Molecular Forms Of Adrenomedullin Research Articles

Increased plasma levels of the mature and intermediate forms of adrenomedullin in obesity

Adrenomedullin (AM) is a cardiovascular protective peptide produced in various organs and tissues including adipose tissue. In the present study, we measured the plasma AM levels of subjects with or without obesity by two assay methods to separately evaluate the biologically active AM–NH 2 and the intermediate form of AM–glycine (AM–Gly). We measured the total AM and AM–NH 2 levels of plasma in 52 obese and 172 non-obese residents of a Japanese community, who received regular health check-ups and had no overt cardiovascular disease. AM–Gly values were obtained by subtracting AM–NH 2 levels from those of total AM. Both the AM–NH 2 and AM–Gly levels of the subjects with obesity were higher than those without obesity, and significant relationships were noted between body mass index (BMI) and the plasma levels of the two molecular forms of AM in a simple regression analysis. Moreover, the significant factors identified by multivariate analyses were BMI and serum triglyceride for AM–NH 2 and diastolic blood pressure, insulin, high-density lipoprotein-cholesterol, and plasma renin activity for AM–Gly. These results suggest active roles for the two molecular forms of AM in metabolic disorders associated with obesity in subjects without overt cardiovascular disease.

Role of adrenomedullin system in lipid metabolism and its signaling mechanism in cultured adipocytes

We investigated the levels of adrenomedullin (AM) system during the process of preadipocyte differentiation and its role in lipid metabolism and cellular signaling mechanism in differentiated adipocytes. We cultured rat preadipocytes and measured the following during the process of differentiation: two molecular forms of AM in the culture medium using a specific immunoradiometric assay and gene expression of AM and its receptor component using RT-PCR analysis. In differentiated adipocytes, we measured the effects of AM on the intracellular cAMP level, lipolysis, glucose incorporation, and the protein levels. Two molecular forms of AM were secreted into the medium, and the AM-mature/AM-total ratio was increased after 6 days of differentiation. Cultured rat preadipocytes highly expressed the genes of AM and its receptor components at day 1, and they increased at day 10. Administration of AM to preadipocytes increased the number of Oil Red O-positive adipocytes and spectrophotometric absorbance of Oil Red O. AM dose dependently increased cAMP level and lipolysis, and its effect was blocked by CGRP(8-37). Isoproterenol increased lipolysis, and AM had additive effects on isoproterenol-induced lipolysis. KT5720 and U0126 significantly inhibited the AM-induced lipolysis, whereas KT5720, but not U0126, significantly inhibited the isoproterenol-induced lipolysis. AM increased glucose incorporation and its effect was blocked by wortmannin. Western blot analysis revealed that AM increased phospho PKA, ERK, and Akt. These results indicate that AM and its receptor component are highly expressed in cultured adipocytes and may play a role in lipid metabolism via a different signaling pathway.

Adrenomedullin in the Kidney-Renal Physiological and Pathophysiological Roles

Adrenomedullin (AM) is a potent vasodilatory peptide originally discovered in human pheochromocytoma tissue. AM and AM gene expression are widely distributed in the cardiovascular system, including the kidney. The co-localization of AM and its receptor components such as calcitonin receptor-like receptor (CRLR), receptor activity modifying protein (RAMP)2 and RAMP3 in the kidney, heart, and vasculature suggests an important role for the peptide as a regulator of renal, cardiac, and vascular function. Indeed, in addition to its cardiovascular effects, AM has renal vasodilatory, natriuretic, and diuretic actions. Consistent with these observations, immunohistochemical studies revealed that AM is stained in the collecting duct, distal convoluted tubules, vessels, and glomerular mesangial cells, endothelial cells and podocytes. Plasma AM levels are increased in patients with renal impairment in proportion to the severity of the disease. Previously we and other investigators showed that two molecular forms of AM, AM-glycine, an inactive form, and AM-mature, an active form, circulate in human plasma. Urine also contains both forms of AM; however, the AM-mature/AM-glycine ratio is higher in urine than in plasma. Interestingly, plasma AM-glycine and AM-mature levels are increased in renal failure, whereas urinary AM-glycine and AM-mature are decreased in this condition. These results indicate that the origin of urinary AM is different from that of plasma AM. Experimental studies showed that the renal tissue AM-mature/AM-glycine ratio is higher than that in plasma and urine. In addition, renal tissue concentrations of AM are increased in severely hypertensive rats. Considering that AM has antiapoptotic, antifibrotic, and antiproliferative effects, the increase of AM in renal disease may be a protective mechanism. In fact, AM gene delivery or long-term AM infusion significantly improved glomerular sclerosis, interstitial fibrosis, and renal arteriosclerosis in several malignant hypertensive models. This review describes the biochemistry, physiology, and circulating levels of AM and also discusses what is known about the pathophysiological role of AM in renal disease.

