Strain Measurement within an Intact Swine Periodontal Ligament.

The periodontal ligament (PDL) provides support, proprioception, nutrition, and protection within the tooth-PDL-bone complex (TPBC). While understanding the mechanical behavior of the PDL is critical, current research has inferred PDL mechanics from finite element models, from experimental measures on complete TPBCs, or through direct measurement of isolated PDL sections. Here, transducers are used in an attempt to quantify ex vivo PDL strain. In-fiber Bragg grating (FBG) sensors are small flexible sensors that can be placed within an intact TPBC and yield repeatable strain measurements from within the PDL space. The objective of this study was to determine: 1) if the FBG strain measured from the PDL space of intact swine premolars ex vivo was equivalent to physical PDL strains estimated through finite element analysis and 2) if a change in FBG strain could be linearly related to a change in finite element strain under variable tooth displacement, applied to an intact swine TPBC. Experimentally, individual TPBCs were subjected to 2 displacements (n = 14). The location of the FBG was determined from representative micro-computed tomography images. From a linear elastic finite element model of a TPBC, the strain magnitudes at the sensor locations were recorded. An experimental ratio (i.e., FBG strain at the first displacement divided by the FBG strain at the second displacement) and a finite element ratio (i.e., finite element strain at the first displacement divided by the finite element strain at the second displacement) were calculated. A linear regression model indicated a statistically significant relationship between the experimental and finite element ratio (P = 0.017) with a correlation coefficient (R2) of 0.448. It was concluded that the FBG sensor could be used as a measure for a change in strain and thus could be implemented in applications where the mechanical properties of an intact PDL are monitored over time.

Distinct post-translational features of type I collagen are conserved in mouse and human periodontal ligament.

BACKGROUND AND OBJECTIVE: Specifics of the biochemical pathways that modulate collagen cross-links in the periodontal ligament (PDL) are not fully defined. Better knowledge of the collagen post-translational modifications that give PDL its distinct tissue properties is needed to understand the pathogenic mechanisms of human PDL destruction in periodontal disease. In this study, the post-translational phenotypes of human and mouse PDL type I collagen were surveyed using mass spectrometry. PDL is a highly specialized connective tissue that joins tooth cementum to alveolar bone. The main function of the PDL is to support the tooth within the alveolar bone while under occlusal load after tooth eruption. Almost half of the adult population in the USA has periodontal disease resulting from inflammatory destruction of the PDL, leading to tooth loss. Interestingly, PDL is unique from other ligamentous connective tissues as it has a high rate of turnover. Rapid turnover is believed to be an important characteristic for this specialized ligament to function within the oral-microbial environment. Like other ligaments, PDL is composed predominantly of type I collagen. Collagen synthesis is a complex process with multiple steps and numerous post-translational modifications including hydroxylation, glycosylation and cross-linking. The chemistry, placement and quantity of intermolecular cross-links are believed to be important regulators of tissue-specific structural and mechanical properties of collagens. MATERIAL AND METHODS: Type I collagen was isolated from several mouse and human tissues, including PDL, and analyzed by mass spectrometry for post-translational variances. RESULTS: The collagen telopeptide cross-linking lysines of PDL were found to be partially hydroxylated in human and mouse, as well as in other types of ligament. However, the degree of hydroxylation and glycosylation at the helical Lys87 cross-linking residue varied across species and between ligaments. These data suggest that different types of ligament collagen, notably PDL, appear to have evolved distinctive lysine/hydroxylysine cross-linking variations. Another distinguishing feature of PDL collagen is that, unlike other ligaments, it lacks any of the known prolyl 3-hydroxylase 2-catalyzed 3-hydroxyproline site modifications that characterize tendon and ligament collagens. This gives PDL a novel modification profile, with hybrid features of both ligament and skin collagens. CONCLUSION: This distinctive post-translational phenotype may be relevant for understanding why some individuals are at risk of rapid PDL destruction in periodontal disease and warrants further investigation. In addition, developing a murine model for studying PDL collagen may be useful for exploring potential clinical strategies for promoting PDL regeneration.

Eruptive and functional changes in periodontal ligament fibroblast orientation in CD44 wild-type vs. knockout mice.

