Detection and identification of endothelin-1 immunoreactivity in rat and porcine thyroid follicular cells.

Endothelin-1 immunoreactivity (irET-1) was observed in rat and porcine thyroid glands. Using a radioimmunoassay for endothelin-1, the mean concentration in extracts of rat and porcine thyroid glands were 0.75 pg/mg +/- 0.03 (n = 4) and 1.5 pg/mg +/- 0.2 (n = 8) (mean +/- SE) respectively. Gel-filtration and reverse-phase HPLC showed that ir ET-1 eluted in a position identical to synthetic endothelin-1. In addition, immunohistochemical study showed that irET-1 is located within epithelial follicular cells. No immunostaining was seen in parafollicular C-cells nor in parathyroid.

Cardiovascular and endocrine effects of endothelin-1 at pathophysiological and pharmacological plasma concentrations in conscious dogs.

BACKGROUND: Increased plasma concentrations of endothelin-1, a potent vasoconstrictor produced by the endothelium, have been reported in various pathological conditions. This study was conducted to evaluate effects of endothelin-1 at pathophysiological and pharmacological plasma concentrations. METHODS AND RESULTS: Endothelin-1 was infused at increasing doses (2.5, 5, 10, and 20 ng/kg.min for 1 hour each) in nine conscious dogs. During endothelin-1 infusion, plasma endothelin-1 rose from a basal value of 1.8 +/- 0.4 pmol/l to 5.8 +/- 1.1 (pathophysiological), 20.8 +/- 3.9 (pathophysiological), 85.4 +/- 18.9 (pharmacological), and 311.4 +/- 55.7 (pharmacological) pmol/l at each dose, respectively. Heart rate increased at 2.5 ng/kg.min (from 129 +/- 7 to 146 +/- 12 beats/min) but decreased at 20 ng/kg.min (97 +/- 7 beats/min) (p less than 0.001). Such a biphasic response was also observed for peak (+)dP/dt and (dP/dt)/DP40 (p less than 0.005). Left ventricular systolic pressures, mean aortic pressure, and left atrial pressure increased over time (p less than 0.05, p less than 0.005, and p less than 0.001, respectively). The time constant of early isovolumic relaxation rose progressively (p less than 0.001). The percent systolic shortening decreased at 10 and 20 ng/kg.min (p less than 0.005). Pressure-segment length loops showed a reduction in systolic shortening associated with an increase in left ventricular systolic pressure at 20 ng/kg.min. Atrial natriuretic factor rose after 5 ng/kg.min from 28.5 +/- 6.5 to 92.0 +/- 18.2 pmol/l (p less than 0.005). Angiotensin II and catecholamines did not change significantly. Serum urea and creatinine rose progressively (p less than 0.05), whereas glucose decreased (p less than 0.05). The above results differed significantly from measurements obtained in a time-control group of six dogs. CONCLUSIONS: A fourfold increase of plasma endothelin-1 obtained after doubling the infusion rate suggests a reduction in endothelin-1 clearance or endothelin-1 endogenous production. The biphasic response of heart rate is consistent with baroreflex-mediated effects resulting from vasodilation at the pathophysiological level and vasoconstriction at the pharmacological level. Hemodynamic data suggest an increase followed by a decrease in contractility at both levels, respectively. Finally, endothelin-1 is a stimulator of atrial natriuretic factor.

The effects of glucose ingestion and fasting on plasma immunoreactive beta-endorphin, adrenocorticotropic hormone and cortisol in obese subjects.

It has been demonstrated that opioid peptides are involved in the stimulation of food intake in rats and that the circulating beta-endorphin levels are increased in genetically obese rodents. Therefore, to assess whether the changes in food intake may influence circulating beta-endorphin levels in obese subjects, plasma beta-endorphin, ACTH and cortisol concentrations were determined in obese patients after an oral glucose load and during a 7-day total starvation. Baseline plasma beta-endorphin concentrations were significantly higher in obese patients than in control normal-weight subjects, while ACTH and cortisol levels were similar in both groups. Plasma beta-endorphin, ACTH and cortisol concentrations were not affected by the ingestion of 75 g glucose, neither were plasma beta-endorphin concentrations modified during prolonged starvation. Moreover, the lack of nycthemeral variations in beta-endorphin levels, documented before and during starvation while plasma ACTH and cortisol were significantly reduced in the evening, suggests that some extra anterior pituitary sources or some obesity-related changes in beta-endorphin metabolism may contribute to the pool of circulating beta-endorphin in obese subjects. On the other hand, even the extreme changes in nutritional conditions, such as total food deprivation or glucose ingestion, are devoid of any detectable influence on circulating beta-endorphin levels.

