Ciprofloxacin Decreases Collagen in Mouse Tympanic Membrane Fibroblasts

Abstract

Objectives To determine how collagen production by tympanic membrane fibroblasts is affected by ciprofloxacin at levels found in eardrops. Study Design Prospective, controlled, and blinded cell culture study. Setting Academic tertiary medical center. Subjects Cell culture of mouse fibroblasts. Methods A primary fibroblast culture was established from mouse tympanic membranes. Fibroblasts were cultured until they were 75% confluent, then treated with dilute hydrochloric acid (control) or ciprofloxacin (0.01% or 0.3%) for 24 or 72 hours for Western blotting and for 24 or 48 hours for cytotoxicity assay. Cells were observed with phase-contrast microscope. Western blotting was performed for collagen type 1 α1 (collagen 1A1) and α-tubulin. Results Fibroblasts treated with 0.01% and 0.3% ciprofloxacin for 24 hours had lower levels of collagen 1A1 ( P = .0005 and P < .0001, respectively) and α-tubulin (both P < .0001) than control fibroblasts. Collagen 1A1 and α-tubulin levels were lower in fibroblasts treated with 0.3% than with 0.01% ciprofloxacin ( P = .02 and P = .014). After 72 hours, 0.3% ciprofloxacin completely eliminated collagen 1A1 and α-tubulin ( P < .001). Cells treated with 0.01% ciprofloxacin for 72 hours also had lower collagen 1A1 ( P < .0001) and α-tubulin ( P = .005) as compared with the control. Seventy-two-hour incubation in 0.01% or 0.3% ciprofloxacin resulted in lower levels of collagen 1A1 ( P = .009 and P < .0001, respectively) and α-tubulin ( P = .007 and P < .0001, respectively) than 24-hour incubation. Cytotoxicity assay and phase-contrast microscopy mirrored these findings. Conclusions Treatment of tympanic membrane fibroblasts with 0.3% ciprofloxacin, as found in eardrops, reduces fibroblast viability and collagen and α-tubulin protein levels. These findings could explain tympanic membrane healing problems associated with quinolone eardrops.