In vitro and in vivo degradation assessment and preventive measures of biodegradable Mg alloys for biomedical applications.

Magnesium (Mg) and its alloys have been widely explored as a potential biodegradable implant material. However, the fast degradation of Mg-based alloys under physiological environment has hindered their widespread use for implant applications till date. The present review focuses on in vitro and in vivo degradation of biodegradable Mg alloys, and preventive measures for biomedical applications. Initially, the corrosion assessment approaches to predict the degradation behavior of Mg alloys are discussed along with the measures to control rapid corrosion. Furthermore, this review attempts to explore the correlation between in vitro and in vivo corrosion behavior of different Mg alloys. It was found that the corrosion depends on experimental conditions, materials and the results of different assessment procedures hardly matches with each other. It has been demonstrated the corrosion rate of magnesium can be tailored by alloying elements, surface treatments and heat treatments. Various researches also studied different biocompatible coatings such as dicalcium phosphate dihydrate (DCPD), ß-tricalcium phosphate (ß-TCP), hydroxyapatite (HA), polycaprolactone (PCL), polylactic acid (PLA), and so on, on Mg alloys to suppress rapid degradation and examine their influence on new bone regeneration as well. This review shows the need for a standard method of corrosion assessment to predict the in vivo corrosion rate based on in vitro data, and thus reducing the in vivo experimentation.

Investigation of Patient-Specific Maxillofacial Implant Prototype Development by Metal Fused Filament Fabrication (MF3) of Ti-6Al-4V.

Additive manufacturing (AM) and related digital technologies have enabled several advanced solutions in medicine and dentistry, in particular, the design and fabrication of patient-specific implants. In this study, the feasibility of metal fused filament fabrication (MF3) to manufacture patient-specific maxillofacial implants is investigated. Here, the design and fabrication of a maxillofacial implant prototype in Ti-6Al-4V using MF3 is reported for the first time. The cone-beam computed tomography (CBCT) image data of the patient's oral anatomy was digitally processed to design a 3D CAD model of the hard tissue and fabricate a physical model by stereolithography (SLA). Using the digital and physical models, bone loss condition was analyzed, and a maxillofacial implant initial design was identified. Three-dimensional (3D) CAD models of the implant prototypes were designed that match the patient's anatomy and dental implant requirement. In this preliminary stage, the CAD models of the prototypes were designed in a simplified form. MF3 printing of the prototypes was simulated to investigate potential deformation and residual stresses. The patient-specific implant prototypes were fabricated by MF3 printing followed by debinding and sintering using a support structure for the first time. MF3 printed green part dimensions fairly matched with simulation prediction. Sintered parts were characterized for surface integrity after cutting the support structures off. An overall 18 ± 2% shrinkage was observed in the sintered parts relative to the green parts. A relative density of 81 ± 4% indicated 19% total porosity including 11% open interconnected porosity in the sintered parts, which would favor bone healing and high osteointegration in the metallic implants. The surface roughness of Ra: 18 ± 5 µm and a Rockwell hardness of 6.5 ± 0.8 HRC were observed. The outcome of the work can be leveraged to further investigate the potential of MF3 to manufacture patient-specific custom implants out of Ti-6Al-4V.

Influence of Ultrasound and Magnetic Field Treatment Time on Carcinoma Cell Inhibition with Drug Carriers: An in Vitro Study.

The influence of exposing carcinoma cells to a static magnetic field (SMF) and low-intensity pulsed ultrasound (LIPUS), for different durations (15-45 min/d), in the presence of magnetic and non-magnetic drug carriers, on their in vitro inhibition is examined. Increasing the exposure time by 15 min/d decreased the culture duration by 24 h to achieve the same level of inhibition in colon (HCT116) and hepatocellular (HepG2) cells. Cell cycle analysis revealed enhanced cellular blockage in G1 and S phases with SMF + LIPUS exposure, and exposure for 45 min/d completely suppressed the S → G2 transition. Apoptosis of both types of cells increased with SMF + LIPUS treatment time, and HepG2 cells exhibited elevated necrosis with >30 min/d exposure. HepG2 cells also had higher amounts of reactive oxygen species (seven- to eightfold) than HCT116 cells (two- to sixfold), suggesting treatment effectiveness is cell and drug carrier dependent. The accelerated cellular activities are attributed to the enhanced internalization of drug carriers as a consequence of destabilized cellular membranes caused by the SMF + LIPUS-generated mechanical and electrical stimuli.

