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Biomedical subjects

D A Walsh

Publications and source records attributed to D A Walsh.

At least 109 records · Page 6Linked to original sources
PubMed1990

Long term continuous domiciliary oxygen therapy by transtracheal catheter.

Long term continuous oxygen therapy improves prognosis in patients with hypoxaemic chronic airflow limitation. Transtracheal delivery of oxygen permits adequate oxygenation of such patients at lower flow rates than are required for delivery by nasal cannulas, thus increasing the time for which portable oxygen cylinders can be used and improving the efficacy of domiciliary oxygen concentrators in patients with refractory hypoxaemia. In a study of the long term acceptability and risks of transtracheal oxygen in 37 patients, with a total follow up of 595 patient-months, most patients reported symptomatic benefit and four have used transtracheal oxygen successfully for more than four years. Problems have included surgical emphysema (4 patients), catheter fracture (9), local infection (34), and catheter dislodgement (21); but these have been relatively infrequent and in general have not caused important clinical problems. Transtracheal oxygen therapy was discontinued in 10 patients before death for various reasons, including infection (2 patients), catheter fracture (1), and surgical emphysema (1). Accumulation of mucus balls has not been a problem. It is concluded that transtracheal oxygen therapy is a safe and acceptable alternative to nasal cannulas in well motivated patients with hypoxaemia due to chronic airflow limitation who are keen to pursue an active life.

Aged↗
PubMed1990

Bronchoscopic cryotherapy for advanced bronchial carcinoma.

A prospective study was carried out to assess the value of bronchoscopic cryotherapy for palliation of inoperable bronchial carcinoma with bronchial obstruction. Symptoms, lung function, and chest radiographic and bronchoscopic findings were recorded serially before and after 81 cryotherapy sessions in 33 consecutive patients. Most patients improved in terms of overall symptoms, stridor, and haemoptysis and they had an overall improvement in dyspnoea. Objective improvement in lung function was seen in 58% of patients and the changes in lung function correlated with symptoms. Bronchoscopic evidence of relief of bronchial obstruction was seen in 77% of patients and 24% showed improvement in degree of collapse on the radiograph. There were no important complications. These results compare favourably with the results in published series of patients having laser therapy. It is concluded that bronchoscopic cryotherapy is valuable for the palliation of inoperable bronchial carcinoma.

Aged↗
PubMed1990

AHR-14310C: a potent, long-acting, nonsedating H1-antihistamine that prevents antigen-induced mucus formation in sensitive rats.

AHR-14310C(5-[2-[4-[bis(4-fluorophenyl)hydroxymethyl- 1-piperidinyl]ethyl]-3-methyl]-2-oxazolidinone ethanedioate, hydrochloride salt) displays potent and long-acting antihistaminic activity in guinea pigs and in dog and guinea pig models of immediate hypersensitivity. Given orally 1, 5 or 24 hr before an i.v. histamine challenge, AHR-14310C produced ED50 values of 0.76, 0.22 and 0.58 mg/kg, respectively, in protecting naive guinea pigs from the lethal effects of the histamine challenge. AHR-14310C was also effective when the histamine was administered as an aerosol (1-, 5- and 24-hr ED50 values = 0.69, 0.38 and 1.08 mg/kg, respectively). AHR-14310C also attenuated the anaphylactic responses to aerosolized antigen in sensitive guinea pigs and the skin response to antigen in naturally sensitive dogs. AHR-14310C, at doses in vastly excess of those required to block histamine-induced hypotension, did not alter the electroencephalogram of cats, as did the sedating antihistamine, diphenhydramine. AHR-14310C did not affect the autonomic responses to acetylcholine, isoproterenol, epinephrine or 1,1-dimethyl-4-phenylpiperazinium chloride in dogs. At low doses (0.316-1.0 mg/kg p.o.), AHR-14310C blocked antigen-induced tracheal mucous changes in sensitive rats. AHR-14310C has therapeutic potential in allergic individuals, particularly in asthmatics, where bronchorrhea or mucus plugging is a problem.

Acetylcholine↗
PubMed1990

Ethics, jurisprudence, and economics in the medical school curriculum.

