Faculty Bibliography

Cells are transplanted to regenerate an organs' parenchyma, but how transplanted parenchymal cells induce stromal regeneration is elusive. Despite the common use of a decellularized matrix, little is known as to the pivotal signals that must be restored for tissue or organ regeneration. We report that Alx3, a developmentally important gene, orchestrated adult parenchymal and stromal regeneration by directly transactivating Wnt3a and vascular endothelial growth factor. In contrast to the modest parenchyma formed by native adult progenitors, Alx3-restored cells in decellularized scaffolds not only produced vascularized stroma that involved vascular endothelial growth factor signalling, but also parenchymal dentin via the Wnt/β-catenin pathway. In an orthotopic large-animal model following parenchyma and stroma ablation, Wnt3a-recruited endogenous cells regenerated neurovascular stroma and differentiated into parenchymal odontoblast-like cells that extended the processes into newly formed dentin with a structure-mechanical equivalency to native dentin. Thus, the Alx3-Wnt3a axis enables postnatal progenitors with a modest innate regenerative capacity to regenerate adult tissues. Depleted signals in the decellularized matrix may be reinstated by a developmentally pivotal gene or corresponding protein.

The face is the most distinctive feature used to identify others. Modern humans have a short, retracted face beneath a large globular braincase that is distinctively different from that of our closest living relatives. The face is a skeletal complex formed by 14 individual bones that houses parts of the digestive, respiratory, visual and olfactory systems. A key to understanding the origin and evolution of the human face is analysis of the faces of extinct taxa in the hominin clade over the last 6 million years. Yet, as new fossils are recovered and the number of hominin species grows, the question of how and when the modern human face originated remains unclear. By examining key features of the facial skeleton, here we evaluate the evolutionary history of the modern human face in the context of its development, morphology and function, and suggest that its appearance is the result of a combination of biomechanical, physiological and social influences.

Store-operated Ca²⁺ entry (SOCE) channels are highly selective Ca²⁺ channels activated by the endoplasmic reticulum (ER) sensors STIM1 and STIM2. Their direct interaction with the pore-forming plasma membrane ORAI proteins (ORAI1, ORAI2, and ORAI3) leads to sustained Ca²⁺ fluxes that are critical for many cellular functions. Mutations in the human ORAI1 gene result in immunodeficiency, anhidrotic ectodermal dysplasia, and enamel defects. In our investigation of the role of ORAI proteins in enamel, we identified enamel defects in a patient with an ORAI1 null mutation. Targeted deletion of the Orai1 gene in mice showed enamel defects and reduced SOCE in isolated enamel cells. However, Orai2^-/- mice showed normal enamel despite having increased SOCE in the enamel cells. Knockdown experiments in the enamel cell line LS8 suggested that ORAI2 and ORAI3 modulated ORAI1 function, with ORAI1 and ORAI2 being the main contributors to SOCE. ORAI1-deficient LS8 cells showed altered mitochondrial respiration with increased oxygen consumption rate and ATP, which was associated with altered redox status and enhanced ER Ca²⁺ uptake, likely due to S-glutathionylation of SERCA pumps. Our findings demonstrate an important role of ORAI1 in Ca²⁺ influx in enamel cells and establish a link between SOCE, mitochondrial function, and redox homeostasis.

Despite their importance for understanding the evolutionary foundations of modern human senescence, available data on aging processes in nonhuman primates from well-studied natural environments are rare, with notable exceptions. We examined patterns of skeletal aging in Virunga mountain gorillas from Volcanoes National Park, Rwanda. Virunga gorillas are distinctive in their herbivory, increased terrestriality, and accelerated life history compared to other great apes. Data were collected from naturally accumulated skeletons [N=50 M, 39 F, 5 Unk], including 53 known-age individuals (0-43 years). Virunga gorillas do not show the dental senescence reported in some other primates. While molar percent dentine exposure significantly increases with age, and occlusal topography metrics significantly decrease with age, 3D relative shearing crest length is maintained, even in heavily worn teeth. Periodontal disease, evidenced by alveolar bone loss, increases significantly with age in molars but not premolars. Degenerative bone changes are also common. Degenerative joint disease (eburnation, osteophytes, porosity) significantly increases with age for all joints (forelimb, hindlimb, vertebrae). However, the strength of correlations differs across body compartments and sexes. Long bone diaphyses show medullary expansion and cortical thinning with age, and periosteal expansion that preserves bone strength despite endosteal bone loss. Fore-tohindlimb strength decreases with age in Virunga gorilla females, possibly associated with behavioral changes, but they lack the rapid bone loss characteristic of post-menopausal human females. Skeletal aging processes in Virunga gorillas and other primates may be influenced by local ecology and behavior, and provide insights into the unique features of human aging

