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Biomedical subjects

M Howard

Publications and source records attributed to M Howard.

At least 91 records · Page 5Linked to original sources
PubMed1996

[Surgical myocardial revascularization during the 1st 15 days of evolution of acute myocardial infarction].

Revascularization significantly improves early and late prognosis in acute myocardial infarction and has prompted substantial changes in therapeutic stategies. We report 140 patients aged 60.3 years old (123 male) operated within 15 days of sustaining an acute myocardial infarction, between January 1984 and December 1989. Coronary angiogram showed single vessel disease in 8 (6%), double vessel disease in 32 (23%), triple vessel disease in 85 (61%) and left main vessel disease in 13 (9%). Indications for surgery were postinfarction angina in 92 patients (66%), multiple severe coronary stenoses in 18 (13%), infarction of less than six hours from onset in 16 (11%), acute angioplasty failure in 7 (5%) and cardiogenic shock in 7 (5%). Thirty one patients were operated during the initial 24 h of infarction (16 with less than 6 h), 14 between the second and third day and 95 between the fourth and fifteenth day. Overall mortality was 4.3% (6/140). Among patients with failed angioplasty and cardiogenic shock, mortality was 23% (7/140), among patients with postinfarction angina this figure was 2.1% (2/92). No patient operated within 6 hours of infarction onset or due to severe coronary stenosis, died. Ninety seven percent of patients were followed during mean of 49 months. Three patients had a new acute myocardial infarction, two had sudden death and two died of unrelated causes. One required angioplasty and none was reoperated. Five years actuarial survival was 95% and the actuarial probability of being free of acute myocardial infarction, angioplasty or reoperation at five years was 99 and 100% respectively. It is concluded that early surgical revascularization in cute myocardial infarction is safe and has excellent long term results.

Adult↗
PubMed1995

Methods for separation and deglycosylation of mucin subunits.

Mucins (density 1.35-1.50 g/ml) prepared from human cervical, gastric, respiratory, and salivary secretions were reduced and alkylated to generate reduced mucin subunits. These subunits were separated into different populations analytically by using agarose gel electrophoresis and preparatively by using anion-exchange chromatography. Both techniques separate the molecules primarily on the basis of their inherent charge. A procedure has been developed for the efficient deglycosylation of small amounts of these reduced mucin subunits immobilized on polyvinylidene difluoride. The combination of these procedures has allowed the isolation and identification of glycoforms of a specific mucin gene product in a respiratory mucous secretion.

Blotting, Western↗
PubMed1995

Asymmetric arc technique for posterior pharyngeal wall and retropharyngeal space tumors.

PURPOSE: Tumors of the posterior pharyngeal wall and nasopharyngeal cancer with retropharyngeal extension can partly encircle the cervical vertebrae. Treating the patient within spinal cord tolerance can cause a geographic miss. A simple technique has been developed to avoid this problem. METHODS AND MATERIALS: The standard fields for posterior pharyngeal wall and nasopharyngeal tumors are used up to 36-40 Gy. A planning computed tomography (CT) scan is taken during the second or third week of treatment with the patient fitted in a new shell ensuring that the cord is straight and parallel to the treatment couch. The asymmetric arc technique consists of two posterior arcs with closure of one jaw beyond the central axis. Each arc delivers the total dose to each ipsilateral side, while the median region of the U-shaped volume is treated by the summation of both arcs. RESULTS: We have treated 10 patients using asymmetric arcs in the last 3 years. This technique proved to be a versatile way of treating targets wrapped around the spine. The technique allows better individualization for target volume irregularities than the partial rotation with a central bar.

Humans↗
PubMed1995

Association of protein kinase A and protein phosphatase 2B with a common anchoring protein.

Specificity of protein kinases and phosphatases may be achieved through compartmentalization with preferred substrates. In neurons, adenosine 3', 5'-monophosphate (cAMP)-dependent protein kinase (PKA) is localized at postsynaptic densities by association of its regulatory subunit with an A kinase anchor protein, AKAP79. Interaction cloning experiments demonstrated that AKAP79 also binds protein phosphatase 2B, or calcineurin (CaN). A ternary complex of PKA, AKAP, and CaN was isolated from bovine brain, and colocalization of the kinase and the phosphatase was established in neurites of cultured hippocampal neurons. The putative CaN-binding domain of AKAP79 is similar to that of the immunophilin FKBP-12, and AKAP79 inhibited CaN phosphatase activity. These results suggest that both PKA and CaN are targeted to subcellular sites by association with a common anchor protein and thereby regulate the phosphorylation state of key neuronal substrates.

A Kinase Anchor Proteins↗
PubMed1995

A synthetic peptide corresponding to a critical intracellular signaling region of the human IL-4 receptor inhibits IL-4-induced proliferation.

We have previously identified a critical region for growth signal transduction in the cytoplasmic domain of the human IL-4 receptor (hIL-4R). Since the entire cytoplasmic domain of this receptor lacks known catalytic activities such as the tyrosine kinase domain, it is likely that the IL-4R associates with other signal-transducing molecules through this critical cytoplasmic region. We test here whether a synthetic peptide corresponding to this critical cytoplasmic region, designated SP-1, interferes with IL-4-induced proliferation by competing with the IL-4R for binding to intracellular signal-transducing molecules. Our data indicated that 100 micrograms/ml SP-1 peptide completely inhibits human IL-4 (hIL-4)-induced proliferation of Ba/F3 transfectants expressing the full-length hIL-4R (hIL-4R-Ba/F3 transfectants). In contrast, a wide concentration range of an unrelated synthetic peptide, designated SP-2, did not affect hIL-4-induced proliferation of hIL-4R-Ba/F3 transfectants. This difference between SP-1 and SP-2 peptides was not due to their differential uptake by cell, since approximately 100 times more SP-2 peptide could be found in cytoplasmic extracts than SP-1 peptide in experiments using radiolabeled peptides. The specificity of SP-1-mediated inhibition of IL-4-induced proliferation was supported by the fact that the SP-1 peptide had no effect on IL-3-induced proliferation of the same hIL-4-Ba/F3 transfectants. In addition, the SP-1 peptide did not affect either IL-2-induced proliferation of Ba/F3 transfectants expressing the human IL-2 receptor beta chain (hIL-2R beta) or hIL-4-induced proliferation of Ba/F3 transfectants expressing a chimeric receptor consisting of the hIL-4R extracellular domain and the hIL-2R beta cytoplasmic domain. SP-1 was unable to inhibit IL-4-induced proliferation of other IL-4-responsive cell lines such as human erythroleukemic cell line TF-1 and mouse T cell lines HT2 and CTLL-2. In addition, SP-1 caused only a 50% inhibition of Ba/F3 cell proliferation induced by mouse IL-4. The failure of SP-1 to inhibit IL-4-induced proliferation in these various cell lines while producing excellent inhibition of hIL-4-induced proliferation of hIL-4R-Ba/F3 transfectants appeared to be related to the number of IL-4Rs expressed on each cell type.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1995

Murine CD38: an immunoregulatory ectoenzyme.

CD38 is an ectoenzyme that utilizes NAD+ and is expressed by many cells of hematopoietic origin. Antibodies to CD38 potentiate many biological activities on lymphocytes, including induction of murine B-cell proliferation. In this article, Frances Lund and colleagues summarize information concerning the expression, enzymatic activity and signal transduction pathway utilized by murine CD38.

ADP-ribosyl Cyclase↗
PubMed1995

CD38 unresponsiveness of xid B cells implicates Bruton's tyrosine kinase (btk) as a regular of CD38 induced signal transduction.

CD38 is a 42 kDa membrane-associated ectoenzyme expressed by a large proportion of human and mouse lymphocytes. Agonistic antibodies to CD38 induce a strong proliferative response in lymphocytes additionally co-stimulated with other growth co-factors such as IL-4, IL-2 plus accessory cells or sub-mitogenic doses of endotoxin. We show here that B lymphocytes from unstimulated X-linked immunodeficient (xid) mice are unresponsive to CD38 stimulation, both in terms of proliferative response and surface antigen modulation. This CD38 unresponsiveness is evident in the presence of excess quantities of, and normal responses to, the accessory growth co-stimulants required for this response. CD38 molecules expressed on xid B cells are normal in terms of expression levels, size and enzymatic activity, suggesting that CD38 unresponsiveness reflects a down-stream signaling defect. In light of the recent proposal that the xid gene encodes a tyrosine kinase called Bruton's tyrosine kinase (btk), these data suggest that btk is either an integral component or an indirect regulator of the CD38-induced signal transduction pathway.

ADP-ribosyl Cyclase↗
PubMed1995

Epitope tagging permits cell surface detection of functional CFTR.