Cardiac Adrenomedullin: Its Role in Cardiac Hypertrophy and Heart Failure

Co-localization of adrenomedullin (AM) and its receptor components such as calcitonin receptor like receptor (CRLR), receptor activity modifying protein (RAMP)2 and RAMP3 in peripheral tissues, including the heart, kidney, and vasculature, suggests an important role for the peptide as a regulator of cardiovascular function. Indeed, we previously reported that AM gene expression and / or immunoreactivity are increased in the ventricles of cardiac hypertrophy and heart failure. Recently, we also found that not only levels of AM peptide and AM gene expression, but also mRNA levels of CRLR, RAMP2 and RAMP3 are increased in cardiac hypertrophy and failing heart. Cardiac myocytes and fibroblast produce and secrete two molecular forms of AM and express CRLR, RAMP2 and RAMP3, and AM is known to have inhibitory effect of collagen synthesis and antiproliferative effect in cardiac fibroblasts. Stimulation by IL-1beta significantly increased gene expression of AM and its receptor components in cardiac fibroblasts. Preincubated IL-1beta elevated the intracellular cAMP response to exogenous administered AM. AM antisense oligodeoxynucleotide treatment significantly lowered AM levels in cultured medium. IL-1beta significantly increased (3)H-proline incorporation and AM antisense oligodeoxynucleotide treatment further increased (3)H-proline incorporation. Collectively, these results support a protective role for increased AM in the cardiac hypertrophy and heart failure. Then, we tested the effects of acute administration of AM in experimental and human heart failure, because AM has hemodynamic effects including vasodilation, increases in cardiac contractility, cardiac output, diuresis, and natriuresis. We observed profound and sustained cardiovascular, hormonal and renal effects. These effects may incorporate many of the therapeutic goals of heart failure management.

Adrenomedullin in Hypertension

Adrenomedullin (AM) is a potent vasodilatory peptide originally discovered in human pheochromocytoma tissue. The gene expression of AM is widely distributed in the cardiovascular system and many physiological actions of AM on cardiovascular system have been reported. Previous studies demonstrated that the plasma AM level is increased in patients with hypertension and further increased in patients with organ damage. We and other groups recently showed that two molecular forms of AM, AM-glycine, an inactive form, and AM-mature, an active form, circulate in human plasma and that the major molecular form in plasma is AM-glycine. Recent studies using AM antagonist or AM knockout mice confirmed the role of AM in the regulation of blood pressure. Indeed, administration of AM is beneficial in patients with hypertension, renal disease, and heart failure. In addition, recent studies revealed that AM and its receptor components such as calcitonin receptor like receptor (CRLR), receptor activity modifying protein (RAMP) 2, and RAMP3 are colocalized in peripheral tissues, including the heart, kidney, and vasculature, suggesting a role of AM as a regulator of local function. In fact, tissue AM levels and the gene expression levels of AM and its receptor are altered in hypertension. We recently showed that cardiac tissue AM levels and the gene expression levels of AM, CRLR, RAMP2, and RAMP3 are increased in left ventricular hypertrophy and further increased in hypertensive heart failure. These findings imply that increased expression of the AM system including the ligand and receptor components may modulate the pathophysiology in hypertension via the effects of AM not only as a circulating factor, but also as a local regulator. This review describes the structure, structure-activity relationships, tissue distribution, receptors, physiological action, and circulating levels of AM and also discusses what is known about the pathophysiological role of AM in hypertension. Keywords: vasodilatory peptide, pheochromocytoma, blood vessels, cardiovascular disease, carboxypeptidase, calcitonin generelated peptide (cgrp), receptor activity modifying protein (ramp)

