Inappropriate use of proton pump inhibitors.

BACKGROUND: Proton pump inhibitors (PPIs) are widely used expensive medications. AIMS: We performed a cross-sectional study to determine the extent and indication of PPI use in Irish acute medical wards. METHODS: Fifty-five medical charts were reviewed at the beginning and end of 1 month. RESULTS AND CONCLUSIONS: Thirty-three patients were prescribed PPIs; 26 prior to admission. The prescribing of PPIs was concordant with guideline recommendations in only 30% of cases. Two-thirds of PPI use was unlicensed.

Methadone induced torsade de pointes in a patient receiving antiretroviral therapy.

Adverse drug reactions account for approximately 5% of acute medical admissions. A 34-year-old male patient receiving antiretroviral therapy, methadone and flurazepam presented to the emergency room following collapse with associated loss of consciousness. Cardiac monitoring demonstrated marked Q-T prolongation followed by the cardiac arrhythmia, torsade de pointes. The patient made a full recovery following withdrawal of the antiretroviral therapy and a reduction in methadone dose. Methadone is a recognised cause of this potentially fatal cardiac arrhythmia which is more likely to occur when methadone metabolism is inhibited by drugs such as HIV tease inhibitors.

MMR vaccine uptake rates: a data validation study.

As part of our investigation into the decrease in the measles, mumps and rubella (MMR) vaccine uptake rates, we validated MMR vaccination records of all children born between 01/09/1998 and 31/08/1999 in our area (North Cheshire, South Cheshire, and Wirral). A significant number of children had received their MMR vaccine but were not recorded as such by the Child Health Computer System (CHCS). Reported COVER (cover of vaccination evaluated rapidly) data uptake (combined) for North Cheshire, South Cheshire, and Wirral Health Authorities for the period covered by the data validation study was 90.5%, the corrected uptake following the validation was 92.6%, 2.1% higher than the reported coverage. If the coverage data were to continue to form part of the NHS indicators of PCT performance, action by all PCTs to improve accuracy of immunisation data would be highly desirable. Electronic transfer of information from practices to the CHCS and between CHCSs, i.e. across boundaries, could improve data accuracy.

Decreased bone mineral density is common after autologous blood or marrow transplantation.

Survivors of autologous blood or marrow transplantation (ABMT) are predisposed to decreased bone mineral density (BMD), but data are lacking on the incidence and risk factors for this condition. Therefore, we measured BMD in 64 of 68 consecutive ABMT survivors (35 men and 29 women) attending the University of Toronto ABMT long-term follow-up clinic. Patients were evaluated a median of 4.2 years (range: 4.9 months-11.4 years) after ABMT. Median age at evaluation was 49.6 years (range: 23.5-68.2 years). At the L1-L4 vertebrae, 17 (26%) patients (eight men and nine women) had osteopenia and one male (2%) had osteoporosis. Mean BMD at L1-L4 did not differ from healthy young adults or age and sex matched controls. At the femoral neck, 30 patients (46%) (18 men and 12 women) had osteopenia and five (8%) (two men and three women) had osteoporosis. Mean BMD at the femoral neck was significantly lower than in healthy young adults and age- and sex-matched controls. By regression analysis, patients with decreased BMD were older than those with normal BMD (P = 0.02). Gender, body mass index, time from BMT to evaluation and presence of hypogonadism were not associated with decreased BMD. Treatment of decreased bone density was instituted and follow-up data were obtained 1 year after treatment in 22 of 39 patients with reduced BMD. Nineteen (86%) patients had stabilization or improvement of their bone density at follow-up. We conclude that, after ABMT, over half of the patients have evidence of osteopenia or osteoporosis. Men and women were equally affected. In our study, only older age at evaluation was predictive for loss of BMD. We recommend the measurement of BMD as an integral component to the follow-up of ABMT patients.

An exploration of the effects of weighted garments on balance and gait of stroke patients with residual disability.

OBJECTIVE: To explore the effects of weighted garments on the balance and gait of stroke patients. DESIGN: A pilot randomized controlled study with blinded measurement. SETTING: Weighted garments were worn by patients living in the community and measurement was made in a hospital-based gait laboratory. SUBJECTS: Twenty-four adults who were at least six months post stroke and were able to walk 10 metres with or without assistance or a walking aid. INTERVENTION: The six-week treatment-phase subjects were given a set of weighted garments which they were shown how to apply and instructed to wear on their paretic side. Subjects randomly allocated to the six-week control phase were not given any weighted garments. MAIN OUTCOME MEASURES: Balance was measured with the Berg Balance Scale. Gait was measured using GaitMat II, an instrumented walkway. Gait parameters of interest were velocity and symmetry of: step length; single support time; double support time; and support base width. Measures were made at baseline before randomization (baseline) and at the end of the six weeks of intervention (outcome). RESULTS: No statistically significant differences were found between the treatment and control groups at outcome for balance (Mann-Whitney U-test; p = 0.74), gait velocity (p = 0.68) or symmetry of gait parameters (p = 0.33 to p = 0.75). CONCLUSIONS: We found no evidence to support the clinical use of these weighted garments for stroke survivors.

Excision of nasopharyngeal angiofibroma facilitated by intra-operative 3D-image guidance.

The latest 3D-image guidance systems to assist surgeons have greatly improved over earlier models. We describe the use of an optical infra-red system to assist in the removal of a juvenile nasopharyngeal angiofibroma. The specific advantages of this system in pre-operative assessment, intra-operative evaluation and excision of the angiofibroma are discussed.

Variation in proteinase activities in the rumen.

Rumen fluid was taken from fistulated sheep and a cow receiving various diets based on grass hay or grass cubes with and without cereal-based concentrates. Proteinases in the extracellular fluid, and extracted from particulate material by Triton X-100, were visualized using SDS-PAGE in which gelatin was co-polymerized with the gels. Each animal sampled had a different pattern of proteinase activity. No single proteinase band predominated, although a few appeared in several samples, indicating that some microbial species were commonly involved in proteolysis but none was dominant. The banding patterns in samples taken from the same animals two weeks apart were fairly similar, indicating some stability within animals. Patterns obtained with extracellular fluid and Triton X100 extracts of small and large particulate material from the same sample were similar for the most part, although the relative intensity of the bands differed. The serine protease inhibitor, phenyl methyl sulphonyl fluoride, had little influence on the banding pattern. Concentrate in the diet appeared to increase inter-animal variation, and sheep in adjacent pens and consuming the same grass hay:concentrate diet had different banding patterns. Thus, no microbial proteolytic enzyme was predominant in protein digestion in the rumen.

Altered sensitivity of posttranslationally modified microtubules to methylmercury in differentiating embryonal carcinoma-derived neurons.

The effects of methylmercury (MeHg) on microtubules (MTs) in differentiating neurons derived from retinoic acid-induced embryonal carcinoma (EC) cells in culture were examined by immunofluorescence microscopy. Undifferentiated EC cells contained mostly kinetically labile tyrosinated (TYR) MTs which extended from the centrosome and a small population of stable acetylated (ACT) MTs usually (but not exclusively) associated with the centrosome. TYR MTs of undifferentiated cells underwent concentration- and time-dependent disassembly upon exposure to low concentrations of MeHg (1-2 microM), whereas ACT MTs were resistant to MeHg at low concentrations, with many remaining intact even in 5 microM MeHg. After 2 days in retinoic acid the appearance of short neuritic processes was indicative of early differentiation. TYR MTs predominated both in the short neurites and cell soma and remained susceptible to low concentrations of MeHg. ACT MTs also extended into the short neurites but were most often found in small bundles within the perikarya. ACT MTs remained more resistant to MeHg than TYR MTs. The neuron-specific tubulin isotype betaIII was first detected during the second day of differentiation. MTs labeled with antibodies to betaIII showed similar sensitivity to MeHg as TYR MTs, suggesting that MTs containing betaIII were largely tyrosinated. After 4 and 6 days of differentiation a greater number of betaIII-positive neurons were present with progressively longer and often branching neurites. TYR MTs were present in cell soma and neurites while ACT MTs were found almost exclusively in neurites. TYR MTs remained highly susceptible to MeHg (most notably in perikarya) in comparison to ACT MTs at all stages of differentiation; however, with increasing time in culture, even TYR MTs gained appreciable stability to MeHg. These data indicate that microtubules in developing neurons become progressively more resistant to disassembly by MeHg, suggesting that the most critical period of MT susceptibility occurs very early in development in vivo.

The effect of aglycosylation on the immunogenicity of a humanized therapeutic CD3 monoclonal antibody.

