Effects of Lymphocyte Isolation and Timing of Processing on Detection of CD127 Expression on T Cells in Human Immunodeficiency Virus-Infected Patients

Decreases in the detection of CD127 expression on T cells of human immunodeficiency virus-infected patients by flow cytometry can occur by delayed processing or by peripheral blood mononuclear cell isolation and cryopreservation. These observations should be considered in the interpretation of functional studies and the planning of multicenter clinical trials.     

Viral protein production in homogeneous and mixed infections of cytopathic and noncytopathic BVD virus

Summary Experiments were conducted to examine dual infection of cultured cells with cytopathic and noncytopathic bovine viral diarrhea virus (BVDV). Cell monolayers infected with a noncytopathic BVDV isolate and subsequently superinfected with a cytopathic BVDV isolate were refractive to the cytopathic effects of the cytopathic BVDV isolate, as reported in the literature. Immunofluorescence staining of superinfected cultures with monoclonal antibodies specific for the cytopathic or the noncytopathic viral isolate, demonstrated that cells in superinfected cultures contained both viral biotypes. Immunoprecipitation was used to compare the temporal detection of viral induced polypeptides in superinfected cultures to that of cultures infected with a single viral biotype. In single cytopathic viral infections, viral induced polypeptides of 80 kDa and 53–56 kDa are detected simultaneously, but in superinfections a 4 h gap occurred between detection of the 53–56 kDa polypeptide and detection of the 80 kDa polypeptide.     

Adenoviral vectors expressing siRNAs for discovery and validation of gene function

RNA interference is a powerful tool for studying gene function and for drug target discovery in diverse organisms and cell types. In mammalian systems, small interfering RNAs (siRNAs), or DNA plasmids expressing these siRNAs, have been used to down-modulate gene expression. However, inefficient transfection protocols, in particular, for primary cell types, have hampered the use of these tools in disease-relevant cellular assays. To be able to use this technology for genome-wide function screening, a more robust transduction protocol, resulting in a longer duration of the knock-down effect, is required. Here, we describe the validation of adenoviral vectors that express hairpin RNAs that are further processed to siRNAs. Infection of cell lines, or primary human cells, with these viruses leads to an efficient, sequence-specific, and prolonged reduction of the corresponding target mRNA, resulting in a reduction of the encoded protein level in the cell. For knock-down of one of the targets, GalphaS, we have measured inhibition of ligand-dependent, G-protein-coupled signaling. It is expected that this technology will prove to be of great value in target validation and target discovery efforts.     

Effects of enzyme replacement therapy on pain and health related quality of life in patients with Fabry disease: data from FOS (Fabry Outcome Survey)

Background: Fabry disease is an X linked lysosomal storage disease caused by deficiency of the lysosomal enzyme α-galactosidase A. This leads to accumulation of globotriaosylceramide in nearly all tissues, including the blood vessels, kidney, myocardium, and nervous system. Symptoms often begin in childhood and include acroparaesthesia, with burning or tingling pain that spreads from the extremities to more proximal sites.

Aims: This study set out to evaluate pain and its influence on quality of life in patients with Fabry disease receiving enzyme replacement therapy (ERT) with agalsidase alfa.

Methods: Data were obtained from the Fabry Outcome Survey. Pain was measured using the Brief Pain Inventory (BPI), and health-related quality of life (HRQoL) was documented with the European Quality of Life Questionnaire (EQ-5D).

Results: The mean (SD) score for "pain at its worst" on the BPI prior to ERT was 5.1 (2.7). One year after commencement of ERT, this had improved by 0.5, and improved by a further 0.6 after 2 years (p<0.05). Similar statistically significant improvements were seen for "pain on average" and "pain now" after 2 years of ERT. The mean HRQoL utility score prior to ERT was 0.66 (0.32). After 12 months of treatment with agalsidase alfa, this had improved to 0.74 (0.26; p<0.05); this improvement was maintained after 2 years.

Conclusions: ERT with agalsidase alfa significantly reduces pain and improves quality of life in patients with Fabry disease.

     

The Mikulicz Cell in Rhinoscleroma: Light, Fluorescent and Electron Microscopic Studies

The stages in the development of the Mikulicz cell in human rhinoscleroma were studied in biopsy specimens obtained from 10 patients using light, immunofluorescent and electron microscopy. The Mikulicz cell was identified morphologically as a macrophage, not a plasma cell. Acutely inflamed areas of rhinoscleroma presented abundant bacteria with a slime layer. The microorganism was infrequent and the mucopolysaccharide was scanty in rhinoscleromal tissue, where plasma cells predominated, and in cicatricial fibrous tissue. In the granulomatous stage of rhinoscleroma, the mucopolysaccharide was found within the Mikulicz cells. The vacuoles observed in the Mikulicz cells were considered to be phagosomes containing, principally, bacterial mucopolysaccharide and few bacteria and, to a lesser extent, swollen mitochondria. It was concluded that the slime layer of Klebsiella rhinoscleromatis plays an important role in the pathogenesis of the disease. It is postulated that this material is a nondigestible mucopolysaccharide that resides in the phagosomes of macrophages, increases the osmotic pressure and forms multiple hydropic vacuoles that rupture not only the phagosomes but also the cells, resulting in the liberation of the mucopolysaccharide. This would initiate a cycle that would prolong the disease in the absence of the bacteria.     

A comparison of research general practices and their patients with other practices--a cross-sectional survey in Trent.

BACKGROUND: When interpreting results of studies undertaken by research networks we need to know how representative volunteer practices and their registered patients are of the total population of practices and patients in their locality. AIM: To compare the following in research and non-research general practices in one region: practice and population demography, morbidity and mortality, selected performance indicators, and health outcomes. DESIGN OF STUDY: Cross-sectional survey. SETTING: Sixty-six Trent Focus Collaborative Research Network general practices and 749 other general practices in Trent, United Kingdom. METHOD: Practice characteristics and GP contract data were obtained from the NHS Executive, Quarry House, Leeds. The Trent Regional NHS Hospital Admission Database was searched to identify all relevant admissions to hospital from all practices between 1 April 1993 and 31 March 1997. Ward-linked data on cancer were obtained from the Trent Cancer Registry. RESULTS: Of the 815 general practices in Trent Region in the study period, 66 (8%) were in the Trent Focus network. They were more likely to be involved in training GPs and to have a female partner. They tended to be larger, with fewer single-handed doctors and younger GPs. Network practices prescribed a higher proportion of generics (median % prescribed/practice = 70%, versus 51%, Mann-Whitney U = 1615, P<0.0001). There were no clinically important differences between hospital admission rates between the two groups or waiting times for surgical procedures. There was no difference in the incidence of cancer and standardised mortality ratios related to the electoral wards of the GP surgery. CONCLUSION: Although there were differences in practice structure and some aspects of performance, we found no important differences in the demography of registered patients, nor in morbidity, mortality, or access to or use of secondary care.     

Retinal projection to the nucleus of the optic tract in the cat as revealed by retrograde transport of horseradish peroxidase

Retrograde transport of horseradish peroxidase injected iontophoretically into the nucleus of the optic tract of cats revealed that the direction-selective cells in this pretectal nucleus receive direct retinal projections from small retinal ganglion cells, the so-called gamma-cells. These cells from a horizontal band on the contralateral retina. Few labeled cells are found in the ipsilateral temporal retina. The input from the contralateral retina is 10 times more numerous than from the ipsilateral one. In both retinae the highest concentration of labeled cells is near the area centralis.