Alpha 2-macroglobulin capture allows detection of mast cell chymase in serum and creates a reservoir of angiotensin II-generating activity.
J Immunol
; 182(9): 5770-7, 2009 May 01.
Article
em En
| MEDLINE
| ID: mdl-19380825
ABSTRACT
Human chymase is a highly efficient angiotensin II-generating serine peptidase expressed by mast cells. When secreted from degranulating cells, it can interact with a variety of circulating antipeptidases, but is mostly captured by alpha(2)-macroglobulin, which sequesters peptidases in a cage-like structure that precludes interactions with large protein substrates and inhibitors, like serpins. The present work shows that alpha(2)-macroglobulin-bound chymase remains accessible to small substrates, including angiotensin I, with activity in serum that is stable with prolonged incubation. We used alpha(2)-macroglobulin capture to develop a sensitive, microtiter plate-based assay for serum chymase, assisted by a novel substrate synthesized based on results of combinatorial screening of peptide substrates. The substrate has low background hydrolysis in serum and is chymase-selective, with minimal cleavage by the chymotryptic peptidases cathepsin G and chymotrypsin. The assay detects activity in chymase-spiked serum with a threshold of approximately 1 pM (30 pg/ml), and reveals native chymase activity in serum of most subjects with systemic mastocytosis. alpha(2)-Macroglobulin-bound chymase generates angiotensin II in chymase-spiked serum, and it appears in native serum as chymostatin-inhibited activity, which can exceed activity of captopril-sensitive angiotensin-converting enzyme. These findings suggest that chymase bound to alpha(2)-macroglobulin is active, that the complex is an angiotensin-converting enzyme inhibitor-resistant reservoir of angiotensin II-generating activity, and that alpha(2)-macroglobulin capture may be exploited in assessing systemic release of secreted peptidases.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Alfa-Macroglobulinas
/
Angiotensina II
/
Soro
/
Quimases
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Mastócitos
Tipo de estudo:
Diagnostic_studies
Limite:
Adult
/
Child
/
Humans
Idioma:
En
Revista:
J Immunol
Ano de publicação:
2009
Tipo de documento:
Article