Human conjunctival mast cell responses in vitro to various secretagogues

Human conjunctiva was enzymatically digested into monodispersed cell preparations using collagenase and hyaluronidase. The preparations were enriched for mast cells using Percoll gradient centrifugation prior to challenge with various secretagogues. Mast cell percentage (9.7 ± 2.9%) and viability (96 ± 3%) were determined using toluidine blue staining and vital dye exclusion, respectively. The mast cell enriched preparations were challenged with calcium ionophore A(23187') compound 48/80, concanavalin A, anti-IgE, morphine and substance P. Mediator release was quantified by radioimmunoassay. The calcium ionophore A(23187), compound 48/80, concanavalin A and anti-IgE stimulated histamine release from the conjunctival mast cell preparations, with EC(30) values of 0.028, 62.5, 24.4 and 1.9 ϵg/ml, respectively. Morphine but not substance P challenge induced significant but low levels of histamine release. Tryptase, sulfidopeptide leukotriene, and prostaglandin D(2) levels in cell supernatants were also increased following stimulation. The profile of mediator release observed with these cells compared to that reported from human lung and human skin mast cells suggest that human conjunctival mast cells may have unique biological responses.