title: Prorenin accumulation and activation in human endothelial cells: importance of mannose 6-phosphate receptors.
abstract: ACE inhibitors improve endothelial dysfunction, possibly by blocking endothelial angiotensin production. Prorenin, through its binding and activation by endothelial mannose 6-phosphate (M6P) receptors, may contribute to this production. Here, we investigated this possibility as well as prorenin activation kinetics, the nature of the prorenin-activating enzyme, and M6P receptor-independent prorenin binding. Human umbilical vein endothelial cells (HUVECs) were incubated with wild-type prorenin, K/A-2 prorenin (in which Lys42 is mutated to Ala, thereby preventing cleavage by known proteases), M6P-free prorenin, and nonglycosylated prorenin, with or without M6P, protease inhibitors, or angiotensinogen. HUVECs bound only M6P-containing prorenin (K(d) 0.9+/-0.1 nmol/L, maximum number of binding sites [B(max)] 1010+/-50 receptors/cell). At 37 degrees C, because of M6P receptor recycling, the amount of prorenin internalized via M6P receptors was >25 times B(max). Inside the cells, wild-type and K/A-2 prorenin were proteolytically activated to renin. Renin was subsequently degraded. Protease inhibitors interfered with the latter but not with prorenin activation, thereby indicating that the activating enzyme is different from any of the known prorenin-activating enzymes. Incubation with angiotensinogen did not lead to endothelial angiotensin generation, inasmuch as HUVECs were unable to internalize angiotensinogen. Most likely, therefore, in the absence of angiotensinogen synthesis or endocytosis, M6P receptor-mediated prorenin internalization by endothelial cells represents prorenin clearance.
authors.affiliation: Cardiovascular Research Institute COEUR, Department of Pharmacology, Erasmus University Rotterdam, Rotterdam, the Netherlands.
authors.forename: M M
authors.initials: MM
authors.lastname: van den Eijnden
authors.forename: J J
authors.initials: JJ
authors.lastname: Saris
authors.forename: R J
authors.initials: RJ
authors.lastname: de Bruin
authors.forename: E
authors.initials: E
authors.lastname: de Wit
authors.forename: W
authors.initials: W
authors.lastname: Sluiter
authors.forename: T L
authors.initials: TL
authors.lastname: Reudelhuber
authors.forename: M A
authors.initials: MA
authors.lastname: Schalekamp
authors.forename: F H
authors.initials: FH
authors.lastname: Derkx
authors.forename: A H
authors.initials: AH
authors.lastname: Danser
mesh_terms: D000809:Angiotensins / Q000096:biosynthesis; D002478:Cells, Cultured; D003080:Cold Temperature; D004705:Endocytosis; D004730:Endothelium, Vascular / Q000378:metabolism*; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D009154:Mutation; D017527:Receptor, IGF Type 2 / Q000502:physiology*; D012083:Renin / Q000235:genetics / Q000378:metabolism*
chemical_list: D000809:Angiotensins; D004792:Enzyme Precursors; D017527:Receptor, IGF Type 2; D012083:Renin