The factors affecting the immunogenicity of a humanized gamma 1 CD3 monoclonal antibody (mAb) were investigated in transgenic mice that express the human CD3 antigen epsilon polypeptide (the mAb target antigen). Two derivatives of the mAb were employed, one with a normal, glycosylated Fc region (gamma 1 CD3 mAb), and the other with an aglycosylated Fc region (aglycosyl gamma 1 CD3 mAb). Comparisons of the antiglobulin responses elicited by the two derivatives in transgenic and nontransgenic mice demonstrated that Fab-mediated cell binding activity, dependent on target antigen expression, was a major positive determinant of CD3 mAb immunogenicity. A second positive factor was mAb Fc region glycosylation. At low dose levels the gamma 1 CD3 mAb consistently produced a higher antiglobulin response than the aglycosyl gamma 1 CD3 mAb. This was probably a result of the nonspecific, in vivo T cell activating property of the gamma 1 CD3 mAb, a consequence of its ability to cross-link T cells to Fc gamma receptor-bearing cells. (The aglycosyl gamma 1 CD3 mAb has a reduced Fc binding affinity for Fc gamma receptors and so does not activate T cells in vivo.) In support of this hypothesis, the gamma 1 CD3 mAb was able to nonspecifically enhance humoral immunity to an unrelated, coadministered antigen, whereas the aglycosyl gamma 1 CD3 mAb was not. The lower immunogenicity of the aglycosyl gamma 1 CD3 mAb correlated with a longer in vivo half-life and an improved capacity to block the target CD3 antigen. These results suggest that, as well as reducing the cytokine-induced side effects normally associated with CD3 mAb therapy, the nonactivating aglycosyl gamma 1 CD3 mAb will be less likely than the activating gamma 1 CD3 mAb to stimulate a neutralizing antiglobulin response.

Rotavirus-induced fusion from without in tissue culture cells.

We present the first evidence of fusion from without induced in tissue culture cells by a nonenveloped virus. Electron micrographs of two strains of rotavirus, bovine rotavirus C486 and rhesus rotavirus, show that virally mediated cell-cell fusion occurs within 1 h postinfection. Trypsin activation is necessary for rotavirus to mediate cell-cell fusion. The extent of fusion is relative to the amount of virus used, and maximum fusion occurs between pHs 6.5 and 7.5. Fusion does not require virus-induced protein synthesis, as virus from both an empty capsid preparation and from an EDTA-treated preparation, which is noninfectious, can induce fusion. Incubation of rotavirus with neutralizing and nonneutralizing monoclonal antibodies before addition to cells indicates that viral protein 4 (VP4; in the form of VP5* and VP8*) and VP7 are involved in fusion. Light and electron micrographs document this fusion, including the formation of pores or channels between adjacent fused cells. These data support direct membrane penetration as a possible route of infection. Moreover, the assay should be useful in determining the mechanisms of cell entry by rotavirus.

The molecular basis of microtubule stability in neurons.

Microtubule (Mt) populations show large differences in dynamic properties (i.e., turnover rates) among cell types, and even within the same cell type at different stages of the cell cycle or stages of differentiation. These differences in dynamic properties are correlated with altered sensitivity to Mt-disassembling drugs (e.g. colchicine) which bind specifically to the Mt protein tubulin and to certain toxic metals which also interact with tubulin (e.g. methylmercury) and result in Mt disassembly. Mts in neurons become progressively more stable and more resistant to such compounds during differentiation. We are using the P19 embryonal carcinoma cell line, which can be induced to differentiate along the neural pathway by retinoic acid, as a model system in which to analyze the development of stable Mts. Our results show that during differentiation there is an evolution in the sorting of tubulin isotypes into the stable Mts. This appears related both to the expression of specific Mt-associated proteins and to concomitant posttranslational modifications of tubulin.

Differential sorting of beta tubulin isotypes into colchicine-stable microtubules during neuronal and muscle differentiation of embryonal carcinoma cells.

