Ancient vertebrate dermal armor evolved from trunk neural crest.

Bone is an evolutionary novelty of vertebrates, likely to have first emerged as part of ancestral dermal armor that consisted of osteogenic and odontogenic components. Whether these early vertebrate structures arose from mesoderm or neural crest cells has been a matter of considerable debate. To examine the developmental origin of the bony part of the dermal armor, we have performed in vivo lineage tracing in the sterlet sturgeon, a representative of nonteleost ray-finned fish that has retained an extensive postcranial dermal skeleton. The results definitively show that sterlet trunk neural crest cells give rise to osteoblasts of the scutes. Transcriptional profiling further reveals neural crest gene signature in sterlet scutes as well as bichir scales. Finally, histological and microCT analyses of ray-finned fish dermal armor show that their scales and scutes are formed by bone, dentin, and hypermineralized covering tissues, in various combinations, that resemble those of the first armored vertebrates. Taken together, our results support a primitive skeletogenic role for the neural crest along the entire body axis, that was later progressively restricted to the cranial region during vertebrate evolution. Thus, the neural crest was a crucial evolutionary innovation driving the origin and diversification of dermal armor along the entire body axis.

The stem osteichthyan Andreolepis and the origin of tooth replacement.

The teeth of gnathostomes (jawed vertebrates) show rigidly patterned, unidirectional replacement that may or may not be associated with a shedding mechanism. These mechanisms, which are critical for the maintenance of the dentition, are incongruently distributed among extant gnathostomes. Although a permanent tooth-generating dental lamina is present in all chondrichthyans, many tetrapods and some teleosts, it is absent in the non-teleost actinopterygians. Tooth-shedding by basal hard tissue resorption occurs in most osteichthyans (including tetrapods) but not in chondrichthyans. Here we report a three-dimensional virtual dissection of the dentition of a 424-million-year-old stem osteichthyan, Andreolepis hedei, using propagation phase-contrast synchrotron microtomography, with a reconstruction of its growth history. Andreolepis, close to the common ancestor of all extant osteichthyans, shed its teeth by basal resorption but probably lacked a permanent dental lamina. This is the earliest documented instance of resorptive tooth shedding and may represent the primitive osteichthyan mode of tooth replacement.

Proteasome inhibitor MG132 enhances the sensitivity of human OSCC cells to cisplatin via a ROS/DNA damage/p53 axis.

Cis-diamine-dichloroplatinum II (cisplatin, CDDP) is a key chemotherapeutic regimen in the treatment of oral squamous cell carcinoma (OSCC). However, the therapeutic efficacy of cisplatin in OSCC may be hampered by chemoresistance. Therefore, the development of novel combination therapy strategies to overcome the limitations of CDDP is of great importance. The proteasome inhibitor MG132 exhibits anti-cancer properties against various types of cancer. However, our knowledge of its anti-cancer effects in combination with CDDP in OSCC cells remains limited. In the current study, the synergetic effects of MG132 and CDDP were evaluated in the human CAL27 OSCC cell line. CAL27 cells were treated with CDDP alone or in combination with MG132. The results showed that MG132 significantly reduced cell viability in a dose-dependent manner. Additionally, cell viability was significantly reduced in CAL27 cells treated with 0.2 µM MG132 and 2 µM CDDP compared with cells treated with MG132 or CDDP alone. In addition, MG132 significantly enhanced the CDDP-induced generation of intracellular reactive oxygen species and DNA damage in OSCC cells. Furthermore, treatment with CDDP or MG132 alone notably inhibited colony formation and proliferation of OSCC cells. However, co-treatment of OSCC cells with MG132 and CDDP further hampered colony formation and proliferation compared with cells treated with either MG132 or CDDP alone. Finally, in cells co-treated with MG132 and CDDP, the expression of p53 was markedly elevated and the p53-mediated apoptotic pathway was further activated compared with cells treated with MG132 or CDDP alone, as shown by the enhanced cell apoptosis, Bax upregulation, and Bcl-2 downregulation. Overall, the results of the current study support the synergistic anti-cancer effects of a combination of MG132 and CDDP against OSCC, thus suggesting that the combination of MG132 and CDDP may be a promising therapeutic strategy for the management of OSCC.

