Immunization with recombinant Helicobacter pylori urease in specific-pathogen-free rhesus monkeys (Macaca mulatta)

Immunization with urease can protect mice from challenge with Helicobacter pylori, though results vary depending on the particular vaccine, challenge strain, and method of evaluation. Unlike mice, rhesus monkeys are naturally colonized with H. pylori and so may provide a better estimate of vaccine efficacy in humans. The purpose of this study was to examine the effectiveness of H. pylori urease as a vaccine in specific-pathogen (H. pylori)-free rhesus monkeys. Monkeys raised from birth and documented to be free of H. pylori were vaccinated with orogastric (n = 4) or intramuscular (n = 5) urease. Two control monkeys were sham vaccinated. All monkeys were challenged with a rhesus monkey-derived strain of H. pylori, and the effects of vaccination were evaluated by use of quantitative cultures of gastric tissue, histology, and measurement of serum immunoglobulin G (IgG) and salivary IgA. Despite a humoral immune response, all monkeys were infected after H. pylori challenge, and there were no differences in the density of colonization. Immunization with urease therefore does not fully protect against challenge with H. pylori. An effective vaccine to prevent H. pylori infection will require different or more likely additional antigens, as well as improvements in the stimulation of the host immune response.     

Cryptococcus neoformans var. gattii in the koala (Phascolarctos cinereus): serological evidence for subclinical cryptococcosis.

Cryptococcus neoformans var. gattii has been shown to have a strong association with eucalypts frequently used by koalas and, not surprisingly, it has been shown to colonize the nasal cavities of koalas. The progression from nasal colonization to tissue invasion is critical to understanding the pathogenesis of cryptococcosis in this species and provides a model for pathogenesis of cryptococcosis in other species. Cryptococcal antigenaemia was detected in twenty-eight healthy koalas from three different regions. This was interpreted as representing limited subclinical disease. One koala developed cryptococcal pneumonia 6 months after leaving the study, whereas another developed cryptococcal meningoencephalitis during the course of the study. Opportunistic necropsies on ten antigen-positive koalas resulted in discovery of small cryptococcal lesions in two (paranasal sinus and lung, respectively). Our data suggest that cryptococcal antigenaemia occurs commonly in koalas, especially in areas with a high environmental presence of C n. var. gattii. Subclinical disease appears most likely to manifest as a small focal lesion in the respiratory tract. Possible outcomes include elimination by an effective immune response, quiescence with possibility of later re-activation or direct progression to overt disease. Symptomatic and subclinical cases showed differences in levels of antigenaemia. The data presented have significant implications for koalas in captivity.     

The involvement of matrix glycoproteins in vascular calcification and fibrosis: an immunohistochemical study.

Calcification and fibrointimal proliferation are associated with advanced complicated atherosclerosis in large arteries but may also occur in smaller vessels, resulting in ischaemic tissue necrosis. This study investigates whether the mechanisms of calcification and intimal fibrosis are similar in vessels of different sizes. The localization of osteopontin (OPN), matrix Gla protein (MGP), thrombospondin-1 (TSP-1), and cartilage oligomeric matrix protein (COMP) was investigated in three types of human vascular lesions: atherosclerosis, chronic vascular rejection (CVR) in renal allografts, and calcific uraemic arteriolopathy (calciphylaxis). These lesions were chosen as they affect different sized blood vessels and they exhibit a fibroproliferative intimal reaction, with or without calcification, resulting in luminal obliteration and ischaemic complications. OPN, MGP, TSP-1, and COMP were not detected in normal blood vessels. However, OPN and MGP were expressed at sites of calcification within atherosclerotic lesions and in microvessels in calciphylaxis, suggesting that calcification in different sized vessels may occur by a common mechanism. These proteins were not detected in areas of fibrointimal proliferation. In contrast, TSP-1 was localized primarily within the fibrous tissue of atherosclerotic lesions and was also expressed in the expanded fibrous intima of arteries showing CVR. COMP was localized primarily within the fibrous tissue under the lipid core of the majority of advanced atherosclerotic lesions. TSP-1 and COMP were also detected in areas of microcalcification in atherosclerotic lesions and TSP-1 was detected adjacent to areas of calcification in calciphylaxis. However, neither TSP-1 nor COMP was localized to calcific foci within these lesions. The localization of OPN, MGP, TSP-1, and COMP to pathological, but not normal arterial intima supports a pathogenetic role for these proteins in the development of vascular fibrosis and calcification. Modulation of their production and activity may offer a novel approach to the therapy of a number of vascular diseases.     

Identification and characterization of aquaporin-2 water channel mutations causing nephrogenic diabetes insipidus with partial vasopressin response

Congenital nephrogenic diabetes insipidus (NDI) is a rare disease caused most often by mutations in the vasopressin V2 receptor (AVPR2). We studied a family which included a female patient with NDI with symptoms dating from infancy. The patient responded to large doses of desmopressin (dDAVP) which decreased urine volume from 10 to 4 I/day. Neither the parents nor the three sisters were polyuric. The patient was found to be a compound heterozygote for two novel recessive point mutations in the aquaporin-2 (AQP2) gene: L22V in exon 1 and C181W in exon 3. Residue Cys181 in AQP2 is the site for inhibition of water permeation by mercurial compounds and is located near to the NPA motif conserved in all aquaporins. Osmotic water permeability (Pf) in Xenopus oocytes injected with cRNA encoding C181W-AQP2 was not increased over water control, while expression of L22V cRNA increased the Pf to approximately 60% of that for wild-type AQP2. Co-injection of the mutant cRNAs with the wild-type cRNA did not affect the function of the wild-type AQP2. Immunolocalization of AQP2-transfected CHO cells showed that the C181W mutant had an endoplasmic reticulum-like intracellular distribution, whereas L22V and wild-type AQP2 showed endosome and plasma membrane staining. Water permeability assays showed a high Pf in cells expressing wild-type and L22V AQP2. This study indicates that AQP2 mutations can confer partially responsive NDI.      

