Rapid degradation of neurotensin by stimulated rat mast cells.
A RIA towards neurotensin (NT) using C-terminal- and N-terminal-specific antisera was used to study degradation of this tridecapeptide by isolated rat mast cells. Incubation of NT (10 microM) with peritoneal or pleural mast cells resulted in a rapid loss of NT immunoreactivity (iNT), as measured by C-terminal-directed antiserum, with little effect on N-terminal iNT. The rate of the reaction was faster with pleural cells (T1/2, 30 s) than with peritoneal cells (T1/2, 180 s) and was greater than 10-fold slower in the presence of metabolic poisons. The enzyme(s) involved is most likely released from the cells during secretion, as NT was degraded by media conditioned by compound 48/80-stimulated mast cells 40-60 times faster than by media from unstimulated cells. This degradation by conditioned media was concentration dependent, pH dependent, and temperature sensitive. HPLC analyses indicated a near stoichiometric conversion of NT to NT(1-12) (66%) and NT(1-11) (34%) after incubation for 10-30 s with conditioned media. By 30 min only NT(1-11) and NT(1-10) were present. Phenanthroline (1 mM), an inhibitor of carboxypeptidase, prevented the loss of C-terminal iNT and the generation of NT(1-12) and NT(1-11). While NT(1-12) was effective in releasing histamine from mast cells in vitro and increasing vascular permeability in vivo, NT(1-11) was not. These results suggest that carboxypeptidase-like enzyme(s) could modulate the level and form of NT-related peptides in various states involving activation of mast cells.
Cochrane
D E
DE
Department of Biology, Tufts University, Medford, MA 02155.
Carraway
R E
RE
Boucher
W
W
Feldberg
R S
RS
D000595:Amino Acid Sequence; D000818:Animals; D006636:Histamine Release / Q000187:drug effects; D066298:In Vitro Techniques; D008297:Male; D008407:Mast Cells / Q000187:drug effects / Q000378:metabolism*; D008969:Molecular Sequence Data; D009496:Neurotensin / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains
D010446:Peptide Fragments; D009496:Neurotensin; C059582:neurotensin (1-11)