On magnesium-modified titanium coatings and magnesium alloys for oral orthopaedic applications: in vitro investigation

Abstract: In dentistry and orthopaedic surgery, research to find and develop improved biomaterials is progressing rapidly. Of specific interest is to accelerate bone formation around the implant surface, which could improve the reliability of the implant even in patients with compromised situations. Although the surface modification of the implant has been proven to certain extent to promote osseointegration, the lack of bone in the patient remains a major issue and bone augmentation is commonly conducted prior to implant insertion. Synthetic and naturally derived resorbable materials are commonly used. However, problems such as the lack of optimal mechanical properties or the undesirable material resorption kinetics still exist and there still remain possibility for improvement. Clinical approaches for orthopaedic trauma require the use of non-resorbable screws, plates and pins made of metallic materials such as titanium, cobalt-chrome and stainless steel alloys. The major drawback of these materials is the need of implant removal at re-entry. Therefore, the research of bioresorbable materials that could withstand the mechanical stresses is an ongoing topic.Based on this clinical reality, the aim of this thesis was to investigate the suitability of magnesium (Mg) as a biomaterial for regenerative bone applications. Namely, Mg as a doping material for engineered mesoporous titanium implant surfaces (Studies I, II and III), and as a bioresorbable metal alloy for bone regeneration in bone trauma and bone defects conditions (Study IV).Study I, II, III Mesoporous titania films produced with evaporation-induced selfassembly (EISA) technique and applied as implant surface coatings are under investigation as a release system for the controlled administration of several substances, such as osteoporotic drugs, to enhance early bone anchorage to the implant. Modulating the pore size of such films though the selection of EISA parameters permits to control the adsorption of such substances into the mesoporous matrix and their subsequent release into the peri-implant region. Studies I, II and III analysed the effect of Mg incorporation into mesoporous titania coatings towards two cellular models during early and later stages of cell activity.Study I characterized the morphology, chemistry, and topography of mesoporous titania coatings and the effects of Mg-loading on surface micro- and nano-structures. Mg release was determined and its effect was evaluated on human foetal osteoblast populations. It was shown that mesoporous films possessed a smooth surface with pores that faced outward. Mg adsorption did not substantially alter the mesoporous surface roughness both at micro- and nano- levels. Mg was released within 24 hours of incubation in cell culture conditions, thus its bioactive effect only occurred during initial osteoblasts activity. Study II evaluated the ability of Mg-loaded mesoporous coatings to modulate multipotent adipose-derived stromal cell differentiation toward the osteoblast phenotype. The results demonstrated that Mg release had a strong impact on this cellular model, promoting osteoblast marker expression in standard cell culture conditions. Interestingly, Mg significantly promoted the expression of osteopontin, a protein that is essential for early biomaterial-cell osteogenic interaction.In study III, the reagents and EISA parameters in the mesoporous deposition were varied to generate three mesoporous titania coatings with 2-, 6- and 7-nm average pore size, to increase Mg content in the interconnected porosity of the films. The effect of various Mg contents released from the three mesoporous structureswas tested on human foetal osteoblasts populations with pre-designed osteogenic PCR arrays and real-time polymerase chain reaction. It was shown that Mg release affected osteogenesis and was controlled by tuning the pore dimensions of the mesoporous films. Increasing pore size by 1 nm (from 6 nm to 7 nm) significantly enhanced the bioactivity of the film without altering the surface roughness.Study IV In orthopaedics Mg alloys has received increasing attention as bioresorbable metals for bone regeneration. However, localized material degradation is too fast and provokes the premature loss of mechanical properties, preventing correct cellular development and bone healing in vivo . For this reason, various alloying elements are combined with high-purity Mg to modulate and optimize degradation behaviour. Study IV of this thesis investigated the degradation parameters of Mg2Ag, Mg10Gd, and Mg4Y3RE alloys and how the alloys differently affect human umbilical cord perivascular cell adhesion and spreading. Mg4Y3RE showed the highest degradation rate and, thereby, the highest trend in increases in pH and osmolality of the surrounding fluid. However, both Mg4Y3RE and Mg10Gd allowed cells to better adhere and spread across their degraded surfaces; in comparison, surface degradation of Mg2Ag was more aggressive with weak or no visible cellular structures on it. Conclusions In summary, the results of the present thesis explored the potential of Mg for its application in bone tissue regeneration. Titanium implant surfaces coated with mesoporous TiO2 thin films and further loaded with Mg enhanced bone cell activity and osteoprogenitor development into mature osteoblasts. Thus, mesoporous deposition followed by Mg loading may be a suitable alternative to existing implant surface treatments. Bioresorbable materials must degrade slowly and uniformly in order to simulate the tissue healing process. Mg10Gd possesses reduced content of alloying element and a suitable homogenous degradation pattern in vitro that allows proper adhesion of undifferentiated cells. Mg10Gd thus represents a biodegradable Mg-based material with promising mechanical and biological properties for use in dental and orthopaedic fields.