RESUMO
The revolution in genetics has rapidly increased our knowledge of human and mouse genes that are critical for the formation of dental enamel and helps us understand how enamel evolved. In this graphical review we focus on the roles of 41 genes that are essential for the secretory stage of amelogenesis when characteristic enamel mineral ribbons initiate on dentin and elongate to expand the enamel layer to the future surface of the tooth. Based upon ultrastructural analyses of genetically modified mice, we propose a molecular model explaining how a cell attachment apparatus including collagen 17, α6ß4 and αvß6 integrins, laminin 332, and secreted enamel proteins could attach to individual enamel mineral ribbons and mold their cross-sectional dimensions as they simultaneously elongate and orient them in the direction of the retrograde movement of the ameloblast membrane.
Assuntos
Ameloblastos/metabolismo , Amelogênese/genética , Proteínas do Esmalte Dentário/genética , Esmalte Dentário/metabolismo , Modelos Genéticos , Ameloblastos/citologia , Ameloblastos/ultraestrutura , Animais , Colágeno/genética , Colágeno/metabolismo , Esmalte Dentário/citologia , Proteínas do Esmalte Dentário/metabolismo , Humanos , Integrinas/genética , Integrinas/metabolismo , Laminina/genética , Laminina/metabolismo , Camundongos , Microscopia Eletrônica de Varredura/métodosRESUMO
Dentin matrix protein 1 (DMP1) is found abundantly in the extracellular matrices of bone and dentin. Secretory DMP1 begins with a tripeptide of leucine-proline-valine (LPV) after the endoplasmic reticulum (ER)-entry signal peptide is cleaved. The goal of this study was to determine the role of the LPV motif in the secretion of DMP1. A series of DNA constructs was generated to express various forms of DMP1 with or without the LPV motif. These constructs were transfected into a preosteoblast cell line, the MC3T3-E1 cells, and the subcellular localization and secretion of various forms of DMP1 were examined by immunofluorescent staining and Western-blotting analyses. Immunofluorescent staining showed that the LPV-containing DMP1 variants were primarily localized in the Golgi complex, whereas the LPV-lacking DMP1 variants were found abundantly within the ER. Western-blotting analyses demonstrated that the LPV-containing DMP1 variants were rapidly secreted from the transfected cells, as they did not accumulate within the cells, and the amounts increased in the conditioned media over time. In contrast, the LPV-lacking DMP1 variants were predominantly retained within the cells, and only small amounts were secreted out of the cells over time. These results suggest that the LPV motif is essential for the efficient export of secretory DMP1 from the ER to the Golgi complex.
Assuntos
Motivos de Aminoácidos/genética , Retículo Endoplasmático/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Complexo de Golgi/metabolismo , Células 3T3 , Animais , Dentina/metabolismo , Retículo Endoplasmático/genética , Proteínas da Matriz Extracelular/genética , Complexo de Golgi/genética , Humanos , Leucina/genética , Camundongos , Osteoblastos/metabolismo , Prolina/genética , Valina/genéticaRESUMO
STATEMENT OF PROBLEM: Various techniques are used to cement implant-supported restorations. Excess residual cement is a concern. PURPOSE: The purpose of this in vitro study was to evaluate 3 techniques for cementing implant-supported restorations. MATERIAL AND METHODS: Twelve crowns and implant-abutment complexes (IAC) were cemented with 3 techniques: technique 1, a certain amount of cement evenly placed and excess cement removed with an explorer; technique 2, a smaller amount of cement without removal of excess cement; technique 3, a resin abutment replica used for excess cement removal after using a large amount of cement. Each specimen was treated with each technique 3 times. Precementing discrepancies (predis) and postcementing discrepancies (postdis) between IACs and crowns and the postcementing linear roughness (Ra and Rz) on designated junction areas of each specimen were measured. Tensile strength of the specimens was recorded with a universal testing machine at a crosshead speed of 0.5 mm/min. Repeated measures analysis with mixed models was used for differences among the 3 cementing techniques (α=.05). RESULTS: Compared with techniques 1 and 2, the specimens with technique 3 showed significantly lower mean differences between postdis and predis and Ra and Rz and higher mean tensile strength (P<.05). CONCLUSIONS: The application of a resin abutment replica for the cementation of implant-supported restorations decreased the discrepancy between the restoration and abutment, reduced cement residue, and increased restoration retention.