The cystic fibrosis transmembrane conductance regulator (CFTR) is a phosphorylation-activated Cl channel responsible for adenosine 3',5'-cyclic monophosphate (cAMP)-induced Cl secretion across the apical membranes of epithelial cells. To optimize its detection for membrane localization studies, we tagged CFTR with epitope sequences at the carboxy terminus or in the fourth external loop. When epitopes were added to the fourth external loop, the N-linked glycosylation sites in that loop were either preserved or they were mutated to produce a deglycosylated CFTR (dgCFTR). Tagged CFTRs were expressed in HeLa cells, and their cAMP-sensitive Cl permeability was assayed using the halide-sensitive fluorophore SPQ. CFTRs containing the M2 epitope showed halide permeability responses to cAMP, whereas cells expressing CFTR with the hemagglutinin (HA) tag showed little or no cAMP response. Xenopus oocytes expressing dgCFTR, with or without the M2 epitope, showed Cl conductance responses that were 20% of the wild-type response, whereas M2-tagged constructs retaining the glycosylation sites responded like wild-type CFTR. External M2-tagged CFTR was detected in the surface membrane of nonpermeabilized cells. The surface expression of the mutant M2-tagged CFTRs correlated with processing of these mutants (Gregory et al. Mol. Cell. Biol. 11:3886-3893, 1991). M2-901/CFTR is a useful reporter for the trafficking of wild-type and mutant CFTRs to the cell surface.

Animals↗
PubMed1995

Analysis of respiratory mucus glycoproteins in asthma: a detailed study from a patient who died in status asthmaticus.

Airway mucus from asthmatics is often unusually solid. The death of a patient in status asthmaticus allowed the collection of 28 g of abnormal airway mucus at autopsy. Its chemical and physical properties were studied to reveal differences from more normal airway mucus. The gel plug taken from the airways could be dispersed in 6 M guanidinium chloride, but it took > 1 wk and 700 ml of extractant to disperse 3 g of exudate completely. In contrast, treatment with 10 mM dithiothreitol, which reduces disulfide bonds, dispersed the gel within seconds. Mucins accounted for 25% of the non-dialyzable material in the gel, while DNA constituted < 1% and proteoglycans could not be detected. The mucins were similar in architecture and general composition to other respiratory mucins and were present at a high concentration (approximately 40 mg/ml). The majority of mucins were of extreme size (mean M(r) 30-40 x 10(6)) and slow to dissolve, but sequential extraction experiments on the gel exudate demonstrated a proportion of mucins (15%), the most readily extracted, which had a higher density, 1.45-1.55 g/ml, a lower M(r) (11.5 x 10(6)) and were markedly more acidic than the bulk of the mucins. Both major and minor mucin populations were extremely heterogeneous in mass distribution. Electron microscopy of the major mucin species demonstrated extensive networks of molecules many microns in length. The major mucin species was distinctly less acidic than mucins previously described from either normal or diseased airways. Amino acid analysis of fractions across the charge distribution suggested the presence of at least two different mucin proteins occurring as distinct glycoforms.

Gels↗
PubMed1995

It's about time.

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Continuity of Patient Care↗
PubMed1995

Cationic lipids for reporter gene and CFTR transfer to rat pulmonary epithelium.

Increasing evidence indicates that cationic liposomes are capable of safely transferring foreign genes to pulmonary epithelium in vitro and in vivo. To transfer reporter genes and the cystic fibrosis transmembrane conductance regulator (CFTR) to mammalian respiratory epithelium we used two cationic lipid formulations: N-[1-(2,3-dioleoyloxy)propyl] N,N,N-triethylammonium chloride (DOTMA), and 1,2-dimyristyloxy-propyl-3-dimethylhydroxyethylammonium bromide (DMRIE) at a 1:1 molar ratio with dioleoyl phosphatidylethanolamine (DOPE). Lipid-DNA conjugates containing either CFTR or LacZ were instilled directly into the airways of Sprague-Dawley rats. Rats treated with LacZ cDNA in vivo demonstrated expression in 30-50% of the large and medium-sized airways, with some airways showing high efficiency gene transfer and expression (in the most proximal airways, 70-80% of surface epithelial cells were positive for expression of a nuclear targeted LacZ). While control and LacZ treated tracheas mounted in Ussing chambers showed minimal stimulation of transepithelial chloride (Cl)-currents by cAMP (suggesting low levels of endogenous rat CFTR activity), tracheas taken from animals receiving CFTR exhibited significant forskolin-stimulated currents at 72 h after gene transfer. Human CFTR gene expression was also detected by polymerase chain reaction (PCR) analysis of reverse transcribed lung RNA. These results, together with previous studies using lipid-mediated gene transfer in mice, help confirm the potential for cationic lipid-mediated gene transfer in the gene therapy of cystic fibrosis in humans.