A Core-Shell MWCNT-Pt Nanowire Electron Source with Anomalously Long-Term Stable Field Emission.

A hybrid core-shell structured nanowire is proposed for a long-term stable electron source based on an isolated platinum/multi-walled carbon nanotube (Pt/MWCNT). This hybrid nanowire is prepared by growing a Pt shell on a metallic MWCNT through a field-emission-induced deposition (FEID) method. An in situ field emission (FE) platform was constructed inside a scanning electron microscope (SEM) equipped with two nanorobotic manipulators (NRMs) for the preparation and testing of the hybrid nanowire. An in situ fatigue test was conducted with high current intensity (500 nA) to show the influence of the Pt shell. Compared with the pristine bare MWCNT, our hybrid-nanowire-based electron source has a lifetime of hundreds of times longer and can work continuously for up to 48 h under relatively high pressure (3.6×10-3 Pa) without having an apparent change in its structure or emission currents, demonstrating good stability and tolerance to poor working conditions. The anomalous long-term stability is attributed mainly to the shielding of oxygen by Pt from the carbon shells and less heating due to the work function lowering by Pt.

In Situ Device-Level TEM Characterization Based on Ultra-Flexible Multilayer MoS2 Micro-Cantilever.

Current state-of-the-art in situ transmission electron microscopy (TEM) characterization technology has been capable of statically or dynamically nanorobotic manipulating specimens, affording abundant atom-level material attributes. However, an insurmountable barrier between material attributes investigations and device-level application explorations exists due to immature in situ TEM manufacturing technology and sufficient external coupled stimulus. These limitations seriously prevent the development of in situ device-level TEM characterization. Herein, a representative in situ opto-electromechanical TEM characterization platform is put forward by integrating an ultra-flexible micro-cantilever chip with optical, mechanical, and electrical coupling fields for the first time. On this platform, static and dynamic in situ device-level TEM characterizations are implemented by utilizing molybdenum disulfide (MoS2 ) nanoflake as channel material. E-beam modulation behavior in MoS2  transistors is demonstrated at ultra-high e-beam acceleration voltage (300 kV), stemming from inelastic scattering electron doping into MoS2  nanoflakes. Moreover, in situ dynamic bending MoS2  nanodevices without/with laser irradiation reveals asymmetric piezoresistive properties based on electromechanical effects and secondary enhanced photocurrent based on opto-electromechanical coupling effects, accompanied by real-time monitoring atom-level characterization. This approach provides a step toward advanced in situ device-level TEM characterization technology with excellent perception ability and inspires in situ TEM characterization with ultra-sensitive force feedback and light sensing.

The developmental relationship between teeth and dermal odontodes in the most primitive bony fish Lophosteus.

The ontogenetic trajectory of a marginal jawbone of Lophosteus superbus (Late Silurian, 422 Million years old), the phylogenetically most basal stem osteichthyan, visualized by synchrotron microtomography, reveals a developmental relationship between teeth and dermal odontodes that is not evident from the adult morphology. The earliest odontodes are two longitudinal founder ridges formed at the ossification center. Subsequent odontodes that are added lingually to the ridges turn into conical teeth and undergo cyclic replacement, while those added labially achieve a stellate appearance. Stellate odontodes deposited directly on the bony plate are aligned with the alternate files of teeth, whereas new tooth positions are inserted into the files of sequential addition when a gap appears. Successive teeth and overgrowing odontodes show hybrid morphologies around the oral-dermal boundary, suggesting signal cross-communication. We propose that teeth and dermal odontodes are modifications of a single system, regulated and differentiated by the oral and dermal epithelia.