Assuntos
Cimentação/métodos , Prótese Dentária Fixada por Implante/métodos , Restauração Dentária Permanente/métodos , Coroas , Dente Suporte , Cimentos Dentários/efeitos adversos , Cimentos Dentários/uso terapêutico , Humanos , Técnicas In VitroRESUMO
The drug encapsulation efficiency, release rate and time, sustained release, and stimulus-response of carriers are very important for drug delivery. However, these always cannot obtained for the carrier with a single component. To improve the comprehensive performance of chitosan-based carriers for 5-Fu delivery, diatomite-incorporated hydroxypropyl cellulose/chitosan (DE/HPC/CS) composite aerogel microspheres were fabricated for the release of 5-fluorouracil (5-Fu), and the release performance was regulated with the content of diatomite, pH value, and external coating material. Firstly, the 5-Fu loaded DE/HPC/CS composite aerogel microspheres and Eudragit L100 coated microspheres were prepared with cross-linking followed by freeze-drying, and characterized by SEM, EDS, FTIR, XRD, DSC, TG, and swelling. The obtained aerogel microspheres have a diameter of about 0.5 mm, the weight percentage of F and Si elements on the surface are 0.55 % and 0.78 % respectively. The glass transition temperature increased from 179 °C to 181 °C and 185 °C with the incorporation of DE and coating of Eudragit, and the equilibrium swelling percentage of DE/HPC/CS (1.5:3:2) carriers are 101.52 %, 45.27 %, 67.32 % at pH 1.2, 5.0, 7.4, respectively. Then, the effect of DE content on the drug loading efficiency of DE/HPC/CS@5-Fu was investigated, with the increase of DE content, the highest encapsulation efficiency was 82.6 %. Finally, the release behavior of DE incorporated and Eudragit L100 Coated microspheres were investigated under different pH values, and evaluated with four kinetic models. The results revealed that the release rate of 5-Fu decreased with the increase of DE content, sustained release with extending time and pH-responsive were observed for the Eudragit-coated aerogel microspheres.
Assuntos
Celulose , Celulose/análogos & derivados , Quitosana , Preparações de Ação Retardada , Terra de Diatomáceas , Portadores de Fármacos , Liberação Controlada de Fármacos , Fluoruracila , Microesferas , Ácidos Polimetacrílicos , Quitosana/química , Celulose/química , Fluoruracila/química , Fluoruracila/administração & dosagem , Terra de Diatomáceas/química , Ácidos Polimetacrílicos/química , Portadores de Fármacos/química , Concentração de Íons de Hidrogênio , Géis/químicaRESUMO
Dentin sialophosphoprotein (DSPP) is primarily expressed by differentiated odontoblasts (dentin-forming cells), and transiently expressed by presecretory ameloblasts (enamel-forming cells). Disease-causing DSPP mutations predominantly fall into two categories: 5' mutations affecting targeting and trafficking, and 3' - 1 frameshift mutations converting the repetitive, hydrophilic, acidic C-terminal domain into a hydrophobic one. We characterized the dental phenotypes and investigated the pathological mechanisms of DsppP19L and Dspp-1fs mice that replicate the two categories of human DSPP mutations. In DsppP19L mice, dentin is less mineralized but contains dentinal tubules. Enamel mineral density is reduced. Intracellular accumulation and ER retention of DSPP is observed in odontoblasts and ameloblasts. In Dspp-1fs mice, a thin layer of reparative dentin lacking dentinal tubules is deposited. Odontoblasts show severe pathosis, including intracellular accumulation and ER retention of DSPP, strong ubiquitin and autophagy activity, ER-phagy, and sporadic apoptosis. Ultrastructurally, odontoblasts show extensive autophagic vacuoles, some of which contain fragmented ER. Enamel formation is comparable to wild type. These findings distinguish molecular mechanisms underlying the dental phenotypes of DsppP19L and Dspp-1fs mice and support the recently revised Shields classification of dentinogenesis imperfecta caused by DSPP mutations in humans. The Dspp-1fs mice may be valuable for the study of autophagy and ER-phagy.
