OBJECTIVE; To evaluate the adhesion, proliferation and osteogenic differentiation of rabbit BMSCs after cultured on freeze-dried demineralized bone matrix (FDBM) modified with type II cadherin ectodomain (Cad-II).
Methods: BMSCs isolated from 10 Japanese white rabbits (male and female, 4-week-old, 0.61-0.88 kg) were cultured. The second generation of BMSCs (cell density 1 x 10(6)/mL) were seeded onto the Cad-II modified allogenic FDBM (experimental group) and only FDBM (control group) respectively, and then cocultured in vitro. The densities of seeded cells, the adhesion rate and their ALP activity were measured. The complex was observed through inverted phase contrast microscope and scanning electron microscope to evaluate the interaction between cells and FDBM. Another group of second generation of BMSCs (cell density 5 x 10(5)/mL) were seeded onto the Cad-II modified FDBM (experimental group) and only FDBM (control group) respectively, and then cocultured in vitro too. The ALP activity and osteocalcin immunohistochemical was measured.
Results: There was no significant difference in cell proliferation between experimental group and control group. The adhesion rate of cells in the experimental group was 87.41% +/- 5.19%, higher than that in the the control group 35.56% +/- 1.75% (P < 0.01); the densities of seeded cells reached 5.0 x 10(5), showing significant difference compared with the control group (2.6 x 10(4), P < 0.05). Inverted phase contrast microscope showed that in the experimental group, more cultured BMSCs pasted in the hole and edge of the scaffold than that in the control group. HE staining showed the densities of seeded cells in the experimental group was higher than that in the control group. Scanning electron microscope showed that in the experimental group, a lot of cultured BMSCs adhered, spreaded in the scaffold, in the control group only a few BMSCs unevenly distributed in the scaffold. After 7 days of culture, the cultured BMSCs on modified FDBM expressed higher ALP activity; after 14 days of culture, the ALP activity (29.33 +/- 1.53) was higher than that cultured on unmodified FDBM (18.31 +/- 1.32), the positive rates of osteocalcin were 83% +/- 7% in the experimental group and 56% +/- 7% in the control group, showing significant difference (P < 0.01).
Conclusion: Cad-II enhanced cell adhesion to FDBM and promoted BMSCs differentiate to osteoblast, but no obvious effects were observed in cell proliferation.