Surface Reactivity of Calcium Phosphate Based Ceramics in a Cell Culture System

Abstract

Surface reactivity of Calcium Phosphate materials - Hydroxyapatite (HA), Tricalcium Phosphate (β-TCP), Hydroxyapatite-Tricalcium Phosphate (HA-TCP) were elucidated in a cell culture system. MG-63 osteoblast-like cells were seeded onto the ceramic discs to evaluate changes in the cell morphology and functionality with respect to the different substrates. The dissolution and re-precipitation of calcium phosphate phases on the surface of the discs in the culture medium was found to be prominent on β-TCP when compared with HA. Low calcium (Ca), magnesium (Mg) and alkaline phosphatase (ALP) levels and high phosphorous (P) levels in the medium of β-TCP were observed. This indicated that P must have leached out into the medium from β-TCP and Ca in turn deposited from the medium onto β-TCP resulting in the apatite phase transformation. The low ALP activity in β-TCP medium is however an indication of low osteoblastic activity. Under the phase contrast microscope, the osteoblast cells around HA material were found to be confluent and viable, while in the vicinity of β-TCP only cellular debris was observed. In the case of HA-TCP, only a few viable cells surrounded the material amidst the debris. Scanning electron microscopy revealed numerous cells on the surface of HA showing different cell behaviour like anchorage, attachment, adhesion and spreading in the early time period as the surface was only slightly disturbed with re-crystallisation. But with time the entire surface of HA had changed due to precipitation and re-crystallization which did not support cell behaviour while the cells surrounding the material showed normal growth. On the contrary, cells were scarcely observed on the entirely changed surface of β-TCP and HA-TCP even from the earlier days of the culture and the morphology of cells surrounding the material too started changing. These results establish that HA promoted the activity of osteoblast cells. HA surface remained unaltered for some time, while the surface of β-TCP underwent dissolution of surface ions and resulted in the re-crystallization of apatite over the surface. The resulting changes in the surrounding milieu of β-TCP with high phosphate and low Ca levels probably was responsible for the death of the cells.