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Biomedical subjects

J W Ryan

Publications and source records attributed to J W Ryan.

At least 19 recordsLinked to original sources
PubMed1992

A radioassay for aminoacylproline hydrolase (aminopeptidase P) activity.

A radioassay was developed in which aminoacylproline hydrolase acts on Arg-Pro-Pro-[3H]benzylamide to yield arginine plus Pro-Pro-[3H]benzylamide. By stopping the reaction with base (0.1 M NaOH), the radioactive product is deprotonated to an organophilic form and is separable from the hydrophilic substrate by extraction of the alkaline aqueous solution with an organic solvent. When scintillants are included in the organic solvent, the enzyme:substrate reaction, extraction and quantification of Pro-Pro-[3H]benzylamide can all be conducted using a single liquid scintillation vial. Thus, aminoacylproline hydrolase activity is measured in terms of the rate of release of Pro-Pro-[3H]benzylamide. The substrate is obtainable at greater than 20 Ci/mmol, which enables its use under conditions of first-order enzyme kinetics. Conditions of near-zero order kinetics are readily attained by adding unlabeled substrate (Km 0.7 microM). The substrate is highly reactive (a 1:2000 dilution of guinea pig plasma hydrolyzed greater than 10% of the substrate during a 10 min incubation at 37 degrees C) and specific in that it is not degraded by leucine aminopeptidase, aminopeptidase A or N, dipeptidyl peptidase IV nor prolyl endopeptidase. The assay was used to measure aminoacylproline hydrolase specific activities in tissues of rat and guinea pig. Activity was found in virtually all major tissues of both species, and some guinea pig tissues (e.g. kidney and plasma) were found to be notably rich sources of the enzyme.

Amino Acid Sequence↗
PubMed1992

Purification and characterization of guinea pig serum aminoacylproline hydrolase (aminopeptidase P).

Aminoacylproline hydrolase (EC 3.4.11.9) of guinea pig serum has been obtained as two apparently homogeneous isoforms. Dialyzed serum was chromatographed successively on Affi-gel blue, hydroxyapatite, DE-cellulose, phenyl-Sepharose, an affinity matrix for angiotensin converting enzyme and concanavalin-Sepharose. On the latter matrix, 68% of the enzyme activity was eluted with alpha-methyl mannoside at 10 and 100 mM, and 29% was eluted with alpha-methyl glucoside, 500 mM, at 56 degrees C. The two fractions ('biantennary' and 'high mannose' fractions, respectively) were concentrated and then chromatographed separately on Sephacryl S-200HR. Both fractions were eluted as expected for a globular protein of Mr 217,000. On SDS-PAGE, under reducing and non-reducing conditions, each of the concanavalin-Sepharose fractions was separated into two protein bands, Mr 89,000 and Mr 81,500. Each of the bands was found to be N-blocked when N-terminal amino acid sequencing was attempted. The reaction of the 'biantennary' fraction with the synthetic substrate Arg-Pro-Pro-[3H]benzylamide was characterized in part: Km 0.7 microM, kcat 124.6 min-1, kcat/Km 1.78.10(8) M-1 min-1. Hydrolysis of the substrate was strongly inhibited by bradykinin and those of its lower homologs that contain two adjacent proline residues. Cu2+ was strongly inhibitory. Co2+ at 30 microM activated the enzyme, as did Mn2+, Mg2+ and Ca2+ at higher concentrations. Sulfhydryl compounds, including captopril, inhibited the enzyme as did 1,10-phenanthroline. Iodoacetamide and N-ethylmaleimide had no effects, but 4-hydroxymercuribenzoate conferred a partial inhibition over a remarkably wide concentration range: 0.34-1400 microM. Amastatin and bestatin did not inhibit the enzyme. Aminoacylproline hydrolase of guinea pig serum appears to be a heterogeneous, glycosylated metallo-enzyme with a high affinity for bradykinin and related peptides in which the sequence Pro-Pro, Xaa-Pro-Pro or Xaa-Pro-Hyp is N-terminal.

Amino Acid Sequence↗
PubMed1992

Pulmonary angiotensin-converting enzyme substrate hydrolysis during exercise.