Atrial natriuretic factor, cardiac volumes and filling pressures during exercise in congestive heart failure.

To study the mechanism of atrial natriuretic factor (ANF) release in heart failure, we measured plasma ANF concentrations, cardiac volumes and filling pressures at rest and during three graded exercise levels (E1, E2, E3) in six male patients with congestive heart failure (CHF) and compared them with 13 normal male subjects. At rest, ANF concentrations were sixfold higher in patients with CHF than in normal subjects (at rest: 53 +/- 12 vs 8 +/- 1 pmol.l-1; P less than 0.02). End-systolic ventricular volumes were increased threefold in patients with CHF (P less than 0.02) despite normal mean central venous pressure, pulmonary artery pressure (PAP) and pulmonary wedge pressure (PWP). A positive correlation was found between end-systolic ventricular volumes and plasma ANF (r = 0.93, P less than 0.001). During exercise, ANF rose by 120% over basal values both in patients with CHF and in normal subjects (P less than 0.01). Volumes higher than normal in patients with CHF increased further at E2 (P less than 0.05) in contrast to a decrease of systolic volumes recorded in normal subjects (P less than 0.05). Filling pressures rising abnormally in patients with CHF correlated with plasma ANF during exercise (r = 0.53, P less than 0.02 for PAP; r = 0.51, P less than 0.05 for PWP). In conclusion, our data suggest that ANF release in CHF is regulated at rest by cardiac volumes when filling pressures are still normal. During exercise, ANF release is not impaired in CHF with normal rest filling pressures and is regulated during exercise by left filling pressures.

Plasma atrial natriuretic factor and catecholamines before and during excision of pheochromocytoma.

We studied changes of atrial natriuretic factor (ANF) and catecholamines in three patients with pheochromocytoma occurring in the familial syndrome of multiple endocrine neoplasia type IIa. Previous studies have suggested a stimulating effect of catecholamines on ANF release. In pheochromocytoma, we observed normal basal ANF levels despite increased catecholamine secretion. In contrast, a rise in plasma ANF was observed when a hypertensive paroxysm occurred. Also during surgery, dissection of pheochromocytoma led to a rise in plasma ANF and catecholamines associated with an increase in blood pressure (in the 3 cases) and in pulmonary artery pressure (in 2 cases). We concluded that chronic elevation of basal catecholamines are without effect on plasma ANF but that manipulation of pheochromocytoma leads to a stimulation of ANF release, possibly mediated by either a direct effect of endogenously released catecholamines and/or an increase in atrial pressure.

Atrial natriuretic factor during percutaneous transluminal coronary angioplasty.

To study the release of plasma atrial natriuretic factor (ANF) and to explain the mechanism underlying its increase during myocardial ischemia, we measured plasma ANF and mean pulmonary capillary wedge pressure (PCW) before, during and after percutaneous transluminal coronary angioplasty (PTCA) in eight patients. All patients were free of calcium channel antagonists and beta-blocking drugs. Evidence of myocardial ischemia was observed in all patients with an increase of PCW from 3.2 +/- 1.2 to 10.6 +/- 2.9 mm Hg (mean +/- SD; p less than 0.001). Heart rate and mean blood pressure did not change significantly. We observed an increase of plasma ANF during PTCA, from 53 +/- 24 to 100 +/- 37 pmol/L (mean +/- SD; p less than 0.005). There was a correlation between absolute values of ANF and PCW before and during PTCA (r = 0.64, p less than 0.01). After PTCA, ANF levels remained increased for at least twenty minutes (p less than 0.005 vs basal state) despite a decrease in PCW. Thus, increase of PCW during this very short-term left ventricular ischemic dysfunction induces an increase of plasma ANF, which persists during a certain time when PCW returns to normal.