Up-regulated synthesis of mature-type adrenomedullin in coronary circulation immediately after reperfusion in patients with anterior acute myocardial infarction

Objective Levels of adrenomedullin (AM), a potent vasodilatory peptide, have been shown to increase in the early stage of acute myocardial infarction (AMI). The purpose of this study was to determine whether coronary sinus-aortic step-up of mature forms of AM is accelerated in patients with AMI after reperfusion. Methods The subjects were 29 consecutive patients with a first episode of anterior AMI and 10 normal controls. All patients with AMI underwent balloon reperfusion therapy within 24 h after symptom onset. Plasma levels of two molecular forms of AM (an active, mature form [AM-m] and an intermediate, inactive glycine-extended form [AM-Gly]) in the aorta and coronary sinus (CS) were measured by specific immunoradiometric assay after reperfusion. Results Plasma levels of AM-m and AM-Gly in the aorta and CS were higher in AMI patients than in controls. CS-aortic step-up of AM-m, which is an index of myocardial production of AM-m, was significantly greater in AMI patients than in controls (1.7 ± 1.4 vs. 0.4 ± 0.3 pmol/L, P < 0.01). However, there was no significant difference in CS-aortic step-up of AM-Gly ( P = 0.30). AMI patients with left ventricular dysfunction ( n = 10) had a significantly higher CS-aortic AM-m step-up than AMI patients without left ventricular dysfunction ( n = 19). AM-m in the aorta and CS negatively correlated with the left ventricular ejection fraction ( r = -0.50, r = -0.48, P < 0.01). Conclusions Myocardial synthesis of AM-m is accelerated in patients with reperfused AMI, especially in patients with critical left ventricular dysfunction. Increased myocardial synthesis of active AM may protect against cardiac dysfunction, myocardial remodeling, or both after the onset of AMI.

Response of adrenomedullin system to cytokine in cardiac fibroblasts?role of adrenomedullin as an antifibrotic factor

The adrenomedullin system acts as an autocrine or paracrine factor (or both) in the development of cardiac hypertrophy and in the regulation of cardiac function. However, several aspects of the local action of adrenomedullin remain unclear. We studied the effects of interleukin 1-beta (IL-1beta) on the adrenomedullin system in cardiac fibroblasts and also examined the pathophysiological significance of such effects. We cultured rat neonatal cardiac fibroblasts with or without IL-1beta and measured (1) two molecular forms of adrenomedullin in culture medium by specific immunoradiometric assay; (2) gene expression of adrenomedullin, calcitonin receptor like receptor (CRLR), receptor activity modifying protein2 (RAMP2), and RAMP3, components of the adrenomedullin receptor, by Northern blot analysis or RT-PCR analysis; (3) intracellular cAMP levels in response to exogenously administered adrenomedullin; and (4) (3)H-proline incorporation with and without a specific adrenomedullin antisense oligodeoxynucleotide. (1) IL-1beta time-dependently increased the levels of two molecular forms of adrenomedullin, adrenomedullin-mature and adrenomedullin-glycine (P<0.01). In contrast to known levels in plasma (about 10%), adrenomedullin-mature was a major molecular form in the culture medium of cardiac fibroblasts and myocytes (65-80%). (2) IL-1beta significantly increased gene expression of adrenomedullin and its receptor components (adrenomedullin: +46%, CRLR: +460%, RAMP2: +32%, RAMP3: +350%, all P<0.01). (3) Preincubated IL-1beta elevated the intracellular cAMP response to exogenous adrenomedullin administered at a concentration of 10(-7) M (+26%, P<0.05). (4) Adrenomedullin antisense oligodeoxynucleotide treatment significantly lowered adrenomedullin-mature levels in culture medium (-50%). Adrenomedullin nonsense oligodeoxynucleotide treatment did not change (3)H-proline incorporation or mRNA levels of collagen I and III, whereas adrenomedullin antisense oligodeoxynucleotide treatment significantly increased (3)H-proline incorporation and mRNA levels of collagen I and III in IL-1beta-treated cardiac fibroblasts. These results provide evidence that the adrenomedullin system acts as an autocrine antifibrotic factor in the regulation of collagen synthesis in cardiac fibroblasts exposed to higher cytokine levels. This may beneficially modulate the pathophysiology of certain cardiac diseases.

