Endothelin increases [Ca2+]i, protein phosphorylation, and O2-. production in human alveolar macrophages.
The goal of the present study was to investigate whether endothelin (ET) increases O2-.production in alveolar macrophages and to establish which second messengers are activated by ET. We measured the effects of ET on cytosolic free calcium concentration [Ca2+]i, protein phosphorylation, and O2-.production in human alveolarmacrophages (HAM). Human macrophages were obtained by bronchoalveolar lavage. [Ca2+]i was measured by the fura-2 method both in cell suspensions and in isolated single macrophages. Protein phosphorylation was assessed by gel electrophoresis and autoradiography after labeling the cells with 32P. ET increased [Ca2+]i in a dose-dependent manner (50% effective concentration = 1 x 10(-7)M). At a concentration of 10(-6)M ET, [Ca2+]i rose from basal values of 121 +/- 23 to 456 +/- 41 nM. This increase was comparable with the increase of [Ca2+]i induced by N-formyl-L-methionyl-L-leucyl-L-phenylalanine and platelet-activating factor (PAF; 1-O-alkyl-2-acetyl-sn-glyceryl-3-phosphorylcholine). The initial rise in [Ca2+]i and the sustained phase were significantly reduced in calcium-free medium (1 mM ethyleneglycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid) and after preincubation of HAM with the calcium channel blocker nitrendipine (10(-7)M). Exposure of 32P-labeled HAM to ET for 1 min induced the phosphorylation of a group of 48-kDa proteins and of a 35-kDa protein. The ET-induced 32P incorporation into the 48-kDa proteins was less pronounced than the 12-O-tetra-decanoylphorbol-13-acetate or PAF-induced response (168 +/- 24 vs. 356 +/- 68 and 278 +/- 61% of control).(ABSTRACT TRUNCATED AT 250 WORDS)
Haller
H
H
Division of Nephrology, Freie Universität Berlin, Klinikum Steglitz, Federal Republic of Germany.
Schaberg
T
T
Lindschau
C
C
Lode
H
H
Distler
A
A
D000328:Adult; D001992:Bronchoalveolar Lavage Fluid; D002118:Calcium / Q000378:metabolism*; D016232:Endothelins / Q000502:physiology*; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D008262:Macrophage Activation; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D010766:Phosphorylation; D010972:Platelet Activating Factor / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D016232:Endothelins; D010972:Platelet Activating Factor; D011506:Proteins; D013481:Superoxides; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium