Antiviral action of the tumor suppressor ARF

Oncogenic viruses frequently target the pathways controlled by tumor suppressor genes, suggesting an extra function for these proteins as antiviral factors. The control exerted by the tumor suppressor Arf on cellular proliferation is crucial to restrict tumor development; however, a potential contribution of Arf to prevent viral infectivity has remained unexplored. In the present study, we investigated the consequences of loss or increased expression of Arf on viral infection. Our results reveal that ARF expression is induced by interferon and after viral infection. Furthermore, we show that ARF protects against viral infection in a gene dosage-dependent manner, and that this antiviral action is mediated in part by PKR through a mechanism that involves ARF-induced release of PKR from nucleophosmin complexes. Finally, Arf-null mice were hypersensitive to viral infection compared to wild-type mice. Together, our results reveal a novel and unexpected role for the tumor suppressor ARF in viral infection surveillance.     

Bacterial Analysis of Breast Milk: A Tool to Differentiate Raynaud’s Phenomenon from Infectious Mastitis During Lactation

Lactational Raynaud’s syndrome may be misdiagnosed as infectious mastitis on the basis of the breast pain. The objective of this work was to elucidate if microbiological analysis of milk may contribute to the differentiation of both conditions. Ten lactating women clinically diagnosed by Spanish lactation consultants were included in the study. Of these, five suffered from mastitis and the remaining five suffered from Raynaud’s syndrome. Breast milk samples were inoculated on diverse culture media. Seventy isolates were selected and identified by 16S rDNA PCR sequencing. Parallel, PCR-DGGE and quantitative real-time PCR were used to assess the presence of bacterial DNA in the samples. Neither bacteria nor yeasts could be detected in the milk samples provided by the women suffering from Raynaud’s syndrome. In contrast, large numbers of bacteria were isolated from those with infectious lactational mastitis. Globally, the levels of bacterial DNA were significantly higher in the milk of mastitis-suffering women. Bacteriological analysis of milk can be an useful tool to facilitate the differential diagnosis between the infectious mastitis and Raynaud’s syndrome during lactation.     

Polychaetes and oligochaetes associated with intertidal rocky shores in a semi-enclosed industrial and urban embayment,with the description of two new species

The species composition and relative abundance of the annelid benthic macrofauna (Polychaeta and Oligochaeta) inhabiting the rocky intertidal zone of the ria of Ferrol (Galicia, NW Spain) were studied during field collections, from 2000 to 2002. A total of 14,619 specimens (11,585 polychaetes belonging to 76 species and 24 families and 3,034 oligochaetes belonging to 18 species and two families) were collected from 98 quantitative samples taken from 13 sampling sites. The general spatial distribution of the annelid fauna reflects an increase of the diversity from the inner to the outer part of the ria. The general patterns found in the annelid composition suggest that the assemblages were dominated by oligochaetes in the inner sheltered sampling sites and polychaetes in the outer more exposed sites. Several faunistical and taxonomical remarks on selected species are presented. Two new species of oligochaetes are described: Coralliodrilus artabrensis sp. n. and Pirodrilus fungithecatus sp. n. (Naididae, Phallodrilinae). A new biological index, based on the oligochaete/polychaete ratio (O/P), is proposed as tool to evaluate environmental quality and to monitor future changes in the environment.     

Diketopiperazines produced by endophytic fungi found in association with two Asteraceae species

Diketopiperazine (DKP) derivatives, named colletopiperazine, fusaperazine C and E as well as four known DKPs were isolated from cultures of Colletotrichum gloeosporioides, Penicillium crustosum, both endophytic fungi isolated from Viguiera robusta, and a Fusarium spp., an endophyte of Viguiera arenaria, respectively. Their structures were established on the basis of their spectroscopic data. Conformational analysis of two known DKPs showed that folded conformations were as energetically stable as the extended one.     

Velopharyngeal motion after sphincter pharyngoplasty: a videonasopharyngoscopic and electromyographic study

Sphincter pharyngoplasty is a surgical procedure for managing velopharyngeal insufficiency after palatal closure. This procedure is intended to create an active diaphragm for velopharyngeal closure. The purpose of this study was to evaluate velopharyngeal motion after sphincter pharyngoplasty, by using selective electromyography and simultaneous videonasopharyngoscopy. Twenty-five patients who were subjected to sphincter pharyngoplasty from 1985 to 1996 were reviewed. All conditions were evaluated by using electromyography with simultaneous videonasopharyngoscopy. The following velopharyngeal muscles were examined: superior constrictor pharyngeus, palatopharyngeus, and levator veli palatini. The palatopharyngeus was included in the superiorly based surgical flaps inserted at the posterior pharyngeal wall. Twenty-three patients (92 percent) showed complete velopharyngeal closure. The two patients with residual velopharyngeal insufficiency showed a defect size of 20 and 25 percent. None of the patients showed electromyographic activity at the superiorly based flaps, indicating absence of activity of the palatopharyngeus muscles. However, all patients showed normal electromyographic activity at the superior constrictor pharyngeus and the levator veli palatini. Videonasopharyngoscopy demonstrated that lateral pharyngeal wall movements, which ranged from 25 to 40 percent, were related to strong electromyographic activity at the superior constrictor pharyngeus. It is concluded that the superiorly based pharyngeal flaps of the sphincter pharyngoplasty do not seem to create an active diaphragm for velopharyngeal closure. Moreover, the observed sphinctering seems to be passive, caused by the contraction of the superior constrictor pharyngeus.     