Human teeth are an example of odontodes: hard structures made of a material called dentine that are sometimes coated in enamel. Teeth are the only odontodes humans have, but other vertebrates (animals with backbones) have tooth-like scales on their skin. These structures are called dermal odontodes, and sharks and rays, for example, are covered with them. How these structures evolved, and whether teeth or dermal odontodes developed first, continues to spark great discussion among palaeontologists. Some researchers think that teeth evolved from dermal odontodes, a theory known as the 'scales-to-teeth' hypothesis. Others think dermal odontodes are distinct from teeth because they lack the same spatial organization. To investigate this further, palaeontologists are looking at the earliest examples of odontodes they can find: fossils of early vertebrates that carry both teeth and dermal odontodes. Here, Chen et al. have studied Lophosteus, one of the earliest bony fishes that lived more than 400 million years ago, to explore early tooth evolution and growth patterns. Chen et al. digitally dissected a fossilized Lophosteus jawbone using submicron X-ray imaging, a technique with resolution to less than one millionth of a metre. Imaging thin sections of the specimen, found in Estonia, Chen et al. reconstructed the entire sequence of odontode development in the bony fish in 3D. The analysis showed that teeth and dermal odontodes initially take shape together but differentiate as they grow, presumably instructed to do so by various developmental signals. However, at a later stage, the two types of odontodes become similar in appearance again, suggesting that they respond to each other's signals. For example, as the jawbone grows, dermal odontodes overgrow the earliest formed teeth. These younger odontodes resemble teeth, while the new teeth developing near the dermal odontodes take after dermal odontodes. These findings suggest that teeth and dermal odontodes are not wholly separate systems but, instead, are closely related on a molecular level. The results also show that contrary to the 'scale-to-teeth' hypothesis, teeth do not evolve from fully formed dermal odontodes, rather the two types of odontodes form out of one founder. This research builds on our knowledge from modern sharks and points to a previously unrecognised evolutionary relationship between teeth and dermal odontodes. It also furthers our understanding of how molecular regulation controls development.

Marginal dentition and multiple dermal jawbones as the ancestral condition of jawed vertebrates.

The dentitions of extant fishes and land vertebrates vary in both pattern and type of tooth replacement. It has been argued that the common ancestral condition likely resembles the nonmarginal, radially arranged tooth files of arthrodires, an early group of armoured fishes. We used synchrotron microtomography to describe the fossil dentitions of so-called acanthothoracids, the most phylogenetically basal jawed vertebrates with teeth, belonging to the genera Radotina, Kosoraspis, and Tlamaspis (from the Early Devonian of the Czech Republic). Their dentitions differ fundamentally from those of arthrodires; they are marginal, carried by a cheekbone or a series of short dermal bones along the jaw edges, and teeth are added lingually as is the case in many chondrichthyans (cartilaginous fishes) and osteichthyans (bony fishes and tetrapods). We propose these characteristics as ancestral for all jawed vertebrates.

Periostin promotes migration and osteogenic differentiation of human periodontal ligament mesenchymal stem cells via the Jun amino-terminal kinases (JNK) pathway under inflammatory conditions.

