Unmasking and redistribution of lysosomal sulfated glycoconjugates in phagocytic polymorphonuclear leukocytes

Rabbit heterophil and human neutrophil primary granules contain sulfated glycosaminoglycans (GAGs) and acid phosphatase, which can be readily stained in immature but not mature lysosomes. To determine whether this loss of staining represents masking of reactive components or removal of these components, we examined rabbit heterophils to see if high-iron diamine (HID)-reactive sulfate and acid phosphatase staining reappears in phagocytic vacuoles. Rabbit heterophils, obtained by peritoneal lavage, were incubated in vitro with latex beads or Pseudomonas aeruginosa for 15-60 min. Pre-embedment HID staining was enhanced in thin sections of unosmicated specimens with thiocarbohydrazide and silver proteinate (TCH-SP). Phagocytosis of latex beads or bacteria was progressively more prominent with time. Primary granules that were degranulated or in the process of degranulating into phagocytic vacuoles demonstrated intense sulfate staining with large (13 +/- 7 nm) HID-TCH-SP stain deposits. Smaller (6 +/- 1 nm) HID-TCH-SP stain deposits were present in tertiary granules, which were less frequently observed degranulating into phagosomes. Acid phosphatase staining was most intense during early phagolysosome formation. HID-TCH-SP staining was also observed in extracellular degranulated lysosomal matrices and on the surface of many peritoneal heterophils. These results indicate that loss of sulfate staining in mature heterophil granules is the result of masking by intragranular substances rather than of removal, and that these components may be unmasked during phagocytosis and/or redistributed to the cell surface after exocytosis.