Tooth cementum, the connective tissue anchoring mammalian teeth within their alveolar socket, grows continuously, throughout the individual's life. Studies have observed its biannual increments of contrasting brightness, which have been used to estimate season and age at death in multiple species. Moreover, reproductive events have been inferred from the relative thickness of the bands in two mammalian species. The aim of this project is to test the hypothesis that life history variables affect cementum microstructure in a way that can be discerned via light microcopy and that can be correlated with the age of the individual. Thin sections of donated male and female H. sapiens teeth with known age and life history parameters were prepared for circularly polarized light (CPL) imaging. Digital micrographs were acquired. In all samples a small number of bands with contrasting brightness were observed in CPL. Previous work suggests that these correspond to changes in crystal orientation and/or size. Using FIJI we measured the thickness of each band and estimated the age of the individual corresponding to the time when the bands were formed by establishing proportional relationships with the total thickness of the cementum. We confirmed that the bands correspond to reproductive events and menopause (with an average error of +/- 2 years) in females, and observed that in males they correlate with skeletal maturity and, perhaps, somatic/physiological aging (~50 years of age). This methodology has the potential to inform us on the timing of life-history events in archaeological and paleoanthropological contexts

Bone tissue is an inorganic-organic composite made of hydroxyapatite mineral, collagen fibers, and cement (non-collagenous proteins). Although the mineral component hydroxyapatite is negatively birefringent and represents over 65% of bone weight, undecalcified bone sections imaged by polarization microscopy show a predominant positive birefringence. The analysis of bone mineral thus relies on other methods, such as electron microscopy or imaging methods based on X-rays that have shown the hydroxyapatite phase to consist of nanometric-sized platelets arranged parallel to the collagen fibers. In this study we attempt to visualize the mineral phase using polarization microscopy. Samples from human cortical bone were rendered anorganic by: i) heating the samples at 200degreeC for 24 hours, or ii) treating with (0.5%) NaOCl solution at 50degreeC for 3 days. Samples were then prepared for and observed by polarization microscopy, which demonstrated a birefringence specific for the mineral phase. To better characterize the mineral, we focused on osteonal lamellae of very thin sections (< 10mum-thick) using polarization microscopy configured with several compensators (1/4 -lambda, 3/4- lambda, 1- lambda, 1 1/4 lambda). The high spatial resolution images identified mineral grains of micrometric size (0.43 +/-0.07 mum diameter). As a result of small changes in the relative retardation of the O-and E-rays, these grains were optically stained, exhibiting different interference colors. These grains are interpreted as mineral-packets in which the crystals present the same orientation of their optic axis. They exhibited the original fibrous texture and were arranged in an orderly fashion relating to the lamellar microarchitecture

Linear enamel hypoplasia (LEH) appears as pronounced horizontal grooves on the outer surface of teeth. LEH defects are understood to represent episodes of nonspecific stress in early life, but little is known about their etiology in nonhuman primates. Researchers have suggested that more severe stressors result in deeper LEH defects, while others argue that depth is related to enamel geometry, i.e., larger or smaller striae angles, reflecting differences in growth rates. Here we ask whether inter- and intraspecific variation in enamel growth corresponds with documented differences in defect depth among great apes. Enamel extension rate (EER), or the rate at which teeth grow in height, was assessed using histological methods in canines of four taxa (Gorilla beringei, Gorilla gorilla, Pan troglodytes, Pongo sp., N=16). While sample sizes are too small for statistical comparisons, mountain gorillas have faster EER, and show no overlap with other taxa in the middle 3/5ths of crown height, where LEH defects most commonly occur. This matches our previously reported pattern of defect depth, with mountain gorillas having shallower defects than other taxa in the same crown region. In general, males have faster extension rates than females, but there is some overlap, particularly outside the midcrown. We found that EER is negatively correlated with defect depth (r2=0.66, p<0.001). These results suggest that enamel growth variation influences LEH defect morphology, with faster-growing crowns having shallower defects. However, stress severity may also play an important role in defect formation and help to explain the appearance of particularly deep defects

