Susmita Bose

Recent advances in bone tissue engineering scaffolds.

Bone disorders are of significant concern due to increase in the median age of our population. Traditionally, bone grafts have been used to restore damaged bone. Synthetic biomaterials are now being used as bone graft substitutes. These biomaterials were initially selected for structural restoration based on their biomechanical properties. Later scaffolds were engineered to be bioactive or bioresorbable to enhance tissue growth. Now scaffolds are designed to induce bone formation and vascularization. These scaffolds are often porous, made of biodegradable materials that harbor different growth factors, drugs, genes, or stem cells. In this review, we highlight recent advances in bone scaffolds and discuss aspects that still need to be improved.

Calcium phosphate ceramic systems in growth factor and drug delivery for bone tissue engineering: A review

Calcium phosphates (CaPs) are the most widely used bone substitutes in bone tissue engineering due to their compositional similarities to bone mineral and excellent biocompatibility. In recent years, CaPs, especially hydroxyapatite and tricalcium phosphate, have attracted significant interest in simultaneous use as bone substitute and drug delivery vehicle, adding a new dimension to their application. CaPs are more biocompatible than many other ceramic and inorganic nanoparticles. Their biocompatibility and variable stoichiometry, thus surface charge density, functionality, and dissolution properties, make them suitable for both drug and growth factor delivery. CaP matrices and scaffolds have been reported to act as delivery vehicles for growth factors and drugs in bone tissue engineering. Local drug delivery in musculoskeletal disorder treatments can address some of the critical issues more effectively and efficiently than the systemic delivery. CaPs are used as coatings on metallic implants, CaP cements, and custom designed scaffolds to treat musculoskeletal disorders. This review highlights some of the current drug and growth factor delivery approaches and critical issues using CaP particles, coatings, cements, and scaffolds towards orthopedic and dental applications.

Development of controlled porosity polymer-ceramic composite scaffolds via fused deposition modeling

This research is focused on development and fabrication of controlled porosity polymer-ceramic composite scaffolds, with 3-D interconnectivity designed to promote richer supply of blood, oxygen and nutrients for healthy in-growth of bone cells. Particulate-reinforced polymer-ceramic composites were developed by high shear mixing of polypropylene (PP) polymer and tricalcium phosphate (TCP) ceramic. Processing aids were used to improve plasticity and processibility to the composites. Controlled porosity scaffolds were fabricated via the fused deposition process, one of the commercially available rapid prototyping (RP) techniques. These porous scaffolds were characterized for their use as bone grafts in terms of physical, mechanical and biological properties. Hg-porosimetry was performed to determine pore size and their distribution. Scaffolds with different complex internal architectures were also fabricated using this composite material. Tensile properties of neat PP (as received), PP with processing aids (without TCP) and PP-TCP composite (with processing aids) were evaluated and compared using standard dog bone samples. Uniaxial compression tests were performed on cylindrical porous samples with an average pore size of 160 μm and varying vol.% porosity (36%, 48% and 52%). Samples with 36 vol.% porosity showed the best compressive strength of 12.7 MPa. Cytotoxicity and cell proliferation studies were conducted with a modified human osteoblast cell-line (HOB). Results showed that these samples were non-toxic with excellent cell growth during the first two weeks of in vitro testing.

Porous tantalum structures for bone implants: Fabrication, mechanical and in vitro biological properties

