mode of action of microcin B17 and the induction of the SOS system are discussed.     

Quantitative measurement of smooth muscle shortening in isolated pig trachea

Matched porcine tracheal rings were exposed to theophylline and increasing doses of carbachol in Krebs solution. Histological sections of each ring were traced and each of the following dimensions measured: the external perimeter (Pe) and external area (Ae) defined by the outer border of smooth muscle and inner surface of cartilage, and the internal perimeter (Pi) and internal area (Ai) defined by the luminal surface of the epithelium and the muscle length (L) along its outer border. Absolute wall area (WA = Ae - Ai) and relative wall area (PW = WA/Ae) were calculated. Carbachol-treated tracheal ring dimensions were compared with those of their matched theophylline-treated rings. In tracheal rings with intact cartilage, maximal smooth muscle shortening of 44% was achieved with 10(-2) M carbachol. In tracheal rings in which anterior and posterior segments of cartilage were excised, the trachealis muscle passively shortened by 20% and maximal shortening (10(-3) M carbachol) was 57%. Although Ai decreased with maximal smooth muscle shortening, there were no changes in the length of Pi or in WA. These data show that the cartilage in the porcine trachea exerts both a preload that passively stretches the trachealis muscle and an afterload that limits maximal smooth muscle shortening.     

CORRELATED BIOCHEMICAL AND ULTRASTRUCTURAL CHANGES IN NITROGEN-STARVED EUGLENA GRACILIS1

Growth of Euglena gracilis Z Pringsheim under photoheterotrophic conditions in a nitrogen-deprived medium resulted in progressive loss of chloroplastic material until total bleaching of the cells occurred. Biochemical analysis and ultrastructural observation of the first stages of the starvation process demonstrated an early lag phase (from 0 to 9 h) in which cells increased in size, followed by a period of cell division, apparently supported by the mobilization of some chloroplastic proteins such as the photosynthetic CO₂-fixing enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase. The degradation of the enzyme started after 9 h of starvation and was preceded by a transient concentration of this protein in pyrenoidal structures. Protein nitrogen and photosynthetic pigments as well as number of chloroplasts per cell decreased during proliferation through mere distribution among daughter cells. However, after 24 h, when cell division had almost ceased, there was a slow but steady decline of photosynthetic pigments. This was paralleled by observable ultrastructural changes including progressive loss of chloroplast structure and accumulation of paramylon granules and lipid globules in the cytoplasm. These findings reinforce the role of chloroplastic materials as a nitrogen source during starvation of E. gracilis in a carbon-rich medium. The excess of ribulose-1,5-bisphosphate carboxylase/oxygenase acts as a first reservoir that, once exhausted, is superseded by the generalized disassembly of the photosynthetic structures, if the adverse environment persists more than 24 h.