Regulation of Human Osteoblast-to-Osteocyte Differentiation by Direct-Write 3D Microperiodic Hydroxyapatite Scaffolds

Abstract

Culturing osteocytes on planer surfaces on Petri dish in vitro fails to recapitulate their natural orientation as well as biological functionality in human bone. Ability to recapitulate spatial arrangement of osteoblasts can govern signaling cascades for osteoblast-to-osteocyte differentiation and osteogenesis. Therefore, in this study, we explored how direct-write microperiodic hydroxyapatite (HAP) three-dimensional (3D) scaffolds can modulate osteocyte differentiation as compared to two-dimensional (2D) HAP scaffolds. Increased level of osteocalcin expression and relatively early mineralization on 2D HAP were indicative of osteoblast maturation. On the other hand, unique features of 3D HAP direct-write scaffolds, in terms of the architecture and subtle difference in chemical composition, surface roughness, and stiffness, play a pivotal role in governing the complex mechanism of differentiation of osteoblast to osteocyte-like phenotype. This was characterized by sequential events involving modulation in proliferation rate, time-dependent downregulation in the expression of osteoblast markers, collagen type I and alkaline phosphatase, followed by the unregulated expression of matrix metalloproteinase, osteocytic terminal differentiation markers (podoplanin, sclerostin), and dendritic extension formation. Thus, establishment of a simple in vitro model to study osteoblast differentiation with respect to specific cues would offer potential as an investigative platform and versatile tool for developing a 3D in vitro model for regenerative medicine.