Role of endogenous adrenomedullin system in rat cardiac fibroblasts

Background: We investigated the effects of interleukin 1-beta (IL-1b) on the adrenomedullin system in cardiac fibroblasts and also examined the role of it. Methods: With or without the presence of IL-1b in cultured rat neonatal cardiac fibroblasts, we measured; (1) two molecular forms of adrenomedullin in cultured medium by specific immunoradiometric assay, (2) gene expression of adrenomedullin, calcitonin receptor like receptor (CRLR), receptor activity modifying protein2 (RAMP2), and RAMP3, which are components of adrenomedullin receptor, by northern blot analysis or RT-PCR, (3) cAMP levels in response to exogenous administered adrenomedullin, (4) 3H-proline incorporation with and without a specific antisense oligodeoxynucleotide. Results: (1) IL-1b dose-dependently increased the levels of two molecular forms of adrenomedullin, adrenomedullin-mature and adrenomedullin-glycine (P < 0.01). Adrenomedullin-mature form was a major molecular form in culture medium (85%). (2) IL-1b significantly increased gene expression of adrenomedullin and its receptor components (adrenomedullin: +30%, CRLR: +460%, RAMP2: +25%, RAMP3: +350%, all P < 0.01). (3) Preincubated IL-1b elevated the intracellular cAMP response to exogenous administered adrenomedullin at 10 −7 M (+26%, P < 0.05). (4) AM antisense oligodeoxynucleotide treatment significantly lowered adrenomedullin levels in cultured medium (-50%). IL-1b significantly increased 3H-proline incorporation and adrenomedullin antisense oligodeoxynucleotide treatment further increased 3H-proline incorporation (+22%, P < 0.05). Conclusion: These results support a physiological role for adrenomedullin in the regulation of cell growth as an autocrine factor in cardiac fibroblasts.

Different secretion patterns of two molecular forms of cardiac adrenomedullin in pressure- and volume-overloaded human heart failure

Background In the final step of production of adrenomedullin (AM), an inactive intermediate form of glycine-extended AM (AM-glycine) is converted to the active mature form of adrenomedullin (AM-mature) by enzymatic amidation. Recent studies have revealed that AM-mature and AM-glycine circulate in human plasma. In this study, we investigated the differences of the concentrations of cardiac AM between pressure-overloaded (PO) heart failure (HF) and volume-overloaded (VO)-HF in humans. Methods and results We measured AM-mature and AM-glycine by immunoradiometric assays in pericardial fluid and plasma in 38 patients who underwent valve replacement surgery (PO-HF: aortic stenosis, n = 14; VO-HF: aortic or mitral regurgitation, n = 24). Stable coronary artery disease with normal left ventricular function served as the control (n = 24). Plasma AM-mature (VO-HF: +59%, PO-HF: +65%, P<.05) and AM-glycine (VO-HF: +43%, PO-HF: +50%, P<0.05) were similarly higher in the 2 HF groups than in the control group. Interestingly, pericardial fluid AM-mature was markedly higher than that in plasma (control: +789%, VO-HF: +1050%, PO-HF: +1745%, all P<.001). Pericardial fluid AM-mature was higher in VO-HF (+106%, P<.01) than in controls and they were further increased in PO-HF (+243%, P<.05). Pericardial fluid molecular forms of AM correlated with left ventricular systolic pressure, but not with left ventricular end-diastolic volume index in PO-HF. In contrast, they correlated with left ventricular end-diastolic volume index, but not with left ventricular systolic pressure in VO-HF. Conclusion These results suggest that cardiac AM is differently regulated from plasma AM and that cardiac AM production is upregulated in both types of HF in response to each different stimulus.

Two molecular forms of adrenomedullin in congenital heart disease.

