High resolution DLP stereolithography to fabricate biocompatible hydroxyapatite structures that support osteogenesis

Current methods for fabricating bone scaffolds face limitations in achieving the precise geometries and biocompatibility needed for successful orthopaedic applications. While additive manufacturing techniques like digital light processing (DLP) show promise, their suitability for biocompatible materials such as hydroxyapatite (HA) remains underexplored. Prior research has shown that HA supports bone regeneration, but its use in 3D printing is still limited by concerns about material compatibility and cell response. This gap motivated a study to assess HA printed via DLP for its ability to support cell viability, proliferation, and osteogenic differentiation. No prior work had resolved whether DLP-printed HA could meet the mechanical and biological requirements of bone scaffolds. The study aimed to test whether HA coupons printed using DLP could serve as a feasible platform for future scaffold development. This uncertainty drove the need to evaluate surface properties and biological performance of the printed material. The lack of data on leachable and extractable effects from the printing process also required investigation. The study addressed these questions by combining material characterization with in vitro cell culture experiments.

The primary aim of the study was to assess the biocompatibility and osteogenic potential of hydroxyapatite (HA) coupons fabricated using digital light processing (DLP) 3D printing. The specific problem addressed was whether DLP-printed HA could support cell viability, proliferation, and osteogenic differentiation of human cell lines and mesenchymal stem cells (MSCs). The motivation for this work stemmed from the need to develop advanced scaffolds for bone regeneration that combine complex geometries with favorable biological properties. The study sought to determine if DLP HA coupons could serve as a reliable platform for future scaffold designs. By evaluating cell behavior on printed coupons, the researchers aimed to establish the feasibility of using DLP for orthopaedic applications. The study also aimed to investigate the impact of the printing process on material leachables and extractables. The researchers hypothesized that DLP HA coupons would support cell growth and osteogenesis comparable to conventional substrates. This hypothesis was tested using a combination of material analysis and cell culture experiments.

The study employed digital light processing (DLP) to fabricate hydroxyapatite (HA) coupons using a LithaBone HA400 slurry. Surface properties of the coupons were analyzed using contact angle measurements, profilometry, and scanning electron microscopy (SEM) to assess hydrophilicity, porosity, and surface roughness. Cell viability and proliferation were evaluated using the U2OS human cell line and human mesenchymal stem cells (MSCs) cultured on the coupons for up to 35 days. Cell proliferation was measured using direct cell counting and the RealTime-Glo™ MT Cell Viability Assay. Osteogenic differentiation was assessed using alkaline phosphatase activity, Alizarin Red S staining, and von Kossa staining. Morphological changes in MSCs cultured in osteogenic and adipogenic media were observed over time. The study compared cell behavior on DLP HA coupons with that on conventional tissue culture polystyrene plates. Leachable and extractable effects from the printing process were also evaluated. The experimental design ensured that all findings were directly tied to the material and cell interactions under investigation.

The DLP-printed hydroxyapatite (HA) coupons exhibited favorable surface properties, including hydrophilicity, porosity, and micro-scale roughness suitable for cell culture. No significant leachable or extractable effects were observed from the printing process. Cell viability and proliferation on HA coupons were comparable to those on conventional tissue culture polystyrene plates, as measured by direct cell counts and the RealTime-Glo™ MT Cell Viability Assay. Human mesenchymal stem cells (MSCs) cultured on HA coupons showed osteogenic differentiation confirmed by alkaline phosphatase activity, Alizarin Red S staining, and von Kossa staining. Morphological analysis revealed that MSCs cultured in osteogenic medium for 14 to 35 days exhibited geometric, cuboidal morphology with dark nodules on HA coupons, similar to those on tissue culture plates. Adipogenic features, such as lipid vesicles and deposits, were also observed. The study found no evidence of cytotoxic effects from the DLP printing process. The results demonstrated that HA coupons printed using DLP are biocompatible and suitable for osteogenic applications.

The study concluded that hydroxyapatite (HA) coupons fabricated using digital light processing (DLP) are biocompatible and support osteogenic differentiation of human mesenchymal stem cells (MSCs). The HA coupons exhibited favorable surface properties, including hydrophilicity, porosity, and micro-scale roughness. No leachable or extractable effects from the printing process were observed. Cell viability and proliferation on HA coupons were comparable to those on conventional tissue culture polystyrene plates. Osteogenic differentiation of MSCs was confirmed using multiple staining techniques. The morphology of MSCs cultured in osteogenic medium was similar on HA coupons and tissue culture plates. The findings suggest that DLP HA coupons serve as an effective platform for evaluating cell responses on 3D-printed materials. The results support the feasibility of using DLP for future development of complex scaffold geometries that meet mechanical, chemical, and porosity requirements for artificial bone scaffolds.