Volume 53, Issue 4, Pages (April 1998)

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Volume 53, Issue 4, Pages 979-985 (April 1998) Autocrine/paracrine role of adrenomedullin in cultured endothelial and mesangial cells Hideo Michibata, Masashi Mukoyama, M.D., Ph.D., Issei Tanaka, Shin-ichi Suga, Masayo Nakagawa, Rieko Ishibashi, Masahisa Goto, Kenichi Akaji, Yoichi Fujiwara, Yoshiaki Kiso, Kazuwa Nakao Kidney International Volume 53, Issue 4, Pages 979-985 (April 1998) DOI: 10.1111/j.1523-1755.1998.00855.x Copyright © 1998 International Society of Nephrology Terms and Conditions

Figure 1 A typical standard curve of adrenomedullin (AM) and cross-reactivity profiles of its related peptides (A) and dilution curves of culture media (B) in the radioimmunoassay (RIA) for AM with a monoclonal antibody KY-AM-I. Abbreviations and symbols in A are: (•) hAM, human AM; (○) rAM, rat AM; (▴) hAM (15-31); (▵) hAM (1-15); (▪) hAM (27-52); (□) hCGRP-I. In B, symbols and abbreviations are: (○) bEC, bovine endothelial cells; (▵) rVSMC, rat vascular smooth muscle cells; (□) rMC, rat mesangial cells. Kidney International 1998 53, 979-985DOI: (10.1111/j.1523-1755.1998.00855.x) Copyright © 1998 International Society of Nephrology Terms and Conditions

Figure 2 Typical high performance-gel permeation chromatography (HP-GPC) profiles of adrenomedullin-like immunoreactivity (AM-LI) in the culture supernatant of bovine endothelial cells (EC; A), rat vascular smooth muscle cells (VSMC; B), and rat mesangial cells (MC; C). Arrows denote elution positions of a series of myoglobins of a polypeptide molecular weight calibration kit (Pharmacia), void volume (Vo) and total volume (Vt). The elution position of synthetic AM is also indicated. Kidney International 1998 53, 979-985DOI: (10.1111/j.1523-1755.1998.00855.x) Copyright © 1998 International Society of Nephrology Terms and Conditions

Figure 3 Adrenomedullin (AM)-stimulated increase of cAMP production in cultured bovine endothelial cells (EC; A), rat vascular smooth muscle cells (VSMC; B), and rat mesangial cells (MC; C). Values are expressed as the mean ±SEM (N = 4). B is basal. Kidney International 1998 53, 979-985DOI: (10.1111/j.1523-1755.1998.00855.x) Copyright © 1998 International Society of Nephrology Terms and Conditions

Figure 4 Effects of pretreatment with the monoclonal antibody KY-AM-I (▪) or control IgG () on basal and AM-stimulated cAMP production in cultured bovine endothelial cells (EC; □). Values are expressed as the mean ±SEM (N = 6). *P < 0.01, **P < 0.001. Kidney International 1998 53, 979-985DOI: (10.1111/j.1523-1755.1998.00855.x) Copyright © 1998 International Society of Nephrology Terms and Conditions

Figure 5 Inhibitory effects of adrenomedullin (AM) on platelet-derived growth factor (PDGF)-stimulated3H-thymidine incorporation into cultured bovine endothelial cells (EC; A), rat vascular smooth muscle cells (VSMC; B), and rat mesangial cells (MC; C). Values are expressed as the mean ±SEM (N = 6). *P < 0.05, **P < 0.01, as compared to the levels with PDGF alone. Kidney International 1998 53, 979-985DOI: (10.1111/j.1523-1755.1998.00855.x) Copyright © 1998 International Society of Nephrology Terms and Conditions

Figure 6 Effects of pretreatment with the monoclonal antibody (MAb) on basal 3H-thymidine incorporation into cultured endothelial cells (EC). Symbols are: (□) basal level (B) with vehicle; () mouse IgG (50 μg/ml); () low dose of MAb KY-AM-I (1 μg/ml); (▪) high dose of KY-AM-I (50 μg/ml). Values are expressed as the mean ±SEM (N = 6). *P < 0.01, **P < 0.001, as compared to basal. Kidney International 1998 53, 979-985DOI: (10.1111/j.1523-1755.1998.00855.x) Copyright © 1998 International Society of Nephrology Terms and Conditions