To investigate the pathophysiological role of two forms of adrenomedullin (AM), a mature AM (AM-m) and a glycine-extended AM (AM-Gly), in congenital heart disease, we measured plasma levels of AM in patients with cyanotic heart disease, high pulmonary blood flow without pulmonary hypertension (PH), high pulmonary blood flow with PH, Fontan procedure, intracardiac repair without complication, and intracardiac repair with PH and control subjects. Plasma AM-m and AM-Gly were increased only for cyanotic heart disease (2.5 +/- 1.3 pmol/L, p < 0.001; 13.1 +/- 6.2 pmol/L, p < 0.05) and intracardiac repair with PH (2.3 +/- 1.5 pmol/L, p < 0.01; 13.0 +/- 7.0 pmol/L, p < 0.05) compared with control (1.0 +/- 1.4 and 8.6 +/- 1.3 pmol/L, respectively). They were similarly correlated with mean systemic arterial pressure (r = -0.40 and -0.37 respectively; p < 0.001), mixed venous oxygen saturation (r = -0.60 and -0.50; p < 0.0001), systemic arterial oxygen saturation (SA(sat)) (r = -0.56 and -0.46; p < 0.0001), and pulmonary arterial resistance (Rp) (r = 0.41 and 0.38; p < 0.005). Multiple regression analysis revealed that SA(sat) and Rp were independently correlated with AM. Interestingly, the venous AM-m level was significantly higher than the arterial AM-m, suggesting that the mature form is extracted in pulmonary circulation, whereas there were no venoarterial differences in AM-Gly. These results suggest that plasma AM-m and AM-Gly are similarly regulated and the main clearance site of AM-m is the lung in patients with congenital heart disease.

Altered gene expression of adrenomedullin and its receptor system and molecular forms of tissue adrenomedullin in left ventricular hypertrophy induced by malignant hypertension

To investigate the pathophysiological role of adrenomedullin (AM) in left ventricular hypertrophy (LVH) in hypertension, we measured the plasma level, left ventricle (LV) tissue level, and mRNA abundance of AM and the mRNA abundance of the AM receptor system in the LV. We also analyzed the molecular forms of AM in the plasma and LV tissue and investigated the relationships between AM and the degree of LVH. We studied the following three groups: control Wistar Kyoto rats (WKY), control spontaneously hypertensive rats (SHR), and deoxycorticosterone acetate (DOCA)-salt SHR (D-SHR). We measured AM-mature, active form, and AM-total (active form+inactive form) in plasma and the LV by a newly developed immunoradiometric assay. Gene expression of AM was measured by Northern blot analysis and gene expression of AM receptor system components, such as calcitonin receptor-like receptor (CRLR), receptor activity modifying protein 2 (RAMP2), and RAMP3 was measured by the reverse transcription polymerase chain reaction method. After 3 weeks of DOCA treatment, D-SHR was characterized by higher blood pressure, LV weight, and plasma atrial natriuretic peptide levels compared with those in the other two groups. Plasma AM-mature and AM-total levels were significantly higher in D-SHR than in the other two groups, whereas there were no significant differences in the AM-mature/AM-total ratio among the three groups. On the other hand, LV tissue AM-mature and AM-total levels were also significantly higher in D-SHR than in the other two groups, and the AM-mature/AM-total ratio was significantly higher in LV tissues than in plasma. Furthermore, the LV tissue AM-mature/AM-total ratio was significantly higher in D-SHR compared with the other two groups. The LV tissue AM-mature/AM-total ratio was significantly correlated with LV weight/body weight ( r=0.92, p<0.001). The gene expression levels of AM, CRLR, RAMP2, and RAMP3 in the LV were significantly higher in D-SHR than in the other two groups. These results suggest that the AM amidating enzyme activity, ligand, and receptor system are all upregulated in the LV hypertrophy in this malignant hypertensive rat model. Considering that AM serves as a local antihypertrophic autocrine and/or paracrine factor, the induction of AM system observed here may modulate the pathophysiology of LV hypertrophy in certain forms of malignant hypertension.