Animals↗
PubMed1995

Gut permeability measured by polyethylene glycol absorption in abnormal gut fermentation as compared with food intolerance.

Gut permeability has been studied in patients with either food intolerance or abnormal gut fermentation as well as in normal subjects. Permeability was measured by polyethylene glycol absorption, and the reasons for this choice of probe are discussed. Results show that both symptomatic groups have statistically very highly significant deviations from the normal (P < 0.01), consisting of over-adsorption, significant at molecular weights 242, 286, 330 and 374. Whilst both study groups were different from the normal they were not different from each other. The implications for these findings in the diagnosis and management of food intolerance and abnormal gut fermentation are discussed.

Fermentation↗
PubMed1995

[Bacterial translocation in a model of small intestine autotransplant in dogs].

BACKGROUND: Sepsis is the commonest complication of small bowel transplantation. These infections are presumably caused by bacterial translocation, due to splanchnic ischemia. AIM: To study bacterial translocation in the immediate postoperative period after a small bowel transplantation in dogs and to relate it to splanchnic ischemia. METHODS: Three groups of dogs were studied. In group A (n = 6) spontaneous episodes of splanchnic ischemia were monitored in the first 18 h of the postoperative period. In group B (n = 5), a 60 min ischemia was induced by superior mesenteric artery occlusion, two hours after small bowel transplantation. In group C (n = 5) a 60 min ischemia was induced by occlusion of mesenteric vein, two hours after transplantation. Bacterial translocation was assessed through bacterial cultures from the mesenteric vein and splanchnic ischemia with intramucosal pH measurement (a pH < 7.2 was considered indicative of ischemia). RESULTS: Twenty eight of 83 cultures were positive, specially for Gram negative bacilli. The incidence of positive cultures was 14% for group A, 17% for group B and 79% for group C (p < 0.01 compared to groups A and B). The higher incidence of bacterial translocation occurred during the first two hours after transplantation, when the lower intramucosal pH recordings were obtained. The percentage of positive cultures was 39% during periods of ischemia, compared to 24% during periods without ischemia (p = NS). CONCLUSIONS: Bacterial translocation occurs during the first two hours after intestinal transplantation, in concomitance with the lower intramucosal pH readings.

Animals↗
PubMed1994

Immunoglobulin signal transduction guides the specificity of B cell-T cell interactions and is blocked in tolerant self-reactive B cells.

The specificity of antibody (Ab) responses depends on focusing helper T (Th) lymphocyte signals to suitable B lymphocytes capable of binding foreign antigens (Ags), and away from nonspecific or self-reactive B cells. To investigate the molecular mechanisms that prevent the activation of self-reactive B lymphocytes, the activation requirements of B cells specific for the Ag hen egg lysozyme (HEL) obtained from immunoglobulin (Ig)-transgenic mice were compared with those of functionally tolerant B cells isolated from Ig-transgenic mice which also express soluble HEL. To eliminate the need for surface (s)Ig-mediated Ag uptake and presentation and allow the effects of sIg signaling to be studied in isolation, we assessed the ability of allogeneic T cells from bm12 strain mice to provide in vivo help to C57BL/6 strain-transgenic B cells. Interestingly, non-tolerant Ig-transgenic B cells required both allogeneic Th cells and binding of soluble HEL for efficient activation and Ab production. By contrast, tolerant self-reactive B cells from Ig/HEL double transgenic mice responded poorly to the same combination of allogeneic T cells and soluble HEL. The tolerant B cells were nevertheless normally responsive to stimulation with interleukin 4 and anti-CD40 Abs in vitro, suggesting that they retained the capacity to respond to mediators of T cell help. However, the tolerant B cells exhibited a proximal block in the sIg signaling pathway which prevented activation of receptor-associated tyrosine kinases in response to the binding of soluble HEL. The functional significance of this sIg signaling defect was confirmed by using a more potent membrane-bound form of HEL capable of triggering sIg signaling in tolerant B cells, which markedly restored their ability to collaborate with allogeneic Th cells and produce Ab. These findings indicate that Ag-specific B cells require two signals for mounting a T cell-dependent Ab response and identify regulation of sIg signaling as a mechanism for controlling self-reactive B cells.

Animals↗