A late Neanderthal tooth from northeastern Italy.

The site of Riparo Broion (Vicenza, northeastern Italy) preserves a stratigraphic sequence documenting the Middle-to-Upper Paleolithic transition, in particular the final Mousterian and the Uluzzian cultures. In 2018, a human tooth was retrieved from a late Mousterian level, representing the first human remain ever found from this rock shelter (Riparo Broion 1). Here, we provide the morphological description and taxonomic assessment of Riparo Broion 1 with the support of classic and virtual morphology, 2D and 3D analysis of the topography of enamel thickness, and DNA analysis. The tooth is an exfoliated right upper deciduous canine, and its general morphology and enamel thickness distribution support attribution to a Neanderthal child. Correspondingly, the mitochondrial DNA sequence from Riparo Broion 1 falls within the known genetic variation of Late Pleistocene Neanderthals, in accordance with newly obtained radiocarbon dates that point to approximately 48 ka cal BP as the most likely minimum age for this specimen. The present work describes novel and direct evidence of the late Neanderthal occupation in northern Italy that preceded the marked cultural and technological shift documented by the Uluzzian layers in the archaeological sequence at Riparo Broion. Here, we provide a new full morphological, morphometric, and taxonomic analysis of Riparo Broion 1, in addition to generating the wider reference sample of Neanderthal and modern human upper deciduous canines. This research contributes to increasing the sample of fossil remains from Italy, as well as the number of currently available upper deciduous canines, which are presently poorly documented in the scientific literature.

3D fiber deposited polymeric scaffolds for external auditory canal wall.

The external auditory canal (EAC) is an osseocartilaginous structure extending from the auricle to the eardrum, which can be affected by congenital, inflammatory, and neoplastic diseases, thus reconstructive materials are needed. Current biomaterial-based approaches for the surgical reconstruction of EAC posterior wall still suffer from resorption (biological) and extrusion (synthetic). In this study, 3D fiber deposited scaffolds based on poly(ethylene oxide terephthalate)/poly(butylene terephthalate) were designed and fabricated to replace the EAC wall. Fiber diameter and scaffold porosity were optimized, leading to 200 ± 33 µm and 55% ± 5%, respectively. The mechanical properties were evaluated, resulting in a Young's modulus of 25.1 ± 7.0 MPa. Finally, the EAC scaffolds were tested in vitro with osteo-differentiated human mesenchymal stromal cells (hMSCs) with different seeding methods to produce homogeneously colonized replacements of interest for otologic surgery. This study demonstrated the fabrication feasibility of EAC wall scaffolds aimed to match several important requirements for biomaterial application to the ear under the Tissue Engineering paradigm, including shape, porosity, surface area, mechanical properties and favorable in vitro interaction with osteoinduced hMSCs. This study demonstrated the fabrication feasibility of outer ear canal wall scaffolds via additive manufacturing. Aimed to match several important requirements for biomaterial application to ear replacements under the Tissue Engineering paradigm, including shape, porosity and pore size, surface area, mechanical properties and favorable in vitro interaction with osteo-differentiated mesenchymal stromal cells.

3D enamel thickness in Neandertal and modern human permanent canines.

Enamel thickness figures prominently in studies of human evolution, particularly for taxonomy, phylogeny, and paleodietary reconstruction. Attention has focused on molar teeth, through the use of advanced imaging technologies and novel protocols. Despite the important results achieved thus far, further work is needed to investigate all tooth classes. We apply a recent approach developed for anterior teeth to investigate the 3D enamel thickness of Neandertal and modern human (MH) canines. In terms of crown size, the values obtained for both upper and lower unworn/slightly worn canines are significantly greater in Neandertals than in Upper Paleolithic and recent MH. The 3D relative enamel thickness (RET) is significantly lower in Neandertals than in MH. Moreover, differences in 3D RET values between the two groups appear to decrease in worn canines beginning from wear stage 3, suggesting that both the pattern and the stage of wear may have important effects on the 3D RET value. Nevertheless, the 3D average enamel thickness (AET) does not differ between the two groups. In both groups, 3D AET and 3D RET indices are greater in upper canines than in lower canines, and overall the enamel is thicker on the occlusal half of the labial aspect of the crown, particularly in MH. By contrast, the few early modern humans investigated show the highest volumes of enamel while for all other components of 3D enamel, thickness this group holds an intermediate position between Neandertals and recent MH. Overall, our study supports the general findings that Neandertals have relatively thinner enamel than MH (as also observed in molars), indicating that unworn/slightly worn canines can be successfully used to discriminate between the two groups. Further studies, however, are needed to understand whether these differences are functionally related or are the result of pleiotropic or genetic drift effects.

