A. M. Kappelgaard

Measurement of angiotensin II in human plasma: technical modifications and practical experience.

This paper presents our experience in measurement of plasma angiotensin II concentration according to the method described by Düsterdieck, G. and McElwee, G. (1971) Eur. J. Clin. Invest. 2, 32-38. Certain technical modifications of the procedure have been worked out. For each individual plasma sample, the recovery across extraction and elution steps is determined by addition of radio-iodinated angiotensin II. A time-saving recycling elution procedure is described. Evidence for the clinical applicability of the modified method is presented.

Plasma noradrenaline concentration in hypertensive and normotensive forty-year-old individuals: Relationship to plasma renin concentration

Forty-year-old individuals with labile and with mild sustained essential hypertension, identified during a survey of a population born in 1936, were investigated. None had ever received antihypertensive treatment. In thirty-three individuals (26 M, 7F) with diastolic blood pressure (DBP) greater than or equal to 95 mmHg at the very first examination and in thirty-one (14 M, 17 F) randomly selected normotensive controls plasma noradrenaline concentration (PNAC) was measured at rest supine. In twenty-two patients (16 M, 6 F), with sustained diastolic hypertension (diastolic blood pressure greater than or equal to 95 mmHg on at least three different occasions) and in twenty-four (14 M, 10 F) normotensive controls PNAC and plasma renin concentration (PRC) were measured supine at rest and again 2 h after furosemide and ambulation. Basal and acutely stimulated values for PNAC and PRC were identical in hypertensive and normotensive individuals. A close correlation between PNAC and PRC after acute stimulation (r = 0.77, P < 0.001) as well as between the absolute changes from resting to acutely stimulated values (r = 0.72, P < 0.001) were found in the hypertensive individuals. It is concluded that sympathetic nerve activity, as defined from measurements of plasma noradrenaline concentration, is similar in young patients with mild hypertension and in normotensive controls. We propose that the discrepancies found in the literature might be related to a lack of comparability between hypertensive and normotensive individuals studied, as far as the source of study populations is concerned.

Solid-phase double-antibody radioimmunoassay of pepsinogen I in serum

A solid-phase radioimmunoassay for the determination of pepsinogen I in serum has been developed. The antibody was raised in rabbits with pepsinogen I isolated from urine as previously described. Radioiodination was carried out with a chloramine-T procedure resulting in a tracer with excellent shelf life. In the standard procedure with a 24-h incubation time, followed by 2-h incubation with a second antibody coupled to a solid phase, 50 microliter serum was analyzed, standard range 1.88-60 ng PG I. An eight times more sensitive method was also developed using sequential saturation techniques. Specificity studies demonstrated 0.6% crossreactivity with PG II. The immunoreactivity of PG I purified from urine was nearly identical with the immunoreactivity of PG I purified from gastric mucosa. The levels of PG I in serum from 121 control subjects were similar to those obtained with conventional phase separation methods. It is concluded that the method is simple, precise and free from non-specific serum interference.

The effect of angiotensin II blockade by saralasin (1-Sar-8-Sla-angiotensin II) in normal man

SummaryThe effects of the angiotensin II analogue saralasin were investigated in 6 normal individuals. Blood pressure, plasma renin (PRC), plasma aldosterone (PAC) and plasma saralasin were measured before and during infusion of saralasin (0.54–5.4 nmol/kg/min) with the subjects supine. Plasma angiotensin II concentration (PA II) was measured before the infusion. In the sodium replete state, PA II averaged 11 pmol/1 (range 5 to 17). Saralasin infusion produced an increase in mean arterial pressure (MAP) and PAC, and a slight fall in PRC, which is consistent with an angiotensin II-like effect or a so-called agonistic effect. After sodium depletion, induced by hydrochlorothiazide 50–100 mg/day for 5 days, PA II was high −91 pmol/l on average (range 41 to 217). Angiotensin II blockade produced a fall in MAP in the supine position. The agonistic effect of saralasin on adrenal receptors during sodium depletion was less pronounced or absent. PRC increased sharply during the infusion. Infusion of saralasin at the rate of 5.4 nmol/kg/min produced a plasma saralasin concentration of about 220 nmol/l, i. e. in molar terms the plasma concentration of the analogue was 2000 to 10000 times higher than that of the endogenous octapeptide. The relationship between changes in MAP and basal PA II prior to infusion showed that saralasin exhibited a shift from agonistic to antagonistic properties on vascular receptors when pre-infusion PA II changed from approximately 20 to 40 pmol/l. A shift from agonistic to antagonistic effect on aldosterone secretion was not consistently seen. It is concluded that angiotensin II does not have a decisive role in the maintenance of normal blood pressure during normal sodium balance. However, after sodium depletion the renin-angiotensin system contributes to blood pressure control, even in the supine position. In addition to its antagonistic properties, saralasin possesses a weak agonistic effect on vascular, as well as on renal and adrenal receptors. This has to be taken into consideration when saralasin infusion is used to define “angiotensin II dependency” in patients with hypertension.

Double-antibody solid-phase radioimmunoassay for blood bradykinin

A solid phase radioimmunoassay for the determination of blood bradykinin has been developed. Highly specific antibodies against bradykinin were raised in rabbits after coupling the peptide to thyroglobulin. Iodination of [Tyr8]-bradykinin was carried out with a chloramine-T procedure resulting in a tracer with high specific activity. Bradykinin was isolated in the following way: blood was sampled directly into acetone, and lipids were removed by extraction with petroleum either (40-60 degrees C). The final purification was made on QAE-Sephadex A-25 at pH 7.4. The mean recovery of added [125I-Tyr8]-bradykinin was 28% with a sample volume of 6 ml whole blood. The sensitivity of the radioimmunoassay was 1.25 pg/tube or 3 pg/ml blood. The reproducibility of the method is satisfactory with a between-assay coefficient of variation below 16%. Levels found in venous blood were below 3 pg bradykinin/ml in normal persons.

Radioimmunoassay for saralasin by means of anti-angiotensin II sera

A radioimmunoassay for the angiotensin II-inhibitor saralasin has been developed. The assay is based upon an exploitation of the cross-reactivity of antisera raised against angiotensin II, with radioiodinated saralasin as the tracer peptide. Performance data for the assay and results obtained in clinical infusion studies are presented.

The use of an angiotensin II antagonist (saralasin) as an adjunct during renal vein catheterization

Renal vein catheterization was performed in fifteen hypertensive patients with unilateral renal disease. Samples for measurement of plasma renin concentration were obtained from each of the two renal veins and from the femoral artery (or the inferior caval vein)-before and during saralasin infusion. Saralasin infusion induced a significant decrease in blood pressure. In ten patients with lateralization of renin secretion before infusion, saralasin induced a 2-fold increase of the renin gradient across the diseased kidney, whereas there was no significant renin gradient across the contralateral kidney neither before nor after saralasin infusion. Thus, the renal venous renin ratio (diseased/contralateral) increased from a mean value of 2.10 to 4.13. In five patients without lateralization of renin secretion prior to infusion, saralasin induced a significant increase of renin gradient across both kidneys. In consequence, evidence for lateralization did not emerge and the renal vein renin ratio remained unchanged at 1.10. In cases with lateralization of renin secretion, the use of saralasin provides confirmatory evidence for strictly unilateral renin secretion with suppression of renin output from the contralateral kidney. In patients without obvious lateralization of renin secretion before saralasin, the administration of this angiotensin II inhibitor can serve to demonstrate a potential renin for renin secretion, shared by both kidneys.