Specific orientations of regenerated ligaments are crucially required for mechanoresponsive properties and various biomechanical adaptations, which are the key interplay to support mineralized tissues. Although various 2D platforms or 3D printing systems can guide cellular activities or aligned organizations, it remains a challenge to develop ligament-guided, 3D architectures with the angular controllability for parallel, oblique or perpendicular orientations of cells required for biomechanical support of organs. Here, we show the use of scaffold design by additive manufacturing for specific topographies or angulated microgroove patterns to control cell orientations such as parallel (0°), oblique (45°) and perpendicular (90°) angulations. These results demonstrate that ligament cells displayed highly predictable and controllable orientations along microgroove patterns on 3D biopolymeric scaffolds. Our findings demonstrate that 3D printed topographical approaches can regulate spatiotemporal cell organizations that offer strong potential for adaptation to complex tissue defects to regenerate ligament-bone complexes.
Currently, various tissue engineering strategies have been developed for multiple tissue regeneration and integrative structure formations as well as single tissue formation in musculoskeletal complexes. In particular, the regeneration of periodontal tissues or tooth-supportive structures is still challenging to spatiotemporally compartmentalize PCL (poly-ε-caprolactone)-cementum constructs with micron-scaled interfaces, integrative tissue (or cementum) formations with optimal dimensions along the tooth-root surfaces, and specific orientations of engineered periodontal ligaments (PDLs). Here, we discuss current advanced approaches to spatiotemporally control PDL orientations with specific angulations and to regenerate cementum layers on the tooth-root surfaces with Sharpey’s fiber anchorages for state-of-the-art periodontal tissue engineering.
At present, various tissue engineering strategies have been developed for multiple tissue regeneration and integrative structure formations. However, the regenerations of tooth-supportive structures are still limited and challenging due to the micro-interfacial compartmentalization of multiple tissues, their integrations for systematic responses, and spatiotemporal organizations of engineered tissues. Here, we investigated the scaffold prototype as the regeneration platform of the periodontal complex (cementum-periodontal ligament (PDL)-bone). Based on the tooth image dataset, the prototype scaffold was designed with individual periodontal tissues while using the three-dimensional (3D) printing technique and solvent-casting method with poly-ε-caprolactone (PCL). The architecture was characterized by scanning electron microscope (SEM) and biological assessments were performed with human periodontal ligament (hPDL) cells by confocal microscope. In particular, the angulations and deformations of hPDL cells on PDL architectures were analyzed while using nuclear aspect ratio (NAR = 2.319 ± 0.273) and nuclear shape index (NSI (circularity) = 0.546 ± 0.0273). In in-vitro, designed surface microgroove patterns facilitated angular organizations of hPDL cells (frequency of 0–10° angulations = 75 ± 9.54 out of 97.3 ± 2.52) for seven days. The prototype scaffolding system showed geometric adaptation to the digitized image dataset, hPDL orientations on microgroove-patterned surface, and architectural compartmentalizations for periodontal tissue regeneration.
Specific orientations of periodontal ligaments (PDLs) to tooth-root surface play an important role in offering positional stabilities of teeth, transmitting and absorbing various stresses under masticatory/occlusal loading conditions, or promoting tissue remodeling by mechanical stimulations to periodontal cells. However, it is still challenging to spatially control PDL orientations and collective PDL cell alignments using 3D scaffold architectures. Here, we investigated the optimization of scaffold topographies in order to control orientations of human PDL cells with predictability in in vitro. The 3D PDL-guiding architectures were designed by computer-aided design (CAD) and microgroove patterns on the scaffold surfaces were created with four different slice intervals such as 25.40 µm (μG-25), 19.05 µm (μG-19), 12.70 µm (μG-12), and 6.35 µm (μG-6) by the digital slicing step. After scaffold design and 3D wax printing, poly-ε-caprolactone (PCL) was casted into 3D printed molds and human PDL cells were cultured for 7 days. In the results, μG-25 with low vertical resolution can angularly organize seeded cells predictably rather than μG-6 created by the highest resolution for high surface quality (or smooth surface). Moreover, nuclear orientations and deformability were quantitatively analyzed and a significant correlation between microgroove pattern intervals and cell alignments was calculated for the topographic optimization. In conclusion, controllable microgroove intervals can specifically organize human PDL cells by 3D printing, which can create various surface topographies with structural consistence. The optimal surface topography (μG-25) can angularly guide human PDL cells, but 6.35 µm-thick patterns (μG-6) showed random organization of cell collectivity.
The mineralized tissues (alveolar bone and cementum) are the major components of periodontal tissues and play a critical role to anchor periodontal ligament (PDL) to tooth-root surfaces. The integrated multiple tissues could generate biological or physiological responses to transmitted biomechanical forces by mastication or occlusion. However, due to periodontitis or traumatic injuries, affect destruction or progressive damage of periodontal hard tissues including PDL could be affected and consequently lead to tooth loss. Conventional tissue engineering approaches have been developed to regenerate or repair periodontium but, engineered periodontal tissue formation is still challenging because there are still limitations to control spatial compartmentalization for individual tissues and provide optimal 3D constructs for tooth-supporting tissue regeneration and maturation. Here, we present the recently developed strategies to induce osteogenesis and cementogenesis by the fabrication of 3D architectures or the chemical modifications of biopolymeric materials. These techniques in tooth-supporting hard tissue engineering are highly promising to promote the periodontal regeneration and advance the interfacial tissue formation for tissue integrations of PDL fibrous connective tissue bundles (alveolar bone-to-PDL or PDL-to-cementum) for functioning restorations of the periodontal complex.
The periodontal ligaments (PDLs) with specific orientations to tooth-root surfaces play a key role in generating biomechanical responses between the alveolar bone and cementum as a tooth-supporting tissue. However, control of angulations and regeneration of the ligamentous tissues within micron-scaled interfaces remains challenging. To overcome this limitation, this study investigated surface fabrications with microgroove patterns to control orientations of rat PDL cells in vitro and fibrous tissues in vivo. After being harvested, rat PDL cells were cultured and three different microgroove patterns (∠PDL groove = 0°, ∠PDL groove = 45°, and ∠PDL groove = 90°) were created by the digital slicing step in 3D printing. Cell-seeded scaffolds were subcutaneously transplanted at 3 and 6 weeks. In histology images, rat PDL cells were spatially controlled to angularly organize following the microgroove patterns and fibrous tissues were formed in scaffolds with specific angulations, which were reflected by additively manufactured microgroove topographies. Based on the results, specifically characterized surface topographies were significant to directly/indirectly organizing rat PDL cell alignments and fibrous tissue orientations. Therefore, interactions between surface topographies and tissue organizations could be one of the key moderators for the multiple tissue complex (bone-ligament-cementum) neogenesis in periodontal tissue engineering.
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