A Neanderthal deciduous human molar with incipient carious infection from the Middle Palaeolithic De Nadale cave, Italy.

OBJECTIVES: The aim of the study is the assessment of Nadale 1, a Neanderthal deciduous tooth recently discovered in Northeastern Italy in the De Nadale cave (Middle Palaeolithic). Together with the clear archaeological context of the site, this study brings new insight on Neanderthal behavior and dental morphological variability. MATERIALS AND METHODS: We used microCT data to provide a morphological description and morphometric analysis (diameter measurements and dental tissue volumes) of the Nadale 1 human tooth. Microwear analysis, taphonomical investigation and caries identification were performed using a stereomicroscope and Scanning Electron Microscope. RESULTS: In terms of morphology (i.e., incipient tuberculum molare, marked mesial marginal ridge and well-developed mid-trigonid crest connecting the protoconid and the metaconid, deep anterior fovea) and size, Nadale 1 presents features frequently observed in Neanderthal lower first deciduous molars. Microscope investigations reveal the presence of a small pit which could be correlated to an incipient caries. CONCLUSION: Nadale 1 expands the Italian Middle Palaeolithic fossil record and provides further information on Neanderthal dm1s in terms of dimensional and morphological variability. Furthermore, the presence of an incipient caries brings further data on Neanderthal diet.

The dawn of dentistry in the late upper Paleolithic: An early case of pathological intervention at Riparo Fredian.

OBJECTIVES: Early evidence for the treatment of dental pathology is found primarily among food-producing societies associated with high levels of oral pathology. However, some Late Pleistocene hunter-gatherers show extensive oral pathology, suggesting that experimentation with therapeutic dental interventions may have greater antiquity. Here, we report the second earliest probable evidence for dentistry in a Late Upper Paleolithic hunter-gatherer recovered from Riparo Fredian (Tuscany, Italy). MATERIALS AND METHODS: The Fredian 5 human consists of an associated maxillary anterior dentition with antemortem exposure of both upper first incisor (I1 ) pulp chambers. The pulp chambers present probable antemortem modifications that warrant in-depth analyses and direct dating. Scanning electron microscopy, microCT and residue analyses were used to investigate the purported modifications of external and internal surfaces of each I1 . RESULTS: The direct date places Fredian 5 between 13,000 and 12,740 calendar years ago. Both pulp chambers were circumferentially enlarged prior to the death of this individual. Occlusal dentine flaking on the margin of the cavities and striations on their internal aspects suggest anthropic manipulation. Residue analyses revealed a conglomerate of bitumen, vegetal fibers, and probable hairs adherent to the internal walls of the cavities. DISCUSSION: The results are consistent with tool-assisted manipulation to remove necrotic or infected pulp in vivo and the subsequent use of a composite, organic filling. Fredian 5 confirms the practice of dentistry-specifically, a pathology-induced intervention-among Late Pleistocene hunter-gatherers. As such, it appears that fundamental perceptions of biomedical knowledge and practice were in place long before the socioeconomic changes associated with the transition to food production in the Neolithic.

Middle paleolithic human deciduous incisor from Grotta del Cavallo, Italy.

OBJECTIVES: In this contribution, we present a morphological description and comparative morphometric analysis of Cavallo D, a human tooth unearthed from the Mousterian FIII sublayer of Grotta del Cavallo (Apulia, Italy). MATERIALS AND METHODS: We used microCT data to provide a detailed morphological description and morphometric analysis of the Cavallo D human tooth based on traditional diameter measurements and 3D enamel thickness. Moreover, new AMS radiocarbon dating of charcoals from layers FII was carried out. RESULTS: Morphological features observed in Cavallo D align the tooth to Neandertals. Similarly, the large size of the tooth (e.g., BL diameter) and the relatively thinner enamel thickness are typical Neandertal traits. 14 C datings of layer FII attribute the tooth to a time range of 45,600-42,900 cal BP (at 68% level of probability). DISCUSSION: Up to now, the Rdi1 Cavallo D represents the most recent Neandertal human remain in southern Italy related to a radiocarbon dated stratigraphy. Moreover, since deciduous teeth have been less investigated than the permanent ones, this contribution brings new data to increase our knowledge on the variability of the Neandertal deciduous dentition.

