Tetracycline-Based MMP Inhibitors Can Prevent Fibroblast-Mediated Collagen Gel Contraction In Vitro

Abstract

Collagen gels in vitro can be contracted by fibroblasts. The role of matrix metalloproteinases (MMPs) in the contraction of collagen lattices by human neonatal foreskin fibroblasts (HuFFs) was investigated in tissue culture media supplemented by various doses of known gelatinase inhibitors. Fluorescent assays with model gelatinase substrates and media conditioned by fibroblasts apparently confirmed the ability of chemically modified tetracyclines (CMTs) to act as inhibitors of MMP2, and zymography demonstrated that this was the major cell-derived MMP activity. There were no observable effects on the rate of contraction of attached FPCLs containing 6 x 104 HuFFs (passages 18-25) with either CMT-5 or CMT-2 at all concentrations tested (0-100 μg/mL). However, at greater than 20 μg/mL doxycycline and greater than 5 μg/mL CMT-3, FPCL contraction was completely abolished. Quantitative assessment of cell viability by means of the MTT assay in monolayer and qualitatively within the FPCLs with CalceinAM suggested that differences were not due to cytotoxic effects. Seeding FPCLs with lower-passage fibroblasts produced identical trends. These results may implicate the involvement of MMPs in the process of gel contraction, although tetracyclines have effects additional to their ability to inhibit MMPs directly.