Bioactive Hydroxyapatite Microspheres Obtained from Bone Cement for Biomedical Application

Abstract

With increasing demand of bone substitute, bioactive ceramics have been widely used in dentistry and orthopedics. In particular, hydroxyapatite (HA) has been increasingly getting attention as a promising implant material for bone defect due to its great biocompatibility. Despite many superior properties of HA, dense HA implant is limited owing to low biodegradability and lack of enough space for new bone to ingrow. Microsphere-type materials allow injectability which is minimally invasive procedure and bone ingrowth through the interspacing built among spheres. To accelerate bone healing rate, drugs or bioactive proteins are usually incorporated to ceramics, but thermal treatment of ceramic processing limited it. To overcome the problems, bone cement is introduced to make HA microspheres since room temperature processing is possible for bone cement. Researchers have already made microspheres from bone cement, but the products have low efficiency in fabrication time and drug loading. Previous studies using α-tricalcium phosphate (α-TCP) needed one week of incubation in simulated body fluid (SBF) solution to achieve mechanical and chemical stability. This is problematic for two reasons. First, it takes too long for researchers. Second, if drugs were loaded in the microspheres, a large amount of drugs can be released away during one week of incubation. In the first study, to get mechanical stability without SBF incubation, tetracalcium phosphate(TTCP) and citric acid were added to previous system as bone cement powder and hardening additive respectively. To get chemical stability within only a few hours, 10x SBF-like solution was used to coat the surface of microspheres with apatite. So, the final product is surface modified calcium phosphate microspheres with mechanical and chemical stability. TTCP played an important role in making difference of hardness. TTCP dissolved quick and filled the space among undissolved α-TCP particles since it is 10x more soluble than α-TCP. Furthermore, in vitro cell test using pre-osteoblast cells, the apatite coating layer formed in 10x SBF like solution showed enhanced biocompatibility compared to bare surface. It was meaningful to achieve better product within significantly recued time. Even though in the first study fabrication time was noticeably saved, it could be not adequate in drug delivery applications because there are a few hours of immersion, which causes drug loss to the medium. So, it is hard to approximate the amount of drug loaded and inappropriate for biomedical industry which requires accurate dose of drug. To avoid incubation in hydrophilic medium and achieve chemical stability at the same time, microspheres were kept in the oil, allowing them to be converted to HA with moisture originated from hardening liquid. After 3days, large amount of reactants were consumed and phase was transformed mainly to HA. For the advantage of visualization, green fluorescent protein (GFP) was used instead of drugs or growth factors. GFP was well distributed in the microspheres and its release behavior seemed to be sustained at least one month. To verify the potential use in the biomedical field, preliminary in vivo animal test was performed with a rat. BMP-2 was loaded in the microspheres and it showed better osteoconduction and osteoinduction compared to microspheres without BMP-2. In conclusion, these two experiments were focused on fabricating stable calcium phosphates microspheres with high efficiency in time and in drug loading. In vitro cell test showed rapidly-formed apatite coating layer had effectively enhanced biocompatibility. Also, preliminary in vivo animal test verified that drug loaded calcium phosphate microspheres converted in oil have great potential to be used as a bone filler.