Piscivory in a Miocene Cetotheriidae of Peru: first record of fossilized stomach content for an extinct baleen-bearing whale.

Instead of teeth, modern mysticetes bear hair-fringed keratinous baleen plates that permit various bulk-filtering predation techniques (from subsurface skimming to lateral benthic suction and engulfment) devoted to various target prey (from small invertebrates to schooling fish). Current knowledge about the feeding ecology of extant cetaceans is revealed by stomach content analyses and observations of behavior. Unfortunately, no fossil stomach contents of ancient mysticetes have been described so far; the investigation of the diet of fossil baleen whales, including the Neogene family Cetotheriidae, remains thus largely speculative. We report on an aggregate of fossil fish remains found within a mysticete skeleton belonging to an undescribed late Miocene (Tortonian) cetotheriid from the Pisco Formation (Peru). Micro-computed tomography allowed us to interpret it as the fossilized content of the forestomach of the host whale and to identify the prey as belonging to the extant clupeiform genus Sardinops. Our discovery represents the first direct evidence of piscivory in an ancient edentulous mysticete. Since among modern mysticetes only Balaenopteridae are known to ordinarily consume fish, this fossil record may indicate that part of the cetotheriids experimented some degree of balaenopterid-like engulfment feeding. Moreover, this report corresponds to one of the geologically oldest records of Sardinops worldwide, occurring near the Tortonian peak of oceanic primary productivity and cooling phase. Therefore, our discovery evokes a link between the rise of Cetotheriidae; the setup of modern coastal upwelling systems; and the radiation of epipelagic, small-sized, schooling clupeiform fish in such highly productive environments.

Technical Note: Guidelines for the digital computation of 2D and 3D enamel thickness in hominoid teeth.

The study of enamel thickness has received considerable attention in regard to the taxonomic, phylogenetic and dietary assessment of human and non-human primates. Recent developments based on two-dimensional (2D) and three-dimensional (3D) digital techniques have facilitated accurate analyses, preserving the original object from invasive procedures. Various digital protocols have been proposed. These include several procedures based on manual handling of the virtual models and technical shortcomings, which prevent other scholars from confidently reproducing the entire digital protocol. There is a compelling need for standard, reproducible, and well-tailored protocols for the digital analysis of 2D and 3D dental enamel thickness. In this contribution we provide essential guidelines for the digital computation of 2D and 3D enamel thickness in hominoid molars, premolars, canines and incisors. We modify previous techniques suggested for 2D analysis and we develop a new approach for 3D analysis that can also be applied to premolars and anterior teeth. For each tooth class, the cervical line should be considered as the fundamental morphological feature both to isolate the crown from the root (for 3D analysis) and to define the direction of the cross-sections (for 2D analysis).

A historical case of amelogenesis imperfecta: Giovanna of Austria, Grand Duchess of Tuscany (1547-1578).

The skeletal remains of Giovanna of Austria (1547-1578), daughter of the Emperor Ferdinand I of Habsburg (1503-1564) and first wife of the Grand Duke of Tuscany, Francesco I (1541-1587), exhumed from the Basilica of San Lorenzo in Florence, were submitted to paleopathological study. Examination of the dentition, which was in a good state of preservation, showed maxillary retrognathism, together with a caries lesion, moderate periodontal disease, malposition of the upper second premolars and tooth wear. Furthermore, several horizontal grooves were observed in both the buccal and the lingual crown surfaces of almost all teeth, especially the anterior ones. The orthopantomogram showed hypomineralized enamel and alveolar bone loss. Two third-molar teeth were investigated using micro-computed tomography (micro-CT) analysis, revealing highly irregular enamel caps with reduced average thickness. The observed features suggest a diagnosis of hypoplastic amelogenesis imperfecta, a developmental condition affecting enamel formation.

Alveolar Bone Remodeling with or without Collagen Filling of the Extraction Socket: A High-Resolution X-ray Tomography Animal Study.