Degradation, wettability and surface characteristics of laser surface modified Mg-Zn-Gd-Nd alloy.

This work evaluates the effects of laser surface modification on Mg-Zn-Gd-Nd alloy which is a potential biodegradable material for temporary bone implant applications. The laser surface melted (LSM) samples were investigated for microstructure, wettability, surface hardness and in vitro degradation. The microstructural study was carried out using scanning and transmission electron microscopes (SEM, TEM) and the phases present were analyzed using X-ray diffraction. The in vitro degradation behaviour was assessed in hank's balanced salt solution (HBSS) by immersion corrosion technique and the effect of LSM process parameters on the wettability was analyzed through contact angle measurements. The microstructural examination showed remarkable grain refinement as well as uniform redistribution of intermetallic phases throughout the matrix after LSM. These microstructural changes increased the hardness of LSM samples with an increase in energy density. The wetting behaviour of processed samples showed hydrophilic nature when processed at lower (12.5 and 17.5 J/mm2) and intermediate energy density (22.5 and 25 J/mm2), which can potentially improve cell-materials interaction. The corrosion rate of as cast Mg-Zn-Gd-Nd alloy decreased by ~83% due to LSM.

Mechanical behaviour of additively manufactured bioactive glass/high density polyethylene composites.

Bioactive glass (BAG) is a well-known biomaterial that can form a strong bond with hard and soft tissues and can also aid in bone regeneration. In this study, BAG is added to a polymer to induce bioactivity and to realize fused filament fabrication (FFF) based printing of polymer composites for potential orthopaedic implant applications. BAG (5, 10, and 20 wt%) is melt compounded with high density polyethylene (HDPE) and subsequently extruded into feedstock filament for FFF-printing. Tensile tests on developed filaments reveal that they are stiff enough to resist forces exerted during the printing process. Micrography of printed HDPE/BAG reveals perfect diffusion of raster interface indicating proper selection of printing parameters. Micrography of freeze fractured prints shows the homogeneous distribution and good dispersion of filler across the matrix. The tensile, flexural, and compressive modulus of FFF-printed HDPE/BAG parts increases with filler addition. BAG addition to the HDPE matrix enhances flexural and compressive strength. The tensile and flexural behaviour of FFF-prints is comparable to injection molded counterparts. Property maps exhibit the merits of present study over the existing literature pertaining to desired bone properties and polymer composites used in biomedical applications. It is envisioned that the development of HDPE/BAG composites for FFF-printing can lead to possible orthopaedic implants and scaffolds to mimic the bone properties in customised anatomical sites or injuries.

Probing the Influence of γ-Sterilization on the Oxidation, Crystallization, Sliding Wear Resistance, and Cytocompatibility of Chemically Modified Graphene-Oxide-Reinforced HDPE/UHMWPE Nanocomposites and Wear Debris.

Osteolysis and aseptic loosening due to wear at the articulating interfaces of prosthetic joints are considered to be the key concerns for implant failure in load-bearing orthopedic applications. In an effort to reduce the wear and processing difficulties of ultrahigh-molecular-weight polyethylene (UHMWPE), our research group recently developed high-density polyethylene (HDPE)/UHMWPE nanocomposites with chemically modified graphene oxide (mGO). Considering the importance of sterilization, this work explores the influence of γ-ray dosage of 25 kGy on the clinically relevant performance-limiting properties of these newly developed hybrid nanocomposites in vitro. Importantly, this work also probes into the cytotoxic effects of the wear debris of different compositions and sizes on MC3T3 murine osteoblasts and human mesenchymal stem cells (hMSCs). In particular, γ-ray-sterilized 1 wt % mGO-reinforced HDPE/UHMWPE nanocomposites exhibit an improvement in the oxidation index (16%), free energy of immersion (-12.1 mN/m), surface polarity (5.0%), and hardness (42%). Consequently, such enhancements result in better tribological properties, especially coefficient of friction (+13%) and wear resistance, when compared with UHMWPE. A spectrum of analyses using transmission electron microscopy (TEM) and in vitro cytocompatibility assessment demonstrate that phagocytosable (0.5-4.5 µm) sterilized 1 mGO wear particles, when present in culture media at 5 mg/mL concentration, induce neither significant reduction in MC3T3 murine osteoblast and hMSC growth nor cell morphology phenotype, during 24, 48, and 72 h of incubation. Taken together, this study suggests that γ-ray-sterilized HDPE/UHMWPE/mGO nanocomposites can be utilized as promising articulating surfaces for total joint replacements.

