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Adrenomedullin

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Mid-regional pro-adrenomedullin as a novel predictor of mortality in patients with chronic heart failure

von Haehling et al. Eur J Heart Fail. 2010 May;12(5):484-91. Epub 2010 Mar 9.

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Adrenomedullin: a new target for the design of small moleculemodulators with promising pharmacological activities

Julian et al. Eur J Med Chem. 2005 Aug;40(8):737-50.

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Levels of plasma total adrenomedullin are related with two acute phase inflammatory reactants (fibrinogen and sialic acid) but not with markers of endothelial dysfunction in Type 1 diabetes Adrenomedullin and vascular risk factors in Type 1 DM.

Garcia-Unzueta et al. J Diabetes Complications. 2005 May-Jun;19(3):147-54.

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Molecular and cellular basis of the aetiology and management of diabetic cardiomyopathy: a short review

Adeghate El. Mol Cell Biochem. 2004 Jun;261(1-2):187-91.

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Two molecular forms of adrenomedullin in congenital heart disease

Watanabe et al. Pediatr Cardiol. 2003 Nov-Dec;24(6):559-65. Epub 2003 Sep 4.

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Increased oxidative stress is associated with elevated plasma levels of adrenomedullin in hypertensive patients with type 2 diabetes

Katsuki et al. Diabetes Care. 2003 May;26(5):1642-3.

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

Nishikimi et al. Clin Sci (Lond). 2002 Jun;102(6):669-77

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

Nishikimi et al. Peptides. 2001 Nov;22(11):1867-72.

Immunocytochemical detection of ADM production

Immunocytochemistry-- Rabbit anti-rat ADM antibodies were from Phoenix Pharmaceuticals Inc. and were used at a 1/200 dilution. After fixation with 4% paraformaldehyde, cells were permeabilized with 0.2% Triton X-100 (v/v). Nonspecific binding sites were saturated using 4% normal goat serum (v/v). Anti-ADM antibodies were detected with a goat anti-rabbit antibody coupled to rhodamine (TRITC) (Jackson Immunoresearch).

Mapping in Human Pancreas and Lung Cancer Tissue (H-010-37)

     

Tissue Sample

Human pancreas and lung cancer tissues

Fixative

10% formalin

Embedding

Paraffin

Negative Control

No primary antibody

Pretreatment

N/A

Blocking

3% H2O2, 2% Normal Goat Serum

Primary Antibody

Rabbit anti-Adrenomedullin Gly (Human) antibody (Cat. No.: H-010-37)

Optimal Dilution

1:500

Secondary Antibody

Goat Anti-Rabbit IgG, Biotinylated (1:400), 30 min

Amplification

Streptavidin-HRP (Vector), 1:400, 30 min

Detection System

HRP

Substrate

DAB (Sigma), 3 min

Counterstained

Hematoxylin, 30 sec

 

Mapping in Human Pancreas and Lung Cancer Tissue (H-010-28)

Tissue Sample

Human/Rat lung and lung cancer tissues

Fixative

10% formalin

Embedding

Paraffin

Negative Control

No primary antibody

Pretreatment

N/A

Blocking

3% H2O2, 2% Normal Goat Serum

Primary Antibody

Rabbit anti-Adrenomedullin (1-44) (Human) antibody (Cat. No.: H-010-28)

Optimal Dilution

1:500

Secondary Antibody

Goat Anti-Rabbit IgG, Biotinylated (1:400), 30 min

Amplification

Streptavidin-HRP (Vector), 1:400, 30 min

Detection System

HRP

Substrate

DAB (Sigma), 3 min

Counterstained

Hematoxylin, 30 sec

 

Mapping in Human Pancreas and Lung Cancer Tissue (H-010-10)

Tissue Sample

Rat lung tissues

Fixative

10% formalin

Embedding

Paraffin

Negative Control

No primary antibody

Pretreatment

N/A

Blocking

3% H2O2, 2% Normal Goat Serum

Primary Antibody

Rabbit anti-PAMP-20 (Rat) antibody (Cat. No.: H-010-10)

Optimal Dilution

1:500

Secondary Antibody

Goat Anti-Rabbit IgG, Biotinylated (1:400), 30 min

Amplification

Streptavidin-HRP (Vector), 1:400, 30 min

Detection System

HRP

Substrate

DAB (Sigma), 3 min

Counterstained

Hematoxylin, 30 sec

 

Mapping in Rat Hear Tissue by Anti-Adrenomedullin(1-50) (Rat) Antiserum (H-010-08)

Tissue Sample

Rat Heart Tissue

Fixative

10% formalin

Embedding

Paraffin

Negative Control

No primary antibody

Pretreatment

Target retrieval 25 min (Steam)

Blocking

3% H2O2, 2% Normal Goat Serum

Primary Antibody

Anti-ADM (rat) Antibody (Catalog No.: H-010-08)

Optimal Dilution

1:200, 1 hour at RT

Secondary Antibody

Goat Anti-Rabbit IgG, Biotinylated (1:400), 30 min

Amplification

Streptavidin-HRP (Vector), 1:400, 30 min

Detection System

HRP

Substrate

DAB (Sigma), 3 min

Counterstained

Hematoxylin, 30 sec

 

Detection of Pro-Adrenomedullin in Rat Tissues by Western Blot Kit

(Cat No.: WBK-010-10)

Detection of Pro-Adrenomedullin in Rat Tissues by Western Blot Kit

Accumulation of ADM in the culture medium of CAECs. indicates spontaneous secretion of ADM from CAECs; , ADM secretion from CAECs stimulated with sarafotoxin S6c (10-7 mol/L). *P<0.05 between groups. Hypertension. 1998;32:917-922

