Influences of calcium phosphate microstructure on pulpal and bone cells response
Date Issued
2015
Date
2015
Author(s)
Yang, Jyun-Wei
Abstract
Calcium phosphate is the main inorganic component of mammals’ hard tissues and hydroxyapatite (Meleti et al.) is its primary structure. There are several substructures of hydroxyapatite in different kinds of hard tissues and amorphous calcium phosphate (ACP) is regarded as the primer of hydroxyapatite. Both biological HAP and ACP are in nano-scale. According to previous researches, artificial hydroxyapatite was biocompatible and amorphous calcium phosphate presented good solubility, biodegradability and bioactivity. Hence, ACP and HAP are widely used as biomaterials. In addition, the design of scaffold to mimic the collagen matrix for precipitation of mineral contents in biological hard tissue also play important role in tissue engineering. Since chitosan (CS) has good biocompatibility and similar structure to protein matrix, calcium phosphate combined with chitosan is considered as a potential biomaterial in bone regeneration. Therefore, our research team has successfully developed a new biomaterials composed of chitosan membrane and nano-sized ACP (CS-ACP) and HAP (CS-HAP) via direct precipitation method. However, the disperse pattern of the calcium phosphate products on the CS membrane, the biocompatibility of these materials and their interaction with cells are unclear. The purpose of this study is to investigate the distribution of nano-scaled ACP and HAP in CS membrane, and to evaluate the biocompatibility of CS-ACP and CS-HAP membranes and their influences on the behavior of bone and pulpal cells. The calcium phosphates with different morphology were produced in 2% chitosan (CS) membrane via direct precipitation method. The synthetic process of P-Ca was the CS membrane soaking in solution with phosphate ions first and then in solution with calcium ions. The synthetic process of Ca-P was the reaction via different order of calcium and phosphate solutions. Using XRD, FTIR and SEM as tools, we found ACP (P-Ca) and HAP (P-Ca) particles with the size of 100~200 nm were produced and distributed evenly on CS membrane surface with the depth of 5~6m in close and open system via P-Ca synthetic process, respectively. By Ca-P synthetic process, only HAP (Ca-P) particles were produced but with deeper in depth (7~9m) on CS membrane surface, which may be due to the smaller calcium ions have better ability to penetrate through CS membrane. All of Cs, CS-ACP and CS-HAP membranes presented no cytotoxity and good biocompatibility. In addition, U2OS, MRPC-1 and DPC all presented bested cell adhesion and proliferation behaviors in CS-HAP group and worst in CS group. Compared to the cells growing on culture plate, U2OS, with slow growth in culture plate, presented a higher cell growth rate in CS-HAP group. However, MRPC-1 and DPC, with faster growth in culture plate, presented different behavior with a lower cell growth rate in CS-HAP group. These findings implies that HAP regulates cells functions differently according to their ability in growth. By RT-PCR, we found CS-HAP and CS-ACP could enhance ALP and OCN expression of U2OS cells, which indicates both HAP and ACP may enhance differentiation and mineralization of bone cells. In general, both CS-ACP and CS-HAP presented good biocompability and could enhance the cell adhesion, proliferation and differentiation of pulpal and bone cells. Furthermore, HAP presented better effects on cell functions of pulpal and bone cells than ACP in vitro.
Subjects
Hydroxyapatite
amorphous calcium phosphate
chitosan
cell adhesion
cell division
cell differentiation
U2OS
MRPC-1
DPC
Type
thesis