Diagnostic value of gas exchange tests in patients with clinical suspicion of pulmonary embolism.

OBJECTIVE: To assess the value of parameters derived from arterial blood gas tests in the diagnosis of pulmonary embolism. METHOD: We measured alveolar-arterial partial pressure of oxygen [P(A-a)O2] gradient, PaO2 and arterial partial pressure of carbon diaxide (PaCO2) in 773 consecutive patients with suspected pulmonary embolism who were enrolled in the Prospective Investigative Study of Acute Pulmonary Embolism. DIAGNOSIS: The study design required pulmonary angiography in all patients with abnormal perfusion scans. RESULTS: Of 773 scans, 270 were classified as normal/near-normal and 503 as abnormal. Pulmonary embolism was diagnosed by pulmonary angiography in 312 of 503 patients with abnormal scans. Of 312 patients with pulmonary embolism, 12, 14 and 35% had normal P(A-a)O2, PaO2 and PaCO2, respectively. Of 191 patients with abnormal scans and negative angiograms, 11, 13 and 55% had normal P(A-a)O2, PaO2 and PaCO2, respectively. The proportions of patients with normal/near-normal scans who had normal P(A-a)O2, PaO2 and PaCO2 were 20, 25 and 37%, respectively. No differences were observed in the mean values of arterial blood gas data between patients with pulmonary embolism and those who had abnormal scans and negative angiograms. Among the 773 patients with suspected pulmonary embolism, 364 (47%) had prior cardiopulmonary disease. Pulmonary embolism was diagnosed in 151 (41%) of 364 patients with prior cardiopulmonary disease, and in 161 (39%) of 409 patients without prior cardiopulmonary disease. Among patients with pulmonary embolism, there was no difference in arterial blood gas data between patients with and those without prior CPD. CONCLUSION: These data indicate that arterial blood gas tests are of limited value in the diagnostic work-up of pulmonary embolism if they are not interpreted in conjunction with clinical and other laboratory tests.

Mitochondria act as a reservoir for the basic amine HIPDM in the lung.

The lungs are a site for the uptake, accumulation, and storage of exogenous basic amines. The compound N-N-N'-trimethyl-N'-(2-hydroxy-3-methyl-5-iodobenzyl)-1,3 propanediamine (HIPDM) is a basic amine that can be labelled with radioactive iodine and detected by external counting. Intravenously injected 123I-HIPDM is extracted by the human lung, where it is retained in a slowly effluxable pool. In the present study, we measured HIPDM lung kinetics and subcellular distribution in rabbits given i.v. 125I-HIPDM. Rabbits were killed from 2 min to 5 h after injection, and the radioactivity retained in their lungs was measured. Subcellular lung fractions (nuclear, mitochondrial, lysosomal, microsomal and postmicrosomal supernatant) were assayed for HIPDM radioactivity, protein content, and distribution of specific marker enzymes. HIPDM lung clearance in rabbits was nearly identical to that of humans. Virtually all the HIPDM radioactivity in lungs (98+/-1%) was associated with subcellular membranous structures. The highest HIPDM specific radioactivity was found in the mitochondrial fraction, and the subcellular distribution profile closely resembled that of the mitochondrial marker enzyme succinate cytochrome c reductase. No redistribution of HIPDM among subcellular compartments was observed over a 5 h period after injection. The data indicate that mitochondria act as reservoir for HIPDM in the lungs and contribute to the pulmonary persistence of this compound. HIPDM can be used to investigate the pulmonary uptake of basic amines in health and in lung disease.

Lazaroid U-74389F attenuates phorbol ester-induced lung injury in rabbits.

