Nicardipine inhibits human conjunctival fibroblastic proliferation, migration and attachment in vitro

Abstract

Abstract Purpose: To evaluate the effect of the calcium channel blocker nicardipine (NIC) on fibroblastic proliferation, migration, attachment and cell viability on human conjunctival fibroblasts in vitro Methods: Human conjunctival fibroblasts were incuvated in a humidified incubator (37ºC‐5%CO2), plus the doses of NIC to be tested. Fibroblastic proliferation at 18, 24 and 48 h was evaluated with a colorimetric assay based on the cleavage of the tetrazolium salt WST‐1 by mitochondrial dehydrogenases in viable cells. Cell viability was evaluated at 4 h with the WST‐1 asssay and with trypan blue (TB) staining. Migration was determined in a artificial wound made in a confluent fibroblast monolayer. Cell free areas were monitored with digitallized images at 0, 18, 24 and 48 h. Fibroblastic attachment was evaluated at 24 h with the WST‐1 asssay. Groups: 9 controls and 9 NIC concentrations (10‐4, 7.5x10‐5, 5x10‐5, 2.5x10‐5, 10‐5, 7.5x10‐6, 10‐6, 10‐7 and 10‐8 M). Each experiment was carried out in triplicate and repeated 4 times Results: NIC < or = 10‐5 M showed cell viability >98% with TB and >99% with WST‐1 at 4 h. DI50=1.018x10‐4 M. NIC > or = 10‐5 M at 18 h, and NIC > or = 7.5x10‐6 M at 24 and 48 h showed significantly less proliferation than control groups. DI50 in 7.5x10‐5‐5x10‐5 M rank. NIC > or = 10‐5 M showed significantly larger cell‐free areas at 18, 24 and 48 h. NIC < or = 7.5x10‐6 M didn´t showed differences among the control groups. NIC > or = 7.5x10‐6 M inhibited fibroblastic attachment in >23% at 24 h. DI50=1.107x10‐3 M Conclusions: NIC have a significant inhibitory effect in vitro on conjunctival fibroblastic wound healing process with potential applicability in ophthalmic surgery