Receptor-mediated endocytosis of Neisseria gonorrhoeae into primary human urethral epithelial cells: the role of the asialoglycoprotein receptor

Urethral epithelial cells are invaded by Neisseria gonorrhoeae during gonococcal infection in men. To understand further the mechanisms of gonococcal entry into host cells, we used the primary human urethral epithelial cells (PHUECs) tissue culture system recently developed by our laboratory. These studies showed that human asialoglycoprotein receptor (ASGP-R) and the terminal lactosamine of lacto-N-neotetraose-expressing gonococcal lipooligosaccharide (LOS) play an important role in invasion of PHUECs. Microscopy studies showed that ASGP-R traffics to the cell surface after gonococcal challenge. Co-localization of ASGP-R with gonococci was observed. As ASGP-R-mediated endocytosis is clathrin dependent, clathrin localization in PHUECs was examined after infection. Infected PHUECs showed increased clathrin recruitment and co-localization of clathrin and gonococci. Preincubating PHUECs in 0.3 M sucrose or monodansylcadaverine (MDC), which both inhibit clathrin-coated pit formation, resulted in decreased invasion. N. gonorrhoeae strain 1291 produces a single LOS glycoform that terminates with Gal(beta1-4)Glc-Nac(beta1-3)Gal(beta1-4)Glc (lacto-N-neotetraose). Invasion assays showed that strain 1291 invades significantly more than four isogenic mutants expressing truncated LOS. Sialylation of strain 1291 LOS inhibited invasion significantly. Preincubation of PHUECs in asialofetuin (ASF), an ASGP-R ligand, significantly reduced invasion. A dose-response reduction in invasion was observed in PHUECs preincubated with increasing concentrations of NaOH-deacylated 1291 LOS. These studies indicated that an interaction between lacto-N-neotetraose-terminal LOS and ASGP-R allows gonococcal entry into PHUECs.

The urethral pressure profile and ultrasound imaging of the lower urinary tract

In a prospective study 105 patients with symptoms of stress incontinence underwent video-urodynamic testing, including resting urethral pressure profilometry and translabial ultrasound. The urethral pressure profile (UPP) included maximum urethral closure pressure (MUCP), functional length (FL) and area under the curve (AUC). Ultrasound parameters included urethral thickness, urethral rotation and bladder neck descent, as well as funneling/opening of the internal urethral meatus on Valsalva maneuver. Levator contraction strength was assessed measuring the cranioventral displacement of the internal meatus. Negative correlations between UPP data and age, parity and previous surgery were observed which were consistent with literature data. There was a positive correlation :between the urethral AP diameter on ultrasound and the MUCP, which agrees with reports showing reduced sphincter thickness or volume in stress-incontinent women. Hypermobility on ultrasound did not correlate with UPP data. However, a lower MUCP correlated with extensive opening of the bladder neck. Finally, there was a trend towards poorer pelvic floor function with lower MUCP measurements.

Multi-stage evolution of a sub-aerial volcanic ridge over the last 1.3 Myr: S. Jorge Island, Azores Triple Junction

New K/Ar dating and geochemical analyses have been carried out on the WNW-ESE elongated oceanic island of S. Jorge to reconstruct the volcanic evolution of a linear ridge developed close to the Azores triple junction. We show that S. Jorge sub-aerial construction encompasses the last 1.3 Myr, a time interval far much longer than previously reported. The early development of the ridge involved a sub-aerial building phase exposed in the southeast end of the island and now constrained between 1.32 +/- 0.02 and 1.21 +/- 0.02 Ma. Basic lavas from this older stage are alkaline and enriched in incompatible elements, reflecting partial melting of an enriched mantle source. At least three differentiation cycles from alkaline basalts to mugearites are documented within this stage. The successive episodes of magma rising, storage and evolution suggest an intermittent reopening of the magma feeding system, possibly due to recurrent tensional or trans-tensional tectonic events. Present data show a gap in sub-aerial volcanism before a second main ongoing building phase starting at about 750 ka. Sub-aerial construction of the S. Jorge ridge migrated progressively towards the west, but involved several overlapping volcanic episodes constrained along the main WNW-ESE structural axis of the island. Malic magmas erupted during the second phase have been also generated by partial melting of an enriched mantle source. Trace element data suggest, however, variable and lower degrees of partial melting of a shallower mantle domain, which is interpreted as an increasing control of lithospheric deformation on the genesis and extraction of primitive melts during the last 750 kyr. The multi-stage development of the S. Jorge volcanic ridge over the last 1.3 Myr has most likely been greatly influenced by regional tectonics, controlled by deformation along the diffuse boundary between the Nubian and the Eurasian plates, and the increasing effect of sea-floor spreading at the Mid-Atlantic Ridge.

Avaliação radiológica do refluxo vesico – ureteral em crianças

O Refluxo Vesico – Ureteral (RVU) é uma patologia frequente na idade pediátrica. A detecção precoce de RVU é fundamental na orientação terapêutica, de modo a permitir um crescimento renal adequado e prevenir infecções urinárias recorrentes. A investigação radiológica de RVU baseia-se na ecografia e cistouretrografia miccional seriada (CUMS), sendo este último procedimento, o método de eleição, efectuado sob controlo fluoroscópico. Este trabalho tem como objectivo dar a conhecer o papel CUMS na avaliação de RVU e abordar os parâmetros técnicos de aquisição, posicionamento e critérios de qualidade de imagem.

Detection of mycoplasmas in urethral swabs from HIV-1 infected patients and control individuals using culture techniques and polymerase chain reaction

The objective of the present study was to determine the prevalence of certain mycoplasma species, i.e., Mycoplasma hominis, Ureaplasma urealyticum and Mycoplasma penetrans, in urethral swabs from HIV-1 infected patients compared to swabs from a control group. Mycoplasmas were detected by routine culture techniques and by the Polymerase Chain Reaction (PCR) technique, using 16SrRNA generic primers of conserved region and Mycoplasma penetrans specific primers. The positivity rates obtained with the two methods were comparable. Nevertheless, PCR was more sensitive, while the culture techniques allowed the quantification of the isolates. The results showed no significant difference (p < 0.05) in positivity rates between the methods used for mycoplasma detection.

Molecular analysis of the dengue virus type 1 and 2 in Brazil based on sequences of the genomic envelope-nonstructural protein 1 junction region

The genomic sequences of the Envelope-Non-Structural protein 1 junction region (E/NS1) of 84 DEN-1 and 22 DEN-2 isolates from Brazil were determined. Most of these strains were isolated in the period from 1995 to 2001 in endemic and regions of recent dengue transmission in São Paulo State. Sequence data for DEN-1 and DEN-2 utilized in phylogenetic and split decomposition analyses also include sequences deposited in GenBank from different regions of Brazil and of the world. Phylogenetic analyses were done using both maximum likelihood and Bayesian approaches. Results for both DEN-1 and DEN-2 data are ambiguous, and support for most tree bipartitions are generally poor, suggesting that E/NS1 region does not contain enough information for recovering phylogenetic relationships among DEN-1 and DEN-2 sequences used in this study. The network graph generated in the split decomposition analysis of DEN-1 does not show evidence of grouping sequences according to country, region and clades. While the network for DEN-2 also shows ambiguities among DEN-2 sequences, it suggests that Brazilian sequences may belong to distinct subtypes of genotype III.

Disfunção Vesical, Infecção Urinária e Refluxo Vesico-Ureteral na Criança

Tem sido descrita uma correlação estreita entre infecção urinária, refluxo vesico-ureteral e disfunção miccional na criança. A obstrução funcional causada pela disfunção vesical/uretral representa um elevado risco de recorrência de infecção urinária, indução e perpetuação do refluxo (mesmo após correcção cirúrgica) é de lesão renal permanente. A normalização da alteração da micção como problema primário, é crítica na resolução de problemas secundários tais como a infecção urinária e o refluxo vesico-ureteral. Trinta e sete crianças com refluxo vesicoureteral secundário a disfunção miccional foram detectadas, avaliadas e tratadas entre 1990 e 1995 (5 anos). Foram estudados 49 ureteres. A infecção urinária foi o sintoma revelador em todas as crianças, ocorrendo entre 1 mês e os 13 anos de idade (mediana de 3,5 anos). Todas as crianças eram neurologicamente e estruturalmenle normais, detectando-se sintomatologia sugestiva de instabilidade ou imaturidade vesical em 34 (91,9%) e sugestiva de obstrução esfiocteriana funcional em três (8,1 %). Os estudos ecográfico e cistográfico efectuado em todas as crianças, com o apoio do estudo urodinâmico em 17 (45,9%) confirmaram o diagnóstico clínico. Em 29 (78,4%) das crianças foi efectuada cintigrafia com DMSA (Addo Dimercaptosuccínico), revelando cicatriz renal em 26 (89,6%) dos exames. Foi incentivado um programa de reeducação vesical e regularização dos hábitos intestinais em todas as crianças, associado a terapêutica anticolinérgica em 23 (62,2%) e/ou relaxantes musculares em três (8,1%) e fenoxibenzamina e algaliação intermitente (1,5 mês) em uma (2,7%), para além da quimioprofilaxia da infecção urinária instituida em 34 (91,9%) das crianças. Houve resolução completa da infecção urinária em 35 (94,6%) com redução da sua frequência nas outras duas (5,4%), cura do RVU em 32 (86,5%) e melhoria em quatro (10,8%). Verificou-se desaparecimento dos sinais de disfunção vesical em 22 (59,5%) casos com redução na intensidade e frequência em 14 (37,8%), mantendo-se uma criança (2,7%) com síndrome de urgência e refluxo vesico-ureteral inalterado. Estes dados implicam que a detecção e traramento da disfunção vesical/esfincteriana, são essenciais em todas as crianças com o complexo infecção urinária recorrente e refluxo vesicoureteral.

Genetic diversity of dengue virus serotypes 1 and 2 in the State of Paraná, Brazil, based on a fragment of the capsid/premembrane junction region

INTRODUCTION: The precise identification of the genetic variants of the dengue virus is important to understand its dispersion and virulence patterns and to identify the strains responsible for epidemic outbreaks. This study investigated the genetic variants of the capsid-premembrane junction region fragment in the dengue virus serotypes 1 and 2 (DENV1-2). METHODS: Samples from 11 municipalities in the State of Paraná, Brazil, were provided by the Central Laboratory of Paraná. They were isolated from the cell culture line C6/36 (Aedes albopictus) and were positive for indirect immunofluorescence. Ribonucleic acid (RNA) extracted from these samples was submitted to the reverse transcription polymerase chain reaction (RT-PCR) and nested PCR. RESULTS: RT-PCR revealed that 4 of the samples were co-infected with both serotypes. The isolated DENV-1 sequences were 95-100% similar to the sequences of other serotype 1 strains deposited in GenBank. Similarly, the isolated DENV-2 sequences were 98-100% similar to other serotype 2 sequences in GenBank. According to our neighbor-joining tree, all strains obtained in this study belonged to genotype V of DENV-1. The DENV-2 strains, by contrast, belonged to the American/Asian genotypes. CONCLUSIONS: The monitoring of circulating strains is an important tool to detect the migration of virus subtypes involved in dengue epidemics.

Urethral catheter removal 7 or 14 days after radical retropubic prostatectomy: clinical implications and complications in a randomized study

PURPOSE: To evaluate the hypothesis that a 7-day period of indwelling catheter after radical retropubic prostatectomy is effective and safe without the need of performing cystography. METHODS: In the period from January of 2000 to July of 2002, 73 patients underwent radical retropubic prostatectomy, and these patients were prospectively randomized in 2 groups: Group 1-37 patients who had the urethral catheter removed 7 days after the procedure, and Group 2-36 patients who had the catheter removed 14 days after the surgery. The 2 groups were similar, the surgeons and the technique were the same, and no cystography was performed to evaluate the presence of leaks. RESULTS: Two patients in Group 1 had bleeding and clot retention after having the catheter taken out in the seventh postoperative day and were managed by putting the catheter back in for 7 more days. Two patients in Group 2 developed bladder neck stricture and were treated by bladder neck incision with success. The continence rate was the same, with 2 cases of incontinence in each group. About 2 pads a day were used by the patients with incontinence. The average follow-up was 17.5 months (12-36 months). No urinary fistula, urinoma, or pelvic abscesses developed after catheter removal. Two patients were excluded from the analysis of this series: 1 died with a pulmonary embolus in the third postoperative day, and 1 developed a urinary suprapubic fistula before catheter withdrawal, which was maintained for 16 days. CONCLUSION: Withdrawal of the urethral catheter 7 days after radical retropubic prostatectomy, without performing cystography, has a low rate of short-term complications that are equivalent to withdrawal 14 days after the surgery.

Morphofunctional study of the junction between the left atrium and the pulmonary veins in patient with pulmonary hypertension

OBJECTIVE: To study the arrangement of the myocardial fiber bundles at the pulmonary venous left atrial junction in patients with pulmonary hypertension, and to discuss the pathophysiological importance of this element in the etiology of acute pulmonary edema. METHODS: We obtained 12 hearts and their pulmonary vein extremities from postmortem examinations of patients with the anatomicopathological diagnosis of acute pulmonary edema. The specimens, which had no grossly visible morphological cardiac alterations, were fixed in 10% formalin, and the muscular arrangement of the pulmonary venous left atrial junctions was analyzed. This material was then isolated, embedded in paraffin, underwent serial cutting (50 µm of thickness), and was stained with Azam's trichrome. RESULTS: We observed in our specimens that: a) the myocardial fiber bundles that originate in the atrial wall and involve the openings of the pulmonary veins were fewer than those observed in healthy material; b) the myocardial fiber bundles that extend into the pulmonary veins were shorter than those found in material originating from individuals with no pulmonary hypertension. CONCLUSION: Anatomical changes that result in a reduction in the amount of myocardial fiber bundles in the pulmonary venous left atrial junction, isolated or associated with other factors, may be the cause of disorders in pulmonary circulation, leading to an increase in pulmonary venous pressure, and, consequently, to acute pulmonary edema.

Positive Torsional Strain Causes the Formation of a Four-Way Junction at Replication Forks.

Research by L. Postow, C. Ullsperger, R.W. Keller, C. Bustamante, A.V. Vologodskii, and N.R. Cozzarelli, J. Biol. Chem. 2001, 276, 2790 Condensation and commentary by Alexander Bucka and Andrzej Stasiak, Universite ´ de Lausanne, Switzerland Purpose of the Study To demonstrate that positive torsional strain generated during DNA replication can lead to reversals of replication forks and, consequently can result in the formation of four-way DNA junctions

Modulation of the gap junction protein Connexin36 in neurons in a mouse model of transient focalcerebral ischemia

Intercellular communication is achieved at specialized regions of the plasma membrane by¦gap junctions. Gap junctions are transmembrane channels allowing direct contacts between¦the cytoplasms of neighboring cells. Each cell participates with one hemichannel, or¦connexon, made of six protein subunits named connexins. Thanks to these junctions, cells¦potentially share a pool of small molecules and metabolites, such as nucleotides, amino acids¦and second messengers.¦In an ischemic (i.e. non-perfused) territory of the brain, irreversible damage progresses over¦time from the centre of the most severe flow reduction to the periphery with less disturbed¦perfusion. Functionally impaired tissue can survive and recover if sufficient reperfusion is reestablished¦within a limited time period, which depends on various factors and mechanisms¦modulating the signaling pathways leading to cell death.¦Observations were made indicating the presence of electrical coupling between neurons which¦resist better to an ischemic insult. This electrical coupling is likely to be mediated by¦Connexin36 (Cx36), a neuron specific connexin isoform. It was demonstrated in the past that¦global ischemia induces a selective upregulation of Cx36 expression in regions with neurons¦that survive the insult whereas others undergo apoptosis and die. These observations raise the¦possibility that the neuronal gap junction Cx36 might play a role in the destiny of neurons¦after cerebral ischemia.¦The aim of this work was to characterize the regulation of Connexin36 in a mouse model of¦transient focal cerebral ischemia by immunofluorescence and Western blot analysis. Our¦immunofluorescence results suggest a specific increase in Cx36 in the penumbral region of¦the ischemic hemisphere.