Relationship between natriuresis and changes in plasma atrial natriuretic factor, renin activity and aldosterone levels in fasting obese subjects.

This study was conducted to assess whether changes in atrial natriuretic factor (ANF) secretion could account for the natriuresis of the early phase of fasting. To this end, 8 AM (supine) and 10 AM (standing) plasma ANF concentrations were determined daily and compared with plasma renin activity and aldosterone levels in 8 obese subjects submitted to a 7-day total fast. Depiste constant daily sodium intake (51 mmol), urinary sodium excretion increased from 35 +/- 7 to 109 +/- 8 mmol/day after 4 days of fast (p less than 0.001) and declined thereafter. Urinary ketone excretion progressively increased over the whole period of fasting (p less than 0.001). Interestingly, fasting induced a decrease in plasma ANF concentrations (p less than 0.05). A contrast analysis revealed no significant change in ANF during the initial natriuretic phase of fasting but a decrease at the end of fasting averaging 36% (p less than 0.05) and 18% (p less than 0.05) at 8 and 10 AM respectively. In contrast, plasma aldosterone rose during fasting (p less than 0.05), the difference being significant at the end of fasting (p less than 0.01). Plasma renin activity and cortisol did not change significantly over the fasting period. Postural and/or diurnal changes of ANF, aldosterone, renin and cortisol were preserved during fasting (p less than 0.01). Postural changes of ANF were, however, attenuated at the end of fasting (p less than 0.05). These data indicate that the fasting natriuresis cannot be explained by changes in ANF levels but that the loss of sodium may contribute to a decline of basal ANF levels, with an attenuation of their physiological postural changes, and to a stimulation of the aldosterone secretion.

Exercise and posture-related changes of atrial natriuretic factor and cardiac function in diabetes.

To study whether the release of atrial natriuretic factor (ANF) was altered in diabetic cardiac autonomic neuropathy (CAN), we determined plasma ANF concentrations during exercise and changes of posture in three groups of age- and sex-matched subjects (9 healthy subjects, 7 diabetic patients with CAN, and 7 diabetic patients without CAN). During exercise, plasma ANF concentrations rose threefold (P less than .001), and this increase was similar in the three groups. However, heart-rate response to exercise was impaired in the two groups of diabetic patients (P less than .004 vs. healthy subjects) but was more severely impaired in patients with CAN (P less than .03 vs. patients without CAN). In healthy subjects and patients without CAN, the increases of ANF during exercise correlated significantly with those of heart rate, systolic blood pressure, and rate-pressure product (P less than .01). In patients with CAN, the correlation was found exclusively with heart rate (P less than .01). An increase of ventricular ejection fraction occurred in all groups (P less than .001) but without showing statistical differences between groups. After 30 min of standing, a similar postural drop of plasma ANF concentrations (P less than .002) was observed in all subjects, reflecting preserved sympathetic control of vessels. In conclusion, exercise induces an increase of plasma ANF in diabetic patients with CAN. This increase, occurring similarly to healthy subjects, indicates that autonomic activation plays a minor role in ANF release during exercise. Impaired heart-rate response to exercise in patients without CAN suggests early damage of autonomic function, undetected by conventional rest tests.

Evidence that atrial natriuretic factor is the humoral factor by which volume loading or mannitol infusion produces an improved renal function after acute ischemia. An experimental study in dogs.