Characterization of lymphokine fibronectin from guinea pig lymphoid cell culture supernatants.

Fibronectins (FN) in guinea pig lymphoid cell culture supernatants have been studied using a panel of polyclonal and monoclonal anti-FN antibodies to clarify their relationship with macrophage agglutination factor (MAggF), an inflammatory lymphokine sharing many properties with this family of high molecular weight glycoproteins. MAggF contained cellular FN epitopes, and was reversibly bound by antibodies specific for cellular FN. Enzyme-linked immunoassay and inhibition of MAggF activity by monoclonal anti-plasma FN antibodies revealed immunoreactive FN in guinea pig lymphoid cell culture supernatants to share three epitopes with plasma FN and to lack a fourth epitope present in plasma FN. Immunoreactive FN in gelatin-affinity purified lymph node cell culture supernatants was polydisperse; MAggF activity (Mr 410 kD) was associated with only 13% of total immunoreactive FN. Although a low molecular weight FN fragment (Mr 67 kD) was associated with MAggF activity in salt-fractionated peritoneal exudate culture supernatants, it was not possible to generate MAggF activity by limited proteolysis of MAggF-inactive, high molecular weight FN in lymph node cell culture supernatants. We conclude that MAggF is a cellular FN containing a number of epitopes in common with plasma FN and suggest it may be a unique species of cellular FN produced by T lymphocytes involved in initiating delayed hypersensitivity reactions.     

Key Steps in Conducting Systematic Reviews for Underpinning Clinical Practice Guidelines: Methodology of the European Association of Urology

Context

The findings of systematic reviews (SRs) and meta-analyses (MAs) are used for clinical decision making. The European Association of Urology has committed increasing resources into the development of high quality clinical guidelines based on such SRs and MAs.

Detection of desialylated forms of human chorionic gonadotropin

Urinary forms of human chorionic gonadotropin (hCG) with oligosaccharides deficient in sialic acid content (ashCG) have been reported to be excreted by patients with choriocarcinoma in greater amounts than by healthy, pregnant women. Although ashCG potentially could be a useful marker for the diagnosis and management of gestational trophoblastic neoplasia, the methods previously used for its detection were not suitable for routine clinical application. Therefore, we have devised a simpler method which can provide specific and sensitive measurements of ashCG in urine. This method, which is designated as a lectin-immunoradiometric assay (LIRMA), employs an agarose-coupled lectin to selectively extract the ashCG, which is then quantified directly with a purified and radiolabelled rabbit antibody. The LIRMA has been applied to demonstrate that there is an increased excretion of ashCG by choriocarcinoma patients. It is also applicable, in principle, for the study of any glycoprotein which has a reduced content of sialic acid in its carbohydrate side chains.     

Benzocaine-associated methemoglobinemia following bronchoscopy in a healthy research participant

Benzocaine (ethyl aminobenzoate) is a local anesthetic commonly used to achieve topical anesthesia of the skin and mucous membranes prior to endoscopic procedures. Methemoglobinemia, a condition in which hemoglobin cannot bind and deliver oxygen normally, has been associated with benzocaine use in various patient populations. This is the first report of benzocaine-associated methemoglobinemia occurring in a healthy research participant. The research participant developed a methemoglobin level of 27% and marked cyanosis. No adverse sequelae other than cyanosis were identified. This report extends the population in which benzocaine-associated methemoglobinemia has been described. Additionally, this report supports the observation that methemoglobin levels approaching 30% may be tolerated in otherwise healthy individuals, producing few clinically important effects. Finally, this case also indicates that, in obtaining informed consent for a procedure in which benzocaine will be administered, patients and research participants should be specifically informed of the risk of benzocaine-induced methemoglobinemia. This information is especially important in those settings in which the manufacturer-recommended dose of benzocaine may either intentionally or inadvertently be exceeded.     

Gross, histological and immunohistochemical features of mucormycosis in the platypus

Nine male and five female adult free-living platypuses, obtained in a prospective capture-release study from northern Tasmania, exhibited gross features of cutaneous mycosis caused by Mucor amphibiorum. The lesions were present on the hind limbs (six cases), front limbs (four), tail (five), dorsal trunk (three) and ventral trunk (one). They varied in size, and ranged from raised red nodules or plaques, which sometimes exuded purulent material, to ulcerated lesions with central cavitation, red exuding centres and raised epidermal margins. Older lesions were covered either partly or fully by thickened and irregular epidermis. Histological examination of skin biopsies revealed discrete, poorly encapsulated granulomas, or more commonly a diffuse granulomatous or pyogranulomatous inflammation. Inflammatory cells consisted of neutrophils or eosinophils, sparse plasma cells and lymphocytes, many macrophages and occasional multinucleated giant cells. Fibrovascular tissue was diffusely and irregularly scattered in the granulomatous regions. Sphaerules characteristic of M. amphibiorum infection were observed in all lesions. The cutaneous distribution of the lesions and the natural history of the platypus indicated that entry of M. amphibiorum may have been via superficial skin wounds. T cells were the predominant infiltrating lymphoid cells in the diffuse lesions, indicating the importance of the cell-mediated response to infection.