Elongated Polyproline Motifs Facilitate Enamel Evolution through Matrix Subunit Compaction

How does proline-repeat motif length in the proteins of teeth and bones relate to the evolution of vertebrates? Counterintuitively, longer repeat stretches are associated with smaller aggregated subunits within a supramolecular matrix, resulting in enhanced crystal length in mammalian versus amphibian tooth enamel

Periodontal Bioengineering: A Discourse in Surface Topographies, Progenitor Cells and Molecular Profiles.

Periodontal Bioengineering: A Discourse in Surface Topographies, Progenitor Cells and Molecular Profiles

Successful Periodontal Ligament Regeneration by Periodontal Progenitor Preseeding on Natural Tooth Root Surfaces

The regeneration of lost periodontal ligament (PDL) and alveolar bone is the purpose of periodontal tissue engineering. The goal of the present study was to assess the suitability of 3 odontogenic progenitor populations from dental pulp, PDL, and dental follicle for periodontal regeneration when exposed to natural and synthetic apatite surface topographies. We demonstrated that PDL progenitors featured higher levels of periostin and scleraxis expression, increased adipogenic and osteogenic differentiation potential, and pronounced elongated cell shapes on barren root chips when compared with dental pulp and dental follicle cells. When evaluating the effect of surface characteristics on PDL progenitors, natural root surfaces resulted in elongated PDL cell shapes, whereas PDL progenitors on synthetic apatite surfaces were rounded or polygonal. In addition, surface coatings affected PDL progenitor gene expression profiles: collagen I coatings enhanced alkaline phosphatase and osteocalcin expression levels and laminin-1 coatings increased epidermal growth factor (EGF), nestin, cadherin 1, and keratin 8 expression. PDL progenitors seeded on natural tooth root surfaces in organ culture formed new periodontal fibers after 3 weeks of culture. Finally, replantation of PDL progenitor-seeded tooth roots into rat alveolar bone sockets resulted in the complete formation of a new PDL and stable reattachment of teeth over a 6-month period. Together, these findings indicate that periodontal progenitor cell type as well as mineral surface topography and molecular environment play crucial roles in the regeneration of true periodontal anchorage

Apatite Microtopographies Instruct Signaling Tapestries for Progenitor-Driven New Attachment of Teeth

Dimension and structure of extracellular matrix surfaces have powerful influences on cell shape, adhesion, and gene expression. Here we show that natural tooth root topographies induce integrin-mediated extracellular matrix signaling cascades in tandem with cell elongation and polarization to generate physiological periodontium-like tissues. In this study we replanted surface topography instructed periodontal progenitors into rat alveolar bone sockets for 8 and 16 weeks, resulting in complete reattachment of tooth roots to the surrounding alveolar bone with a periodontal fiber apparatus closely matching physiological controls along the entire root surface. Displacement studies and biochemical analyses confirmed that progenitor-based engineered periodontal tissues were similar to control teeth and uniquely derived from preimplantation green fluorescent protein (GFP)-labeled progenitors. Together, these studies illustrate the capacity of natural extracellular surface topographies to instruct progenitor cell populations to fully regenerate complex cellular and structural morphologies of tissues once lost to disease. We suggest that our strategy could be used for the replantation of teeth lost due to trauma or as a novel approach for tooth replacement using tooth-shaped replicas

Platelet-Rich Fibrin Promotes Periodontal Regeneration and Enhances Alveolar Bone Augmentation

In the present study we have determined the suitability of platelet-rich fibrin (PRF) as a complex scaffold for periodontal tissue regeneration. Replacing PRF with its major component fibrin increased mineralization in alveolar bone progenitors when compared to periodontal progenitors, suggesting that fibrin played a substantial role in PRF-induced osteogenic lineage differentiation. Moreover, there was a 3.6-fold increase in the early osteoblast transcription factor RUNX2 and a 3.1-fold reduction of the mineralization inhibitor MGP as a result of PRF application in alveolar bone progenitors, a trend not observed in periodontal progenitors. Subcutaneous implantation studies revealed that PRF readily integrated with surrounding tissues and was partially replaced with collagen fibers 2 weeks after implantation. Finally, clinical pilot studies in human patients documented an approximately 5 mm elevation of alveolar bone height in tandem with oral mucosal wound healing. Together, these studies suggest that PRF enhances osteogenic lineage differentiation of alveolar bone progenitors more than of periodontal progenitors by augmenting osteoblast differentiation, RUNX2 expression, and mineralized nodule formation via its principal component fibrin. They also document that PRF functions as a complex regenerative scaffold promoting both tissue-specific alveolar bone augmentation and surrounding periodontal soft tissue regeneration via progenitor-specific mechanisms

Platelet-Rich Fibrin Promotes Periodontal Regeneration and Enhances Alveolar Bone Augmentation

In the present study we have determined the suitability of platelet-rich fibrin (PRF) as a complex scaffold for periodontal tissue regeneration. Replacing PRF with its major component fibrin increased mineralization in alveolar bone progenitors when compared to periodontal progenitors, suggesting that fibrin played a substantial role in PRF-induced osteogenic lineage differentiation. Moreover, there was a 3.6-fold increase in the early osteoblast transcription factor RUNX2 and a 3.1-fold reduction of the mineralization inhibitor MGP as a result of PRF application in alveolar bone progenitors, a trend not observed in periodontal progenitors. Subcutaneous implantation studies revealed that PRF readily integrated with surrounding tissues and was partially replaced with collagen fibers 2 weeks after implantation. Finally, clinical pilot studies in human patients documented an approximately 5 mm elevation of alveolar bone height in tandem with oral mucosal wound healing. Together, these studies suggest that PRF enhances osteogenic lineage differentiation of alveolar bone progenitors more than of periodontal progenitors by augmenting osteoblast differentiation, RUNX2 expression, and mineralized nodule formation via its principal component fibrin. They also document that PRF functions as a complex regenerative scaffold promoting both tissue-specific alveolar bone augmentation and surrounding periodontal soft tissue regeneration via progenitor-specific mechanisms

Combinatorial Design of Hydrolytically Degradable, Bone-like Biocomposites Based on PHEMA and Hydroxyapatite.

With advantages such as design flexibility in modifying degradation, surface chemistry, and topography, synthetic bone-graft substitutes are increasingly demanded in orthopedic tissue engineering to meet various requirements in the growing numbers of cases of skeletal impairment worldwide. Using a combinatorial approach, we developed a series of biocompatible, hydrolytically degradable, elastomeric, bone-like biocomposites, comprising 60 wt% poly(2-hydroxyethyl methacrylate-co-methacrylic acid), poly(HEMA-co-MA), and 40 wt% bioceramic hydroxyapatite (HA). Hydrolytic degradation of the biocomposites is rendered by a degradable macromer/crosslinker, dimethacrylated poly(lactide-b-ethylene glycol-b-lactide), which first degrades to break up 3-D hydrogel networks, followed by dissolution of linear pHEMA macromolecules and bioceramic particles. Swelling and degradation were examined at Hank's balanced salt solution at 37 °C in a 12-week period of time. The degradation is strongly modulated by altering the concentration of the co-monomer of methacrylic acid and of the macromer, and chain length/molecular weight of the macromer. 95% weight loss in mass is achieved after degradation for 12 weeks in a composition consisting of HEMA/MA/Macromer = 0/60/40, while 90% weight loss is seen after degradation only for 4 weeks in a composition composed of HEMA/MA/Macromer = 27/13/60 using a longer chain macromer. For compositions without a co-monomer, only about 14% is achieved in weight loss after 12-week degradation. These novel biomaterials offer numerous possibilities as drug delivery carriers and bone grafts particularly for low and medium load-bearing applications

Combinatorial Design of Hydrolytically Degradable, Bone-like Biocomposites Based on PHEMA and Hydroxyapatite.