BACKGROUND AND OBJECTIVE: Periodontal ligament (PDL) fibroblasts establish principal fibers of the ligament during tooth eruption, and maintain these fibers during occlusion. PDL development and occlusal adaptation includes changes in the orientation of PDL fibroblasts; however, the mechanism for these changes in orientation is unclear. The objective of this study was to compare PDL fibroblast orientation in different stages corresponding with first molar eruption and occlusion in CD44 wild-type (WT) and knockout (KO) mice. MATERIAL AND METHODS: CD44 WT and KO mice were raised to six postnatal stages corresponding with first molar (M1 ) eruption (postnatal day 8, 11, 14 and 18) and occlusion (postnatal day 26 and 41). Coronal sections of the first mandibular molar (M1 ) were prepared and the orientation of fibroblasts in the cervical root region was measured. Angle measurements were compared across developmental stages and between strains using Watson-Williams F-test (oriana software) and ANCOVA. RESULTS: PDL fibroblast orientation increased significantly in CD44 WT (9-87°) and KO mice (14-93°; p ≤ 0.05) between intraosseous eruption (day 11), mucosal penetration (day 14) and preocclusal eruption (day 18); however, the PDL fibroblast orientation did not change significantly with the onset of occlusion (day 26) or continued function (day 41). Within each strain, the variance in fibroblast orientation during preocclusal eruption (day 18) was significantly higher than the variance of all other time points (p < 0.0005). CD44 WT and KO mice showed a similar pattern of PDL development and eruption with a significant difference in CD44 WT vs. KO fibroblast orientations only during early function (day 26, 92° vs 116°; p = 0.05). CONCLUSIONS: The development of PDL fibroblast orientation is highly similar between CD44 WT and KO mice. Between early (day 11) and late (day 18) eruptive stages PDL fibroblast orientation increases, corresponding with the upward movement of M1 . The PDL fibroblast orientation established in preocclusal eruption (day 18) is maintained during early (day 26) and late (day 41) stages of occlusal function, suggesting that PDL cells adapt to mechanical loads in the oral cavity before M1 occlusion.

The effects of tooth extraction on alveolar bone biomechanics in the miniature pig, Sus scrofa.

OBJECTIVE: This study investigated the role of occlusion in the development of biomechanical properties of alveolar bone in the miniature pig, Sus scrofa. The hypothesis tested was that the tissues supporting an occluding tooth would show greater stiffness and less strain than that of a non-occluding tooth. DESIGN: Maxillary teeth opposing the erupting lower first molar (M(1)) were extracted on one side. Occlusion developed on the contralateral side. Serially administered fluorochrome labels tracked bone mineralisation apposition rate (MAR). A terminal experiment measured in vivo buccal alveolar bone strain on occluding and non-occluding sides during mastication. Ex vivo alveolar strains during occlusal loading were subsequently measured using a materials testing machine (MTS/Sintech). Whole specimen stiffness and principal strains were calculated. RESULTS: MAR tended to be higher on the extraction side during occlusion. In vivo buccal shear strains were higher in the alveolar bone of the occluding side vs. the extraction side (mean of 471 microvarepsilon vs. 281 microvarepsilon, respectively; p=0.04); however, ex vivo shear strains showed no significant differences between sides. Stiffness differed between extraction and occlusion side specimens, significantly so in the low load range (344 vs. 668 MPa, respectively; p=0.04). CONCLUSIONS: Greater in vivo shear strains may indicate more forceful chews on the occluding side, whereas the similarity in ex vivo bone strain magnitude suggests a similarity in alveolar bone structure and occlusal load transmission regardless of occlusal status. The big overall change in specimen stiffness that was observed was likely attributable to differences in the periodontal ligament rather than alveolar bone.

Functional cues in the development of osseous tooth support in the pig, Sus scrofa.