Source of plasma adrenomedullin in patients with pheochromocytoma

Controversy exists as to the origin of plasma adrenomedullin (AM). To elucidate the source of plasma AM, we measured two molecular forms of AM, an active form of mature AM (AM-m) and an intermediate inactive form of glycine-extended AM (AM-Gly), by immunoradiometric assay using specific kits in two female patients with pheochromocytoma before and 3 weeks after surgery. We also measured plasma AM-m, AM-Gly, and AM-T (AM-m + AM-Gly) levels, in addition to plasma epinephrine (E) and norepinephrine (NE) levels, in bilateral adrenal veins of one patient. Although plasma E and NE levels decreased markedly after surgery in these patients, changes in plasma AM appeared to be confined to the normal range. There were no obvious differences in plasma AM-T, AM-m, or AM-Gly levels in adrenal veins between healthy tissue and tumor sides. Furthermore, plasma AM-T, AM-m, or AM-Gly levels in adrenal veins were comparable with those in the infrarenal inferior vena cavae (IVC) or the suprarenal IVC. In contrast, plasma E and NE levels increased in the adrenal vein of the healthy side and increased further in the adrenal vein of the tumor side compared with those in the infrarenal IVC. These results suggest that the origin of plasma E and NE is the adrenal gland and that elevated plasma levels of E and NE in pheochromocytoma are due to excessive production of E and NE in the adrenal gland of the tumor side. In contrast, it is suggested that neither plasma AM levels in the adrenal vein of the healthy side nor those of the tumor side contribute to the systemic levels of plasma AM. The present results appear to be consistent with the hypothesis that the source of circulating AM is systemic vasculature.

Molecular forms of adrenomedullin in pericardial fluid and plasma in patients with ischaemic heart disease

Molecular forms of adrenomedullin in pericardial fluid and plasma in patients with ischaemic heart disease

Molecular forms of adrenomedullin in pericardial fluid and plasma in patients with ischaemic heart disease

Experimental studies have demonstrated that adrenomedullin (AM) has a positive inotropic action and exerts inhibitory effects against ventricular remodelling as an autocrine and paracrine factor. However, there is no clinical evidence for AM acting as a local regulator in the human heart. We measured the levels of various molecular forms of AM, i.e. an active form of mature AM (AM-m), an intermediate inactive form of glycine-extended AM (AM-Gly) and total AM (AM-T = AM-m+AM-Gly), in plasma and pericardial fluid using our newly developed immunoradiometric assay in consecutive 67 patients undergoing coronary artery bypass graft surgery. Pericardial fluid and plasma cAMP, atrial natriuretic peptide and brain natriuretic peptide levels were also measured. The relationships between pericardial fluid AM levels and ventricular functions and other hormone levels were analysed. The level of each molecular form of AM in pericardial fluid was closely correlated with that of the other molecular forms of AM in the fluid. However, levels were not correlated with those in plasma. AM-T levels were slightly higher in pericardial fluid than in plasma (+72%; P&lt;0.05), whereas AM-m levels and AM-m/AM-T ratios were markedly higher in pericardial fluid than in plasma (AM-m, +994%; AM-m/AM-T ratio, +443%; both P&lt;0.01). AM-m, AM-Gly and AM-T levels in pericardial fluid were correlated with indices of left ventricular function, and with atrial natriuretic peptide and brain natriuretic peptide levels. Interestingly, AM and cAMP levels were positively correlated in plasma, but negatively correlated in pericardial fluid. In addition, AM-m, AM-Gly and AM-T levels in pericardial fluid were higher in patients with acute coronary syndrome than in those with stable ischaemic heart disease (AM-m, +80%; AM-Gly, +96%; AM-T, +83%; all P&lt;0.01). These results suggest that AM in pericardial fluid reflects cardiac synthesis, and that enhanced cardiac secretion of AM is associated with left ventricular dysfunction, ventricular overload and myocardial ischaemia. Considering that AM has positive inotropic, coronary vasodilatory and anti-remodelling actions, increased cardiac AM may play a compensatory role in the ischaemic and failing myocardium.

Two molecular forms of plasma adrenomedullin during tilt test in healthy subjects

