Angiotensin-converting enzyme: Accumulation in medium from cultured endothelial cells

Hayes, Lyle W.; Goguen, Cheryl A.; Ching, Simon; Slakey, Linda L.

doi:10.1016/0006-291x(78)90306-6

Cited by 68 publications

(25 citation statements)

References 17 publications

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“…This difficulty has often not been acknowledged but has recently been highlighted by others (18). There are, however, reports of production of ACE by endothelial cells cultured in medium free of ACE activity (19,20).…”

Section: Resultsmentioning

confidence: 99%

Induction by Glucocorticoids of Angiotensin Converting Enzyme Production from Bovine Endothelial Cells in Culture and Rat Lung In Vivo

Mendelsohn¹,

Lloyd²,

Kachel³

et al. 1982

J. Clin. Invest.

130

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A B S T R A C T The effect of corticosteroids on angiotensin converting enzyme was investigated in endothelial cell cultures and intact rat lung. Cultured endothelial cells from bovine aorta showed net production of angiotensin converting enzyme (ACE) over 2 d culture in serum-free medium. Dexamethasone (DM) increased cell ACE activity six-to sevenfold at 100 nM with a threshold effect at 0.3 nM. The effect of DM on ACE production was completely inhibited by actinomycin D or cycloheximide. Deoxycorticosterone (DOC) and aldosterone were markedly less active, with a threshold near 100 nM and significant (two to threefold) stimulation of ACE activity at 1 ,uM. In cells incubated in the presence of 10 nM DM, DOC (10 ,iM) significantly inhibited ACE production compared with 10 nM DM alone, suggesting that DOC is a partial agonist/partial antagonist in this enzyme system. Protein content of cells or medium was unchanged by steroids at all doses used.In vivo, adrenalectomized rats showed lower pulmonary ACE compared with intact controls, and when injected with DM (40 ,ug/d for 4 d) showed a significant (twofold, P < 0.002) increase in lung ACE over oil-injected, adrenalectomized controls; serum ACE did not change. Injection with DOC (40 jg/d) or aldosterone (10 jg/d) had no effect on lung or serum ACE. Over a range (0.6 to 2,000 jig) of concentrations of DM administered daily for 7 d, the dose-response curve of DM for induction of pulmonary ACE mirrored that for thymolysis; for both, half-maximal effects were seen at -6 jg DM/d, and plateau levels at 60 Ag/d.Received for publication 23 November 1981 and in revised form 18 May 1982. We conclude that glucocorticoids are potent inducers of ACE activity in endothelial cells in culture and in rat lung in vivo, and that the action of aldosterone and DOC reflects occupancy of glucocorticoid receptors. This effect may be of (patho)physiological relevance in regulating levels of ACE in local vascular beds, and thereby modulating local levels of the vasoactive peptides angiotensin II and bradykinin.

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Section: Resultsmentioning

confidence: 99%

Induction by Glucocorticoids of Angiotensin Converting Enzyme Production from Bovine Endothelial Cells in Culture and Rat Lung In Vivo

Mendelsohn¹,

Lloyd²,

Kachel³

et al. 1982

J. Clin. Invest.

130

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“…7 Here we present evidence that 3 H-leucine is incorporated into the cellular converting enzyme, and later appears in the enzyme in the culture medium. The amount of cell-associated enzyme activity remains constant, and the amount of activity in the medium increases linearly during this period.…”

Section: Discussionmentioning

confidence: 68%

“…Cells were collected by trypsinization as described before. 7 The cell pellet from each flask was extracted with 0.5 ml of buffer containing 0.5% NP-40, 10 mM sodium phosphate, pH 7.4, 0.15 M NaCI and 1 mM phenylmethylsulfonylfluoride. The extract was centrifuged at 23,000 x g for 30 minutes and the supernatant solution was used for isolation of the enzyme.…”

Section: Isolation Of Radiolabeled Anglotenslncon Vert Ing Enzyme Fromentioning

confidence: 99%

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Angiotensin-converting enzyme in cultured endothelial cells. Synthesis, degradation, and transfer to culture medium.