Assuntos
Proteínas da Matriz Extracelular , Mutação da Fase de Leitura , Camundongos , Humanos , Animais , Proteínas da Matriz Extracelular/genética , Odontoblastos , Mutação , Fosfoproteínas/genética , Sialoglicoproteínas/genética , Dentina , Autofagia/genéticaRESUMO
OBJECTIVE: To explore the clinical application of lockedge suspension combined with three steel wires vertical fixation in comminuted fracture of inferior pole of patella. METHODS: From August 2016 to May 2019, 23 patients with comminuted fracture of the lower pole of the patella, including 14 males and 9 females, were treated with lockedge suspension combined with three steel wires vertical fixation. The age ranged from 34 to 68 (55.0±1.2) years. One year after operation, the pain and function were evaluated by pain visual analogue scale(VAS) and knee flexion and extension range of motion, and the clinical efficacy was evaluated by Lysholm knee score standard. RESULTS: All 23 patients were followed up for 12 to 14, with a mean of(13.0±0.5) months. One patient had skin irritation by the tail of the steel wire, and the rest had no postoperative complications such as incision infection, internal fixation loosening and fracture displacement. The fractures of 23 patients were healed, and the healing time was 10 to 14 weeks with a mean of(12.0±1.1) weeks. The VAS score decreased from 7.96±0.93 before operation to 0.83±0.65 one year after operation. The range of knee flexion and extension activities increased from(20.30±8.69) ° before operation to 1 year after operation(127.39±6.55) °. Lysholm knee score increased from 18.48±4.00 before operation to 96.09±4.91 one year after operation(P<0.05). CONCLUSION: The treatment of comminuted fracture of the lower pole of patella by lockedge suspension combined with three steel wires vertical fixation has reliable fixation and high fracture healing rate. It can meet the requirements of rapid rehabilitation and functional exercise, and the early clinical effect is satisfactory.
Assuntos
Fraturas Ósseas , Fraturas Cominutivas , Adulto , Idoso , Fios Ortopédicos , Feminino , Fixação Interna de Fraturas , Fraturas Ósseas/cirurgia , Fraturas Cominutivas/cirurgia , Humanos , Masculino , Pessoa de Meia-Idade , Patela/cirurgia , Aço , Resultado do TratamentoRESUMO
Human ACP4 (OMIM*606362) encodes a transmembrane protein that belongs to histidine acid phosphatase (ACP) family. Recessive mutations in ACP4 cause non-syndromic hypoplastic amelogenesis imperfecta (AI1J, OMIM#617297). While ACP activity has long been detected in developing teeth, its functions during tooth development and the pathogenesis of ACP4-associated AI remain largely unknown. Here, we characterized 2 AI1J families and identified a novel ACP4 disease-causing mutation: c.774_775del, p.Gly260Aspfs*29. To investigate the role of ACP4 during amelogenesis, we generated and characterized Acp4R110C mice that carry the p.(Arg110Cys) loss-of-function mutation. Mouse Acp4 expression was the strongest at secretory stage ameloblasts, and the protein localized primarily at Tomes' processes. While Acp4 heterozygous (Acp4+/R110C) mice showed no phenotypes, incisors and molars of homozygous (Acp4R110C/R110C) mice exhibited a thin layer of aplastic enamel with numerous ectopic mineralized nodules. Acp4R110C/R110C ameloblasts appeared normal initially but underwent pathology at mid-way of secretory stage. Ultrastructurally, sporadic enamel ribbons grew on mineralized dentin but failed to elongate, and aberrant needle-like crystals formed instead. Globs of organic matrix accumulated by the distal membranes of defective Tomes' processes. These results demonstrated a critical role for ACP4 in appositional growth of dental enamel probably by processing and regulating enamel matrix proteins around mineralization front apparatus.
