Introduction

It is essential to investigate the tribological maturation of tissue-engineered cartilage that is to be used in medical applications. The frictional performances of tissue engineered cartilage have been measured using flat counter surfaces such as stainless steel, glass or ceramics. However, the measured friction performances were significantly inferior to those of natural cartilage, likely because of cartilage adhesion to the counter surface. Tamura et al. reported that a poly (2- methacryloyloxyethyl phosphoryl-choline (MPC)) grafted surface shows low friction coefficient against cartilage without the adhesion to be equivalent to those for natural cartilage-on-cartilage friction. [1]

On the other hand, Yamamoto et al. reported that applying a relative sliding movement had a potential to alter the expression of tribological function of regenerated cartilage of chondrocytes. [2] In this paper, the effects of the relative sliding movement on the expression of bone marrow stromal cells (BMSC)s were investigated using the poly(MPC) grafted surface as a counter surface.

INTRODUCTION

Several reports suggest that low-intensity pulsed ultrasound stimulation (LIPUS) facilitates chondrogenesis¹⁾. Recently it has been suggested that LIPUS may be transmitted via Integrin: a protein which mediates cellular attachment between cells and extracellular matrix²⁾. In this study, the Arg-Gly-Asp (RGD) amino acid sequence, which is a ligand of Integrin, was induced to the fibroin substrates by either gene transfer or physical mixing, and the variation of chndrocyte response to LIPUS was evaluated.

Physical environments play important roles for maturation of mechanical functions of tissue. In this study, effects of relative tribological movement on the expression of tribological function of regenerated synovial membrane were investigated. Fibroin sponge derived from silk was used as a three-dimension scaffold for the synovial membrane regeneration. Synovial cells were isolated from human synovial membrane, and were seeded onto the fibroin sponge. Magnetic stirring system (named Stirring Chamber) was used for culturing with relative slip motion where the cell-seeded side of the scaffold had been rubbed by a glass culture dish for 24 hours/day.

Histological view of regenerated tissue of the dynamically cultured group (D group) showed extracellular-matrix-like eosinophilic meshwork structure formed continuously on the meshwork structure of the fibroin sponge. The newly formed tissue showed expression of collagen type I, especially on the surface of fibroin sponge. These structures were not seen in the statically incubated group (S group). Each group didn’t show expression of collagen type II.

Frictional force was measured by using leaf spring method under the conditions of the sliding velocity: 0.8 mm/s, the loading time prior to sliding: 1 minute, and the applied load during the experiment: 0.029 N. The counterface for regenerated synovium was a flat stainless steel of which roughness was 0.06 μm Ra. All frictional experiments were performed in the saline solution and at room temperature (25°C). The friction coefficient of tissues cultured statically was 0.6–0.8, and that of tissues cultured with sliding motion was 0.2–0.4 at one week culturing, 0.3–0.5 at two weeks culturing.

Our previous experiment showed that combination of fibroin-sponge scaffold and Stirring-chamber culturing system improved the tribological performance of regenerated cartilage tissue. The present study suggests that this combination have also a possibility for synovial cells to form functional lubricious membrane which can be used as anti-adhesion membrane for knee, ligament, and/or other surgical procedures. However, the deterioration of lubrication properties in the 2 weeks dynamically cultured group would indicate that the too long continuous tribological movement does not provide an optimal condition. More fine tribological loading history should be designed.

Introduction: Fibroin sponge is purified silk protein from which high-strength gel sponge can be produced. The purified fibroin sponge causes no immune response. This study evaluates unique performances of the fibroin sponge for articular-cartilage regeneration, and mechanical properties of regenerated cartilage were also measured.

Methods: Refined silk yarn was dissolved in 9M lithium bromide aqueous solutions, and was frozen in −20& #8451 freezer for 12 hours. Hydrogel sponge was formed under the room temperature. Articular cartilage slices were taken from the proximal humerus, distal femur and proximal tibia of 4-week-old Japanese white rabbits. The cartilage slices cut into small pieces and were digested with 0.25% trypsin in DMEM containing antibiotics for 30 min at 37& #8451. After rinsing with Tyrode’s balanced solution and centrifuging at 180 G for 5 min, the chondrocytes were isolated with 0.25% collagenase for 8 h at 37& #8451. These cells were harvested and inoculated into the fibroin sponge. The constructs of the chondrocytes and the fibroin sponge were cultured in DMEM containing 10& #65285 FCS and 50mL L-ascorbate for 4weeks. Indentation test and dynamic visco-elastic measurement were carried out for these constructs.

Results and discussion: Cell density of the inoculated chondrocytes was increased to about five times as much as initial volume. This regenerated tissue was intensely stained with safranin-O fast green and showed a meta-chromatic reaction. This also stained positively with immunostain for type & #8545 collagen, but negatively with immunostain for type & #8544 collagen. Mechanical tests showed that time constants of compressive creep and E’ values were increased with cultivation days, and the peak value and frequency of tan& #948 shifted to a lower amount. The change in dynamic visco-elastic properties of the regenerated cartilage is caused by synthesis of extracellular matrix.