We examined the effect of the antioxidant lazaroid U-74389F on acute lung injury induced in rabbits by phorbol myristate acetate (PMA). Thirty minutes after receiving either U-74389F (15 mg.kg-1 i.v.) or U-74389F vehicle, rabbits (n = 60) were given PMA (60 micrograms.kg-1 i.v.). PMA vehicle injected rabbits (n = 20) served as controls. Over a 5 h period after PMA or PMA vehicle injection, we measured arterial pH, arterial oxygen tension (Pa,O2), arterial carbon dioxide tension (Pa,CO2), and the plasma concentration of the neutrophil chemoattractant interleukin-8 (IL-8). At postmortem, lungs were inspected for macroscopic injury and examined histologically. Malondialdehyde levels were assayed in lung tissue as an index of lipid peroxidation. In bronchoalveolar lavage (BAL), total and differential cell counts, protein and IL-8 concentrations were measured. Compared to normal controls, rabbits challenged with PMA alone developed arterial acidosis, hypercapnia and hypoxaemia, accompanied by significant rise in plasma IL-8 concentration. U-74389F pretreated animals did not develop significant arterial blood gas abnormalities and had significantly lower IL-8 concentration in plasma. U-74389F did not prevent PMA-induced lipid peroxidation. However, macroscopic signs of lung injury and the degree of alveolar haemorrhage and protein extravasation were significantly less severe in pretreated rabbits than in those given PMA alone. In addition, U-74389F significantly reduced IL-8 concentration and neutrophil number in BAL. By histological assessment, 80% of lung neutrophils were localized in alveolar spaces of animals receiving PMA alone. Conversely, in U-74389F pretreated animals, 75% of neutrophils were distributed within extra-alveolar blood vessels and alveolar septa. We conclude that lazaroid U-74389F attenuates lung injury in rabbits given PMA by preventing neutrophil migration into pulmonary alveoli. This effect may, in part, be related to downregulation of IL-8 production.

Measurement of ATPases in red cells: setting up and validation of a highly reproducible method.

Our aim was to set up and validate a reproducible method to study ATPase family on erythrocyte membranes. We compared several methods for erythrocyte washing and hemolysis and succeeded in preparing completely hemoglobin-free membrane ghosts still bearing intact ATPases. We compared the conventional incubation procedure with the coupled enzyme assay to measure Na-K-Mg, Ca-Mg and Mg ATPase on the membranes. A significant difference was constantly observed between the results by these methods, the values by the incubation procedure being 28, 57 and 58% of the respective values obtained by the linked enzyme assay. By adopting this last one, we obtained uniform and reproducible results in 31 healthy subjects. The following activities of the measured pumps resulted: Na-K-Mg ATPase 0.026 +/- 0.007, mean +/- SD; Ca-Mg ATPase 0.030 +/- 0.010, and Mg ATPase 0.017 +/- 0.003 U/mg protein, respectively. Finally, we investigated the effect of membrane storage time and temperature on ATPase results.

Radioreceptor assay for the measurement of endogenous digitalis-like factors using membrane particulate fraction of human placenta and 125I-digoxin: comparison with RIA method.

We measured purified extracts of serum (plasma) or urine samples of newborns, pregnant women, normal adults, and uremic patients by a radioreceptor assay (RRA), which uses particulate membrane fractions from human placenta as a binding system, and 125I-digoxin as a tracer. We also measured the digoxin-like immunoreactivity by a sensitive RIA, and results were compared with those found by the RRA. Specific 125I-digoxin binding to placental receptors was competitively inhibited by purified plasma and/or urine extracts of newborns, adult subjects, pregnant women and uremic patients. A linear relationship was found between inhibition of binding and volume of plasma and urine assayed. Moreover, a significant correlation was found between the values obtained by RRA and those found by RIA (n = 17, r = 0.699, p = 0.0012). Our data confirm that increased circulating and/or urinary levels of substances with biological and immunological activity similar to cardiac glycoside drugs are present in newborns, pregnant women and uremic patients compared to healthy adult subjects. In addition, our preliminary study indicates that these endogenous factors are able to bind to the specific receptor of digitalis drugs on the placental membranes.

Specific binding of cardiac glycoside drugs and endogenous digitalis-like substances to particulate membrane fractions from human placenta.