This experimental study in dogs was designed to investigate whether maximal loading produces atrial natriuretic factor (ANF) release and whether this physiological peptide is involved in the improvement of the early renal function recovery after acute ischemia. The experimental protocol included a renal artery occlusion for 45 min in uninephrectomized dogs and the measurement of various parameters of renal function over 2-hr period after declamping. There were 3 experimental groups. In the control group (I) (n = 10), the dogs received, after ischemia, an isotonic saline solution infusion at a rate of 0.2 ml/kg/min. In group II, (n = 10) the animals underwent acute volemic expansion (1 ml/kg/min) with whole blood (hematocrit approximately equal to 25%) during the ischemic period, and after declamping, an isotonic saline infusion (NaCl 0.9%) infusion at the same rate as in the control group. In group III, (n = 8) the dogs only received NaCl 0.9% (0.2 ml/kg/min) before ischemia and alpha human ANF (3.6 ng/kg/min) dissolved in saline after ischemia and during the 2 hr of the renal recovery period. Volemic expansion induced a highly significant increase of the cardiac filling pressures concomitant with a prompt but transient 5-6-fold increase in ANF levels (357 +/- 92 pg/ml versus 60 +/- 4.1 pg/ml in controls at the time of declamping [P less than 0.05]). With these higher plasma ANF levels in overloaded animals, we observed, 2 hr after declamping, considerably improved renal function recovery in terms of glomerular filtration rate--37.5% +/- 8.7 versus 11.8 +/- 3.9%; urinary sodium excretion rate--53.89 mu eq/min versus 5.36 +/- 1.2 mu eq/min (P less than 0.01); total Na reabsorption rate--1.2 +/- 0.23 meq/min versus 0.28 +/- 0.09 meq/min (P less than 0.01) (group II vs. controls, respectively). A 1-28 alpha ANF infusion after the ischemic insult allowed a comparable but more significant improved recovery of renal function--indeed, 2 hr after declamping, the GFR reached 73.7 +/- 14% of the preoperative GFR values. The urinary sodium excretion rate was 15-fold higher than in controls, and the total and fractional sodium reabsorption rates followed a similar increase. These beneficial effects of ANF were obtained with low doses of synthetic ANF (3.6 ng/kg/min) inducing plasma levels slightly higher (120 pg/ml) than in controls and comparable to the levels reached in the overloading group. In addition, maximal loading or ANF infusion produces an inhibition of the aldosterone rise occurring after the ischemic insult.(ABSTRACT TRUNCATED AT 250 WORDS)

Late recurrence of operated adrenocortical carcinoma: atrial natriuretic factor before and after treatment with mitotane.

A 62-year-old man arrived at our hospital with recurrence of Cushing's syndrome 14 years after successful surgery for adrenocortical carcinoma. Investigations demonstrated recurrence of a large tumor above the right adrenal area; it was found to be inoperable. The patient was treated initially with a new glucocorticoid antagonist, RU 486, and later with the adrenolytic agent mitotane (o,p'DDD). The latter achieved hypoadrenocorticism and a substantial reduction of tumor size. During the initial period, worsening hyperadrenocorticism resulted in a rise of atrial natriuretic factor and an inhibition of renin activity, consistent with an increase of cortisol and plasma volume. Changes in opposite direction were observed after treatment with mitotane.

Prevention of acute cyclosporine nephrotoxicity by atrial natriuretic factor after ischemia in the rat.

The present experimental study investigates whether the atrial natriuretic factor (ANF) is able to prevent the nephrotoxic effects of cyclosporine infused after 30 min of warm renal ischemia in the rat. At 2 hr after the end of ischemia, the glomerular filtration rate was improved by an ANF infusion: 390 +/- 19 microliters/min/100 g versus 298.3 +/- 31 microliters/min/100 g in ANF and saline-infused rats, respectively (P less than 0.05). Intravenous CsA infusion at a dose of 2.5 mg/kg/day produced a more pronounced fall in GFR when compared with the control: 205.4 +/- 19.7 microliters/min/100 g versus 298.3 +/- 31 microliters/min/100 g in CsA and saline, respectively (P less than 0.05). In contrast, a synthetic rat atriopeptin III (0.5 microgram/kg/min) infusion after ischemia given together with CsA prevented its deleterious effects upon GFR: 316 +/- 22 microliters/min/100 g versus 205.4 +/- 19 microliters/min/100 g in ANF/CsA versus CsA alone (P less than 0.001). Moreover, the natriuretic ANF effects remained unaffected by high plasma CsA peak levels: indeed, other parameters of renal function--urinary flow, urinary sodium concentration and excretion rates, and urinary sodium reabsorption and fractional excretion rates, were significantly increased in ANF alone or CsA/ANF groups. These preliminary results suggest that ANF may be useful in renal transplantation or in the management of patients given large doses of CsA (liver or heart transplant) since, despite nephrotoxic CsA levels (greater than 1500 ng/ml), ANF provides an improved GFR and tubular function after ischemia.