Medical ethics, medical jurisprudence, and medical economics are recognized as important components of a medical school curriculum. These subjects were introduced through a course given at the University of California, Davis, School of Medicine. Four aspects of the format and content of the course were instrumental to its success. Teaching principles of medical ethics within the context of jurisprudence and economics permitted the students to gain an understanding of the institutions and processes that act as positive and negative constraints on physicians' clinical and professional behavior. The course was offered during the fourth year following required clinical rotations so that all aspects of the course could be based on the clinical experiences of the students. It was presented in a continuing medical education format away from the normal teaching environment of first- and second-year classrooms and third-year clerkships. Finally, the course was designed by a multidisciplinary, multidepartmental planning group that included students.

Bioethical Issues↗
PubMed1989

Conformational analysis of PKI(5-22)amide, the active inhibitory fragment of the inhibitor protein of the cyclic AMP-dependent protein kinase.

Fourier-transform i.r. spectroscopy, 1H-n.m.r. spectroscopy and X-ray scattering were used to study the conformation and shape of the peptide PKI(5-22)amide, which contains the active site of the inhibitor protein of the cyclic AMP-dependent protein kinase [Cheng, Van Pattern, Smith & Walsh (1985) Biochem. J. 231, 655-661]. The X-ray-scattering solution studies show that the peptide has a compact structure with Rg 0.9 nm (9.0 A) and a linear maximum dimension of 2.5 nm (25A). Compatible with this, Fourier-transform i.r. and n.m.r. determinations indicate that the peptide contains approx. 26% alpha-helix located in the N-terminal one-third of the molecule. This region contains the phenylalanine residue that is one essential recognition determinant for high-affinity binding to the protein kinase catalytic site.

Amides↗
PubMed1989

Expression of a cDNA for the catalytic subunit of skeletal-muscle phosphorylase kinase in transfected 3T3 cells.

Phosphorylase kinase is a multimeric enzyme of composition (alpha, beta, gamma, delta)4 whose catalytic activity resides in the gamma-subunit. As an approach to understand further its regulation, a cDNA for the gamma-subunit of phosphorylase kinase (gamma PhK) has been cloned into a mammalian expression vector behind the mouse metallothionein-1 promoter. NIH 3T3 cells were co-transfected with this construct (pEV gamma PhK) and pSV2neo, G418-resistant clones were selected, and several were found to have stably incorporated the gamma-subunit cDNA into their genomic DNA. Phosphorylase kinase activity was clearly present in extracts from cultures of pEV gamma PhK-transformed cells and increased several-fold after 24 h of incubation with Zn2+, whereas it was undetectable in the parent 3T3 cells. A significant, but variable, proportion (15-70%) of the activity was Ca2+-dependent. We conclude that the phosphorylase kinase activity expressed by the cells transformed with pEV gamma PhK is due to free gamma-subunit and gamma-subunit associated with cellular calmodulin, which replaces the delta-subunit normally associated with the gamma-subunit in the holoenzyme.

Animals↗
PubMed1989

Protein kinase inhibitor-(6-22)-amide peptide analogs with standard and nonstandard amino acid substitutions for phenylalanine 10. Inhibition of cAMP-dependent protein kinase.

