The stimuli releasing histamine from murine bone marrow-derived mast cells (BMMC). 3. Effect of coculture with 3T3 fibroblasts on the histamine releasability of BMMC

Abstract

Murine bone marrow-derived mast cells at 3 wk in culture were further cultured in the absence (N-BMMC) or presence (F-BMMC) of 3T3 fibroblasts in the medium containing LI-3, and were examined for their functional responses to either histamine releasing stimuli such as compound 48/80 or an inhibitor of histamine release, disodium cromoglycate. After 3 weeks in coculture with 3T3 fibroblasts, the mast cells increased their histamine content greater than 10 fold, and greater than 10% of the cells changed histochemically to become safranin positive. F-BMMC released approximately 10% histamine when challenged with compound 48/80 or substance P, whereas N-BMMC failed to do so. Furthermore, when the sensitized cells were challenged with DNP-HSA antigen, histamine release from F-BMMC but not from N-BMMC was inhibited by preincubation with disodium cromoglycate. We also examined changes in intracellular Ca2+ ([Ca2+]i) in the cells when challenged with compound 48/80. A transient increase in [Ca2+]i was observed on stimulation with the compound in F-BMMC but not in N-BMMC. Taken together, our results indicate that the interleukin 3-dependent cultured murine mast cells change functionally, as well as histochemically, into in vitro counterparts of connective tissue mast cells when cocultured with 3T3 fibroblasts and may be useful tools for analyzing the mechanisms involved in degranulation from connective tissue-type mast cells.