Patterns of enzyme activities in the gills of the catfish Heteropneustes fossilis (Bloch) exposed to the anionactive detergent Na-alkyl-benzenesulphonate (LAS)

Summary The effect of sodium-alkyl-benzenesulphonate (LAS) on the activity of the respiratory enzymes of the gills of Heteropneustes fossilis (Bloch) was investigated. After 48 h exposure, the main injury to gills was the progressive separation of the lamellae from their vascular components. The enzymes of the aerobic part of the metabolism showed a decrease in activity, whereas the activity of lactate dehydrogenase was strongly increased, thus indicating that LAS has a high potential to interfere with aerobic mechanisms; however, the mode of action of it has yet to be clearly defined.     

Mitochondria-rich cells in anuran amphibia: chloride conductance and regional distribution over the body surface

The distribution and density (D(mrc)) of mitochondria-rich cells (MR cells) in skin epithelium, were determined over the whole body surface in nine species of anuran Amphibia that live in a variety of habitats. It was found that the more terrestrial species (beginning with Hyla arborea) have a higher density of MR cells in their pelvic region. In the skin of aquatic (Xenopus laevis) or fossorial (Pelobates syriacus) species, D(mrc) is evenly distributed over the whole body surface. In dorsal skin pieces of H. arborea that lack detectable MR cells, transepithelial voltage activation did not induce Cl(-) conductance as it did in ventral pieces. Skins from Bufo viridis and X. laevis, both have MR cells in their skin, differ markedly in their biophysical properties: a Cl(-) specific current conductance is predominant in the skin epithelium of B. viridis, and is absent in X. laevis. In the latter, anionic conductance is due to glandular secretion. The biophysical properties cannot therefore be related solely to the presence or density of MR cells. Mitochondria-rich cells are sites of Cl(-) conductance across the skin of those amphibians that show this property, but must have different function(s) in other species. It is suggested that the specific zonal distribution of MR cells in the species that were examined in this study could be due to ion exchange activity and water conservation in more terrestrial environments.     

Intestinal passive absorption of water-soluble compounds by sparrows: effect of molecular size and luminal nutrients

We tested predictions that: (1) absorption of water-soluble probes decreases with increasing molecular size, consistent with movement through effective pores in epithelia, and (2) absorption of probes is enhanced when measured in the presence of luminal nutrients, as predicted for paracellular solvent drag. Probes (L-arabinose, L-rhamnose, perseitol, lactulose; MW 150.1-342.3 Da) were gavaged in nonanesthetized House sparrows ( Passer domesticus), or injected into the pectoralis, and serially measured in plasma. Bioavailability was calculated as F=AUC by gavage/AUC by injection, where AUC is the area under the curve of plasma probe concentration vs. time. Consistent with predictions, F declined with probe size by 75% from the smallest to the largest probe, and absorption of probes increased by 40% in the presence of luminal glucose or food compared to a mannitol control. Absorption of water-soluble probes by sparrows is much higher than in humans, which is much higher than in rats. These differences seem mainly attributable to differences in paracellular solvent flux and less to differences in effective paracellular pore size.     

"Side" effects of HAART: decreasing and changing occupational exposure to HIV-infected patients.

To investigate percutaneous exposures to HIV in the highly active antiretroviral therapy (HAART) era, we performed an analysis of all percutaneous exposures reported from January 1994 to December 1998 in 18 Italian acute-care hospitals. Frequency and rate per 100 prevalent AIDS cases of HIV exposures decreased by 40% (from 4.3% to 2.6%, and from 1.0% to 0.6%, respectively; p<0.001), which were mainly those related to the insertion/manipulation of peripheral vascular access devices (from 7.2% to 4.8%; p=0.05). We conclude that the benefits of HAART have changed the complexity of care required and therefore, the number and type of procedures performed on HIV patients that place the HCW at risk of injury.     

