Human Conjunctival Epithelial Cells Express Histamine-1 Receptors Coupled to Phosphoinositide Turnover and Intracellular Calcium Mobilization: Role in Ocular Allergic and Inflammatory Diseases

Abstract

Dispase-dissociated primary cultures of human conjunctival epithelial (HCE) cells were stimulated with histamine and the generation of inositol phosphates ([ 3H]IPs) from [ 3H]phosphoinositide (PI) hydrolysis and the mobilization of intracellular calcium ([Ca 2+] i) were studied using ion exchange chromatography and Fura-2 fluorescence techniques, respectively. Histamine (100μ M) maximally stimulated PI turnover in HCE cells by 210±10% ( n= 21) above basal levels and with a potency (EC 50) of 3.3μ M(n=4). Histamine (EC 50=5.8μ M, N=3) rapidly mobilized [Ca 2+] iwhich peaked within 10sec but which was still significantly elevated 20min after stimulation. The histamine-induced [Ca 2+] iresponses did not desensitize upon repeated applications of histamine. The effects of histamine (100μ M) on PI turnover and [Ca 2+] iwere potently antagonized by the H 1-antagonists, emedastine (IC 50=1.6–2.9n M), triprolidine (IC 50=3.1n M) and levocabastine (IC 50=8n m), but weakly by the H 2-(ranitidine/cimetidine) and H 3-(thioperamide) antagonists (IC 50s=10–100μ M). In conclusion, HCE cells have been shown to possess functional H 1-histamine receptors that couple to inositol phosphates generation which then mobilize intracellular calcium. These intracellular signaling mechanisms may be intimately linked with the process of inflammatory cytokine secretion from the HCE cells after stimulation by histamine released from the conjunctival mast cells. The current results strongly suggest that the HCE cells are active participants in mediating, and perhaps amplifying, the pro-inflammatory and allergic effects of histamine which is released from conjunctival mast cells during ocular allergic and inflammatory reactions.