Bone tissue engineering via nanostructured calcium phosphate biomaterials and stem cells

Tissue engineering is promising to meet the increasing need for bone regeneration. Nanostructured calcium phosphate （CAP） biomaterials/scaffolds are of special interest as they share chemical/crystallographic similarities to inorganic components of bone. Three applications of nano-CaP are discussed in this review： nanostructured calcium phosphate cement （CPC）; nano-CaP composites; and nano-CaP coatings. The interactions between stem cells and nano-CaP are highlighted, including cell attachment, orientation/ morphology, differentiation and in vivo bone regeneration. Several trends can be seen： （i） nano-CaP biomaterials support stem cell attachment/proliferation and induce osteogenic differentiation, in some cases even without osteogenic supplements; （ii） the influence of nano-CaP surface patterns on cell alignment is not prominent due to non-uniform distribution of nano-crystals; （iii） nano-CaP can achieve better bone regeneration than conventional CaP biomaterials; （iv） combining stem cells with nano-CaP accelerates bone regeneration, the effect of which can be further enhanced by growth factors; and （v） cell microencapsulation in nano-CaP scaffolds is promising for bone tissue engineering. These understandings would help researchers to further uncover the underlying mechanisms and interactions in nano-CaP stem cell constructs in vitro and in vivo, tailor nano-CaP composite construct design and stem cell type selection to enhance cell function and bone regeneration, and translate laboratory findings to clinical treatments.

Effects of quaternary ammonium chain length on the antibacterial and remineralizing effects of a calcium phosphate nanocomposite

Composites containing nanoparticles of amorphous calcium phosphate （NACP） remineralize tooth lesions and inhibit caries. A recent study synthesized quaternary ammonium methacrylates （QAMs） with chain lengths （CLs） of 3-18 and determined their effects on a bonding agent. This study aimed to incorporate these QAMs into NACP nanocomposites for the first time to simultaneously endow the material with antibacterial and remineralizing capabilities and to investigate the effects of the CL on the mechanical and biofilm properties. Five QAMs were synthesized： DMAPM （CL3）, DMAHM （CL6）, DMADDM （CL12）, DMAHDM （CL16）, and DMAODM （CL18）. Each QAM was incorporated into a composite containing 20% NACP and 50% glass fillers. A dental plaque microcosm biofilm model was used to evaluate the antibacterial activity. The flexural strength and elastic modulus of nanocomposites with QAMs matched those of a commercial control composite （n = 6; P 〉 0.1）. Increasing the CL from 3 to 16 greatly enhanced the antibacterial activity of the NACP nanocomposite （P 〈 0.05）; further increasing the CL to 18 decreased the antibacterial potency. The NACP nanocomposite with a CL of 16 exhibited biofilm metabolic activity and acid production that were 10-fold lesser than those of the control composite. The NACP nanocomposite with a CL of 16 produced 2-log decreases in the colony-forming units （CFU） of total microorganisms, total streptococci, and mutans streptococci. In conclusion, QAMs with CLs of 3-18 were synthesized and incorporated into an NACP nanocomposite for the first time to simultaneously endow the material with antibacterial and remineralization capabilities. Increasing the C/reduced the metabolic activity and acid production of biofilms and caused a 2-log decrease in CFU without compromising the mechanical properties. Nanocomposites exhibiting strong anti-biofilm activity, remineralization effects, and mechanical properties are promising materials for tooth restorations that inhibit caries.

Primer containing dimethylaminododecyl methacrylate kills bacteria impregnated in human dentin blocks

Antibacterial dimethylaminododecyl methacrylate （DMADDM） was recently synthesized. The objectives of this study were to： （1） investigate antibacterial activity of DMADDM-containing primer on Streptococcus mutans impregnated into dentin blocks for the first time, and （2） compare the antibacterial efficacy of DMADDM with a previous quaternary ammonium dimethacrylate （QADM）. Scotchbond Multi-Purpose （SBMP） bonding agent was used. DMADDM and QADM were mixed into SBMP primer. Six primers were tested： SBMP control primer P, P＋2.5% DMADDM, P＋5% DMADDM, P＋7.5% DMADDM, P＋10% DMADDM, and P＋10% QADM. S. mutans were impregnated into human dentin blocks, and each primer was applied to dentin to test its ability to kill bacteria in dentinal tubules. Bacteria in dentin were collected via a sonication method, and the colony-forming units （CFU） and inhibition zones were measured. The bacterial inhibition zone of P＋10% DMADDM was 10 times that of control primer （P〈0.05）. CFU in dentin with P＋10% DMADDM was reduced by three orders of magnitude, compared with control. DMADDM had a much stronger antibacterial effect than QADM, and antibacterial efficacy increased with increasing DMADDM concentration. Dentin shear bond strengths were similar among all groups （P〉0.1）. In conclusion, antibacterial DMADDM-containing primer was validated to kill bacteria inside dentin blocks, possessing a much stronger antibacterial potency than the previous QADM. DMADDM-containing bonding agent was effective in eradicating bacteria in dentin, and its efficacy was directly proportional to DMADDM mass fraction. Therefore, DMADDM may be promisine for use in bonding agents as well as in other restorative and oreventive materials to inhibit bacteria.

Induced Pluripotent Stem Cell-derived Mesenchymal Stem Cell Seeding on Biofunctionalized Calcium Phosphate Cements

Induced pluripotent stem ceils （iPSCs） have great potential due to their proliferation and differentiation capability. The objectives of this study were to generate iPSC-derived mesenchymal stem cells （iPSC-MSCs）, and investigate iPSC-MSC proliferation and osteogenic differentiation on calcium phosphate cement （CPC） containing biofunctional agents for the first time. Human iPSCs were derived from marrow CD34＋ cells which were reprogrammed by a single episomal vector, iPSCs were cultured to form embryoid bodies （EBs）, and MSCs migrated out of EBs. Five biofunctional agents were incorporated into CPC： RGD （Arg-Gly-Asp） peptides, fibronectin （Fn）, fibronectin-like engineered polymer protein （FEPP）, extracellular matrix Geltrex, and platelet concentrate, iPSC-MSCs were seeded on five biofunctionalized CPCs： CPC-RGD, CPC-Fn, CPC- FEPP, CPC-Geltrex, and CPC-Platelets. iPSC-MSCs on biofunctional CPCs had enhanced proliferation, actin fiber expression, osteogenic differentiation and mineralization, compared to control. Cell proliferation was greatly increased on biofunctional CPCs. iPSC-MSCs underwent osteogenic differentiation with increased alkaline phosphatase, Runx2 and coUagen-I expressions. Mineral synthesis by iPSC-MSCs on CPC-Platelets was 3-fold that of CPC control. In conclusion, iPSCs showed high potential for bone engineering, iPSC- MSCs on biofunctionalized CPCs had cell proliferation and bone mineralization that were much better than traditional CPC. iPSC-MSC-CPC constructs are promising to promote bone regeneration in craniofacial/ orthopedic repairs.