John Annie, Varma H K, Kumari T V
Biomedical Technology Wing, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Poojapura, Thiruvananthapuram-695012, India.
J Biomater Appl. 2003 Jul;18(1):63-78. doi: 10.1177/0885328203018001006.
Surface reactivity of Calcium Phosphate materials--Hydroxyapatite (HA), Tricalcium Phosphate (beta-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 beta-TCP when compared with HA. Low calcium (Ca), magnesium (Mg) and alkaline phosphatase (ALP) levels and high phosphorous (P) levels in the medium of beta-TCP were observed. This indicated that P must have leached out into the medium from beta-TCP and Ca in turn deposited from the medium onto beta-TCP resulting in the apatite phase transformation. The low ALP activity in beta-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 beta-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 beta-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 beta-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 beta-TCP with high phosphate and low Ca levels probably was responsible for the death of the cells.
磷酸钙材料的表面反应性——羟基磷灰石（HA）、磷酸三钙（β-TCP）、羟基磷灰石-磷酸三钙（HA-TCP）在细胞培养系统中得到了阐明。将MG-63成骨样细胞接种到陶瓷圆盘上，以评估相对于不同底物细胞形态和功能的变化。与HA相比，发现培养基中圆盘表面磷酸钙相的溶解和再沉淀在β-TCP上更为显著。在β-TCP培养基中观察到低钙（Ca）、镁（Mg）和碱性磷酸酶（ALP）水平以及高磷（P）水平。这表明P必定从β-TCP中沥出到培养基中，而Ca则从培养基中沉积到β-TCP上，导致磷灰石相转变。然而，β-TCP培养基中低ALP活性表明成骨细胞活性较低。在相差显微镜下，发现HA材料周围的成骨细胞汇合且存活，而在β-TCP附近仅观察到细胞碎片。在HA-TCP的情况下，材料周围仅有少数存活细胞夹杂在碎片中。扫描电子显微镜显示，HA表面有许多细胞，在早期显示出不同的细胞行为，如锚定、附着、黏附和铺展，因为表面仅因再结晶而受到轻微干扰。但随着时间的推移，由于沉淀和再结晶，HA的整个表面发生了变化，不再支持细胞行为，而材料周围的细胞显示正常生长。相反，即使在培养早期，在β-TCP和HA-TCP完全改变的表面上也几乎观察不到细胞，材料周围细胞的形态也开始发生变化。这些结果表明HA促进了成骨细胞的活性。HA表面在一段时间内保持不变，而β-TCP表面经历了表面离子的溶解，并导致表面磷灰石的再结晶。β-TCP周围环境中高磷酸盐和低钙水平的最终变化可能是细胞死亡的原因。