There is accumulating evidence suggesting that adrenomedullin (AM) may participate in the regulation of circulatory homeostasis and pathophysiology of cardiovascular disease. A recent study revealed that two molecular forms of AM, an active form of mature AM (AM-m) and an intermediate inactive form of glycine-extended AM (AM-Gly), circulate in human plasma. The object of the present study was to evaluate the effect of orthostasis on a time course of two molecular forms of plasma AM and to compare them with the behavior of other vasoactive hormones. Twelve healthy male volunteers were studied. The experimental protocol consisted of 20 min of supine rest, tilting at 70° for 20 min, and then 20 min of supine rest. Blood pressure and heart rate were measured every minute. Blood samples were obtained before, at 2 and 18 min during the tilt test, and 2 and 18 min after the test for the measurements of vasoacting hormones and hematocrit. Blood pressure and heart rate were slightly increased earlier during tilting and then remained elevated until the end of the test. The increase in heart rate and blood pressure returned to normal levels early after the tilt test. Plasma epinephrine and norepinephrine significantly increased during the tilt test. These hormones returned to normal levels 18 min after the test. The plasma renin activity, antidiuretic hormone and dopamine were also increased by the end of the tilt test, whereas plasma atrial natriuretic peptide was significantly decreased after the tilt test. Hematocrit increased slightly in the early phase of the tilt test and was further increased by the end of the test. In contrast, plasma AM-Gly or AM-m did not change during the tilt test or the recovery period. Nitric oxide metabolites did not change, either. There were no significant relationships between plasma catecholamines and AM. Plasma brain natriuretic peptide did not change during the tilt test or the recovery period, either. These results suggest that the two molecular forms of AM, AM-m and AM-Gly in plasma, did not respond to the short term tilting stress. These findings may support the hypothesis that plasma AM is secreted in a constitutive manner from the vascular wall.

Molecular forms of plasma and urinary adrenomedullin in normal, essential hypertension and chronic renal failure.

Human adrenomedullin precursor is converted to glycine-extended adrenomedullin (AM-Gly), an intermediate inactive form of adrenomedullin. Subsequently, AM-Gly is converted to active form of mature adrenomedullin (AM-m). The aim of the present study was to investigate (i) whether sex or age influences plasma and urinary AM-m and AM-Gly levels in normal subjects; (ii) the daytime variability of plasma AM-m and AM-Gly levels in normal subjects; (iii) AM-m and AM-Gly levels and its ratio in plasma and urine in normal subjects, individuals with essential hypertension (HT), and chronic renal failure (CRF); and (iv) the ratio of AM-m and AM-total (T) in plasma of various veins and aorta. We measured plasma levels and urinary excretions of AM-m, AM-Gly and AM-T (AM-m + AM-Gly) by recently developed immunoradiometric assay in normal subjects (n = 81), HT (n = 28) and CRF (n = 30). We also determined the molecular forms of plasma adrenomedullin taken from various sites during angiography in patients with suspected renovascular hypertension (n = 9). There were no differences in plasma and urinary excretions of two molecular forms of adrenomedullin among sexes or ages in normal subjects. There was no daytime variation of plasma two molecular forms of adrenomedullin in normal subjects. Plasma AM-m, AM-Gly and AM-T levels were increased in patients with HT and CRF compared with normal subjects, whereas urinary AM-m, AM-Gly and AM-T excretions were decreased in patients with HT and CRF compared with normal subjects. Urinary AM-m: AM-T ratios were significantly higher than plasma AM-m: AM-T ratios. Plasma AM-m and AM-T levels taken from various veins were similar, and they were significantly higher than those of aorta, although there were no differences in plasma AM-Gly levels between aorta and veins. These results suggest that in normal subjects, and individuals with HT and CRF: (i) plasma and urinary excretions of AM-m and AM-Gly are not affected by age or sex; (ii) AM-m in parallel with AM-Gly is increased; (iii) urine contains a higher percentage of active adrenomedullin than plasma; and (iv) plasma AM-m may be partly metabolized in the lung.

Distribution and molecular forms of adrenomedullin and proadrenomedullin N-terminal 20 peptide in the porcine gastrointestinal tract.

Adrenomedullin (AM) and proadrenomedullin N-terminal 20 peptide (PAMP) are potent hypotensive peptides that are generated from the same precursor and expressed in a variety of mammalian tissues. The purpose of this study was to elucidate the distribution and endogenous molecular forms of AM and PAMP. To this end, we used new radioimmunoassays that recognize the ring structure of AM and the C-terminal region of PAMP to compare the distribution of AM and PAMP in porcine gastrointestinal tissues. Immunoreactive AM was abundant in the duodenum (0.334+/-0.132 fmol/mg) and ileum (0.439+/-0.235 fmol/mg), and ubiquitous in other gastrointestinal tissues. The duodenal and ileal concentrations of AM were about 4 to 14 times higher than those in other gastrointestinal tissues. Similarly, immunoreactive PAMP was abundant in the duodenum (0.577+/-0.417 fmol/mg) and ileum (1.575+/-1.445 fmol/mg) and ubiquitous in other gastrointestinal tissues. The concentrations of immunoreactive AM and PAMP were highest in the ileum. Characterization of these peptides in the ileum, using high-performance liquid chromatography, showed that they were authentic. Furthermore, the concentrations of immunoreactive AM and PAMP in the mucosa and submucosa of the ileum were significantly higher than those in whole ileum. These results suggest that AM and PAMP play physiological roles in the porcine small intestine.