Mechanical and in vitro degradation behavior of magnesium-bioactive glass composites prepared by SPS for biomedical applications.

In order to make magnesium (Mg) a successful candidate for fracture fixation devices, it is imperative to control the corrosion rate and enhance its elastic modulus. In the present work, we have prepared bioactive glass (BG) reinforced magnesium composite using spark plasma sintering (SPS). Simultaneous application of heat and pressure during SPS decreased the softening point of BG (600°C), allowing it to coat the Mg particles partially. As a result, BG was found along the Mg particle boundaries, which was confirmed by elemental mapping. Addition of BG improved microhardness and elastic modulus of Mg-BG composites. Corrosion behavior was studied by hydrogen evolution and immersion corrosion in phosphate buffered saline (PBS). After 64 h of immersion, Mg-10 wt % BG composite showed highest corrosion resistance. Quantitative micro-computed tomography (micro-CT) results indicated porosity increase in Mg-BG composites during immersion. The maximum increase in porosity (1.66%) was noticed for pure Mg while the minimum for Mg-10 wt % BG composite. MG63 cell-material interactions, using extract method, showed good cytocompatibility for Mg-10 wt % BG composite. The concentration of Mg ion in cell culture media was measured using atomic absorption spectroscopy after 24 h immersion of Mg/BG composites. The results indicated that using BG as reinforcement and SPS as sintering method; we can prepare corrosion resistant and high modulus Mg-BG composites that can be used for fabricating bone fracture fixation plates. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 107B: 352-365, 2019.

Laser surface modification of 316L stainless steel.

Medical grade 316L stainless steel was laser surface melted (LSM) using continuous wave Nd-YAG laser in argon atmosphere at 1 and 5 mm/s. The treated surfaces were characterized using electron backscatter diffraction to study the influence of top surface crystallographic orientation and type of grain boundaries on corrosion resistance, wettability, and biocompatibility. The laser scan velocity was found to have a marginal influence on the surface roughness and the type of grain boundaries. However, the crystal orientation density was found to be relatively high in 1 mm/s samples. The LSM samples showed a higher concentration of {101} and {123} planes parallel to the sample surface as well as a higher fraction of low-angle grain boundaries. The LSM samples were found to exhibit better surface wettability and enhanced the viability and proliferation of human fetal osteoblast cells in vitro when compared to the untreated samples. Further, the corrosion protection efficiency of 316L stainless steel was improved up to 70% by LSM in as-processed condition. The increased concentration of {101} and {123} planes on surfaces of LSM samples increases their surface energy, which is believed to be responsible for the improved in vitro cell proliferation. Further, the increased lattice spacing of these planes and high concentration of low-energy grain boundaries in LSM samples would have contributed to the better in vitro corrosion resistance than untreated 316L stainless steel. Our results indicate that LSM can be a potential treatment option for 316L stainless steel-based biomedical devices to improve biocompatibility and corrosion resistance. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 569-577, 2018.

Understanding compressive deformation behavior of porous Ti using finite element analysis.

In the present study, porous commercially pure (CP) Ti samples with different volume fraction of porosities were fabricated using a commercial additive manufacturing technique namely laser engineered net shaping (LENS™). Mechanical behavior of solid and porous samples was evaluated at room temperature under quasi-static compressive loading. Fracture surfaces of the failed samples were analyzed to determine the failure modes. Finite Element (FE) analysis using representative volume element (RVE) model and micro-computed tomography (CT) based model have been performed to understand the deformation behavior of laser deposited solid and porous CP-Ti samples. In vitro cell culture on laser processed porous CP-Ti surfaces showed normal cell proliferation with time, and confirmed non-toxic nature of these samples.