The relatively high cost of manufacturing and the inability to produce modular implants have limited the acceptance of tantalum, in spite of its excellent in vitro and in vivo biocompatibility. In this article, we report how to process Ta to create net-shape porous structures with varying porosity using Laser Engineered Net Shaping (LENS) for the first time. Porous Ta samples with relative densities between 45% and 73% have been successfully fabricated and characterized for their mechanical properties. In vitro cell materials interactions, using a human fetal osteoblast cell line, have been assessed on these porous Ta structures and compared with porous Ti control samples. The results show that the Youngs modulus of porous Ta can be tailored between 1.5 and 20 GPa by changing the pore volume fraction between 27% and 55%. In vitro biocompatibility in terms of MTT assay and immunochemistry study showed excellent cellular adherence, growth and differentiation with abundant extracellular matrix formation on porous Ta structures compared to porous Ti control. These results indicate that porous Ta structures can promote enhanced/early biological fixation. The enhanced in vitro cell-material interactions on the porous Ta surface are attributed to its chemistry, its high wettability and its greater surface energy relative to porous Ti. Our results show that these laser-processed porous Ta structures can find numerous applications, particularly among older patients, for metallic implants because of their excellent bioactivity.

TiO2 nanotubes on Ti: Influence of nanoscale morphology on bone cell–materials interaction

Ti being bioinert shows poor bone cell adhesion with an intervening fibrous capsule. Ti could be made bioactive by several methods including growing in situ TiO2 layer on Ti-surface. TiO2 nanotubes were grown on Ti surface via anodization process and the bone cell-material interactions were evaluated. Human osteoblast cell attachment and growth behavior were studied using an osteoprecursor cell line for 3, 7, and 11 days. An abundant amount of extracellular matrix (ECM) between the neighboring cells was noticed on anodized nanotube surface with filopodia extensions coming out from cells to grasp the nanoporous surface of the nanotube for anchorage. To better understand and compare cell-materials interactions, anodized nanoporous sample surfaces were etched with different patterns. Preferential cell attachment was noticed on nanotube surface compare to almost no cells in etched Ti surface. Cell adhesion with vinculin adhesive protein showed higher intensity, positive contacts on nanoporous surface and thin focal contacts on the Ti-control. Immunochemistry study with alkaline phosphatase showed enhanced osteoblastic phenotype expressions in nanoporous surface. Osteoblast proliferation was significantly higher on anodized nanotube surface. Surface properties changed with the emergence of nanoscale morphology. Higher nanometer scale roughness, low contact angle and high surface energy in nanoporous surface enhanced the osteoblast-material interactions. Mineralization study was done under simulated body fluid (SBF) with ion concentration nearly equal to human blood plasma to understand biomimetic apatite deposition behavior. Although apatite layer formation was noticed on nanotube surface, but it was nonuniform even after 21 days in SBF.

Influence of porosity on mechanical properties and in vivo response of Ti6Al4V implants

Metallic biomaterials are widely used to restore the lost structure and functions of human bone. Due to the large number of joint replacements, there is a growing demand for new and improved orthopedic implants. More specifically, there is a need for novel load-bearing metallic implants with low effective modulus matching that of bone in order to reduce stress shielding and consequently increase the in vivo lifespan of the implant. In this study, we have fabricated porous Ti6Al4V alloy structures, using laser engineered net shaping (LENS), to demonstrate that advanced manufacturing techniques such as LENS can be used to fabricate low-modulus, tailored porosity implants with a wide variety of metals/alloys, where the porosity can be designed in areas based on the patients need to enhance biological fixation and achieve long-term in vivo stability. The effective modulus of Ti6Al4V alloy structures has been tailored between 7 and 60 GPa and porous Ti alloy structures containing 23-32 vol.% porosity showed modulus equivalent to human cortical bone. In vivo behavior of porous Ti6Al4V alloy samples in male Sprague-Dawley rats for 16 weeks demonstrated a significant increase in calcium within the implants, indicating excellent biological tissue ingrowth through interconnected porosity. In vivo results also showed that total amount of porosity plays an important role in tissue ingrowth.