The healing process of the tooth extraction socket often leads to significant resorption of the alveolar bone, eventually causing clinical difficulties for future implant-supported rehabilitations. The aim of the present animal study was to evaluate alveolar bone remodeling after tooth extraction in a rabbit model, either with or without the use of a plain collagen plug inside the socket, by means of micro-computed tomography. The study included the micro-tomography analysis of 36 rabbits' incisor extraction sockets, either left empty or filled with a collagen plug. All animals were euthanized in a staggered manner, in order to address molecular, histologic, and radiographic analyses at different time-points, up to 90 days after surgery. The three-dimensional evaluation was carried out using micro-computed tomography technology on excised bone blocks including the alveolus and the contralateral bone. Both linear and volumetric measures were recorded: the percentage of bone volume change (ΔBV) within the region of interest was considered the primary endpoint of the study. The micro-CT analysis revealed mean volumetric changes of -58.1% ± from baseline to 3 months for the control group, and almost no bone loss for the test group, -4.6%. The sockets treated with the collagen plug showed significantly less dimensional resorption, while the natural-healing group showed an evident collapse of the alveolar bone three months after extraction surgery.

Direct evidence that late Neanderthal occupation precedes a technological shift in southwestern Italy.

OBJECTIVES: During the middle-to-upper Paleolithic transition (50,000 and 40,000 years ago), interaction between Neanderthals and Homo sapiens varied across Europe. In southern Italy, the association between Homo sapiens fossils and non-Mousterian material culture, as well as the mode and tempo of Neanderthal demise, are still vividly debated. In this research, we focus on the study of two human teeth by using 3D geometric morphometric approaches for a reliable taxonomical attribution as well as obtaining new radiometric dates on the archeological sequence. MATERIAL AND METHODS: This work presents two lower deciduous molars uncovered at Roccia San Sebastiano (Mondragone-Caserta, Italy), stratigraphically associated with Mousterian (RSS1) and Uluzzian (RSS2) artifacts. To obtain a probabilistic attribution of the two RSS teeth to each reference taxa group composed of Neanderthals and Homo sapiens, we performed and compared the performance of three supervised learning algorithms (flexible discriminant analysis, multiadaptive regression splines, and random forest) on both crown and cervical outlines obtained by virtual morphometric methods. RESULTS: We show that RSS1, whose Mousterian context appears more recent than 44,800-44,230 cal BP, can be attributed to a Neanderthal, while RSS2, found in an Uluzzian context that we dated to 42,640-42,380 cal BP, is attributed to Homo sapiens. DISCUSSION: This site yields the most recent direct evidence for a Neanderthal presence in southern Italy and confirms a later shift to upper Paleolithic technology in southwestern Italy compared to the earliest Uluzzian evidence at Grotta del Cavallo (Puglia, Italy).

Human deciduous teeth from the Middle Stone Age layers of Sibudu Cave (South Africa).

In the African Pleistocene, the fossil evidence for early Homo sapiens populations is still relatively limited. Here we present two additional specimens (two deciduous teeth) recovered from the Middle Stone Age (MSA) deposits of Sibudu Cave (KwaZulu-Natal, South Africa). We describe their morphology and metrics, using three-dimensional models of the teeth obtained from high-resolution micro-CT images. The first specimen is a Ldm1 (HUM. TO 1) recovered in the BS5 layer dated 77.3±2.7 ka, and associated with stone tools assigned to the "pre-Still Bay" assemblage. The other specimen is a Rdi1 (HUM. TO 2) coming from the Pinkish Grey Sand (PGS) layer, dated 64.7±2.3 ka, and associated with a Howieson's Poort industry. Both teeth are well preserved, with minor post mortem cracks not affecting the overall morphology, and they comprise the intact, worn crown and the remnants of the roots, naturally resorbed. A large carious lesion occupies most of the distal face and part of the occlusal surface in the Ldm1; also a chip of enamel is missing from the disto-buccal corner. For both teeth, we compared mesio-distal (MD) and bucco-lingual (BL) diameters with those of other Late Pleistocene deciduous teeth and extant Homo sapiens. The analysis has shown that the teeth are comparable in size with the other MSA specimens described in the literature.

Grotta del Cavallo (Apulia-Southern Italy). The Uluzzian in the mirror.