Ching¹,

Hayes²,

Slakey³

1983

Arteriosclerosis

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Cultured endothelial cells from swine aorta possess the ecto-enzyme angiotensinconverting enzyme (E.C.3.4.15.1) in cell-associated form and also release it in soluble form into the culture medium. Using antibody to purified converting enzyme from swine kidney and incorporation of 3 H-leucine, we examined the synthesis, degradation, and release of enzyme into the medium.3 H-leucine is incorporated into cellular converting enzyme, and later appears in the enzyme in the culture medium. The amount of cell-associated enzyme activity remains constant, and the amount of activity in the medium increases linearly during this period. These data show unequivocally that the appearance of enzyme activity in the cell culture medium is accompanied by synthesis of new enzyme protein.In pulse-chase experiments, the radioactivity disappeared from the cellular enzyme in two kinetic components with apparent half-lives of 1-2 hours and 20 hours, respectively. The appearance of the radiolabel in the medium enzyme corresponds very closely in rate and amount to the slow disappearance from the cells. There was no apparent uptake of labeled medium enzyme by the cells. The data suggest that the pool of active enzyme on the cell surface is the immediate precursor of the medium enzyme. The effects of culture conditions on the turnover of angiotensin-converting enzyme were also examined. The incorporation of 3 H-leucine into both cell and medium enzyme was greater when cells were maintained in medium containing serum than when they were maintained in serum-free medium. The rate of degradation of the cellular enzyme was similar under the two culture conditions. (Arteriosclerosis 3:581-588, November/December 1983)

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“…Factor VIII antigen is associated with cells fromn human veins (12,15) and with cells from aortas of animials (19,21,24). Angiotensin I convertinig enzyme activity is found in cells from umbilical cord veins (15,16) and also in aortic endothelial cells from various species (20,28,29). Cultured endothelial cells can transport serotonin (22) and adenine nucleotides (20), and they can form prostacyclin in response to mechanical or enzymatic stimulation (30)(31)(32)(33).…”

Section: Introductionmentioning

confidence: 99%

Human Pulmonary Endothelial Cells in Culture

Johnson¹

1980

J. Clin. Invest.

204

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A B S T R A C T Encdothelial cells were cutltured from various different human vessels, including aortas, pulmonarv, ovarian, and umbilical arteries, an(l pulmonary, ovarian, an(l umilbilical veins. The culturedl cells were identifie(d as endothelial cells by the presence of Factor VIII antigeni and antiotenisini I convertinig enzyme (kininlase II). They retainedl these markers for at least five passages in culture, and somle cells had them for seven passages or nmore. Endothelial cells from the various vessels were comiiparedl with respect to their ability to metabolize angiotensins I and II and bradykinin. Cells fromi arteries had three to five timles the angiotenisini I converting enzyme activity as cells from veins. The activity of angiotensinase A (aspartyl ami nopeptidlase) had a similar distribution, andl cells from arteries were conisistenitly more active than cells fromn veins. Cultures of endothelial cells from pulmonary and umbilical vessels formed prostacyclin in responise to mechanical stimulationi. MIedia from cell moniolayers that were subjecte(l to a change of medliumii and gentle agitation inhibited aggregation of humilani platelets. This inhibitory activity was generated within 2-5 mnm, and it was not formed by cells that were treated with indomnethacini or tranylcypromiine. Additioni of prostaglanidin (PG)H2 to indomethacin-treated cells restored the ability to form the inhibitor, but cells treated with tranylcypromine were not responsive to PGH2. In experiments where [ 14C ]arachidonic acid was added to the cells before stimulation, the major metabolite identified by thin-layer chromatography was 6-keto PGF1t. Thus, it appears that pulmonary endothelial cells, as well as umbilical cord cells, can form prostacyclin. In experiments comparing the ability of arterial and venous cells to form prostacyclin, arterial cells were more active than venous cells. These studies of cells from various human vessels suggest that the vascular origin ofculturecl endothelial cells determines how they metabolize vasoactive peptides and form prostacyclin.

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Angiotensin-converting enzyme: Accumulation in medium from cultured endothelial cells

Cited by 68 publications

References 17 publications

Induction by Glucocorticoids of Angiotensin Converting Enzyme Production from Bovine Endothelial Cells in Culture and Rat Lung In Vivo

Induction by Glucocorticoids of Angiotensin Converting Enzyme Production from Bovine Endothelial Cells in Culture and Rat Lung In Vivo

Angiotensin-converting enzyme in cultured endothelial cells. Synthesis, degradation, and transfer to culture medium.

Human Pulmonary Endothelial Cells in Culture

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