We studied the characteristics of binding of cardiac glycosides to particulate membrane fractions from human placenta, to demonstrate that placental tissue is a suitable source of receptors for digitalis drugs. Moreover, we performed preliminary experiments with 125I-labeled digoxin and placental particulates to develop a radioreceptor assay for measurement of endogenous substances with activity similar to cardiac glycoside drugs (EDLS). Placental membrane fractions were incubated with [3H]ouabain (10 nmol/L) or 125I-labeled digoxin (50 pmol/L). With both ligands, binding followed a pseudo-first-order reaction kinetics and was saturable. Scatchard analysis revealed a single class of sites [for ouabain, KD = 20.2 +/- 5.8 nmol/L (mean +/- SEM), Bmax = 3.1 +/- 0.9 nmol per gram of protein; for digoxin, KD = 29.7 +/- 1.9 nmol/L, Bmax = 24.3 +/- 1.1 nmol per gram of protein]. As expected, digoxin was less potent than ouabain in displacing both tracers from digitalis drugs receptors; progesterone, cortisone, digitoxose, furosemide, bumetanide, and propranolol had no or little effect. Specific 125I-labeled digoxin binding was competitively inhibited by plasma and (or) urine extracts from newborns, adults, pregnant women, and patients with renal insufficiency. Inhibition of binding and volume of plasma and urine assayed were linearly related. These findings support the hypothesis that cardiac glycosides and EDLS can interact with the human placenta and suggest placental tissue to be a suitable source of receptors for cardiac glycosides.

Binding of 14C-furosemide to isolated human erythrocytes.

The incubation of 14C-furosemide at high specific activity with intact red blood cells at 37 degrees C, pH 7.4, has enabled the furosemide binding sites to be characterized with respect to time course, affinity and specificity. The binding reaction was rapid, reversible and close to thermodynamic equilibrium. Binding was dependent on cell and furosemide concentration and was saturable. At equilibrium, pharmacological doses of furosemide competitively inhibited 14C-furosemide binding with 50% inhibition at 3 x 10(-5) M. The Na+/K+ pump inhibitor ouabain had no effect on the 14C-furosemide binding. Bumetanide, which is more potent than furosemide as inhibitor of Na+/K+ co-transport system and equally effective in inhibiting anion transport, was less effective than furosemide in displacing 14C-furosemide from its binding sites, suggesting a different mechanism of action for the two drugs in the red blood cell. The preincubation of erythrocytes with 4,4'-diisothiocyano-stilbene-2,2'-disulphonic acid (DIDS), the potent and specific inhibitor of anion permeability, reduced specific furosemide binding by more than 80% at a furosemide concentration of 0.1 microM, while it had little effect on the non-specific furosemide binding. Taken together, these data suggest that furosemide interacts with specific binding sites in the human red blood cell, whose nature has not been clarified, but whose location is probably on (or near) the protein in band 3, i.e. the membrane macromolecule-mediating anion transport.

Lung tissue binding of iodobenzyl-propanediamine: involvement of beta-adrenergic receptors.

The basic compound N-N-N'-trimethyl-N'-(2-hydroxy-3-methyl-5-iodobenzyl)-1,3- propanediamine (HIPDM) accumulates in human and rabbit lungs, where it forms a slowly effluxable pool. In isolated perfused rat lung, HIPDM is taken up by a saturable, energy-independent mechanism, which is competitively inhibited by imipramine, chlorpromazine and propranolol. To ascertain whether beta-adrenergic receptors are involved in the binding process of HIPDM to lung tissue, the ability of unlabelled HIPDM to displace the beta-adrenergic receptor ligand [125I]iodocyanopindolol (ICYP) from rabbit lung beta-receptors was examined. Lung microsomal membrane fractions (75 micrograms ml-1) were incubated at 37 degrees C for 3 h with 68 pM ICYP (with or without 1 microM of (+/-)-propranolol) in the presence of HIPDM (10(-10)-10(-3) M). Bound and free radioactivity were separated through glass-fibre filters and the retained radioactivity was counted in a gamma-spectrometer. HIPDM competed with ICYP for beta-adrenoceptors (13% displacement at 10(-5) M. 50% at 5 x 10(-5) M, and 90% at 2 x 10(-4) M). The inhibition curve of ICYP binding by HIPDM was similar to that observed for (-)-noradrenaline. Although the results of the in vitro studies cannot be extrapolated to in vivo conditions, they suggest that beta-adrenergic receptors may be involved in the observed lung uptake of the basic amine HIPDM.