The minimal structure in the heat-stable inhibitor protein of cAMP-dependent protein kinase required for a low nanomolar potency of inhibition is the peptide Thr6-Tyr-Ala-Asp-Phe-Ile-Ala-Ser-Gly-Arg-Thr-Gly-Arg-Arg-Asn-Ala-+ ++Ile22-NH2 (PKI-(6-22)-amide). While primary structural determinants for interaction with the protein kinase are distributed throughout the 17 residues of this peptide, we have previously shown that phenylalanine 10 in the NH2-terminal portion is a particularly important determinant for high affinity binding (Glass, D. B., Cheng, H.-C., Mende-Mueller, L., Reed, J., and Walsh, D. A. (1989) J. Biol. Chem. 264, 8802-8810). To investigate this requirement further, peptide analogs of PKI-(6-22)-amide in which various natural and nonstandard amino acids are substituted for phenylalanine 10 have been synthesized and tested for inhibitory potency against the catalytic subunit of the protein kinase. Consistent with the importance of the hydrophobicity of phenylalanine, an alanine 10 substitution analog exhibited a 270-fold decrease in inhibitory potency, whereas the leucine 10 analog lost only 33-fold in activity as compared to the parent peptide PKI-(6-22)-amide. Peptides containing the spatial conformation analogs D-phenylalanine, homophenylalanine, or phenylglycine were 60-120-fold less potent than the parent peptide. Peptides containing various para-substituted phenylalanines at position 10 were only 5-11-fold less potent. One exception to this was (4'-azidophenylalanine 10)PKI-(6-22)-amide, which was nearly equipotent with the parent inhibitor. The most potent analogs were those peptides containing highly aromatic residues at position 10. The 2'-thienylalanine 10, tryptophan (formyl) 10, tryptophan 10, and the 1'-naphthylalanine 10 analogs were 3-fold less potent, equipotent, slightly more potent, and 4-fold more potent than the parent peptide inhibitor, respectively. We conclude that phenylalanine 10 in PKI-(6-22)-amide, and presumably in the native protein inhibitor, interacts through specific hydrophobic and/or aromatic binding to a hydrophobic pocket or cleft near the active site of the protein kinase.

Amino Acid Sequence↗
PubMed1989

Primary structural determinants essential for potent inhibition of cAMP-dependent protein kinase by inhibitory peptides corresponding to the active portion of the heat-stable inhibitor protein.

PKI-(5-24)-amide is a 20-residue peptide with the sequence, Thr5-Thr-Tyr-Ala-Asp-Phe-Ile-Ala-Ser-Gly-Arg-Thr-Gly-Arg-Arg-Asn-A la-Ile-His- Asp24-NH2, that corresponds to the active portion of the heat-stable inhibitor protein of cAMP-dependent protein kinase (Cheng, H.-C., Kemp, B. E., Pearson, R. B., Smith, A. J., Misconi, L., Van Patten, S. M., and Walsh, D. A. (1986) J. Biol. Chem. 261, 989-992). Amino acid residues in PKI-(5-24)-amide responsible for the potent inhibition (Ki = 2.3 nM) of the catalytic subunit of protein kinase were further investigated using deletion and substitution analogs of the synthetic peptide. Residues 5, 23, and 24 were not required for activity since the 17-residue PKI-(6-22)-amide retained full potency. Sequential removal of the first seven amino acids from the NH2 terminus of PKI-(5-24)-amide caused a progressive 50-fold loss of inhibitory potency. In contrast, substitution of either Thr6, Asp9, or Ile11 with alanine, or Ala8 by leucine, in PKI-(5-22)-amide produced less than 3-fold decreases in potency. Of the 2 aromatic residues in PKI-(5-22)-amide, the individual substitution of Phe10 and Tyr7 by alanine caused, respectively, 90- and 5-fold decreases in inhibitory potency, demonstrating important roles for each. This NH2-terminal portion of the peptide is believed to contain a significant portion of alpha-helix. Many recognition or structural determinants are also essential in the COOH-terminal portion of PKI-(5-22)-amide. In addition to the basic subsite provided by the three arginines, several other of the residues are critical for full inhibitory potency. Substitution of Ile22 by glycine in either PKI-(5-22)-amide or PKI-(14-22)-amide lowered the inhibitory potency by 150- and 50-fold, respectively. Separate replacement of Gly17 or Asn20, in either PKI-(5-22)-amide or PKI-(14-22)-amide, caused 7-15-fold decreases in potency. Substitution of both Gly17 and Asn20 together (in PKI-(14-22)-amide) produced a synergistic loss of inhibitory activity. [Leu13,Ile14]PKI-(5-22)-amide, a doubly substituted analog exhibited a 42-fold increase in Ki value. We conclude that Ser13 and/or Gly14, Gly17, Asn20, and Ile22 each contribute important features to the binding of these inhibitory peptides to the protein kinase, either by providing recognition determinants, inducing structure, and/or allowing essential peptide backbone flexibility.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1989

Regulation of the inducible heat shock 71 genes in early neural development of cultured rat embryos.