Microstructure and corrosion behavior of laser processed NiTi alloy.

Laser Engineered Net Shaping (LENS™), a commercially available additive manufacturing technology, has been used to fabricate dense equiatomic NiTi alloy components. The primary aim of this work is to study the effect of laser power and scan speed on microstructure, phase constituents, hardness and corrosion behavior of laser processed NiTi alloy. The results showed retention of large amount of high-temperature austenite phase at room temperature due to high cooling rates associated with laser processing. The high amount of austenite in these samples increased the hardness. The grain size and corrosion resistance were found to increase with laser power. The surface energy of NiTi alloy, calculated using contact angles, decreased from 61 mN/m to 56 mN/m with increase in laser energy density from 20 J/mm(2) to 80 J/mm(2). The decrease in surface energy shifted the corrosion potentials to nobler direction and decreased the corrosion current. Under present experimental conditions the laser power found to have strong influence on microstructure, phase constituents and corrosion resistance of NiTi alloy.

Converted marine coral hydroxyapatite implants with growth factors: in vivo bone regeneration.

Herein we report rabbit model in vivo bone regeneration of hydrothermally converted coralline hydroxyapatite (HCCHAp) scaffolds without (group I) and with growth factors namely insulin like growth factor-1 (IGF-1) (group II) and bone morphogenetic protein-2 (BMP-2) (group III). All HCCHAp scaffolds have been characterized for phase purity and morphology before implantation. Calcined marine coral was hydrothermally converted using a mineralizer/catalyst to phase pure HAp retaining original pore structure and geometry. After sintering at 1250°C, the HCCHAp found to have ~87% crystallinity, 70-75% porosity and 2±0.5MPa compressive strength. In vitro growth factor release study at day 28 revealed 77 and 98% release for IGF-1 and BMP-2, respectively. The IGF-1 release was more sustained than BMP-2. In vivo bone healing of different groups was compared using chronological radiology, histological evaluations, scanning electron microscopy and fluorochrome labeling up to 90days of implantation. In vivo studies showed substantial reduction in radiolucent zone and decreased radiodensity of implants in group II followed by group III and group I. These observations clearly suggest in-growth of osseous tissue, initiation of bone healing and complete union between implants and natural bone in group II implants. A statistical score sheet based on histological observations showed an excellent osseous tissue formation in group II and group III scaffolds and moderate bone regeneration in group I scaffolds.

Effect of hydroxyapatite particle size, morphology and crystallinity on proliferation of colon cancer HCT116 cells.

The aim of the present work is to chemically and physically characterize the synthesized Hydroxyapatite (HAp) micro and nanoparticles and to explore the inhibitory effect of nano-HAps on the in vitro growth of human colon cancerous cells HCT116. HAp powder was synthesized using three different routes to achieve micro and nanosized powders, with different morphologies and crystallinity. The synthesized powders were characterized using X-ray diffraction, FTIR spectroscopy and scanning electron microscope. The results showed that the average crystallite size of HAp powder varies from 11nm to 177nm and respective crystallinity of powder found to be in the range of 0.12 and 0.92. The effect of these physico-chemical properties of HAp powders on human colon cancer HCT116 cells inhibition was determined in vitro. It was found that decreasing the HAp powder crystallite size between 11nm and 22nm significantly increases the HCT116 cell inhibition. Our results demonstrate that apart from HAp powder size their crystallinity and morphology also play an important role in cellular inhibition of human colon cancer cells.

Microstructure, mechanical and wear properties of laser surface melted Ti6Al4V alloy.