Alveolar bone supports teeth during chewing through a ligamentous interface with tooth roots. Although tooth loads are presumed to direct the development and adaptation of these tissues, strain distribution in the alveolar bone at different stages of tooth eruption and periodontal development is unknown. This study investigates the biomechanical effects of tooth loading on developing alveolar bone as a tooth erupts into occlusion. Mandibular segments from miniature pigs, Sus scrofa, containing M(1) either erupting or in functional occlusion, were loaded in compression. Simultaneous recordings were made from rosette strain gages affixed to the lingual alveolar bone and the M(2) crypt. Overall, specimens with erupting M(1)s were more deformable than specimens with occluding M(1)s (mean stiffness of 246 vs. 944 MPa, respectively, p=0.004). The major difference in alveolar strain between the two stages was in orientation. The vertically applied compressive loads were more directly reflected in the alveolar bone strains of erupting M(1)s, than those of occluding M(1)s, presumably because of the mediation of a more mature periodontal ligament (PDL) in the latter. The PDL interface between occluding teeth and alveolar bone is likely to stiffen the system, allowing transmission of occlusal loads. Alveolar strains may provide a stimulus for bone growth in the alveolar process and crest.

Defining the roots of cementum formation.

Significant progress has been seen in research aimed at regeneration of the disease-damaged periodontium. Our own strategy has been to approach periodontal tissue development (i.e. root, cementum, periodontal ligament, and bone) as a source for the identification of key regulators of cellular processes that may be applicable to periodontal tissue repair. Specifically, enamel-like molecules, bone morphogenetic proteins (BMPs), and phosphates have been investigated for their role in altering gene expression and cell functions in follicle cells, periodontal ligament cells, and cementoblasts. Amelogenin, leucine-rich amelogenin peptide, and tyrosine-rich amelogenin peptide have been found to similarly affect cementoblast gene expression and cementoblast-mediated mineralization in vitro; however, these enamel-like factors do not increase cell proliferation as has been observed in cells treated with Emdogain (Biora AB, Malmö, Sweden), an enamel matrix derivative. BMP-2 has been found to promote differentiation of follicle cells into a cementoblast/osteoblast phenotype, and BMP-3 is being investigated as a negative regulator of mineralization. The increased ratio of phosphate to pyrophosphate in the local region during root development has been found to significantly enhance the extent of cementum formation in animal models. Furthermore, phosphate has been identified as a regulator of cementoblast SIBLING (small integrin-binding ligand N-linked glycoprotein) gene expression in vitro. These investigations of candidate factors for periodontal regeneration have uncovered mechanisms regulating gene expression and cell function in cells controlling the behavior of periodontal tissues (i.e. follicle cells, periodontal cells, and cementoblasts) and offer new directions to consider for clinical repair of periodontal defects.

Enamel microstructure and microstrain in the fracture of human and pig molar cusps.

The role of microstructure in enamel strain and breakage was investigated in human molar cusps and those of the pig, Sus scrofa. Rosette strain gauges were affixed to cusp surfaces (buccal human M3, n=15, and lingual pig M1, n=13), and a compressive load was applied to individual cusps using an MTS materials testing machine. Load and strain data were recorded simultaneously until cusp fracture, and these data were used to estimate enamel stresses, principal strains, and stiffness. Fractured and polished enamel fragments were examined in multiple planes using scanning electron microscopy (SEM). Human cusp enamel showed greater stiffness than pig enamel (P=0.02), and tensile stress at yield was higher (17.9 N/mm2 in humans versus 8.9 N/mm2 in pigs, P=0.06). SEM revealed enamel rod decussation in both human and pig enamel; however, only pig enamel showed a decussation plane between rod and inter-rod crystallites. Human inter-rod enamel was densely packed between rods, whereas in pig enamel, inter-rod enamel formed partitions between rows of enamel rods. Overall, human enamel structure enabled molar cusps to withstand horizontal tensile stress during both elastic and plastic phases of compressive loading. In contrast, pig cusp enamel was less resistant to horizontal tensile stresses, but appeared to fortify the enamel against crack propagation in multiple directions. These structural and biomechanical differences in cusp enamel are likely to reflect species-level differences in occlusal function.

Mechanical properties of the periosteum in the pig, Sus scrofa.