We examined exercise-induced changes in indicator-dilution estimates of the angiotensin-converting enzyme first-order kinetic parameter, the ratio of a normalized maximal enzymatic conversion rate to the Michaelis constant (Amax/Km), which, under stable enzymatic conditions, will vary with the pulmonary vascular surface area accessible to vascular substrate, the extravascular lung water (an index of the proportion of lung tissue perfused), and the central blood volume (from pulmonary trunk to aorta). Experiments were performed in 10 mongrel dogs at rest and through two increasing levels of treadmill exercise, with the use of two vascular space tracers (labeled erythrocytes and albumin), a water space tracer ([1,8-14C]-octanediol), and a vascular endothelium surface area marker, benzoyl-Phe-Gly-Pro ([3H]BPGP), which is a pharmacologically inactive angiotensin-converting enzyme substrate. The exercise-induced increase in cardiac output was accompanied by a linear increase in central blood volume, and dilutional extravascular lung water rapidly increased to an asymptotic proportion close to 100% of postmortem vascular lung water. There was an average 55% [3H]BPGP hydrolysis, which did not vary with flow, and the computed Amax/Km increased linearly with exercise. We conclude that exercise results in complete lung tissue recruitment and increases the pulmonary vascular surface area available for BPGP hydrolysis linearly with flow, so that pulmonary vascular recruitment continues after full tissue recruitment.

Amino Acid Sequence↗
PubMed1992

Influence of vascular smooth muscle heterogeneity on angiotensin converting enzyme activity in chicken embryonic aorta and in endothelial cells in culture.

The smooth muscle of the abdominal region of the chicken aorta derives from locally recruited mesenchyme (mesenchymal smooth muscle), whereas that of the thoracic region derives from the neural crest (ectomesenchymal smooth muscle). We hypothesized that this smooth muscle heterogeneity might affect important enzymatic functions of the vessel wall. Therefore, we measured angiotensin converting enzyme (ACE) activity in homogenates of chicken thoracic and abdominal aorta at different embryonic stages (days 10, 14, and 18 of gestation). ACE activity increased in both regions over the time of gestation (p less than 0.001 in both cases); the increase was steeper and ACE activity was higher in thoracic than in abdominal segments (p less than 0.001). Km values were similar (approximately 7 microM) at all times and between the two segments, whereas changes in Vmax values closely paralleled those in enzyme activity, indicating gestation-dependent increases in the amount of enzyme. Neural crest ablation at an early developmental stage resulted in an increase of ACE activity in thoracic homogenates (p less than 0.001), predictably leaving that in abdominal homogenates unaffected. Bovine pulmonary artery endothelial cell monolayers exposed to media conditioned with cultured mesenchymal or ectomesenchymal smooth muscle cells exhibited elevated ACE activity (46% and 83%, respectively, relative to control medium, with p less than 0.01 in both cases; p less than 0.05 between the two media). Increases in endothelial cell ACE activity corresponded to proportional increases in ACE protein determined by enzyme-linked immunosorbent assay (r = 0.99) and were interpreted as indicative of enhanced enzyme synthesis subsequent to exposure of endothelial cells to smooth muscle-conditioned media.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

The Patient Intensity for Nursing Index: a validity assessment.

The psychometric assessment of a new measure of nursing intensity, The Patient Intensity for Nursing Index (PINI) is reported. Four dimensions of nursing intensity are measured by the PINI: (a) severity of illness, (b) dependency, (c) complexity of care, and (d) time. There are 10 items rated on a 5-point ordinal scale. A purposively drawn sample of 6,445 patients was obtained from five hospitals, with 487 registered nurses (RNs) providing daily patient ratings from 29 medical-surgical and intensive care units. Substantial interrater reliability was demonstrated. Validity testing included confirmatory factor analysis, hypothesis-testing, contrasted groups, and an observational study of time. The factor structure confirmed specified components of severity, dependency, and complexity, with time loading on all three factors. PINI scores were significantly related to medical severity of illness, length of hospital stay, disposition at discharge, number of secondary medical diagnoses and specialty consults, and scores on three different hospital classification systems used for staffing. PINI scores were significantly different for high- and low-intensity DRGs. Observed time spent delivering nursing care to specific patients was significantly correlated with nurse estimates of time.

Dependency, Psychological↗
PubMed1991

Changing how nurses spend their time.

The results of work sampling studies are used to examine how nurses spend their time and to relate nurses' time to the shortage of nursing practice in hospitals. Four types of proposals for improving the delivery of nursing care in hospitals are discussed. The likely impact of these proposals on how nurses spend their time and the consequences of increasing the effectiveness of professional nursing practice are explored.

Career Mobility↗
PubMed1991

Species variation in pulmonary endothelial aminopeptidase P activity.

Pulmonary endothelial aminopeptidase P (AmP) may be an important contributor to the inactivation of circulating bradykinin in certain species. To examine this possibility, we measured AMP activity in vivo and in vitro using Arg-Pro-Pro-[3H]benzylamide (3H-APPB) as substrate under conditions of first order enzyme kinetics. Utilizing multiple indicator dilution techniques, metabolism of 3H-APPB to Arg and Pro-Pro-[3H]benzylamide by AmP was not detectable during a single transpulmonary passage in anesthetized rabbits (n = 4), cats (n = 3) and pigs (n = 4). However, percent metabolism of 3H-APPB ranged from 54 to 63% in anesthetized rats (n = 6). In all experiments, the substrate remained within the vascular space and was thus accessible to endothelial and blood AmP only. At the same time, single-pass transpulmonary percent metabolism of [14C]benzoyl-Ala-Gly-Pro by endothelial-bound angiotensin converting enzyme was remarkably similar among rabbits, cats, rats and pigs (60-65%). In culture, Vmax/Km of AmP was 3 to 10 x 10(-4) min-1 for human basal arterial and rabbit and bovine pulmonary arterial endothelial cell monolayers (2 x 10(5) cells). AmP activity in the supernatant of lung and kidney tissue (homogenized in saline containing 1-o-n-octyl-beta-glucopyranoside) from rabbit, cat, pig and rat expressed as Vmax/Km(min-1) per (g wet tissue/ml) was 0.74, 2.25, 3.91 and 185.8 (lung), and 1.0, 3.7, 8.4 and 438.3 (kidney), respectively. Similarly, Vmax/Km values of AmP in plasmas of cat, dog, rabbit, pig, calf (serum), human and rat were 0, 0.016, 0.025, 0.068, 0.191, 0.237 and 3.53 min-1. These results suggest that 1) there are large interspecies variations in AmP activities of plasma, lung and kidney; 2) of the species studied, the rat contains the largest activities of AmP; and 3) AmP appears to be located on the luminal surface of the rat pulmonary endothelium.

Aminopeptidases↗
PubMed1991

Nuclear imaging analysis of human low-density lipoprotein biodistribution in rabbits and monkeys.

We have evaluated the biodistribution of human low-density lipoprotein (LDL) radiolabeled with 99mTc or with 123I-tyramine cellobiose in rabbits and in rhesus monkeys. Biodistribution was assessed after intravenous injection of radiolabeled LDL by quantitative analysis of scintigrams, counting of excreta, and counting of tissues at necropsy. Both rabbits and monkeys showed lower renal uptake (123I:99mTc approximately 1:3, as regional percent injected activity corrected for physical decay) and excretion (1:2 to 1:4), but higher hepatic (1.5:1 to 2:1) and cardiac (1.7:1 to 4:1) uptake of 123I than of 99mTc. Adrenals were visualized in normolipemic animals with 123I-tyramine cellobiose-LDL but not with 99mTc-LDL. Hyperlipemic animals showed increased cardiac (up to six-fold) and decreased hepatic activity (by 50%-60%) of both radionuclides. We conclude that 123I-tyramine cellobiose-LDL is better suited than 99mTc-LDL for dynamic studies of LDL metabolism in vivo.

Animals↗
PubMed1990

Detection and assessment of severity of tricuspid regurgitation using first-pass radionuclide angiography and comparison with pulsed Doppler echocardiography.

Noninvasive detection and semiquantitative assessment of tricuspid regurgitation (TR) were performed using first-pass radionuclide angiography (RNA), by sampling a region of interest over the right atrium for any tracer entering the right atrium during right ventricular systole. The relative amount of tracer entering the right atrium was quantitated and the results were compared with semiquantitative Doppler echocardiographic grading of TR severity. Using the right ventricular time-activity curve to define end-diastolic and end-systolic frames, the right atrial counts for the 2 or 3 cardiac cycles after the peak right ventricular counts were summed. The right atrial "injection fraction" was calculated using the following formula: [(end-systolic counts - end-diastolic counts)/(end-diastolic counts)] X 100%. The right atrial injection fraction was examined in 51 patients who had good quality RNA and Doppler studies. Of 27 patients with no evidence of TR by Doppler, 26 had a negative right atrial injection fraction. All 24 patients with a positive Doppler for TR had a positive right atrial injection fraction. Comparison of right atrial injection fraction grade ranges with semiquantitative grades of TR severity on Doppler revealed identical grades in 21 of the 24, with a single grade difference in the remaining 3 patients. Thus, right atrial time-activity curve quantitation during routine first-pass RNA allows detection and grading of the severity of TR, with results very similar to pulsed Doppler echocardiography. This simple procedure is easily appended to the evaluation of ventricular performance with first-pass RNA.