OBJECTIVES: Mesenchymal stem cell (MSC)-mediated periodontal tissue regeneration is considered to be a promising method for periodontitis treatment. The molecular mechanism of functional regulation by MSCs remains unclear, thus limiting their application. Our previous study discovered that Periostin (POSTN) promoted the migration and osteogenic differentiation of periodontal ligament mesenchymal stem cells (PDLSCs), but it is still unclear whether POSTN is able to restore the regenerative potential of PDLSCs under inflammatory conditions. In this study, we investigated the effect of POSTN on PDLSCs under inflammatory conditions and its mechanism. MATERIALS AND METHODS: PDLSCs were isolated from periodontal ligament tissue. TNF-α was used at 10 ng/mL to mimic inflammatory conditions. Lentivirus POSTN shRNA was used to knock down POSTN. Recombinant human POSTN (rhPOSTN) was used to stimulate PDLSCs. A scratch assay was used to analyse cell migration. Alkaline phosphatase (ALP) activity, Alizarin Red staining and expression of osteogenesis-related genes were used to investigate the osteogenic differentiation potential. Western blot analysis was used to detect the mitogen-activated protein kinases (MAPK) and AKT signalling pathways. RESULTS: After a 10 ng/mL TNF-α treatment, knockdown of POSTN impeded scratch closure, inhibited ALP activity and mineralization in vitro, and decreased expression of RUNX2, OSX, OPN and OCN in PDLSCs, while 75 ng/mL rhPOSTN significantly accelerated scratch closure, enhanced ALP activity and mineralization in vitro, and increased expression of RUNX2, OSX, OPN and OCN. In addition, knockdown of POSTN inhibited expression of phosphorylated c-Jun N-terminal kinase (p-JNK), while 75 ng/mL rhPOSTN increased expression of p-JNK in PDLSCs with TNF-α treatment. Furthermore, inhibition of JNK by its inhibitor SP600125 dramatically blocked POSTN-enhanced scratch closure, ALP activity and mineralization in PDLSCs. CONCLUSIONS: Our results revealed that POSTN might promote the migration and osteogenic differentiation potential of PDLSCs via the JNK pathway, providing insight into the mechanism underlying MSC biology under inflammatory conditions and identifying a potential target for improving periodontal tissue regeneration.

Electron Beam Irradiation Induced Multiwalled Carbon Nanotubes Fusion inside SEM.

This paper reported a method of multiwalled carbon nanotubes (MWCNTs) fusion inside a scanning electron microscope (SEM). A CNT was picked up by nanorobotics manipulator system which was constructed in SEM with 21 DOFs and 1 nm resolution. The CNT was picked up and placed on two manipulators. The tensile force was 140 nN when the CNT was pulled into two parts. Then, two parts of the CNT were connected to each other by two manipulators. The adhered force between two parts was measured to be about 20 nN. When the two parts of CNT were connected again, the contact area was fused by focused electron beam irradiation for 3 minutes. The tensile force of the junction was measured to be about 100 nN. However, after fusion, the tensile force was five times larger than the tensile force connected only by van der Waals force. This force was 70 percent of the tensile force before pulling out of CNTs. The results revealed that the electron beam irradiation was a promising method for CNT fusion. We hope this technology will be applied to nanoelectronics in the near future.

[A clinical analysis of screw-retained implant-supported casted integrated abutment crowns in the molar region with limited interocclusal space].

PURPOSE: To observe the clinical effects of screw-retained implant-supported casted abutment integrated crowns (IACs) in the molar region with limited interocclusal space. METHODS: This study involved 507 implants in 376 patients with limited interocclusal space in the molar region. All implants were inserted to the jaw by standard one-stage protocol. Screw-retained IACs were used as the final prosthesis. With 6 to 24 months follow-up, the clinical effects of screw-retained IACs were recorded and analyzed using SPSS 17.0 software package. RESULTS: During 6 to 24 months of follow-up, the implant survival rate was 99.61%. 37 patients had ceramic fracture which mainly happened in the group with 3-4 mm interocclusal space. In each group, porcelain fused to metal (PFM) prosthesis had ceramic fracture more easily than PFM prosthesis without porcelain on occlusal surface(P<0.05). As the interocclusal space became smaller, the probability of collapsing porcelain increased(P<0.05).13 patients had screw loosening. 13 patients suffered from gingival swelling and bleeding, they were given periodontal treatment and oral hygiene instruction. Patients were satisfied with the restoration results. CONCLUSIONS: Screw-retained IACs can be used to repair missing teeth in the molar region with limited interocclusal space.

[Effect of miR-21 in promoting angiogenesis differentiation of umbilical cord blood mesenchymal stem cells in vitro].

