Influence of dynamic mechanical stimulation on tissue engineering annulus fibrosus

Abstract

Objective To investigate the influence of dynamic mechanical stimulation on the annulus fibrosus (AF) cells seeded on silk scaffolds. Methods AF cells were isolated from rabbits and were seeded on the scaffold, then cultured for 3, 7, 14 days with different range of dynamic compression. Stereomicroscope and scanning electron microscope (SEM) was used to observe the surface morphology of tissue engineering annulus fibrosus cells (TE-AFs). After fixation, samples were harvested for histological staining. AF cells related extracellular matrix (ECM) was evaluated by the quantitative analysis of total DNA, proteoglycan and collagen I. The mechanical properties were compared within different groups. Results Stereomicroscope and SEM results showed that the colors of TE-AFs in all groups were deepening with time going. SEM showed cell adhesion on the scaffold and the secretion of extracellular matrix. Histological, immunohistochemical staining, biochemical quantitative analysis and total DNA content showed that the AF cells inside scaffolds could support AF cell attachment, proliferation and secretion. As a result, the compressive properties were enhanced with increasing culture time. Stereomicroscope showed that the colors of TE-AFs in all groups were deepening with time going after dynamic compression. HE staining, Safranin O staining and Type I collagen staining showed that cell proliferation and secretion, GAG secretion and collagen secretion were increased with time going within different groups. Quantitation of GAG achieved maximum in 15% strain group, and quantitation of collagen achieved maximum in 10% strain group. The total DNA content achieved maximum in 5% strain group, and compression elastic modulus achieved maximum in 15% strain group. The height of TE-AFs did not change after mechanical stimulation for 14 days. Conclusion Suitable mechanical stimulation is a positive factor for new AF tissue engineering that will tend to the nature tissue. Excessive compression can accelerate the progress of cell apoptosis. Key words: Tissue engineering; Silk; Tissue scaffolds; Biomechanics