Lysosomotropic agents selectively potentiate thrombin-induced acid hydrolase secretion from platelets.
Thrombin induces partial secretion (up to 60%) of beta-N-acetyl-D-hexosaminidase (EC 3.2.1.52) from untreated platelets. Preincubation of platelets with 10 mM NH4Cl for up to 2 hr resulted in a time-dependent and marked stimulation of thrombin-induced secretion of both this enzyme and other acid glycosidases from platelets. The enhancement of the thrombin-induced secretion was not due to cell lysis, and NH4Cl alone did not cause leakage of lysosomal enzymes into the medium. The effect could be reversed by reincubating the platelets in NH4Cl-free medium. Stimulation of thrombin-induced secretion also was produced by a series of aliphatic primary amines from methylamine to butylamine, and by micromolar concentrations of chloroquine. The effect of weak bases on platelets appeared to be quite specific for enhancing lysosomal enzyme secretion. Thrombin-induced secretion of adenine nucleotides from dense granules and of beta-thromboglobulin from alpha granules was slightly enhanced by NH4Cl but was slightly inhibited by methylamine. The only direct effect of the weak bases on platelets was the displacement of serotonin from dense granules. Accumulation of weak bases in acidic pools in the platelets (e.g., lysosomes) might, therefore, be responsible for the enhanced secretion of lysosomal enzymes. By using controlled digitonin-induced platelet lysis, it was found that preincubation of platelets with NH4Cl lowered the digitonin concentration required for enzyme solubilization. We suggest that loading of lysosomes with weak bases dissociates already bound enzyme inside the lysosomes, resulting in a more effective discharge upon stimulation by thrombin.
Van Oost
B A
BA
Smith
J B
JB
Holmsen
H
H
Vladutiu
G D
GD
D000135:Acid Phosphatase / Q000097:blood; D000588:Amines / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000187:drug effects; D005966:Glucuronidase / Q000097:blood; D006596:Hexosaminidases / Q000097:blood; D006801:Humans; D006867:Hydrolases / Q000097:blood*; D066298:In Vitro Techniques; D008247:Lysosomes / Q000187:drug effects / Q000201:enzymology*; D008361:Mannosidases / Q000097:blood; D013917:Thrombin / Q000494:pharmacology*; D001616:beta-Galactosidase / Q000097:blood; D001619:beta-N-Acetylhexosaminidases
D000588:Amines; D000643:Ammonium Chloride; D006867:Hydrolases; D000135:Acid Phosphatase; D006596:Hexosaminidases; D008361:Mannosidases; D001616:beta-Galactosidase; D005966:Glucuronidase; D001619:beta-N-Acetylhexosaminidases; D013917:Thrombin