Adolescent↗
PubMed1990

The process of rater training for observational instruments: implications for interrater reliability.

Although the process of rater training is important for establishing interrater reliability of observational instruments, there is little information available in current literature to guide the researcher. In this article, principles and procedures that can be used when rater performance is a critical element of reliability assessment are described. Three phases of the process of rater training are presented: (a) training raters to use the instrument; (b) evaluating rater performance at the end of training; and (c) determining the extent to which rater training is maintained during a reliability study. An example is presented to illustrate how these phases were incorporated in a study to examine the reliability of a measure of patient intensity called the Patient Intensity for Nursing Index (PINI).

Employee Performance Appraisal↗
PubMed1990

Inhibition of endothelial-bound angiotensin converting enzyme, in vivo.

1. We determined apparent Ki constants of two inhibitors, captopril and CL242,817, for pulmonary endothelial-bound angiotensin converting enzyme (ACE) in anaesthetized rabbits. [3H]-benzoyl-Phe-Ala-Pro was used as the substrate. The apparent kinetic parameters Km and Amax (product of Vmax and microvascular plasma volume) were measured, as was the ratio (Amax/Km) (measured under first order reaction conditions) before and 30s after the i.v. administration of captopril 10 nmol kg-1 or CL242,817, 35 nmol kg-1. 2. Under mixed order reaction conditions, ([S] greater than or equal to Km), apparent Km values increased from 12.2 +/- 1.9 microM to 32.9 +/- 3.3 microM (P less than 0.05) in the captopril-treated rabbits and from 9.3 +/- 2.3 microM to 45.8 +/- 9.8 microM (P less than 0.05) in the CL242,817-treated rabbits, indicative of competitive inhibition. However, apparent Amax values decreased from 10.3 +/- 2.1 to 4.5 +/- 0.8 mumol min-1 (P less than 0.05) and 8.9 +/- 1.7 to 4.8 +/- 0.5 mumol min-1 (P less than 0.05), respectively. 3. Under first order reaction conditions ([S] much less than Km), the Amax/Km ratio decreased from 763 +/- 100 to 125 +/- 38 ml min-1 (P less than 0.05) and 1009 +/- 149 to 126 +/- 44 ml min-1 (P less than 0.05) in the captopril- and CL242,817-treated groups respectively. 4. When the single pass transpulmonary binding of 80pmol [3H]-RAC-X-65 (an ACE inhibitor) was measured in additional rabbits, a significant (P < 0.05) decrease in RAC-X-65 binding was observed 30s after captopril (80% decrease) or CL242,817 (85% decrease), a result expected for a loss of catalytically active enzyme mass due to tightly bound captopril or CL242,817. 5. These results indicate that, in vivo, both captopril and CL242,817 are competitive, tight binding inhibitors of lung ACE. Furthermore, they suggest means for evaluating the interaction of other potential ACE inhibitors with the pulmonary endothelial membrane-bound enzyme, in vivo, possibly in phase I clinical trials.

5'-Nucleotidase↗
PubMed1989

Efficacy of computed tomography of the thorax and upper abdomen and whole-body gallium scintigraphy for staging of lung cancer.

To assess the efficacy of performing both computed tomography (CT) of the thorax and upper abdomen and whole-body gallium scintigraphy for staging lung cancer, the results of each test were compared with those obtained by chest radiography and clinical examination in 100 patients. Clinical efficacy was defined in terms of accuracy in staging the tumor based either on surgery, biopsy, or clinical and radiologic follow-up. The CT provided significantly superior accuracy in 27 patients and minor additional staging information in 17 patients compared with the gallium scan. Whole-body gallium scintigraphy provided important additional information in nine patients and minor additional information in a further eight. The diagnostic yield of CT and gallium scanning was considered equivalent in 39 cases. Of the nine cases in which gallium was significantly superior to CT, clinical findings which suggested the presence of metastases had been noted before the scan in four cases. The authors' results confirm the utility of CT for staging lung cancer and indicate that the additional yield from gallium scintigraphy is relatively low provided a thorough history and physical examination have been performed.

Adult↗
PubMed1989

Localization of angiotensin-converting enzyme in the trout gill.