Demonstration of beta 1-adrenergic receptors in human placenta by (-)[125I]iodocyanopindolol binding.

The highly specific beta-adrenergic radioligand (-)[125I]iodocyanopindolol (ICYP) was used to characterize the beta-adrenergic receptor subtype present in human placenta. Binding of ICYP to membranes from human placenta was saturable with time and ligand concentration, of high affinity, and demonstrated appropriate stereoselectivity and agonist rank order of potency for binding to a beta-adrenergic receptor. From saturation binding curves, the KD and Bmax values for ICYP binding were 233 +/- 51 pM and 690 +/- 139 fmol/mg of proteins, respectively. Analysis of inhibition of ICYP binding by beta 1- and beta 2-selective adrenergic antagonists via Hofstee analysis resulted in linear plots, indicating the existence of a homogeneous population of beta-adrenergic receptors. From the resulting KI-values for the beta 1-selective drugs practolol (4.0 +/- 0.9 microM) and metoprolol (0.19 +/- 0.07 microM) and for the beta 2-selective drug ICI 118,551 (0.30 +/- 0.06 microM) it is concluded that the beta-adrenergic receptor in human placenta is of the beta 1-subtype. This is further supported by the fact that (-)-noradrenaline and (-)-adrenaline were equipotent in inhibiting ICYP binding.

Endogenous cardiac glycoside-like substances in newborns, adults, pregnant women and patients with hypertension or renal insufficiency.

In an attempt to confirm the presence of endogenous substances with cardiac glycoside-like activity, the biological and immunological cardiac glycoside-like activity was measured by a sensitive solid-phase radioimmunological assay (RIA), two radioreceptor assays (RRA), and a 86Rb uptake method in normal subjects and in some pathophysiological conditions characterized by sodium retention and volume expansion. Significant concentrations of digoxin-like immunoreactive substances (DLIS) were measured in plasma (or serum) of normal subjects while significantly higher levels were found in pregnant women, newborns and in patients with renal impairment, and in some with essential hypertension. Concentrations in urine of normal adults or newborns were several times higher than in plasma. The results obtained by RIA correlated with those obtained by RRA and 86Rb uptake methods. In 88 normal subjects, DLIS excretion rates (overnight urine collection) in men were significantly higher than in women (68.6 +/- 23.6 pg/min vs 50.9 +/- 21.0 pg/min, p less than 0.01). The DLIS excretion rates correlated with creatinine, Na and K urinary excretion rates, and also with the subjects' body weight, height, body mass index, and systolic blood pressure. These findings confirm the presence of endogenous substances with immunological and biological activity similar to cardiac glycosides in human body fluids and also confirm the hypothesis that these endogenous factors may be involved in fluid and electrolyte regulation in man. In addition, the present data indicate that urinary excretion of DLIS is dependent on body mass and renal glomerular filtration.

Arterial blood pressure and the renin-angiotensin-aldosterone system during postural changes in hypertensive patients with unilateral renal mobility.

Unilateral renal mobility was identified in 27 out of 100 essential hypertensive patients by examination of renal scintiphotos. The pattern of response to postural changes of blood pressure (BP), plasma renin activity (PRA) and plasma aldosterone concentration (PAC) was investigated in 11 patients with renal mobility and without treatment and compared with that of an age- and sex-matched group of untreated hypertensives without renal mobility. The patients with renal mobility had higher BP levels (X +/- SD mm Hg: supine 185 +/- 39/112 +/- 18 vs. 149 +/- 18/97 +/- 14; upright 167 +/- 38/108 +/- 17 vs. 144 +/- 7/93 +/- 10; p less than 0.05). Significant correlations were obtained in the patients with renal mobility (but not in those without renal mobility) between upright PRA and PAC (p less than 0.001), their postural variations (p less than 0.01) and between upright PRA (and PAC) and BP levels (p less than 0.05). The high prevalence of renal mobility in hypertension and the relationship observed between the activated renin-angiotensin-aldosterone system and BP in this condition suggest the importance of searching for unilateral renal mobility when examining the renin-angiotensin-aldosterone system in hypertensive patients, particularly during postural manoeuvres.