Laser surface melting (LSM) of Ti6Al4V alloy was carried out with an aim to improve properties such as microstructure and wear for implant applications. The alloy substrate was melted at 250W and 400W at a scan velocity of 5mm/s, with input energy of 42J/mm(2) and 68J/mm(2), respectively. The results showed that equiaxed α+ß microstructure of the substrate changes to mixture of acicular α in ß matrix after LSM due to high cooling rates in the range of 2.25×10(-3)K/s and 1.41×10(-3)K/s during LSM. Increasing the energy input increased the thickness of remelted region from 779 to 802µm and 1173 to 1199µm. Similarly, as a result of slow cooling rates under present experimental conditions, the grain size of the alloy increased from 4.8µm to 154-199µm. However, the hardness of the Ti6Al4V alloy increased due to LSM melting and resulted in lowest in vitro wear rate of 3.38×10(-4)mm(3)/Nm compared to untreated substrate with a wear rate of 6.82×10(-4)mm(3)/Nm.

In situ synthesized TiB-TiN reinforced Ti6Al4V alloy composite coatings: microstructure, tribological and in-vitro biocompatibility.

Wear resistant TiB-TiN reinforced Ti6Al4V alloy composite coatings were deposited on Ti substrate using laser based additive manufacturing technology. Ti6Al4V alloy powder premixed with 5wt% and 15wt% of boron nitride (BN) powder was used to synthesize TiB-TiN reinforcements in situ during laser deposition. Influences of laser power, scanning speed and concentration of BN on the microstructure, mechanical, in vitro tribological and biological properties of the coatings were investigated. Microstructural analysis of the composite coatings showed that the high temperature generated due to laser interaction with Ti6Al4V alloy and BN results in situ formation of TiB and TiN phases. With increasing BN concentration, from 5wt% to 15wt%, the Young's modulus of the composite coatings, measured by nanoindentation, increased from 170±5GPa to 204±14GPa. In vitro tribological tests showed significant increase in the wear resistance with increasing BN concentration. Under identical test conditions TiB-TiN composite coatings with 15wt% BN exhibited an order of magnitude less wear rate than CoCrMo alloy-a common material for articulating surfaces of orthopedic implants. Average top surface hardness of the composite coatings increased from 543±21HV to 877±75HV with increase in the BN concentration. In vitro biocompatibility and flow cytometry study showed that these composite coatings were non-toxic, exhibit similar cell-materials interactions and biocompatibility as that of commercially pure titanium (CP-Ti) samples. In summary, excellent in vitro wear resistance, high stiffness and suitable biocompatibility make these composite coatings as a potential material for load-bearing articulating surfaces towards orthopaedic implants.

Laser surface modification of 316 L stainless steel with bioactive hydroxyapatite.

Laser-engineered net shaping (LENS™), a commercial additive manufacturing process, was used to modify the surfaces of 316 L stainless steel with bioactive hydroxyapatite (HAP). The modified surfaces were characterized in terms of their microstructure, hardness and apatite forming ability. The results showed that with increase in laser energy input from 32 J/mm(2) to 59 J/mm(2) the thickness of the modified surface increased from 222±12 µm to 355±6 µm, while the average surface hardness decreased marginally from 403±18 HV0.3 to 372±8 HV0.3. Microstructural studies showed that the modified surface consisted of austenite dendrites with HAP and some reaction products primarily occurring in the inter-dendritic regions. Finally, the surface-modified 316 L samples immersed in simulated body fluids showed significantly higher apatite precipitation compared to unmodified 316 L samples.

Microwave-sintered 3D printed tricalcium phosphate scaffolds for bone tissue engineering.

This study reports the manufacturing process of 3D interconnected macroporous tricalcium phosphate (TCP) scaffolds with controlled internal architecture by direct 3D printing (3DP), and high mechanical strength obtained by microwave sintering. TCP scaffolds with 27%, 35% and 41% designed macroporosity with pore sizes of 500 µm, 750 µm and 1000 µm, respectively, were manufactured by direct 3DP. These scaffolds were then sintered at 1150 °C and 1250 °C in conventional electric muffle and microwave furnaces, respectively. Total open porosity between 42% and 63% was obtained in the sintered scaffolds due to the presence of intrinsic micropores along with designed pores. A significant increase in compressive strength between 46% and 69% was achieved by microwave compared to conventional sintering as a result of efficient densification. Maximum compressive strengths of 10.95 ± 1.28 MPa and 6.62 ± 0.67 MPa were achieved for scaffolds with 500 µm designed pores (~ 400 µm after sintering) sintered in microwave and conventional furnaces, respectively. An increase in cell density with a decrease in macropore size was observed during in vitro cell-material interactions using human osteoblast cells. Histomorphological analysis revealed that the presence of both micro- and macropores facilitated osteoid-like new bone formation when tested in femoral defects of Sprague-Dawley rats. Our results show that bioresorbable 3D-printed TCP scaffolds have great potential in tissue engineering applications for bone tissue repair and regeneration.