Effects of silica and zinc oxide doping on mechanical and biological properties of 3D printed tricalcium phosphate tissue engineering scaffolds

OBJECTIVES To evaluate the effects of silica (SiO(2)) (0.5 wt%) and zinc oxide (ZnO) (0.25 wt%) dopants on the mechanical and biological properties of tricalcium phosphate (TCP) scaffolds with three dimensionally (3D) interconnected pores. METHODS Scaffolds were created with a commercial 3D printer. Post sintering phase analysis was determined by X-ray diffraction. Surface morphology of the scaffolds was examined by field emission scanning electron microscopy (FESEM). Mechanical strength was evaluated with a screw driven universal testing machine. MTT assay was used for cellular proliferation characteristics and cellular morphology was examined by FESEM. RESULTS Addition of dopants into TCP increased the average density of pure TCP from 90.8 ± 0.8% to 94.1 ± 1.6% and retarded the β to α phase transformation at high sintering temperatures, which resulted in up to 2.5 fold increase in compressive strength. In vitro cell-materials interaction studies, carried out using hFOB cells, confirmed that the addition of SiO(2) and ZnO to the scaffolds facilitated faster cell proliferation when compared to pure TCP scaffolds. SIGNIFICANCE Addition of SiO(2) and ZnO dopants to the TCP scaffolds showed increased mechanical strength as well as increased cellular proliferation.

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. Copyright

Antibacterial and biological characteristics of silver containing and strontium doped plasma sprayed hydroxyapatite coatings.

Infection in primary total joint prostheses is estimated to occur in up to 3% of all surgery. As a measure to improve the antimicrobial properties of implant materials silver (Ag) was incorporated into plasma sprayed hydroxyapatite (HA) coatings. To offset potential cytotoxic effects of Ag in the coatings strontium (Sr) was also added as a binary dopant. HA powder was doped with 2.0 wt.% Ag(2)O, 1.0 wt.% SrO and was then heat treated at 800 °C. Titanium substrates were coated using a 30 kW plasma spray system equipped with a supersonic nozzle. X-ray diffraction confirmed the phase purity and high crystallinity of the coatings. Samples were evaluated for mechanical stability by adhesive bond strength testing. The results show that the addition of dopants did not affect the overall bond strength of the coatings. The antibacterial efficacies of the coatings were tested against Pseudomonas aeruginosa. Samples that contained the Ag(2)O dopant were found to be highly effective against bacterial colonization. In vitro cell-material interactions using human fetal osteoblast cells were characterized by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay for cell viability, field emission scanning electron microscopy for cell morphology and confocal imaging for the important differentiation marker alkaline phosphatase (ALP). Our results showed evidence of cytotoxic effects of the Ag-HA coatings, characterized by poor cellular morphology and cell death and nearly complete loss of functional ALP activity. The addition of SrO to the Ag-HA coatings was able to effectively offset these negative effects and improve performance compared with pure HA-coated samples.

Role of surface charge and wettability on early stage mineralization and bone cell–materials interactions of polarized hydroxyapatite

Our objective was to determine the role of surface charge and wettability on early stage mineralization as well as bone cell adhesion and proliferation on polarized HAp surface. To estimate the surface wettability, contact angles were measured in water, simulated body fluid (SBF) and Dulbeccos modified Eagles medium/nutrient mixture F-12 Ham (DMEM). Experimental results show that HAp surface wettability and surface energy can be tailored by inducing surface charge without introducing any volumetric effects in the material. Increasing the surface charge increased the wettability and also the energy of HAp surfaces in all tested media. A maximum surface energy of 49.47+/-3.76mJ/m(2) was estimated for positively charged HAp surfaces polarized at 400(o)C. The in vitro bioactivity of polarized HAp samples was evaluated by soaking in SBF and DMEM (cell media). Cell-materials interaction was studied by culturing with human fetal osteoblast cells (hFOB). In vitro results show that tailoring the combined effect of wettability and charge polarity on the HAp surface enable differential binding of inorganic ions (e.g., Ca(2+), Cl(-), Na(+), HCO(3)(-) etc) and organic cell adhesive proteins (e.g., fibronectin, vitronectin etc) with different surface properties, which results in accelerated or decelerated mineralization as well as cell adhesion and proliferation on polarized HAp surface.