Immunomorphologic studies of mast cell heterogeneity, location, and distribution in the rat conjunctiva

Mast cells are crucial components of immediate and some delayed-type hypersensitivity reactions. They play a pivotal role in allergic conjunctivitis and other immunoinflammatory disorders of the ocular surface, yet little is known of their distribution and heterogeneity in the conjunctiva of potential animal models, such as the rat. In this study, mast cell types were investigated in histologic sections and corneal-conjunctival-lid whole mounts by using toluidine blue, alcian blue-safranin, and immunohistochemical staining methods (anti-rat mast cell proteinase [RMCP] antibodies). Quantitative analyses were performed on corneal-conjunctival-lid whole mounts by using the optical dissector procedure to obtain the density of mast cells per unit volume in different regions of the conjunctiva. Single and double immunohistochemical analyses revealed that the mast cells in the conjunctiva of the limbus, fornices, and lid margin were strongly RMCP I+, suggesting that they were of the connective tissue phenotype. Mast cells containing the mucosal mast cell proteinase RMCP II were not present in the normal conjunctiva. Histochemical analysis revealed that the maturity of the connective tissue mast cells, as assessed by the presence or absence of safranin (heparin)-positive granules in their cytoplasm varied in different regions. In the lid margin 60% to 78% of the mast cells were solely alcian blue-positive, whereas in the fornices 68% to 78% were safranin-positive. In the limbus the predominant type of mast cell was either safranin-positive or contained mixed granules. Mast cell densities were greatest close to the lid margin (10,000 to 12,000 cells/mm3), followed by the limbus (3400 to 4800 cells/mm3) and were rare in the remainder of the conjunctiva (500 to 1000 cells/mm3), with the exception of the region around the nictitating membrane. This study of rat conjunctival mast cells provides essential baseline data for future studies of the role of mast cells in models of allergic conjunctivitis.