PURPOSE: To investigate the effect of miR-21 on promoting vascular differentiation of umbilical cord blood derived mesenchymal stem cells (UCBMSCs) in vitro. METHODS: UCBMSCs (from Stem Cell Bank of Tongji University) were cultured; Lenti-LacZ-Luciferase and Lenti-miR-21-Luciferase Lentiviral vector were reconstructed; Real-time fluorescence quantification PCR and Western blot were performed to detect the expression of key angiogenic factors at 0, 1, 4, 7 and 14 d after target gene was transduced into UCBMSCs; UCBMSCs of the third generation were transduced with Lenti-LacZ-Luciferase and Lenti-miR-21- Luciferase; matrigel tube assay was performed to observe the tube-like structure under microscope. The data were statistically analyzed using Student's t test and variance analysis with SPSS13.0 software package. RESULTS: UCBMSCs were successfully cultured; The results of qPCR and Western blot showed the expression of HIF-1α and VEGF in miR-21 group significantly increased at 4 d and continued until 7 d and 14 d; Matrigel results showed the tubular structures in Lenti-miR-21-Luciferase group were more than other groups both in amount and in length. CONCLUSIONS: MiR-21 can promote vascular differentiation of UCBMSCs.

Development of cyclic shedding teeth from semi-shedding teeth: the inner dental arcade of the stem osteichthyan Lophosteus.

The numerous cushion-shaped tooth-bearing plates attributed to the stem group osteichthyan Lophosteus superbus, which are argued here to represent an early form of the osteichthyan inner dental arcade, display a previously unknown and presumably primitive mode of tooth shedding by basal hard tissue resorption. They carry regularly spaced, recumbent, gently recurved teeth arranged in transverse tooth files that diverge towards the lingual margin of the cushion. Three-dimensional reconstruction from propagation phase-contrast synchrotron microtomography (PPC-SRµCT) reveals remnants of the first-generation teeth embedded in the basal plate, a feature never previously observed in any taxon. These teeth were shed by semi-basal resorption with the periphery of their bases retained as dentine rings. The rings are highly overlapped, which evidences tooth shedding prior to adding the next first-generation tooth at the growing edge of the plate. The first generation of teeth is thus diachronous. Successor teeth at the same sites underwent cyclical replacing and shedding through basal resorption, producing stacks of buried resorption surfaces separated by bone of attachment. The number and spatial arrangement of resorption surfaces elucidates that basal resorption of replacement teeth had taken place at the older tooth sites before the addition of the youngest first-generation teeth at the lingual margin. Thus, the replacement tooth buds cannot have been generated by a single permanent dental lamina at the lingual edge of the tooth cushion, but must have arisen either from successional dental laminae associated with the individual predecessor teeth, or directly from the dental epithelium of these teeth. The virtual histological dissection of these Late Silurian microfossils broadens our understanding of the development of the gnathostome dental systems and the acquisition of the osteichthyan-type of tooth replacement.

[Application of one-piece implant-supported detachable telescope retained fixed bridge in edentulous cases].

PURPOSE: To evaluate the clinical effectiveness of one-piece implant-supported detachable telescopic fixed bridge in edentulous patients. METHODS: Seventeen patients were treated with one-piece implant-supported detachable telescopic fixed bridge. A total of 18 prostheses were fabricated with 8 in the upper jaws and 10 in the lower jaws.Fixed bridges retained by telescopic crowns were used as final prostheses, with milling titanium or all-ceramic abutments as primary crowns, gold-electroforming crowns as secondary crowns. Surveys about clinical and radiographic examination, satisfaction and prosthetic complications were conducted after 3 months,1 year, 2 years, 3 years after final rehabilitation. Data analysis was performed using SPSS 17.0 software package. RESULTS: Radiography showed stable bone levels for all implants except 2 implants, which were observed slight marginal bone resorption. The results of one-way ANOVA showed that no significant difference in modified plaque index or modified sulcus blooding index was found during the follow-up period (P>0.05). The probing attachment level deteriorated by 1.5 mm during the first 3 years (P<0.05). Eighteen restoration provided sufficient fixation and stability. Two porcelain fractures occured but had no influence on restoration. The patients were highly satisfied with the outcomes. The frequency of prosthetic maintenance per patient per year was 0.11. CONCLUSIONS: One-piece implant-supported detachable telescopic fixed bridge is an effective method with satisfactory long-term aesthetic and stable outcomes in edentulous patients.