Activation of the inducible heat shock 71 genes and their role in the heat shock response was studied in vitro in 9.5 day-old rat embryos at neural tube closure. The transcriptional response of a 71 kilodalton (kD) heat shock gene induced after various regimes of heat shock and acquired thermotolerance was investigated. Expression and accumulation of the heat shock (hs) 71 mRNA in the neuroectoderm was studied by Northern and dot blot analysis. Specific expression in various cell types and regions of the neuroectoderm were examined by in situ hybridization. Exposure of embryos to a heat shock at 43 degrees C for 7.5 min caused high levels of hs mRNA 71 accumulation in the neuroectoderm, pronounced protein synthesis inhibition, and regulated recovery. Specific neuroectoderm cell death followed, resulting in major developmental defects of the eye and forebrain region. A mild heat shock of 42 degrees C for 10 min induced the heat shock response, hsp synthesis, and cell recovery, but produced no cell death or deformities. Preheating the embryo at 42 degrees C resulted in acquired thermotolerance to an otherwise teratogenic 43 degrees C heat shock. Thermotolerance was associated with a rapid recovery of protein synthesis associated with hs 71 mRNA expression. Dot blot analysis showed that after a 42 degrees C heat shock, 71 mRNA was rapidly transcribed and transported into the cytoplasm where it was degraded within 2 hr of the initial response. The results suggest that the heat shock protein (hsp) 71 gene may have a protective rather than a rescuing function.

Animals↗
PubMed1989

Heat shock affects cell cycling in the neural plate of cultured rat embryos: a flow cytometric study.

The effects of heat shock on cell cycling in the mammalian neuroectoderm were determined by applying heat shocks to cultured rat embryos at the neural plate stage, as part of a study on the teratogenic effects of heat shock on neural development. The heat shocks had been characterized previously (Walsh et al.: Teratology 36:181-191, 1987) with respect to their effects on the gross morphological development of the rat embryos. The effects on cell cycling were observed in DNA histograms of neural plate cells recorded in a flow cytometer after staining with DAPI. The mild heat shock (42 degrees C for 10 min) arrested cells at entry to S phase. The teratogenic heat shock (43 degrees C for 7.5 min) arrested cells at entry to S phase also but for a longer time and inhibited cycling through S phase. After each arrest, a synchronized peak of cells later entered S phase and progressed through the cycle. The induced-thermotolerance heat shock, which was the mild heat shock followed after an interval by the teratogenic heat shock, showed that pre-treatment with the mild heat shock reduced the magnitude of the response to the teratogenic heat shock. The cell-cycle inhibitor ICRF 159 showed the effects on cycling rates of the heat-shock treatments. The arrest of cells at entry to S phase by heat shock may function to prevent cells entering DNA synthesis under non-optimal conditions. We report estimates of proportions of non-proliferative cells in the neural plate of the rat embryos.

Animals↗
PubMed1989

Synthesis and antiallergy activity of 4-(diarylhydroxymethyl)-1-[3-(aryloxy)propyl]piperidines and structurally related compounds.

A series of 4-(diarylhydroxymethyl)-1-[3-(aryloxy)propyl]piperidines was synthesized and evaluated for antiallergy activity. Several analogues had potent activity in the passive foot anaphylaxis (PFA) assay, an IgE-mediated model useful in the detection of compounds possessing antiallergic activity. In particular 1-[4-[3-[4-[bis(4-fluorophenyl)hydroxymethyl]-1-piperidinyl] propoxy]-3-methoxyphenyl]ethanone (1, AHR-5333) was more potent than oxatomide and terfenadine in this assay.

Anaphylaxis↗
PubMed1989

Synthesis, characterization and inhibitory activities of (4-N3[3,5-3H]Phe10)PKI(6-22)amide and its precursors: photoaffinity labeling peptides for the active site of cyclic AMP-dependent protein kinase.