The fibrous periosteum forms an intermediary between muscle and ligament forces and the underlying osteoblastic tissue, thus the mechanical properties of the periosteum are critical to understanding osteogenic stimuli. Regional and directional variation in periosteal properties may contribute to the biomechanical regulation of growth in some bones. Periostea of the pig mandibular body, zygomatic arch and metacarpal were loaded to failure under continuous tension. Each tissue type was tested in both the long-axis and transverse orientation. Stiffness, peak stress and peak strain were compared between orientations and among regions. Within the zygomatic periosteum there was little indication of regional difference, and neither zygomatic nor mandibular periosteum showed directional differences. The metacarpal periosteum showed a directional effect only in peak strain, which was greater longitudinally than transversely. There were striking differences, however, among the periostea of the three bones. The zygomatic arch periosteum was the stiffest tissue (91.7+/-30.5 MPa) and showed the highest strength (12.3+/-4.6 MPa). The metacarpal periosteum demonstrated slightly lower stiffness and strength (84.7+/-35.1 and 11.3+/-5.3 MPa), and peak strains in zygomatic and metacarpal periostea were similarly high (17.7+/-3.7 and 17.9+/-3.7 MPa, respectively). The periosteum of the mandibular body was the most deformable tissue (63.0+/-25.4 MPa), with the lowest-peak strain (15.6+/-3.0 MPa) and the least strength (8.2+/-4.1 MPa). These results correspond with those of previous work in long bones, in that periosteum interfacing with ligament or muscle (e.g. zygomatic, metacarpal) demonstrates greater stiffness and strength than periosteum adjacent to loose connective tissue (e.g. mandibular body). Therefore, the degree to which the periosteal tissue serves as a functional interface between bone and muscle is reflected in the different failure properties of periostea from different bones. The structural fortification of the zygomatic arch periosteum relative to other periosteal tissues suggests a role for the periosteum in stabilizing the zygomatic arch-muscle functional complex. On the other hand, the similar failure properties of zygomatic and squamosal periostea from the zygomatic arch mean that the differential growth of these bones cannot be attributed to mechanical stimuli intrinsic to the periosteal tissue.

The fracture behaviour of human and pig molar cusps.

Masticatory efficiency depends upon the ability of the molar cusps to apply concentrated bite forces to food particles and simultaneously to withstand the dental stresses that may cause enamel fracture. This study investigated how low-crowned molar cusps in omnivorous mammals, specifically humans, Homo sapiens, and pigs, Sus scrofa, resist fracture under compressive load. A uniaxial compressive load was applied to individual molar cusps with a materials testing machine. The progressive loading and deformation of the cusps were recorded for interrupted and continuous tests. In interrupted tests, the appearance of progressive cusp fracture was recorded. Stiffness and fracture stresses were calculated from continuous test results. Pig cusps responded to both interrupted and continuous loads with greater deformation; progressive crumbling of the cusp tip resulted in new occlusal contacts on enamel lophs. Conversely, human cusps showed minimal breakage before failure. Continuous compressive tests demonstrated the greater stiffness of human cusps, as well as the capacity to sustain higher cusp tip stresses. The greater stiffness and high fracture resistance of human cusps may be attributed to the thickness of enamel. Test results reflected fundamentally different means of crown stress management that correspond with phylogenetic differences in masticatory function.

Ontogeny of postcanine tooth form in the ferret, Mustela putorius (Carnivora: Mammalia), and the evolution of dental diversity within the Mustelidae.

This study describes dental development within the ferret, Mustela putorius, through study of the form of the carnassial teeth and the upper first molar at progressive growth stages. Primordial teeth were serially sectioned in sagittal and transverse planes and three-dimensional reconstructions of tooth primordia were generated using MacReco software. Regional growth of the crown and asynchronous maturation of the dental tissues were observed in each tooth. The upper carnassial blade develops early and the tooth increases in length rapidly. Lingual growth of the upper carnassial is less pronounced and the protocone and its surrounding region mature late. The lower carnassial blade develops early and the talonid is late to mature. Development of the upper first molar differs from carnassial development in the early emphasis upon transverse growth and reduced lengthwise expansion. The early development of the carnassial blades in the ferret is shared with other carnivores, and may reflect the functional significance of this feature. Later stages of tooth ontogeny differ among carnivoran taxa and the specialized morphology of ferret teeth results from an apparently truncated period of late tooth ontogeny. This suggests that carnivoran species may share a common path of early development that specifies the ontogeny of homologous tooth features and that in later stages developmental differences result in species-specific tooth forms.