Elevation of two molecular forms of adrenomedullin in plasma and urine in patients with acute myocardial infarction treated with early coronary angioplasty

Elevation of two molecular forms of adrenomedullin in plasma and urine in patients with acute myocardial infarction treated with early coronary angioplasty

Elevation of two molecular forms of adrenomedullin in plasma and urine in patients with acute myocardial infarction treated with early coronary angioplasty

Adrenomedullin (AM) has vasodilatory, diuretic and natriuretic actions. Two molecular forms of AM circulate in human plasma: an active, mature form of AM (AM-m) and an intermediate, inactive, glycine-extended form of AM (AM-Gly). In the present study we investigated the pathophysiological significance of the two molecular forms of AM in plasma and urine in patients with acute myocardial infarction. We serially measured venous and arterial plasma levels and urinary excretion of AM-m, AM-Gly and total AM (Am-T; =AM-m+AM-Gly) over 2 weeks using our recently developed immunoradiometric assay in 26 consecutive patients with acute myocardial infarction and in age-matched normal controls, and studied the relationships between AM levels and clinical parameters. Plasma AM-m, AM-Gly and AM-T levels were increased on admission in patients with acute myocardial infarction compared with age-matched normal controls. Levels of AM-m, AM-Gly and AM-T in plasma reached a peak 24 h after the onset of symptoms. Plasma AM-m, AM-Gly and AM-T levels were significantly correlated with plasma levels of brain natriuretic peptide and pulmonary arterial pressure. Plasma AM-Gly levels in the vein were similar to those in the artery, whereas plasma AM-m levels were significantly lower in the artery than in the vein. Urinary excretion of AM-m, AM-Gly and AM-T was also increased on admission, and reached a peak at 12 h after the onset of symptoms. Urinary excretion of AM-m and AM-Gly was significantly correlated with urinary sodium excretion. The AM-m/AM-T ratio was significantly higher in the urine than in the vein or artery. AM-m levels were significantly correlated with AM-Gly levels in both the urine and plasma; however, there were no significant correlations between plasma and urinary AM levels. The results suggest that levels of both molecular forms of AM are increased in the urine as well as in the plasma in the acute phase of myocardial infarction. Since AM exerts potent cardiovascular and renal effects, increased concentrations of AM in plasma and urine in the acute phase of myocardial infarction may be involved in the defence mechanism against further elevations of peripheral and pulmonary vascular resistance and oliguria in acute myocardial infarction.

Production and clearance sites of two molecular forms of adrenomedullin in human plasma

Human adrenomedullin (AM) precursor is converted to glycine-extended AM (AM-Gly), an inactive intermediate form of AM. Subsequently, AM-Gly is converted to active mature AM (AM-m) by enzymatic amidation. A recent study showed that two molecular forms of adrenomedullin (AM) are present in human plasma. In this study we investigated the production and clearance sites of two molecular forms of adrenomedullin in humans. We measured plasma levels of AM-m and AM-Total (T) (AM-m+AM-Gly) by immunoradiometric assay and calculated plasma levels of AM-Gly in blood samples taken from various sites during cardiac catheterization in patients with ischemic heart disease. Plasma AM-m levels were significantly lower in left-sided sites after passing through pulmonary circulation than in right-sided sites, whereas there were no significant differences in AM-Gly levels between left-sided sites and right-sided sites. These results suggest that AM-m produced in many organs is released into veins and that the main clearance sites of AM-m are the lungs. Considering that AM preferentially dilates pulmonary vessels rather than systemic vessels, a possible role of this peptide is suggested in the regulation of pulmonary vascular tonus.