Angiotensin-converting enzyme (ACE) was localized in perfused trout gills by measuring gill extraction of two radiolabeled ACE inhibitors, 125I-351A (an iodinated derivative of lisinopril) and 3H-RAC-X-65, and by autoradiography of gills perfused with 125I-351A. A 125I-351A pulse was preferentially extracted by the arterio-arterial (AA) pathway (61.7% +/- 1.8% extraction; mean +/- SE, N = 4); the arteriovenous (AV) pathway extracted an additional 10%. Extraction by either pathway was reduced by simultaneous perfusion with 10(-5) M unlabeled lisinopril. AA extraction of RAC-X-65 during continuous perfusion was maximal (75% +/- 5%, N = 6) during the first few minutes of perfusion and decreased steadily to 38% +/- 9% by 20 min and to less than 10% by 40 min. AV extraction of RAC-X-65 was negligible. Autoradiography of gills continuously perfused with 125I-351A showed that the radiolabel was concentrated in the respiratory lamellae. The highest grain density was associated with the pillar cells nearest the medial (inner) lamellar margin. Afferent filamental arteries and afferent lamellar arterioles were labeled to a lesser extent. Relatively little label was found on the efferent lamellar arterioles or efferent filamental arteries. 125I-351A binding was not evident in AV vessels. These findings support the hypothesis that the gill is an important site for formation of plasma angiotensin II and they suggest that enzymes associated with mammalian endothelial cells are also common to gill pillar cells.

Animals↗
PubMed1989

Planar positron imaging of rubidium-82 for myocardial infarction: a comparison with thallium-201 and regional wall motion.

Rubidium-82 (Rb-82) is a generator-produced, short half-life (76 seconds) positron emitting potassium analog. Using a mobile gamma camera equipped with a rotating tungsten collimator and high-energy shielding, we examined the use of Rb-82 in the coronary care unit and clinical laboratory for detection of perfusion defects due to myocardial infarction. We studied 31 subjects, 10 patients with acute myocardial infarction, 12 with remote myocardial infarction, and nine controls. Rb-82 images were compared with Tl-201 and regional wall motion for detection of infarct-related arteries. Of the 22 patients with myocardial infarction, 16 were identified with Rb-82 and Tl-201. In nine control subjects, eight were normal with each method. Correlation between Rb-82 and Tl-201 defect scores was excellent. Sensitivity and specificity for infarct-related arteries were similar for Rb-82, Tl-201, and wall motion imaging. Thus planar Rb-82 imaging can detect MI reliably in the coronary care unit and in the clinical laboratory.

Coronary Care Units↗
PubMed1989

Peptidase enzymes of the pulmonary vascular surface.

No fewer than five peptidase enzymes are disposed on the pulmonary vascular surface and are capable of reacting with peptides (including peptide hormones and pro-hormones) within the intravascular space. The review that follows describes these enzymes and their actions and suggests how the enzymes may be assayed in vivo to improve understanding of pulmonary microvascular functions as well as aspects of endocrinology.

Amino Acid Sequence↗
PubMed1989

Difficulties of parathyroidectomy after previous thyroidectomy.

Although the risks of reoperative thyroidectomy and parathyroidectomy have been well studied, the problems associated with parathyroidectomy after prior thyroidectomy have not been emphasized. Among a group of 282 patients who were treated for primary hyperparathyroidism in recent years at the University of Chicago Medical Center, 14 (4.8%) had undergone one or more previous thyroidectomies, and 6 others (2.1%) had undergone thyroid ablation with radioactive iodine as therapy for Graves' disease. Numerous difficulties were encountered during surgery in the postthyroidectomy group of patients as a result of scarring and fibrosis, prior recurrent laryngeal nerve injuries in 13%, the inability to known with certainty how many viable, normal parathyroid glands remained after previous operations, and the need for additional thyroid resection, mostly for associated malignant lesions. Preoperative vocal cord assessment, evaluation of prior operative and pathology reports, and localization studies with thallium-technetium scanning and ultrasonographic techniques were especially helpful. A "lateral approach" was used frequently during surgery. Each of these 14 patients was cured of the hyperparathyroidism. The postthyroid ablation group presented fewer intraoperative challenges, although in some patients the thyroid gland was virtually absent, which obscured the normal landmarks of the surgical field. Five of these six patients were cured of hyperparathyroidism. Parathyroidectomy after thyroidectomy presents many operative challenges to the surgeon and should be approached with the same care and concern that one reserves for a reoperative parathyroid operation.

Adult↗