Assuntos
Amelogênese Imperfeita , Proteínas do Esmalte Dentário , Fosfatase Ácida/metabolismo , Ameloblastos/metabolismo , Amelogênese , Amelogênese Imperfeita/metabolismo , Animais , Proteínas do Esmalte Dentário/genética , Proteínas do Esmalte Dentário/metabolismo , Histidina/metabolismo , Humanos , Camundongos , MutaçãoRESUMO
Dentin sialophosphoprotein (DSPP) is an extracellular matrix protein that is highly expressed in odontoblasts, but only transiently expressed in presecretory ameloblasts during tooth development. We previously generated a knockin mouse model expressing a mouse equivalent (DSPP, p.P19L) of human mutant DSPP (p.P17L; referred to as "DsppP19L/+ "), and reported that DsppP19L/+ and DsppP19L/P19L mice manifested a dentin phenotype resembling human dentinogenesis imperfecta (DGI). In this study, we analyzed pathogenic effects of mutant P19L-DSPP on enamel development in DsppP19L/+ and DsppP19L/P19L mice. Micro-Computed Tomography (µCT) analyses of 7-week-old mouse mandibular incisors showed that DsppP19L/P19L mice had significantly decreased enamel volume and/or enamel density at different stages of amelogenesis examined. Acid-etched scanning electron microscopy (SEM) analyses of mouse incisors demonstrated that, at the mid-late maturation stage of amelogenesis, the enamel of wild-type mice already had apparent decussating pattern of enamel rods, whereas only minute particulates were found in DsppP19L/+ mice, and no discernible structures in DsppP19L/P19L mouse enamel. However, by the time that incisor enamel was about to erupt into oral cavity, distinct decussating enamel rods were evident in DsppP19L/+ mice, but only poorly-defined enamel rods were revealed in DsppP19L/P19L mice. Moreover, µCT analyses of the mandibular first molars showed that DsppP19L/+ and DsppP19L/P19L mice had a significant reduction in enamel volume and enamel density at the ages of 2, 3, and 24weeks after birth. Backscattered and acid-etched SEM analyses revealed that while 3-week-old DsppP19L/+ mice had similar pattern of enamel rods in the mandibular first molars as age-matched wild-type mice, no distinct enamel rods were observed in DsppP19L/P19L mice. Yet neither DsppP19L/+ nor DsppP19L/P19L mice showed well-defined enamel rods in the mandibular first molars by the age of 24weeks, as judged by backscattered and acid-etched SEM. In situ hybridization showed that DSPP mRNA level was markedly reduced in the presecretory ameloblasts, but immunohistochemistry revealed that DSP/DSPP immunostaining signals were much stronger within the presecretory ameloblasts in Dspp mutant mice than in wild-type mice. These results suggest that mutant P19L-DSPP protein caused developmental enamel defects in mice, which may be associated with intracellular retention of mutant DSPP in the presecretory ameloblasts.
RESUMO
The human oral microbiota plays a vital role in maintaining metabolic homeostasis. To explore the relationship between Helicobacter pylori (Hp) and reflux esophagitis, we collected 86 saliva samples from reflux esophagitis patients (RE group) and 106 saliva samples from healthy people (C group) for a high-throughput sequencing comparison. No difference in alpha diversity was detected between the RE and the C groups, but beta diversity of the RE group was higher than the C group. Bacteroidetes was more abundant in the RE group, whereas Firmicutes was more abundant in the C group. The linear discriminant analysis effect size analysis demonstrated that the biomarkers of the RE group were Prevotella, Veillonella, Leptotrichia, and Actinomyces, and the biomarkers of the C group were Lautropia, Gemella, Rothia, and Streptococcus. The oral microbial network structure of the C group was more complex than that of the RE group. Second, to explore the effect of Hp on the oral microbiota of RE patients, we performed the 14C-urea breath test on 45 of the 86 RE patients. We compared the oral microbiota of 33 Hp-infected reflux esophagitis patients (REHpp group) and 12 non-Hp-infected reflux esophagitis patients (REHpn group). No difference in alpha diversity was observed between the REHpn and REHpp groups, and beta diversity of the REHpp group was significantly lower than that of the REHpn group. The biomarkers in the REHpp group were Veillonella, Haemophilus, Selenomonas, Megasphaera, Oribacterium, Butyrivibrio, and Campylobacter; and the biomarker in the REHpn group was Stomatobaculum. Megasphaera was positively correlated with Veillonella in the microbial network of the REHpp group. The main finding of this study is that RE disturbs the human oral microbiota, such as increased beta diversity. Hp infection may inhibit this disorderly trend.
Assuntos
Esofagite Péptica , Infecções por Helicobacter , Helicobacter pylori , Microbiota , Humanos , SalivaRESUMO
Mutations of Odontogenesis-Associated Phosphoprotein (ODAPH, OMIM *614829) cause autosomal recessive amelogenesis imperfecta, however, the function of ODAPH during amelogenesis is unknown. