M-protein gene-type distribution and hyaluronic acid capsule in group A Streptococcus clinical isolates in Chile: association of emm gene markers with csrR alleles.

Streptococcus pyogenes causes a variety of infections because of virulence factors such as capsular hyaluronic acid and M protein. The aim of this study was to determine emm types and capsule phenotype in 110 isolates of S. pyogenes from patients with invasive (sterile sites) and non-invasive (mainly pharyngitis) infections in Chile, and the relationship between both virulence factors. The most abundant types found were emm12, emm1, emm4 and emm28 and their distribution was similar to that seen in Latin America and developed countries, but very different from that in Asia and Pacific Island countries. Ten of 16 emm types identified in pharyngeal isolates were found in sterile-site isolates, and three of nine emm types of sterile-site isolates occurred in pharyngeal isolates; three emm subtypes were novel. The amount of hyaluronic acid was significantly higher in sterile-site isolates but did not differ substantially among emm types. Only three isolates were markedly capsulate and two of them had mutations in the csrR gene that codes for a repressor of capsule synthesis genes. We found a non-random association between emm types and csrR gene alleles suggesting that horizontal gene transfer is not freely occurring in the population.

Stigmoid bodies contain type I receptor proteins SorLA/LR11 and sortilin: new perspectives on their function.

Stigmoid bodies (SBs) are structures in the cytoplasm of neurons. SBs are mostly found in the hypothalamic region of the rat and contain a protein called huntingtin-associated protein 1 (HAP1). In a recent publication, large cytoplasmic structures were shown to be immunoreactive for a type I receptor called SorLA/LR11. By light microscopic analysis, these structures appeared similar to SBs in size and in brain regional and subcellular localization. To determine whether these large puncta correspond to HAP1-containing SBs, we used antibodies specific to various domains of the apolipoprotein receptor LR11 to perform immunocytochemistry in rat and mouse brain tissue. Transfection studies using HeLa cells were conducted to demonstrate the specificity of the antibodies. We found that, in both species, antibodies to the domain II (or VSP10 for vacuolar sorting protein 10 domain) of LR11 immunoreact with large cytoplasmic structures. Co-localization immunolabeling experiments in rat brain tissue sections and in neuron cultures showed that these LR11-immunoreactive structures correspond to HAP1-positive SBs. Electron microscopy was performed in rat hypothalamus and further demonstrated the presence of LR11 in SBs and its co-localization with HAP1. LR11-containing SBs were most abundant in the hypothalamus but were also found in many brainstem nuclei, thalamus, and hippocampus. Our data also show that sortilin, another transmembrane protein containing a VPS10 domain, localizes to large cytoplasmic puncta and is found in LR11-positive and Hap1-positive SBs in hypothalamic neuron cultures.

[AFP-producing adenocarcinoma of the stomach. A rare tumor with poor prognosis]. / Das AFP-positive Adenokarzinom des Magens. Ein seltener Tumor mit früher Disseminierung.

A 27-year-old, previously healthy man with abdominal discomfort was diagnosed with a small gastric tumor of the cardia by means of gastroscopy. Further staging revealed diffuse hepatic metastases and enlarged mediastinal lymph nodes. Serum alpha-fetoprotein (AFP) was grossly increased (7179 micro g/l). Biopsies taken from the gastric tumor and one of the hepatic metastases revealed a poorly differentiated adenocarcinoma (grade 3) with papillary and small solid areas and frequent clear cells. Cytoplasmic hyaline droplets were positive with PAS staining (diastase-resistant). Immunohistochemistry revealed focal tumor cells strongly positive for AFP and there was luminal expression of CEA. The diagnosis of an AFP-producing adenocarcinoma of the stomach was made. In spite of intensive combination chemotherapy the patient succumbed to his disease 3 months after diagnosis. The rare AFP-producing adenocarcinoma of the stomach is characterised by a distinct morphology and immunohistochemistry. A hepatoid differentiation may occur but is not obligatory as this case shows. In differential diagnosis, a metastasising germ cell tumor should be excluded. The prognosis for an AFP-positive adenocarcinoma is poor.

SorLA, a member of the LDL receptor family, is expressed in the collecting duct of the murine kidney.

SorLA/LR11 is a recently identified member of the LDL receptor superfamily, which is broadly expressed in the nervous system, but also in some non-neuronal tissues. To investigate sorLA expression in more detail we generated sorLA-specific antibodies. The kidney-specific expression of sorLA was investigated by in situ hybridization, immunohistochemistry, and immunofluorescence of murine kidney sections. In situ hybridizations of embryonic and adult kidney sections revealed transcripts mainly in the renal medulla. This expression pattern correlated with sorLA immunostainings. By co-staining with various markers sorLA expression was found to be restricted to cortical and medullary collecting tubules. SorLA was detected in intracellular vesicular structures, but not in the plasma membrane. Vesicles were distributed throughout the cytoplasm of cortical collecting tubules, whereas a more apical localization was noted in the medullary part. This intracellular compartment partially overlapped with the trans-Golgi marker gamma-adaptin and the transferrin receptor, a marker for shuttling compartments, but not with the lysosomal marker lamp2. These results demonstrate a highly specific expression of sorLA in collecting tubules. SorLA overlaps with intracellular shuttling compartments, indicating a role of sorLA for intracellular trafficking in the kidney.

Macrolide resistance phenotypes in Streptococcus pneumoniae in Santiago, Chile.

The mechanism of resistance was investigated in 39 macrolide-resistant clinical isolates of Streptococcus pneumoniae isolated from January 1997 to July 1999 in Santiago, Chile. Our results showed that 22 (56.5%) were macrolide-resistant, clindamycin-susceptible isolates (M phenotype) and 17 (43.5%) were macrolide and clindamycin resistant (MLS(B) phenotype). mefE gene was detected in all M phenotype, while ermB gene was detected in all MLS(B)-phenotype strains. Serotype 14 was the most frequent serotype among M-phenotype strains, and serotypes 19 and 23F were the most frequent serotypes in MLS(B) strains. These results demonstrate that both phenotypes of macrolide-resistant S. pneumoniae are found in Santiago, Chile, with the M phenotype predominating.

Characterization of the VPS10 domain of SorLA/LR11 as binding site for the neuropeptide HA.

The single transmembrane receptor sorLA/LR11 contains binding domains typical for the low-density lipoprotein receptors and a VPS10 domain which, in the related receptor sortilin, binds the neuropeptide neurotensin. SorLA is synthesized as a proreceptor which is processed to the mature form by a furin-like propeptidase. Endogenous sorLA and its hydra homologue HAB bind the neuropeptide head activator (HA). Transiently expressed partial sorLA constructs were investigated for ligand binding. We found that HA binds with nanomolar affinity to the VPS10 domain. The sorLA propeptide also bound to the VPS10 domain, whereas the receptor-associated protein RAP interacted both with the class A repeats and the VPS10 domain. The sorLA propeptide inhibited binding of HA to full-length sorLA and to the VPS10 domain. It also interfered with binding of HA to hydra HAB, which is taken as evidence for a highly conserved tertiary structure of the VPS10 domains of this receptor in hydra and mammals. The propeptide inhibited HA-stimulated mitosis and proliferation in the human neuroendocrine cell line BON and the neuronal precursor cell line NT2. We conclude that sorLA is necessary for HA signaling and function.

The neuropeptide head activator induces activation and translocation of the growth-factor-regulated Ca(2+)-permeable channel GRC.

The neuropeptide head activator stimulates cell proliferation of neuronal precursor and neuroendocrine cells. The mitogenic signaling cascade requires Ca(2+) influx for which, as we show in this paper, the growth-factor-regulated Ca(2+)-permeable cation channel, GRC, is responsible. GRC is a member of the transient receptor potential channel family. In uninduced cells only low amounts of GRC are present on the plasma membrane but, upon stimulation with head activator, GRC translocates from an intracellular compartment to the cell surface. Head activator functions as an inducer of GRC translocation in neuronal and neuroendocrine cells, which express GRC endogenously, and also in COS-7 cells after transfection with GRC. Head activator is no direct ligand for GRC, but its action requires the presence of a receptor coupled to a pertussis-toxin inhibitable G-protein. Heterologously expressed GRC becomes activated by head activator, which results in opening of the channel and Ca(2+) influx. SK&F 96365, an inhibitor specific for TRP-like channels, blocks Ca(2+) entry and, consequently, translocation of GRC is prevented. Head activator-induced GRC activation and translocation are also inhibited by wortmannin and KN-93, blockers of the phosphatidylinositol 3-kinase and of the Ca(2+)/calmodulin-dependent kinase, respectively, which implies a role for both kinases in head-activator signaling to GRC.

SorCS1, a member of the novel sorting receptor family, is localized in somata and dendrites of neurons throughout the murine brain.

The expression of the sorCS1 protein in the central nervous system of adult mice was studied by immunohistochemistry. A detailed mapping revealed a distribution of sorCS1 immunoreactivity in a widespread population of neurons throughout the brain. Two different types of cellular localization were observed. Many neurons exhibited a punctate cytoplasmic staining which extended into the dendrites, in other neurons sorCS1 immunoreactivity was associated with the plasma membrane. This suggests variable functions for sorCS1 in the neurons of the brain.

Brenner tumors but not transitional cell carcinomas of the ovary show urothelial differentiation: immunohistochemical staining of urothelial markers, including cytokeratins and uroplakins.

To determine whether Brenner tumors and transitional cell carcinomas (TCCs) of the ovary are urothelial in type, the immunoprofiles of 14 Brenner tumors, including three malignant examples, and eight ovarian TCCs were compared with those of Walthard nests, urothelium, 12 urinary bladder TCCs and 17 ovarian adenocarcinomas (serous, endometrioid, mucinous, and undifferentiated type). The immunohistochemical stains used included those for cytokeratins CKs 5/6, CK7, CK8, CK13, and CK20, vimentin, CA125, and the specific urothelial differentiation marker uroplakin III. CK7 and CK8 were broadly expressed in most tumors of ovary and bladder examined, while vimentin was focally present in some ovarian TCCs and adenocarcinomas. As in normal and neoplastic bladder urothelium, urothelial markers, including uroplakin III, CK13, and CK20, were detected in the epithelial nests of Brenner tumors. Brenner tumor cells also expressed uroplakins Ia and II. CA125 was observed focally in some Brenner tumors. In contrast, TCCs of the ovary and Walthard nests lacked uroplakins and were essentially negative for CK20 and CK13 but quite strongly expressed CA125. This immunophenotype closely resembled that found in ovarian adenocarcinomas. Thus, it appears that the only true urothelial-type ovarian neoplasm is the Brenner tumor, whereas ovarian TCC most likely represents a poorly differentiated adenocarcinoma with a morphologic transitional cell pattern. These results may explain the controversies as expressed in the recent literature concerning TCC of the ovary and establish its place among the ovarian adenocarcinomas of müllerian type.

Organization of uroplakin subunits: transmembrane topology, pair formation and plaque composition.

The apical surfaces of urothelial cells are almost entirely covered with plaques consisting of crystalline, hexagonal arrays of 16 nm uroplakin particles. Although all four uroplakins, when SDS-denatured, can be digested by chymotrypsin, most uroplakin domains in native urothelial plaques are resistant to the enzyme, suggesting a tightly packed structure. The only exception is the C-terminal, cytoplasmic tail of UPIII (UPIII) which is highly susceptible to proteolysis, suggesting a loose configuration. When uroplakins are solubilized with 2% octylglucoside and fractionated with ion exchangers, UPIa and UPII were bound as a complex by a cation exchanger, whereas UPIb and UPIII were bound by an anion exchanger. This result is consistent with the fact that UPIa and UPIb are cross-linked to UPII and UPIII, respectively, and suggests that the four uroplakins form two pairs consisting of UPIa/II and UPIb/III. Immunogold labelling using a new mouse monoclonal antibody, AU1, revealed that UPIII is present in all urothelial plaques, indicating that the two uroplakin pairs are not segregated into two different types of urothelial plaque and that all plaques must have a similar uroplakin composition. Taken together, these results indicate that uroplakins form a tightly packed structure, that the four uroplakins interact specifically forming two pairs, and that both uroplakin pairs are required for normal urothelial plaque formation.

Identification of SorCS2, a novel member of the VPS10 domain containing receptor family, prominently expressed in the developing mouse brain.

We report the identification of a fourth member of the VPS10 domain containing receptor family, SorCS2, highly expressed in the developing and mature murine central nervous system. During early central nervous system development its main site of expression is the floor plate. In addition, high transcript levels were detected transiently in a variety of brain regions including the dopaminergic midbrain nuclei and the dorsal thalamus. Outside the nervous system expression is detected in lung and heart and transiently in a variety of mesodermally derived tissues.

Prevalence and mechanisms of macrolide resistance in Streptococcus pyogenes in Santiago, Chile.

Thirty-two macrolide-resistant Streptococcus pyogenes isolates were found among 594 clinical isolates collected from 1990 to 1998 in Santiago, Chile, for an overall prevalence of 7.2%. Among the 32 resistant isolates, 28 (87.5%) presented the M phenotype and 4 (12. 5%) presented the MLS(B) phenotype. Serotyping and pulsed-field gel electrophoresis analysis showed genetic diversity among the resistant isolates.

Ectodomain shedding, translocation and synthesis of SorLA are stimulated by its ligand head activator.