 

Radioimmunoassay for ADM

After incubation for the indicated time, the culture medium was removed and immediately acidified with acetic acid (final concentration, 1 mol/L), heated at 100°C for 10 minutes to inactivate intrinsic protease, and stored at -20°C until assay. Culture medium (1 mL) was extracted on C-18 Bond Elute cartridges and eluted with 75% methanol containing 1% trifluoroacetic acid. Concentrated eluates were then assayed using a specific and sensitive radioimmunoassay for ADM(1-52) (Phoenix Pharmaceuticals Inc) as previously described. Minimal detectable concentration for the assay is 1 pg per tube, and the half-maximal inhibition dose of radioiodinated ligand binding by ADM is 20 pg per tube. Recovery is 72%, and intra-assay and interassay variations are 10% and 12%, respectively.

Jougasaki M., et al. Hypertension. 1998;32:917-922

 

Adrenomedullin, Pro (45-92), (Human) RIA Kit (RK-010-06)

Adrenomedullin Precusor Schematic (Human)

Adrenomedullin Precusor Schematic (Rat)

Adrenomedullin Sequence Comparisons between Human, Porcine & Rat

PAMP Sequence Comparisons between Human, Porcine, Canine, Rat, Mouse

 

 

Comparison of the Human Adrenoedullin amino acid sequence to the sequences of other relevent peptides

Central and peripheral cardiovascular actions of adrenomedullin 5, a novel member of the calcitonin gene-related peptide family, in mammals

Adrenomedullin 5 (AM5) is a new member of the calcitonin gene-related peptide (CGRP) family identified in teleost fish. Although its presence was suggested in the genome database of mammals, molecular identity and biological function of AM5 have not been examined yet. In this study, we cloned a cDNA encoding AM5 in the pig and examined its cardiovascular and renal effects. Putative mature AM5 was localized in the middle of prohormone and had potential signals for intermolecular ring formation and C-terminal amidation. The AM5 gene was expressed most abundantly in the spleen and thymus. Several AM5 genes were newly identified in the database of mammals, which revealed that the AM5 gene exists in primates, carnivores, and undulates but could not be identified in rodents. In primates, nucleotide deletion occurred in the mature AM5 sequence in anthropoids (human and chimp) during transition from the rhesus monkey. Synthetic mature AM5 injected intravenously into rats induced dose-dependent decreases in arterial pressure at 0.1-1 nmol/kg without apparent changes in heart rate. The decrease was maximal in 1 min and AM5 was approximately half as potent as AM. AM5 did not cause significant changes in urine flow and urine Na+ concentration at any dose. In contrast to the peripheral vasodepressor action, AM5 injected into the cerebral ventricle dose-dependently increased arterial pressure and heart rate at 0.1-1 nmol. The increase reached maximum more quickly after AM5 (5 min) than AM (15-20 min). AM5 added to the culture cells expressing calcitonin receptor-like receptor (CLR) or calcitonin receptor (CTR) together with one of the receptor activity-modifying proteins (RAMPs), the combination of which forms major receptors for the CGRP family, did not induce appreciable increases in cAMP production in any combination, although AM increased it at 10(-)(10)-10(-)(9) M when added to the CLR and RAMP2/3 combination. These data indicate that AM5 seems to act on as yet unknown receptor(s) for AM5, other than CLR/CTR+RAMP, to exert central and peripheral cardiovascular actions in mammals.

Takei et al. J Endocrinol. 2008 May;197(2):391-400.

Peptide Receptor
AM1/ AM4 CLR-RAMP2/3
AM2 / AM3 CLR-RAMP3
AM5 specific AM5 Receptor?
CGRP CLR-RAMP1
Amylin CTR-RAMP2

Amino acid sequence of Pig Adrenomedullin 5

Sequence Comparison between Human Hypothetical protein LCO199800 and Pig Adrenomedullin 5

Sequence Comparison between Human/Porcine ADM, human IMD, Eel AM3, Porcine AM5& human AM5

 

Human Placentra Tissue by Anti-Adrenomedullin-5 (Porcine) Antiserum (H-010-40)

Tissue Sample

Human placenta tissues

Fixative

10% formalin

Embedding

Paraffin

Negative Control

No primary antibody

Pretreatment

N/A

Blocking

3% H2O2, 2% Normal Goat Serum

Primary Antibody

Rabbit anti-Adrenomedullin-5 (Porce) Antiserum (Cat. No.: H-010-40)

Optimal Dilution

1:500

Secondary Antibody

Goat Anti-Rabbit IgG, Biotinylated (1:400), 30 min

Amplification

Streptavidin-HRP (Vector), 1:400, 30 min

Detection System

HRP

Substrate

DAB (Sigma), 3 min

Counterstained

Hematoxylin, 30 sec

Yin Yang Theory of Adrenomedullin

For every force there is an opposing force that balances the overall effect. Such is the basic tenant of Yin and Yang. Adrenomedullin (ADM) eloquently fits this ancient wisdom. Hypotensive effects are elicited by fragments of ADM retaining the C-terminal with ring structure. Hypertensive effects are seen with N-terminal ADM fragments containing the ring structure, or with the ring structure alone. The hypotensive effects can be antagonized with C-terminal fragments lacking the ring structure. Nature has endowed ADM with the ability to regulate itself, however it remains to be seen if the Yin and Yang of ADM is utilized in vivo.

intermedin

%Adrenomedullin%


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