title: In vitro evidence for an intracellular site of angiotensin action.
abstract: To differentiate the relative effects of nuclear and cell surface angiotensin II (Ang II) receptors, we mutated the angiotensinogen cDNA by removing the signal sequence-encoding region to produce a nonsecreted form of angiotensinogen [Ang(-S)Exp]. Rat hepatoma cells (which produce renin and angiotensin-converting enzyme mRNAs) were stably transfected with Ang(-S)Exp/pSVL (or a corresponding control) expression plasmid, and mitotic indices were measured for stably transfected cell lines. Experimental clonal cell lines demonstrate an average of 33+/-4.4% (P<0.001) increase in percentage-labeled nuclei compared with control cell lines. The mitogenic effect is blocked by 10(-6) mol/L losartan and by 1 micromol/L renin antisense phosphorothioate oligomers but not by 10(-6) mol/L candesartan. In addition, phenylarsine oxide, which blocks angiotensin receptor internalization, abolishes the losartan inhibitory effect, suggesting that after cell-surface receptor-mediated endocytosis, losartan blocks Ang II nuclear receptors. PDGF mRNA levels are elevated 2.2-fold in Ang(-S)Exp transfected cell lines; addition of anti-PDGF antibodies to the culture medium partially blocks the mitogenic effect of Ang(-S)Exp, while anti-Ang II antibodies have no effect. These results suggest that the Ang(-S)Exp growth effect is due, in part, to autocrine/paracrine stimulation by secreted PDGF after Ang II/Ang II receptor intracellular interactions. We further demonstrate that these cells produce the alternative renin transcript, renin 1A, which apparently lacks a signal sequence and is maintained intracellularly. Collectively, these studies of cultured cells suggest that some cell types may possess components of the renin-angiotensin system that permit intracellular processing of angiotensinogen to Ang II and that Ang II generated intracellularly may be mitogenic.
authors.affiliation: Division of Research, Ochsner Clinic Foundation, New Orleans, LA, USA. jcook@ochsner.org
authors.forename: J L
authors.initials: JL
authors.lastname: Cook
authors.forename: Z
authors.initials: Z
authors.lastname: Zhang
authors.forename: R N
authors.initials: RN
authors.lastname: Re
mesh_terms: D017398:Alternative Splicing; D057911:Angiotensin Receptor Antagonists; D000808:Angiotensinogen / Q000235:genetics / Q000378:metabolism; D000809:Angiotensins / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D000959:Antihypertensive Agents / Q000494:pharmacology; D001562:Benzimidazoles / Q000494:pharmacology; D001713:Biphenyl Compounds; D001973:Bromodeoxyuridine; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D002999:Clone Cells / Q000378:metabolism; D007424:Intracellular Fluid / Q000378:metabolism*; D019808:Losartan / Q000494:pharmacology; D008940:Mitotic Index; D016297:Mutagenesis, Site-Directed; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D007703:Peptidyl-Dipeptidase A / Q000096:biosynthesis; D016133:Polymerase Chain Reaction; D051381:Rats; D011945:Receptors, Angiotensin / Q000378:metabolism*; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012083:Renin / Q000096:biosynthesis / Q000235:genetics; D017384:Sequence Deletion; D013777:Tetrazoles / Q000494:pharmacology; D014162:Transfection; D014407:Tumor Cells, Cultured; D000092502:Renin Inhibitors
chemical_list: D057911:Angiotensin Receptor Antagonists; D000808:Angiotensinogen; D000809:Angiotensins; D000959:Antihypertensive Agents; D001562:Benzimidazoles; D001713:Biphenyl Compounds; D001973:Bromodeoxyuridine; D019808:Losartan; D016376:Oligonucleotides, Antisense; D007703:Peptidyl-Dipeptidase A; D011945:Receptors, Angiotensin; D018160:Receptors, Cytoplasmic and Nuclear; D011994:Recombinant Proteins; D012083:Renin; D013777:Tetrazoles; D000092502:Renin Inhibitors; C081643:candesartan