Different apatite structures47 seeded with MC3T3-E1 cells showed lower cell number compared with tissue culture Palbociclib price plastic after different time points (4 and 14 d) and Anselme and coworkers43 showed that proliferation of human bone derived cells on plasma sprayed hydroxyapatite (HA) coatings was only possible after prolonged soaking of the coated scaffolds in culture medium. In contrast, PLLA films coated with apatite or collagen/apatite blend showed a significantly higher proliferation of Saos-2 cells compared with bare PLLA films.48 It is therefore difficult to draw general conclusions on the effect of Ca-P on proliferation of MSCs.
In the present study, however, the effect of Ca-P coating on cell number was only visible for hybrid scaffolds, and not for 3DF ones, which indeed suggests that the ��clogging�� effect caused by physical presence of the Ca-P layer may be of bigger importance that the chemical effect of presence of Ca-P or release of calcium and phosphate ions. While in vitro studies on combination of ESP and 3D RP scaffolds have been performed,19,20,42 they have mainly assessed cell proliferation, morphology and biochemical expression of typical markers like ALP and GAG on cell lines or animal derived cells. In order to assess applicability of these technologies in tissue repair and regeneration, experiments with human cells are of importance prior to in vivo testing. Therefore, we seeded our scaffolds with bone marrow derived hMSCs and analyzed the gene expression of various osteogenic markers at two different time points ��day 7 and day 21.
The applied Ca-P coating comprises a mixture of OCP and CA, biologically relevant phases of Ca-P. The bioactivity of Ca-P coatings in a bony environment that is believed to originate in degradation of Ca-P is the main reasons for their use in orthopedic and maxillo-facial implants. This degradation leads to an increase in local ion concentration in the vicinity of the implant, resulting in subsequent precipitation of a bone like carbonated apatite on the substrate.49 Previous studies performed on similar coatings have shown the formation of a carbonated apatitic phase two weeks after an OCP coated Ti plate was placed in ��-MEM49 suggesting that the degradation process starts earlier. In the current experimental set up, the released calcium and/or phosphate ions plausibly affected differentiation of hMSCs.
Tada and coworkers observed increased BMP-2 expression50 in dental pulp cells due to elevated levels of calcium, which is in accordance with our results using hMSCs. Another study51 showed that at calcium concentrations greater than 6 mM, MC3T3E1 Batimastat osteoblasts showed enhanced mineralization and expression of angiopoietin-1 (Ang-1) that promotes the structural integrity of blood vessels and variation in expression of angiopoietin-2 (Ang-2), a naturally occurring antagonist for promoting blood vessel growth.