The Uluzzian techno-complex is commonly considered to be a "transitional industry" mostly on the basis of some inferred characteristics such as a chiefly flake-based production, a small amount of Upper Palaeolithic-like tools and a combination of Middle and Upper Palaeolithic elements both in the toolkit and in the technical systems. Following its discovery, the Uluzzian was identified as the Italian counterpart of the French Châtelperronian and attributed to Neandertals. However, a study issued in 2011 has established the modern character of the two deciduous teeth found in 1964 in the Uluzzian deposit of Grotta del Cavallo, fostering renewed interests to the Uluzzian culture, which real nature is almost unknown to the international scientific community. Here we provide preliminary results of the study on the lithic assemblage from the earliest Uluzzian layer and on backed pieces from the whole Uluzzian sequence of Grotta del Cavallo (Apulia, Italy), the type site of the Uluzzian. Moreover, besides a thorough review on the stratigraphy of Grotta del Cavallo (Supplementary Materials), we provide updated information on the human remains by presenting two unpublished teeth from the reworked deposit of the same cave. We conclude that the early Uluzzians demonstrate original technological behavior and innovations devoid of any features deriving or directly linked with the late Mousterian of Southern Italy. Therefore, the novelty nature of the Uluzzian techno-complex (with respect to the preceding Mousterian) complies with the recent reassessment of the two deciduous teeth from Grotta del Cavallo in suggesting an earliest migration of modern humans in southern Europe around 45,000 years ago.

A Dental Prosthesis from the Early Modern Age in Tuscany (Italy).

BACKGROUND: During archaeological excavation, carried out in the S. Francesco Monastery at Lucca (Tuscany, Italy), a golden dental appliance was discovered. The prosthesis was found, together with commingled human remains, in the collective tomb of the aristocratic family of the Guinigi, a powerful family who governed Lucca from 1392 until 1429. The exact archaeological dating of the prosthesis was not possible, but some elements suggest a dating to the beginning of the 17th century. PURPOSE: Aim of the paper is to study and describe the dental appliance trough a multidisciplinary approach. MATERIALS AND METHODS: Macroscopical and micro-CT examinations were performed to investigate the technics used for the realization of the dental prosthesis. SEM analysis was performed to study alloy composition of the metallic fixing lamina and microstructure of the deposits on the dental surface. RESULTS: The dental prosthesis consists in five mandibular teeth: three central incisors and two lateral canines linked together by a golden band inserted into the dental roots to replace the anterior arch of the jaw. Micro-CT scan revealed the presence of two small golden pins inserted into each tooth crossing the root and fixing the teeth to the internal gold band. SEM examination of the lamina indicated a homogeneous composition, with average contents of 73 wt% gold, 15.6 wt% Ag, and 11.4 wt% Cu. Apposition of dental calculus on the teeth indicated that the prosthesis had been worn for a long period. CONCLUSIONS: This dental prosthesis provides a unique finding of technologically advanced dentistry in this period. In fact, during the Early Modern Age, some authors described gold band technology for the replacement of missing teeth; nevertheless, no direct evidences of these devices have been brought to light up so far.

Evaluation of the effect of a gamma irradiated DBM-pluronic F127 composite on bone regeneration in Wistar rat.

Demineralized bone matrix (DBM) is widely used for bone regeneration. Since DBM is prepared in powder form its handling properties are not optimal and limit the clinical use of this material. Various synthetic and biological carriers have been used to enhance the DBM handling. In this study we evaluated the effect of gamma irradiation on the physical-chemical properties of Pluronic and on bone morphogenetic proteins (BMPs) amount in DBM samples. In vivo studies were carried out to investigate the effect on bone regeneration of a gamma irradiated DBM-Pluronic F127 (DBM-PF127) composite implanted in the femur of rats. Gamma irradiation effects (25 kGy) on physical-chemical properties of Pluronic F127 were investigated by rheological and infrared analysis. The BMP-2/BMP-7 amount after DBM irradiation was evaluated by ELISA. Bone regeneration capacity of DBM-PF127 containing 40% (w/w) of DBM was investigated in transcortical holes created in the femoral diaphysis of Wistar rat. Bone porosity, repaired bone volume and tissue organization were evaluated at 15, 30 and 90 days by Micro-CT and histological analysis. The results showed that gamma irradiation did not induce significant modification on physical-chemical properties of Pluronic, while a decrease in BMP-2/BMP-7 amount was evidenced in sterilized DBM. Micro-CT and histological evaluation at day 15 post-implantation revealed an interconnected trabeculae network in medullar cavity and cellular infiltration and vascularization of DBM-PF127 residue. In contrast a large rate of not connected trabeculae was observed in Pluronic filled and unfilled defects. At 30 and 90 days the DBM-PF127 samples shown comparable results in term of density and thickness of the new formed tissue respect to unfilled defect. In conclusion a gamma irradiated DBM-PF127 composite, although it may have undergone a significant decrease in the concentration of BMPs, was able to maintains bone regeneration capability.