Isolation, structure, and HIV-1-integrase inhibitory activity of structurally diverse fungal metabolites

HIV-1 integrase is a critical enzyme for replication of HIV, and its inhibition is one of the most promising new drug strategies for anti-retroviral therapy, with potentially significant advantages over existing therapies. In this report, a series of HIV-1 inhibitors isolated from the organic extract of fermentations from terrestrial fungi is described. These fungal species, belonging to a variety of genera, were collected from throughout the world following the strict guidelines of Rio Convention on Biodiversity. The polyketide- and terpenoid-derived inhibitors are represented by two naphthoquinones, a biphenyl and two triphenyls, a benzophenone, four aromatics with or without catechol units, a linear aliphatic terpenoid, a diterpenoid, and a sesterterpenoid. These compounds inhibited the coupled and strand-transfer reaction of HIV-1 integrase with an IC₅₀ value of 0.5–120 µM. The bioassay-directed isolation, structure elucidation, and HIV-1 inhibitory activity of these compounds are described.Abbreviations CH ₃ CN Acetonitrile - TFA Trifluoroacetic acid - CD ₃ CN Deuterio acetonitrile - CD ₂ Cl ₂ Deuterio methylene chloride - MEK Methyl ethyl ketone - ZnSe Zinc selenide - UV Ultraviolet - IR Infrared - NMR Nuclear magnetic resonance - COSY Homonuclear H-H correlation spectroscopy - HMQC Heteronuclear multiple-quantum coherence - HMBC Heteronuclear multiple-bond correlation - EIMS Electron-impact mass spectrometry - HREIMS High-resolution electron impact mass spectrometry - ESIMS Electrospray ionization mass spectrometry - HRFABMS High-resolution fast atom bombardment mass spectrometry - MES 2-(N-morpholino)-ethane sulfonic acid     

Domain formation in sphingomyelin/cholesterol mixed membranes studied by spin-label electron spin resonance spectroscopy

Interactions of palmitoylsphingomyelin with cholesterol in multilamellar vesicles have been studied over a wide range of compositions and temperatures in excess water by using electron spin resonance (ESR) spectroscopy. Spin labels bearing the nitroxide free radical group on the 5 or 14 C-atom in either the sn-2 stearoyl chain of phosphatidylcholine (predominantly 1-palmitoyl) or the N-stearoyl chain of sphingomyelin were used to determine the mobility and ordering of the lipids in the different phases. Two-component ESR spectra of the 14-position spin labels demonstrate the coexistence first of gel (L(beta)) and liquid-ordered (L(o)) phases and then of liquid-ordered and liquid-disordered (L(alpha)) phases, with progressively increasing temperature. These phase coexistences are detected over a limited range of cholesterol contents. ESR spectra of the 5-position spin labels register an abrupt increase in ordering at the L(alpha)-L(o) transition and a biphasic response at the L(beta)-L(o) transition. Differences in outer splitting between the C14-labeled sphingomyelin and phosphatidylcholine probes are attributed to partial interdigitation of the sphingomyelin N-acyl chains across the bilayer plane in the L(o) state. In the region where the two fluid phases, L(alpha) and L(o), coexist, the rate at which lipids exchange between phases (<7 x 10(7) s(-)(1)) is much slower than translational rates in the L(alpha) phase, which facilitates resolution of two-component spectra.     

Different effects of long- and short-chain ceramides on the gel-fluid and lamellar-hexagonal transitions of phospholipids: a calorimetric, NMR, and x-ray diffraction study

The effects on dielaidoylphosphatidylethanolamine (DEPE) bilayers of ceramides containing different N-acyl chains have been studied by differential scanning calorimetry small angle x-ray diffraction and (31)P-NMR spectroscopy. N-palmitoyl (Cer16), N-hexanoyl (Cer6), and N-acetyl (Cer2) sphingosines have been used. Both the gel-fluid and the lamellar-inverted hexagonal transitions of DEPE have been examined in the presence of the various ceramides in the 0-25 mol % concentration range. Pure hydrated ceramides exhibit cooperative endothermic order-disorder transitions at 93 degrees C (Cer16), 60 degrees C (Cer6), and 54 degrees C (Cer2). In DEPE bilayers, Cer16 does not mix with the phospholipid in the gel phase, giving rise to high-melting ceramide-rich domains. Cer16 favors the lamellar-hexagonal transition of DEPE, decreasing the transition temperature. Cer2, on the other hand, is soluble in the gel phase of DEPE, decreasing the gel-fluid and increasing the lamellar-hexagonal transition temperatures, thus effectively stabilizing the lamellar fluid phase. In addition, Cer2 was peculiar in that no equilibrium could be reached for the Cer2-DEPE mixture above 60 degrees C, the lamellar-hexagonal transition shifting with time to temperatures beyond the instrumental range. The properties of Cer6 are intermediate between those of the other two, this ceramide decreasing both the gel-fluid and lamellar-hexagonal transition temperatures. Temperature-composition diagrams have been constructed for the mixtures of DEPE with each of the three ceramides. The different behavior of the long- and short-chain ceramides can be rationalized in terms of their different molecular geometries, Cer16 favoring negative curvature in the monolayers, thus inverted phases, and the opposite being true of the micelle-forming Cer2. These differences may be at the origin of the different physiological effects that are sometimes observed for the long- and short-chain ceramides.