Enhanced Collagen Production by Smooth Muscle Cells During Repetitive Mechanical Stretching

Abstract

We examined the effect of repetitive mechanical stretching on smooth muscle cell (SMC) collagen production. Porcine SMCs from passages 3 through 7 were seeded in 35-mm2 flexible-bottomed culture wells at a concentration of 2 x 10(5) cells per well and allowed to attach for 24 hours. The experimental group was placed in a vacuum-operated stress-providing instrument that exerted an average elongation of 25% at maximum downward deflection of the culture plate bottom. The stretched cells (nine wells per day) were subjected to a cyclic force regimen of 10 s of elongation and 10 s of relaxation for five days. The control cells (nine wells per day) were subjected to incubation conditions similar to those in the experimental group but without cyclic stretching. Twenty-four hours before harvesting, serum-free medium containing 50-microCi tritiated proline, an amino acid hydroxylated in collagen (hydroxyproline), and 50 micrograms/mL of ascorbate was added per well. On days 3 and 5 the medium and cells were collected, precipitated with trichloroacetic acid, and then sedimented, lyophilized, and analyzed to separate hydroxyproline and proline. Values for collagen and noncollagen protein were calculated after quantitation of the hydroxyproline and proline concentrations. The results indicate that three-cycle-per-minute stretching coordinately stimulated SMC production of collagen and noncollagen protein. We conclude that pulsatile stretch enhances collagen and noncollagen protein synthesis.