PKI(6-22)amide is a 17 residue peptide corresponding to the active portion of the heat-stable inhibitor of cAMP-dependent protein kinase. The peptide is a potent (Ki = 1.6 nM), competitive inhibitor of the enzyme. The photoreactive peptide analog (4-azidophenylalanine10)PKI(6-22)amide was synthesized in both its non-radiolabeled and tritiated forms by chemical modification of precursor peptides that were prepared by stepwise solid-phase synthesis. (4-Amino[3,5-3H]phenylalanine10)PKI(6-22)amide, the precursor for the radiolabeled arylazide peptide, was obtained by catalytic reduction of the corresponding peptide containing the 3,5-diiodo-4-aminophenylalanine residue at position 10. The purified PKI peptides were analyzed by HPLC, amino acid analysis, and u.v. spectra. In the dark, (4-azidophenylalanine10)PKI(6-22)amide inhibited the catalytic subunit of cAMP-dependent protein kinase with a Ki value of 2.8 nM. The photoreactivity of the arylazide peptide was demonstrated by time-dependent u.v. spectral changes on exposure to light. Photolysis of the catalytic subunit (4-azido[3,5-3H]phenylalanine10)PKI(6-22)amide complex resulted in specific covalent labeling of the enzyme. The data indicate that this peptide is a useful photoaffinity labeling reagent for the active site of the protein kinase.

Affinity Labels↗
PubMed1988

The inhibitor protein of the cyclic AMP-dependent protein kinase-catalytic subunit interaction. Composition of multiple complexes.

It has been previously demonstrated that the combination of pure preparations of the inhibitor protein of the cyclic AMP-dependent protein kinase and the catalytic subunit of this enzyme resulted in the formation of multiple complexes [Van Patten, Fletcher & Walsh (1986) J. Biol. Chem. 261, 5514-5523]. In the present study it is demonstrated that these multiple species occur because the bovine heart protein kinase preparation contains multiple forms of catalytic subunit [Kinzel, Hotz, König, Gagelmann, Pyerin, Reed, Köbler, Hofmann, Obst, Gensheimer, Goldblatt & Shaltiel (1987) Arch. Biochem. Biophys. 253, 341-349].

Adenosine Triphosphate↗
PubMed1988

The catalytic subunit of cAMP-dependent protein kinase induces expression of genes containing cAMP-responsive enhancer elements.

Transcriptional regulation of eukaryotic genes by cyclic AMP requires a cAMP-dependent protein kinase (A kinase). Two hypotheses have been proposed to explain how the holoenzyme of the A kinase induces transcription. The regulatory subunits of the A kinase, which bind cAMP and DNA, and have amino-acid homology with the Escherichia coli catabolite activator protein could directly stimulate gene expression. Alternatively, phosphorylation by the catalytic subunits could induce transcription by activating proteins involved in gene transcription. To distinguish between these models, we microinjected purified preparations of the catalytic and regulatory subunits of A kinase into tissue culture cells and monitored expression of a stably integrated fusion gene containing a cAMP-responsive human promoter fused to a bacterial reporter gene, or of the endogenous c-fos gene. The catalytic subunit stimulated expression of these genes, whereas the regulatory subunit did not. These results indicate that the catalytic subunit of A kinase is sufficient to induce expression of two cAMP-responsive genes, without increasing levels of cAMP.

Animals↗
PubMed1988

Pulmonary tissue volume and blood flow as functions of body surface area and age.

We measured pulmonary tissue volume (Vt) and capillary blood flow (Qc) by rebreathing acetylene as the soluble gas in 94 male and 82 female patients with normal healthy lungs. We found that the standard deviation of Vt was a function of Vt and that the standard deviation of Qc was a function of Qc; logarithm transforms were used to prevent this. A significant correlation exists between log Vt and body surface area (BSA) and log Vt and vital capacity (VC). For both sets of regressions older subjects (greater than or equal to 40 years of age) exhibited greater variability about the regression line than younger subjects; therefore the 2 age groups of subjects were analyzed separately for tissue volume. We describe regression equations of log Vt vs. BSA or VC for younger male, younger female, older male, and older female patients. The normal limits by sex and age groups are also included. A significant correlation was found between log Qc, BSA, and age. Thus 2 equations, for male and female patients, are used to predict log Qc based on these 2 covariates.

Adolescent↗