Laser processed TiN reinforced Ti6Al4V composite coatings.

The purpose of this first generation investigation is to evaluate fabrication, in vitro cytotoxicity, cell-material interactions and tribological performance of TiN particle reinforced Ti6Al4V composite coatings for potential wear resistant load bearing implant applications. The microstructural analysis of the composites was performed using scanning electron microscope and phase analysis was done with X-ray diffraction. In vitro cell-material interactions, using human fetal osteoblast cell line, have been assessed on these composite coatings and compared with Ti6Al4V alloy control samples. The tribological performance of the coatings were evaluated, in simulated body fluids, up to 1000 m sliding distance under 10 N normal load. The results show that the composite coatings contain distinct TiN particles embedded in α+ß phase matrix. The average top surface hardness of Ti6Al4V alloy increased from 394±8 HV to 1138±61 HV with 40 wt% TiN reinforcement. Among the composite coatings, the coatings reinforced with 40 wt% TiN exhibited the highest wear resistance of 3.74×10(-6) mm(3)/Nm, which is lower than the wear rate, 1.04×10(-5) mm(3)/Nm, of laser processed CoCrMo alloy tested under identical experimental conditions. In vitro biocompatibility study showed that these composite coatings were non-toxic and provides superior cell-material interactions compared to Ti6Al4V control, as a result of their high surface energy. In summary, excellent in vitro wear resistance and biocompatibility of present laser processed TiN reinforced Ti6Al4V alloy composite coatings clearly show their potential as wear resistant contact surfaces for load bearing implant applications.

MgO-doped tantalum coating on Ti: microstructural study and biocompatibility evaluation.

Pure and MgO incorporated Ta coatings were prepared on Cp-Ti substrate using laser engineered net shaping (LENS), which resulted in diffuse coating-substrate interface. MgO was found along the Ta grain boundaries in the Ta matrix that increased the coating hardness from 185 ± 2.7 HV to 794 ± 93 HV. In vitro biocompatibility study showed excellent early cellular attachment and later stage proliferation in MgO incorporated coatings. The results indicated that although Ta coatings had higher biocompatibility than Ti, it could further be improved by incorporating MgO in the coating, while simultaneously improving the mechanical properties.

Wear Performance of Laser Processed Tantalum Coatings.

This first generation investigation evaluates the in vitro tribological performance of laser-processed Ta coatings on Ti for load-bearing implant applications. Linear reciprocating wear tests in simulated body fluid showed one order of magnitude less wear rate, of the order of 10(-4)mm(3)(N.m)(-1), for Ta coatings compared to Ti. Our results demonstrate that Ta coatings can potentially minimize the early-stage bone-implant interface micro-motion induced wear debris generation due to their excellent bioactivity comparable to that of hydroxyapatite (HA), high wear resistance and toughness compared to popular HA coatings.

Quasi-static Torsional Deformation Behavior of Porous Ti6Al4V alloy.

Laser processed Ti6Al4V alloy samples with total porosities of 0%, 10% and 20% have been subjected to torsional loading to determine mechanical properties and to understand the deformation behavior. The torsional yield strength and modulus of porous Ti alloy samples was found to be in the range of 185-332 MPa and 5.7-11 GPa, respectively. With an increase in the porosity both the strength and the modulus decreased, and at 20% porosity the torsional modulus of Ti6Al4V alloy was found to be very close to that of human cortical bone. Further, the experiments revealed clear strain hardening and ductile deformation in all the samples, which suggests that the inherent brittleness associated solid-state sintered porous materials can be completely eliminated via laser processing for load bearing metal implant applications.