Pluripotent P19 embryonal carcinoma (EC) cells were differentiated along the neuronal and muscle pathways. Comparisons of class I, II, III, and IV beta tubulin isotypes in total and colchicine-stable microtubule (MT) arrays from uncommitted EC, neuronal, and muscle cells were made by immunoblotting and by indirect immunofluorescence microscopy. In undifferentiated EC cells the relative amounts of these four isotypes are the same in both the total and stable MT populations. Subcellular sorting of beta tubulin isotypes was demonstrated in both neuronal and muscle differentiated cells. During neuronal differentiation, class II beta tubulin is preferentially incorporated into the colchicine-stable MTs while class III beta tubulin is preferentially found in the colchicine-labile MTs. The subcellular sorting of class II into stable MTs correlates with the increased staining of MAP 1B, and with the expression of MAP 2C and tau. Although muscle differentiated cells express class II beta tubulin, stable MTs in these cells do not preferentially incorporate this isotype but instead show increased incorporation of class IV beta tubulin. Muscle cells do not show high levels of MAP 1B and do not express MAP 2C or tau. These results are consistent with the hypothesis that a subcellular sorting of tubulin isotypes is the result of a complex interaction between tubulin isotypes and MT-associated proteins.

Association of acetylated microtubules, vimentin intermediate filaments, and MAP 2 during early neural differentiation in EC cell culture.

Pluripotent P19 embryonal carcinoma cell cultures can be induced to differentiate into neurons and glial cells by the addition of 10(-6) M retinoic acid. During early neural differentiation, a bundle of colchicine-stable, acetylated microtubules is formed. This acetylated microtubule array apparently extends to form neurites during neurogenesis. In this paper, we analyze changes in vimentin and MAP 2 distributions during neural differentiation with respect to the changes in the acetylated microtubule array. During a brief period early in differentiation, indirect immunofluorescence staining shows the colocalization of colchicine-stable acetylated microtubules, vimentin, and MAP 2. Using acrylamide to disrupt the organization of vimentin intermediate filaments and estramustine to disrupt the binding of MAP 2 to microtubules, we show that acetylated microtubules, MAP 2, and vimentin intermediate filaments are arranged in an interdependent cytoskeletal array. We suggest this array may serve to stabilize processes in neural stem cells, before the final decision to differentiate into neurons or glia is made.

Establishment of a stable, acetylated microtubule bundle during neuronal commitment.

Two posttranslational modifications of alpha-tubulin, acetylation and detyrosination, are associated with stable microtubule (MT) populations, including those of neuronal processes. We have used a pluripotent embryonal carcinoma cell line, P19, to investigate changes in MT isotype and stability found in MT arrays during neurogenesis. This cell line has an advantage in that both commitment- and differentiation-related events can be observed. Uncommitted P19 cells have minimal arrays of acetylated and detyrosinated MTs. Following neuronal induction with retinoic acid (RA), indirect immunofluorescence microscopy shows that the first MT modifications occur during commitment and before any morphological change is observed. RA-induced cells initially polymerize a temporarily enlarged population of MTs. Included in this population is a new array of acetylated MTs arranged in a bundle of parallel MTs. This bundle is colchicine-stable, although no MT-associated proteins (MAPs) are detectable using a battery of anti-MAP antibodies. Observation of MT arrays with patterns that are intermediate between the early bundles and short neurites suggests that the acetylated MT bundle subsequently extends to form a neurite. MAP 2 is first detected at about the time of neurite extension. However, at this early stage of differentiation, MAP 2 is not yet limited to dendritic processes. This report provides the first evidence that the stable MTs of mature neurons may be initiated during neuronal commitment.

Microfilaments: dynamic arrays in higher plant cells.

By using fluorescently labeled phalloidin we have examined, at the light microscope level, the three-dimensional distribution and reorganization of actin-like microfilaments (mfs) during plant cell cycle and differentiation. At interphase, mfs are organized into three distinct yet interconnected arrays: fine peripheral networks close to the plasma membrane; large axially oriented cables in the subcortical region; a nuclear "basket" of mfs extending into the transvacuolar strands. All these arrays, beginning with the peripheral network, disappear at the onset of mitosis and reappear, beginning with the nuclear basket, after cytokinesis. During mitotic and cytokinetic events, mfs are associated with the spindle and phragmoplast. Actin staining in the spindle is localized between the chromosomes and the spindle poles and changes in a functionally specific manner. The nuclear region appears to be the center for mf organization and/or initiation. During differentiation from rapid cell division to cell elongation, mf arrays switch from an axial to a transverse orientation, thus paralleling the microtubules. This change in orientation reflects a shift in the direction of cytoplasmic streaming. These observations show for the first time that actin-like mfs form intricate and dynamic arrays in plant cells which may be involved in many as yet undescribed cell functions.