Periodontitis is a leading chronic oral inflammatory disease and primary cause of permanent tooth loss estimated to affect 47.2% of adults in the United States. Damage to the tooth-supporting apparatus, which includes periodontal ligament (PDL) fibers that anchor the tooth root to alveolar bone, subsequently initiates osseous tissue resorption. Multi-tissue morbidity is a significant challenge given lack of predictability in reconstructing tissues with physiologic functionality native to healthy periodontium. Tissue engineering strategies have potential to address existing deficiencies in clinically-induced regeneration through combinational approaches using biomaterials, growth factors, and cell-based therapy. The purpose of this work was to develop scaffolds incorporating micropatterned topography for guidance of cell growth and periodontal tissue formation, in conjunction with localized, spatiotemporally-controlled growth factor delivery via gene therapy vectors.

Micropatterned polycaprolactone (PCL) films were designed to assess PDL cell orientation in vitro, with incorporation of the patterned film into a 3D-printed PCL scaffold for evaluation of varying topography on oriented tissue formation in an ectopic murine model. Specifically, pillars with varying groove depths (30um, 10um) and groove widths (15um, 60um) were used for the “PDL” region of the scaffold in combination with human PDL (hPDL) cell seeding, while the 3D-printed base served as a region for osseous tissue formation via delivery of human gingival fibroblasts (hGFs) transduced with adenoviral bone morphogenetic protein (Ad-BMP7). Micropatterned films with pillars containing deeper grooves (30um) provided greater control over hPDL cell orientation and subsequent alignment of soft collagenous tissue compared to non-grooved pillars or an amorphous PCL film, with significant (p<0.05) differences in percentage of aligned cells in vivo observed at 6 weeks post-implantation.

In order to improve spatially-controlled delivery of BMP7 and platelet-derived growth factor (PDGF-BB) using developed 3D-printed, micropatterned scaffolds, each region of the scaffold was separately immobilized with AdBMP7 and AdPDGF-BB, respectively, using chemical vapor deposition (CVD)-based surface modification prior to cell seeding. A separate scaffold was developed for a rat fenestration defect, with the 3D-printed scaffold region replaced by an amorphous PCL film to accommodate the 0.5mm defect thickness. Evaluation of these cell-seeded scaffolds showed significant (p<0.05) bone formation in regions with immobilized AdBMP7 compared to regions immobilized with empty vectors (Ad-empty) and non-cell seeded regions immobilized with AdBMP7. A more detailed assessment of single (BMP7) and dual (BMP7 and PDGF-BB) growth factor delivery effects in combination with varying scaffold topography (i.e., patterned film versus amorphous film in the “PDL” region) was performed using the fenestration defect model. Micro-CT data showed significantly higher (p<0.05) bone formation in groups with AdBMP7 immobilization and gingival fibroblast cell seeding compared to groups with Ad-empty. Collagen III and periostin expression was higher in groups with dual growth factor delivery, with significantly (p<0.05) higher periostin expression in groups combining a patterned film with single or dual growth factor delivery at week 6. Nanoindentation assessment showed higher elastic moduli for regenerated bone and PDL-like tissue regions at the bone-PDL interface in patterned film groups with dual growth factor delivery compared to amorphous films with Ad-empty at week 9 (p<0.05 and p<0.01, respectively), with bone tissue in the dual delivery group having higher (p<0.001) hardness values compared to the negative control. These data indicate improvement in periodontal tissue regeneration when combining scaffold micro-topography cues with localized growth factor delivery, thereby contributing to the development of next-generation scaffolds specific to periodontal regenerative medicine.