The single transmembrane receptor SorLA is the mammalian orthologue of the head activator-binding protein, HAB, from hydra. The human neuronal precursor cell line NT2 and the neuroendocrine cell line BON produce head activator (HA) and respond to HA by entry into mitosis and cell proliferation. They express SorLA, and bind HA with nanomolar affinity. HA coupled to Sepharose is able to precipitate SorLA specifically proving that SorLA binds HA. Using antisera directed against extra- and intracellular epitopes we find SorLA as membrane receptor and as soluble protein released from cells into the culture medium. Cell lines differ strongly in processing of SorLA, with NT2 cells expressing SorLA mainly as membrane receptor, whereas release predominates in BON cells. Soluble SorLA lacks the intracellular domain and is shed from the transmembrane protein by a metalloprotease. Release from cells and brain slices is stimulated by HA and by phorbol ester, and it is blocked by a metalloprotease inhibitor and by lowering the temperature to 20 degrees C. Blockade of SorLA shedding and treatment of cells with SorLA antisense oligonucleotides lead to a decrease in the rate of cell proliferation. From this we conclude that SorLA is necessary to mediate the mitogenic effect of endogenous HA. HA enhances the translocation of SorLA from internal membranes to the cell surface and its internalization. In addition, HA stimulates SorLA synthesis hinting at an autocatalytic feedback loop in which the ligand activates production, processing, and translocation of its receptor.

Identification and characterization of SorCS, a third member of a novel receptor family.

A novel receptor, SorCS, was isolated from murine brain. It shows homology to the mosaic receptor SorLA and the neurotensin receptor sortilin based on a common VPS10 domain which is the hallmark of this new receptor family. In the N-terminus of SorCS two putative cleavage sites for the convertase furin mark the beginning of the VPS10 domain, followed by a module of imperfect leucine-rich repeats and a transmembrane domain. The short intracellular C-terminus contains consensus signals for rapid internalization. The identified putative binding motifs for SH2 and SH3 domains are unique in the family of VPS10 domain receptors. SorCS is predominantly expressed in brain, but also in heart, liver, and kidney. SorCS transcripts detected by in situ hybridization in the murine central nervous system point to a neuronal expression.

Anticardiolipin antibodies in acute rheumatic fever.

Recent reports describe the association of antiphospholipid antibodies (aPL) with chorea or severe heart valve lesions in systemic lupus erythematosus, lupus-like disease, or the primary antiphospholipid antibody syndrome. We conducted a case series and a case-control investigation of patients with rheumatic fever with Sydenham chorea or other manifestations of rheumatic fever for anticardiolipin antibodies (aCL) during the acute attack and disease remission. Eighty percent of patients were positive for aCL during the rheumatic fever attack vs 40% when inactive (p = 0.035); IgG and IgM aCL increased significantly with disease activity. Individuals with or without Sydenham chorea were equally positive for aCL (76 and 83%, respectively). A significant association was found between IgM aCL and carditis: All patients with valvulitis had IgM aCL (100%) vs 37% of patients without valvular involvement (p = 0.02). aPL may play a role in the pathogenesis of some clinical manifestations of acute rheumatic fever.

[Uncomplicated acute male and female gonorrhea: treatment with a single oral dose of epicillin-probenecid]. / Gonorrea aguda masculina y femenina no complicada: tratamiento con dosis única oral de epicilina-probenecid.

We treated 142 patients with acute uncomplicated gonorrhea with a single oral dose of epiciline-probenecid. All cases were confirmed by culture on Thayer Martin medium at the first day and they were all laboratory tested seven days after treatment. At the control time the patients were asked about drug tolerance. All isolated strains were tested for beta-lactamase producing capacity and susceptibility to epiciline and penicillin. All patients who were culture positive seven days after treatment were considered to be treatment failures. The treatment was effective in 94.4% and we detected 8 Neisseria gonorrhoeae strains resistant to penicillin (5.6%); seven of these were PPNG strains. Tolerance to the drug was excellent in 90% of all cases; 9% presented minor adverse reactions for a short period of time. Therefore, the combination of epiciline-probenecid is a valid alternative for routine treatment of acute uncomplicated gonorrhea.

Glomerulonefritis aguda post-estreptococica: descripcion epidemiologica. / Acute post-streptococcal glomerulonephritis: epidemiological report

Se presentan las caracteristicas epidemiologicas, bacteriologicas y serologicas de 84 pacientes de glomerulonefritis aguda ocurridos en el Servicio de Salud Metropolitano Sur Oriente, entre Enero de 1980 y Diciembre de 1981 con especial enfasis en la localizacion de la infeccion estreptococica que los origino y su vinculacion con dichas caracteristicas. Existen claras diferencias en cuanto a edad y estacion del ano de las GNA segun derivan de una infeccion cutanea o faringea, asi como en la tasa de aislamiento del estreptococo B hemolitico A y sus serotipos. Igualmente la respuesta inmunitaria medida con anticuerpos antiestreptococicos es diferente segun la localizacion y prueba usada. Lo anotado confirma lo hallado por autores extranjeros en el sentido de que la localizacion determina caracteristicas epidemiologicas diferentes. Se estudiaron ademas, los contactos intrafamiliares de los casos y en ellos se observo una actividad estreptococica significativamente superior a la de la poblacion general, medida por aislamientos y titulos de anticuerpos antiestreptococicos