Subhadip Bodhak

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.

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.

Electrically polarized HAp-coated Ti: In vitro bone cell–material interactions

Electrically polarized bulk sintered hydroxyapatite (HAp) compacts have been shown to accelerate mineralization and bone tissue ingrowth in vivo. In this work, a comprehensive study has been carried out to investigate the influence of surface charge and polarity on in vitro bone cell adhesion, proliferation and differentiation on electrically polarized HAp-coated Ti. Uniform and crack free sol-gel derived HAp coatings of 20+/-1.38microm thickness were polarized by application of an external d.c. field of 2.0kVcm(-1) at 400 degrees C for 1h. In vitro bioactivity of polarized HAp coatings was evaluated by soaking in simulated body fluid, and bone cell-material interactions were studied by culturing with human fetal osteoblast cells (hFOB) for a maximum period of 11 days. Scanning electron microscopic observation showed that accelerated mineralization on negatively charged surfaces favored rapid cell attachment and faster tissue ingrowth over non-polarized HAp coating surfaces, while positive charge on HAp coating surfaces restricted apatite nucleation with limited cellular response. Immunochemistry and confocal microscopy confirmed that the cell adhesion and early stage differentiation were more pronounced on negatively charged coating surfaces as hFOB cells expressed higher vinculin and alkaline phosphatase proteins on negatively charged surface compared to cells grown on all other surfaces. Our results in this study are process independent and potentially applicable to any other commercially available coating techniques.

Effect of electrical polarization and composition of biphasic calcium phosphates on early stage osteoblast interactions.

In spite of having excellent biocompatibility and osteogenic property of hydroxyapatite (HAp) and β-tricalcium phosphate (β-TCP), concerns have been raised regarding their degradation kinetics. Complete in vivo degradation of HAp takes years because of its slow degradation rate, and fast degradation rate of β-TCP limits its application. Biphasic calcium phosphates (BCPs) composed of both HAp and β-TCP have controlled degradation to some extent. Here, we have prepared three different BCPs composed of β-TCP and HAp. These BCP composites are successfully electrically polarized to generate surfaces with positive (P-poled) and negative (N-poled) charges. Thermally stimulated depolarized current measurement (TSDC) exhibits increased stored charge density with the increase in HAp percentage in the composites. Our study focuses on understanding the effect of composition variation as well as electrical polarization of these composites on early stage osteoblast-cell adhesion, proliferation, and extracellular matrix (ECM) formation capability. No matter what the composition is, N-poled (Negatively poled) surfaces show early stage osteoblast-cell adhesion, proliferation, and ECM formation when compared with U-poled (unpoled) and P-poled (positively poled) surfaces. Irrespective of the surface charge, an improved cell-material interactions are observed as the percentage of HAp content in the composites is increased. These electrically polarized BCP composites can have potential use in the area of orthopedics and dentistry.

3D Cellular Morphotyping of Scaffold Niches

There is currently no method for assessing the nature of the cell niche provided by 3D biomaterial scaffolds. Analyzing human bone marrow stromal cell (hBMSC) 3D cell shape in response to different biomaterial scaffolds allowed the 3D cell niche promoted by biomaterial scaffolds to be evaluated. Primary hBMSCs (p5) were seeded (5,000 cells/cm2) in 10 different biomaterial scaffolds and cultured for 24 h. Samples were fixed and stained for actin and nucleus, imaged with confocal microscopy to obtain a 3D volume (z-stack), and 3D cell shape was analyzed with computational approaches. Over 100 cells were imaged per scaffold group (10 scaffold groups, ~1250 cells total), resulting in the largest known 3D stem cell dataset (~135,000 files, ~135 GB) and enabling a high degree of statistical rigor. The images were segmented using an automated algorithm and a final dataset of 969 well-segmented cells were analyzed with 79 shape metrics, which enabled 3D cellular morphotyping of scaffold niches. The variety of scaffolds studied promoted different cell morphologies during culture and there were significant differences in shape metrics, particularly for cell depth, surface area, and volume. This study demonstrated a quantitative approach to analyze 3D cell shape and morphotype and is the largest known study analyzing 3D cell shape in response to a variety of biomaterial scaffolds. The dataset is publically accessible with an online 3D viewer. These results could inform the selection of prospective scaffolds for applications based on 3D cell shape in the tissue of interest.