Pyrene exposure influences the craniofacial cartilage development of Sebastiscus marmoratus embryos.

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental contaminants, which are known carcinogens and teratogens. However, the toxicity of PAHs during skeletal development and the mechanism involved are not completely clear. In the present study, rockfish (Sebastiscus marmoratus) embryos were exposed to pyrene (Pyr) for 7 days at 0.5, 5 and 50 nM which resulted in craniofacial skeleton deformities. Pyr exposure for 6 days reduced the expression of PCNA, Col2a1 and Sox9 in the craniofacial skeleton revealed using in situ hybridization. These results suggest that Pyr exposure impairs skeleton development via disrupting the proliferation of the chondrocytes. At the same time, Pyr exposure reduced the expression of lox1 and inhibited the activity of lysyl oxidase, which is the key enzyme controlling the collagen cross-linking, and which might therefore have been one of the reasons for the deformative Meckel's cartilage (lower jaw).

Effects of benzo(a)pyrene on the skeletal development of Sebastiscus marmoratus embryos and the molecular mechanism involved.

Polycyclic aromatic hydrocarbons (PAHs) are widespread environmental contaminants, which have been known to be carcinogenic and teratogenic. However, the skeletal development toxicity of PAHs and the mechanism involved remain unclear. In fishes, the neurocranial and craniofacial skeleton develop as cartilage. The signaling molecules of hedgehog (Hh) family play crucial roles in regulating skeletal development. In the present study, rockfish (Sebastiscus marmoratus) embryos were exposed to benzo(a)pyrene (BaP) for 7 days at environmental levels (0.05, 0.5 and 5nmol/L) which resulted in craniofacial skeleton deformities. BaP exposure reduced the cell proliferation activity in the craniofacial skeleton as detected by quantitative PCR and in situ hybridization. The expression of Sonic hedgehog (Shh), rather than Indian hedgehog (Ihh), was down-regulated in the craniofacial skeleton in the 0.5 and 5nmol/L groups. Consistent with the Shh results, the expression of Ptch1 and Gli2 was decreased by BaP exposure and BMP4 was presented on changes in the 0.5 and 5nmol/L groups. These results suggested that BaP could impair the expression and function of Shh signaling pathway, perturbing the proliferation of chondrocytes and so disturbing craniofacial skeletal development.

Inhibition by polycyclic aromatic hydrocarbons of ATPase activities in Sebastiscus marmoratus larvae: relationship with the development of early life stages.

Sebastiscus marmoratus larvae were exposed to waterborne polycyclic aromatic hydrocarbons (PAHs) containing 3-5 rings, benzo[a]pyrene (BaP), pyrene (Py) or phenanthrene (Phe), respectively at 0.01, 0.1 and 1 µg L⁻¹. Cumulative mortality, frequency of dorsal curvature and rate of pericardial and yolk sac edema in larvae treated for 8 days were significantly increased in a dose-dependent manner. All three PAHs resulted in reduction of the lower jaw, and the extent of reduction increased with increasing concentrations of the PAHs. Na+/K+-ATPase and Ca²+-ATPase activity in larvae exposed to the three PAHs were all significantly inhibited in a dose-dependent manner. Analysis using the Pearson correlation coefficient indicated a significant correlation between the rate of the dorsal curvature and edema and the inhibition of Na+/K+-ATPase and Ca²+-ATPase activity, suggesting that the developmental defects caused by PAHs were related to their inhibition of Na+/K+-ATPase and Ca²+-ATPase activity.