The aim of this study was to determine the effects of 38% silver diamine fluoride (SDF) on carious lesions of human deciduous teeth. Ten extracted deciduous incisors with caries were collected and treated with SDF. After the treatment, the teeth were sectioned through the center of the carious lesion. The extent of sliver precipitation was examined using quantitative backscattered electron scanning electron microscopy (qBSE-SEM), energy-dispersive X-ray spectroscopy (EDX), and micro-computed tomography (micro-CT). The qBSE-SEM images revealed that the silver particles could penetrate through the pellicle complex, along with the rod sheaths into the demineralized enamel rods and the dentinal tubules, and form silver-enriched barriers surrounding the carious lesions at depths up to 2,490.2 μm (mean 744.7 ± 448.7 μm) within the dentinal tubules of the carious lesions, but less likely in the sound enamel. The EDX spectrum analysis revealed that carbon, oxygen, phosphorus, chlorine, silver, and calcium were the main elements detected in the lesions treated with SDF. Additionally, sodium, magnesium, aluminum, silicon, zinc, sulfur, and fluorine were detected as the minor elements within the SDF precipitation "zone." The micro-CT analysis further showed that in the deep cavitated lesions, the silver precipitation could be observed in the pulp chamber. These findings provide new evidence defining the SDF mode of action for arresting caries and suggest that the application of a highly concentrated SDF solution on deciduous teeth should be used with caution for various carious lesions.

BACKGROUND:As silver diamine fluoride (SDF) gains popularity for caries arrest, the authors aimed to investigate the content of fluoride and silver in 38% SDF produced for the US market and its short-term stability. METHODS:Five samples of 38% SDF were evaluated when the bottle was first opened, and at 7 and 28 days. Fluoride concentrations were determined with a fluoride ion-selective electrode, and silver concentrations were determined with a simultaneous inductively coupled plasma mass spectrometer. pH was measured with a pH probe. Weight and volume of individual drops were measured. RESULTS:At day 0, 40% of individual measured values were above the expected fluoride concentration, and at day 28, 93% were above the expected fluoride concentration (P = .005). At day 0, 19% of individual measured values were below the lowest expected silver concentration, and at day 28, 93% were below (P < .001). Acidity (pH 10) was consistent over the 3 periods. Mean (standard deviation) weight of a drop was 40 (4.0) milligrams, and mean (standard deviation) volume was 32.55 (1.89) microliters, 30% more than the reported value of 25 μL. CONCLUSION/CONCLUSIONS:Over 28 days, the product pH is stable, whereas the fluoride content tends to increase and the silver content tends to decrease. Drops were larger than expected when dispensed from the bottle. PRACTICAL IMPLICATIONS/CONCLUSIONS:Drops are larger than expected, so each delivers higher than expected quantities of silver and fluoride. Clinicians should exercise caution when using this product on young children, replace the cap immediately, and use as soon as dispensed.

Various natural patterns-such as terrestrial sand dune ripples, lamellae in vertebrate bones, growth increments in fish scales and corals, aortas and lamellar corpuscles in humans and animals-comprise layers of different thicknesses and lengths. Microstructures in manmade materials-such as alloys, perlite steels, polymers, ceramics, and ripples induced by laser on the surface of graphen-also exhibit layered structures. These layered patterns form a record of internal and external factors regulating pattern formation in their various systems, making it potentially possible to recognize and identify in their incremental sequences trends, periodicities, and events in the formation history of these systems. The morphology of layered systems plays a vital role in developing new materials and in biomimetic research. The structures and sizes of these two-dimensional (2D) patterns are characteristically anisotropic: That is, the number of layers and their absolute thicknesses vary significantly in different directions. The present work develops a method to quantify the morphological characteristics of 2D layered patterns that accounts for anisotropy in the object of study. To reach this goal, we use Boolean functions and an N-partite graph to formalize layer structure and thickness across a 2D plane and to construct charts of (1) "layer thickness vs. layer number" and (2) "layer area vs. layer number." We present a parameter disorder of layer structure (DStr) to describe the deviation of a study object's anisotropic structure from an isotropic analog and illustrate that charts and DStr could be used as local and global morphological characteristics describing various layered systems such as images of, for example, geological, atmospheric, medical, materials, forensic, plants, and animals. Suggested future experiments could lead to new insights into layered pattern formation.