With advantages such as design flexibility in modifying degradation, surface chemistry, and topography, synthetic bone-graft substitutes are increasingly demanded in orthopedic tissue engineering to meet various requirements in the growing numbers of cases of skeletal impairment worldwide. Using a combinatorial approach, we developed a series of biocompatible, hydrolytically degradable, elastomeric, bone-like biocomposites, comprising 60 wt% poly(2-hydroxyethyl methacrylate-co-methacrylic acid), poly(HEMA-co-MA), and 40 wt% bioceramic hydroxyapatite (HA). Hydrolytic degradation of the biocomposites is rendered by a degradable macromer/crosslinker, dimethacrylated poly(lactide-b-ethylene glycol-b-lactide), which first degrades to break up 3-D hydrogel networks, followed by dissolution of linear pHEMA macromolecules and bioceramic particles. Swelling and degradation were examined at Hank's balanced salt solution at 37 °C in a 12-week period of time. The degradation is strongly modulated by altering the concentration of the co-monomer of methacrylic acid and of the macromer, and chain length/molecular weight of the macromer. 95% weight loss in mass is achieved after degradation for 12 weeks in a composition consisting of HEMA/MA/Macromer = 0/60/40, while 90% weight loss is seen after degradation only for 4 weeks in a composition composed of HEMA/MA/Macromer = 27/13/60 using a longer chain macromer. For compositions without a co-monomer, only about 14% is achieved in weight loss after 12-week degradation. These novel biomaterials offer numerous possibilities as drug delivery carriers and bone grafts particularly for low and medium load-bearing applications

Integrative Temporo-Spatial, Mineralogic, Spectroscopic, and Proteomic Analysis of Postnatal Enamel Development in Teeth with Limited Growth

Tooth amelogenesis is a complex process beginning with enamel organ cell differentiation and enamel matrix secretion, transitioning through changes in ameloblast polarity, cytoskeletal, and matrix organization, that affects crucial biomineralization events such as mineral nucleation, enamel crystal growth, and enamel prism organization. Here we have harvested the enamel organ including the pliable enamel matrix of postnatal first mandibular mouse molars during the first 8 days of tooth enamel development to conduct a step-wise cross-sectional analysis of the changes in the mineral and protein phase. Mineral phase diffraction pattern analysis using single-crystal, powder sample X-ray diffraction analysis indicated conversion of calcium phosphate precursors to partially fluoride substituted hydroxyapatite from postnatal day 4 (4 dpn) onwards. Attenuated total reflectance spectra (ATR) revealed a substantial elevation in phosphate and carbonate incorporation as well as structural reconfiguration between postnatal days 6 and 8. Nanoscale liquid chromatography coupled with tandem mass spectrometry (nanoLC-MS/MS) demonstrated highest protein counts for ECM/cell surface proteins, stress/heat shock proteins, and alkaline phosphatase on postnatal day 2, high counts for ameloblast cytoskeletal proteins such as tubulin β5, tropomyosin, β-actin, and vimentin on postnatal day 4, and elevated levels of cofilin-1, calmodulin, and peptidyl-prolyl cis-trans isomerase on day 6. Western blot analysis of hydrophobic enamel proteins illustrated continuously increasing amelogenin levels from 1 dpn until 8 dpn, while enamelin peaked on days 1 and 2 dpn, and ameloblastin on days 1–5 dpn. In summary, these data document the substantial changes in the enamel matrix protein and mineral phase that take place during postnatal mouse molar amelogenesis from a systems biological perspective, including (i) relatively high levels of matrix protein expression during the early secretory stage on postnatal day 2, (ii) conversion of calcium phosphates to apatite, peak protein folding and stress protein counts, and increased cytoskeletal protein levels such as actin and tubulin on day 4, as well as (iii) secondary structure changes, isomerase activity, highest amelogenin levels, and peak phosphate/carbonate incorporation between postnatal days 6 and 8. Together, this study provides a baseline for a comprehensive understanding of the mineralogic and proteomic events that contribute to the complexity of mammalian tooth enamel development