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. BMSCs seeded onto fibroin sponge scaffolds were cultured by using the stirring chamber system (Figure 1), which can apply a relative tribological movement to the surface of the specimens. Three culture conditions were applied (dynamic in stirring chamber as frequency as 40 min [D1], as 40 sec [D2] and static in stirring chamber group [S]). The specimens were set into stirrer on a poly(MPC) grafted surface (MPC polymer coated surface, SANSYO). As a counter surface in friction tests, the poly(MPC) grafted surface was prepared by atom transfer radical polymerization, and the regenerated cartilage was prepared by seeding 5×105 cells (BMSCs from rat bone marrow) onto fibroin sponge scaffolds (8 mm diameter and 1 mm thickness) and by 14 days culture.Introduction
Material and methods
Tamura et al. proposed a new friction test to measure the maturity of surface gel-hydration-like lubrication using MPC-polymer (2-Methacryloyloxyethyl phos -phorylcholine polymer) grafted surface as aãζζcounter surface. They suggested that the MPC-polymer grafted surface makes it possible to mimic in-vivo-like condition. Therefore, we can evaluate a lubricating ability of cartilage surface except for the possible effects of deformation resistance. By the way, reduction of lubricating ability of articular cartilage surface has much to do with pathogenesis of primary osteoarthritis. On the other hand, intraarticular injections of hyaluronic acid (HA) has been reported to have some clinical effect, however, it has not been clearly supported that HA restores a lubricating ability of injured cartilage surface. In the present study, the short-term effect of HA on injured cartilage surface's frictional performance was examined by the friction test using MPC-polymer grafted surface. Articular cartilage specimens were taken from porcine femoral condyle and cut into 5 mm diameter plugs. Their surfaces were wiped with particular papers soaked in saline solution. Thereafter, these specimens were preserved with 1 mL volume of HA and saline solution for 0, 3, 6, 9 hours. The concentration of HA was 1% (w/v) in saline solution (MW=9×105 Daltons; Seikagaku corp., Tokyo, Japan). Friction test was carried out in saline solution under a constant pressure of 1.5 Mpa and a relative sliding velocity of 0.8 mm/s, with MPC-polymer grafted glass as counter surface. Besides, superficial layer of cartilage tissue was histologically observed by two kinds of staining method: Toluidine blue (pH7.0) staining and Toluidine blue (pH2.5) staining Then, the Toluidine blue (pH7.0) staining intensity on superficial tissue was quantitatively analyzed. As follows, images of the stained cartilage specimens were analyzed by ImageJ. Measure RGB program was used to average out luminance values of blue in 2.7 μm square area of superficial layer and middle layer. The ration of the mean value in superficial layer and it in middle layer was defined as Toluidine blue (pH7.0) Index.INTRODUCTION
METHODS