Effect of exercise on plasma atrial natriuretic factor and cardiac function in men and women.

In order to provide an integrated view of the physiology of atrial natriuretic factor (ANF) during exercise, we studied changes of its plasma concentrations in 13 normal subjects (seven males, six females) during three graded exercise levels and two periods of recovery (5 and 30 min), concomitantly with an assessment of cardiac function and ventricular volumes by multigated radionuclide angiography. Mean ANF levels (+/- SEM) increased in all patients at the second (P less than 0.002) and third (P less than 0.002) exercise levels, and after 5-min recovery (P less than 0.01): in males from 16 +/- 7 to 30 +/- 11 pg ml-1 at the third level, in females from 27 +/- 12 to 61 +/- 33 pg ml-1. Normal values were observed after 30-min recovery. Even if mean ANF levels were all higher in females, this difference did not reach statistical significance (P = 0.06). Significant decreases of ventricular volumes, as well as increases of ejection fraction and rate pressure product, were noted during exercise and were similar in both sexes. The kinetics of plasma ANF concentrations, compared with the increase of rate pressure product, was characterized by a latency and a remanence in recovery. This remanence, also present in the changes of ventricular volumes, supports the hypothesis that other factor(s) like catecholamines might still exert their influence after the exercise stops.

Beneficial effect of atrial natriuretic factor on ischemically injured kidneys in the rat. A new approach to improve early renal function.

In a rat experimental study we investigated whether the atrial natriuretic peptide by itself is able to improve early renal function after an ischemic injury. Two groups of Wistar male rats underwent a right nephrectomy and a left renal artery occlusion for 30 min and were infused for 2 hr after ischemia with isotonic saline or rat atrial natriuretic peptides (alpha ANF: 28 amino acids (AP 28) and atriopeptin III (AP 24): 24 amino acids). ANF infusion increased the urinary flow (P less than 0.001), the urinary sodium concentration (P less than 0.001), the sodium excretion rate (P less than 0.0001), and improved the glomerular filtration rate (GFR) recovery (P less than 0.02) determined at the end of the 2-hr infusion period. AP 24 exhibited higher natriuretics activities than AP 28. The effect of both peptides upon GFR recovery was equivalent. These effects of ANF observed after acute ischemia suggest that this peptide may be beneficial on the resumption of renal function in the early phases following transplantation.

Longitudinal study of brucellosis in mice by immunoassay of lipopolysaccharide-related antigens in blood and urine.

Immunoassays based on latex agglutination or enzyme labelling (ELISA) were devised for the detection of lipopolysaccharide (LPS) of Brucella abortus, or its degradation products, in biological fluids of infected mice. The agglutination of latex was measured by counting of the remaining non-agglutinated particles in an automated immunoassay analyser. LPS was assayed by agglutination with antibody-coated latex and by competitive inhibition of agglutination of LPS-coated latex by anti-LPS antiserum. The inhibition system was more sensitive for the detection of degradation products of LPS. Correlation between ELISA and agglutination inhibition immunoassay was excellent (r = 0.96). Degradation of LPS occurred during storage, particularly when the samples contained specific antibodies. It could be prevented by removing cells immediately after collecting blood samples and by heating or alkaline denaturation of plasma. CBA/H mice were infected with various doses [65-(65 x 10(6) cfu] of B. abortus biovar 3 cells and the course of infection followed by immunoassay of LPS-related antigens in serum and urine, and by titration of specific antibodies and non-specific circulating immune complexes. The concentration of LPS degradation products, assayed by the agglutination inhibition assay, was related to the severity of the infection, which was assessed by viable counts of B. abortus in the spleen. A close correlation was observed between the values of antigenaemia, the number of cfu (r = 0.97), and the inoculum size (r = 0.99 at day 28).