Proinflammatory cytokine responses induced by influenza A (H5N1) viruses in primary human alveolar and bronchial epithelial cells

Background

Fatal human respiratory disease associated with influenza A subtype H5N1 has been documented in Hong Kong, and more recently in Vietnam, Thailand and Cambodia. We previously demonstrated that patients with H5N1 disease had unusually high serum levels of IP-10 (interferon-gamma-inducible protein-10). Furthermore, when compared with human influenza virus subtype H1N1, the H5N1 viruses in 1997 (A/Hong Kong/483/97) (H5N1/97) were more potent inducers of pro-inflammatory cytokines (e.g. tumor necrosis factor-a) and chemokines (e.g. IP-10) from primary human macrophages in vitro, which suggests that cytokines dysregulation may play a role in pathogenesis of H5N1 disease. Since respiratory epithelial cells are the primary target cell for replication of influenza viruses, it is pertinent to investigate the cytokine induction profile of H5N1 viruses in these cells.

Methods

We used quantitative RT-PCR and ELISA to compare the profile of cytokine and chemokine gene expression induced by H5N1 viruses A/HK/483/97 (H5N1/97), A/Vietnam/1194/04 and A/Vietnam/3046/04 (both H5N1/04) with that of human H1N1 virus in human primary alveolar and bronchial epithelial cells in vitro.

Results

We demonstrated that in comparison to human H1N1 viruses, H5N1/97 and H5N1/04 viruses were more potent inducers of IP-10, interferon beta, RANTES (regulated on activation, normal T cell expressed and secreted) and interleukin 6 (IL-6) in primary human alveolar and bronchial epithelial cells in vitro. Recent H5N1 viruses from Vietnam (H5N1/04) appeared to be even more potent at inducing IP-10 than H5N1/97 virus.

Conclusion

The H5N1/97 and H5N1/04 subtype influenza A viruses are more potent inducers of proinflammatory cytokines and chemokines in primary human respiratory epithelial cells than subtype H1N1 virus. We suggest that this hyper-induction of cytokines may be relevant to the pathogenesis of human H5N1 disease.     

Carotenoid and ultrastructure variations in plastids of Arum italicum Miller fruit during maturation and ripening

The changes in the pigment pattern and composition occurring in the Arum italicum berry during the various steps of maturation (ivory to deep-green stages) and ripening (yellow and red-orange stages) were studied and correlated to the ultrastructural modifications of plastids. Transmission electron microscopy showed that each stage was characterized by a specific plastidial type following the unusual sequence amyloplast-->chloroplast-->chromoplast. Plastidial transitions were accompanied by profound modifications in the pigmental composition, in particular, in the pattern of carotenoids and their precursors. The HPLC analysis of the carotenoids showed that, besides the two usual all-trans metabolic pathways leading to lutein through alpha-carotene and to auroxanthin through beta-carotene, an additional cis-isomeric biosynthetic pathway leading to cis-neoxanthin through cis-beta-carotene exists. During the pre-ripening stages, the three pathways were present even if with qualitative and quantitative variations. When complete ripening was reached, a block occurred at the cyclization level causing the accumulation of both all-trans (i.e. gamma-carotene and neurosporene) and cis-isomer (i.e. lycopene and zeta-carotene) carotene precursors. Because of the occurrence of unusual pigments and the presence of the three main plastidial types, the fruit of A. italicum may constitute a most instructive model for the study of carotenogenesis.     

Molecular dissection of cytokinesis by RNA interference in Drosophila cultured cells

We have used double-stranded RNA-mediated interference (RNAi) to study Drosophila cytokinesis. We show that double-stranded RNAs for anillin, acGAP, pavarotti, rho1, pebble, spaghetti squash, syntaxin1A, and twinstar all disrupt cytokinesis in S2 tissue culture cells, causing gene-specific phenotypes. Our phenotypic analyses identify genes required for different aspects of cytokinesis, such as central spindle formation, actin accumulation at the cell equator, contractile ring assembly or disassembly, and membrane behavior. Moreover, the cytological phenotypes elicited by RNAi reveal simultaneous disruption of multiple aspects of cytokinesis. These phenotypes suggest interactions between central spindle microtubules, the actin-based contractile ring, and the plasma membrane, and lead us to propose that the central spindle and the contractile ring are interdependent structures. Finally, our results indicate that RNAi in S2 cells is a highly efficient method to detect cytokinetic genes, and predict that genome-wide studies using this method will permit identification of the majority of genes involved in Drosophila mitotic cytokinesis.     

Cholesterol exchange and synthesis in the live rat

The turnover of plasma cholesterol and de novo cholesterol synthesis were measured simultaneously in the live rat, immediately after administration of [3H]water together with a large volume exchange transfusion of whole blood prelabeled with [14C]cholesterol. It was possible to separate the exchange of unesterified cholesterol from the uptake and secretion of lipoprotein cholesteryl ester, and also to assess the impact of plasma cholesterol exchange on the measurement of in vivo rates of cholesterolgenesis by individual tissues. Cholesterol was measured by an HPLC procedure that effectively separated cholesterol from other structurally similar sterols, and synthesis was determined by the incorporation of [3H]water into cholesterol. Plasma unesterified cholesterol turnover was multiphasic and exceedingly rapid (initial T1/2, 4.1 min) in contrast to the near linear and much slower turnover of plasma cholesteryl ester (initial T1/2, 59.4 min). Plasma unesterified cholesterol equilibrated with different tissues at different rates, with the liver and adrenal equilibrating most rapidly. Full equilibration of plasma unesterified cholesterol was not achieved with any tissue during the course of this study. For rapidly exchanging tissues like the liver, which was responsible for about 60% of plasma unesterified cholesterol exchange, unesterified cholesterol appeared to be kinetically compartmentalized into rapidly, and much less rapidly, exchangeable pools. After [3H]water administration, the content of newly synthesized cholesterol was greatest in the liver, adrenal, and intestine, and appreciably lower in all other tissues studied. Hepatectomy and intestinal resection resulted in a profound reduction of newly synthesized cholesterol in the plasma and adrenal, but no certain change in the already low amounts at other sites. Thus, while it is clear that appreciable amounts of newly synthesized cholesterol in the adrenal were derived from the plasma by exchange, it was not possible to make this assessment for other selected individual tissues. When, however, newly synthesized cholesterol was determined in the total mass of all extrahepatic and extraintestinal tissues together, exchange could be calculated to account for close to 50% of the new cholesterol recovered in the carcass (in studies of 60 min duration). After correcting for exchange, the liver accounted for 82% of all newly synthesized cholesterol, the intestine for about 10%, and the remaining tissues of the body for just 9%. These results are in marked contrast to recent findings of others and demonstrate that in the live rat cholesterol synthesis is principally confined to the liver.     

Lectin-binding pattern on the surface epidermis of Ambystoma tigrinum larvae

Summary The surface epidermis of Ambystoma tigrinum larvae was examined at the light- and electron-microscope levels using five different lectin conjugates as probes for the detection of sugar residues on the cell membranes. Concanavalin A (Con-A), wheat-germ agglutinin (WGA), Ricinus communis agglutinin I (RCA-I), Dolichos biflorus agglutinin and soybean agglutinin (SBA) conjugates clearly labelled the surface cells, especially their apical surfaces. At electron microscopy, the labelling on plasma membranes was found to exhibit regional differences. Among the lectins tested WGA displayed a particularly characteristic binding pattern. WGA also bound to basolateral cell surfaces, including the tight-junction zone wich was also stained by the RCA-I conjugate. The different labelling intensity and staining patterns obtained with the conjugates indicated the polarity of the cell surfaces. It is also assumed that the WGA staining of the basolateral membranes and intercellular spaces reflected transcellular transport, which is facilitated by acidic glycoconjugates. Other functional aspects of the polarized distribution of the lectin conjugates were also correlated with the receptor sites of certain sugar residues.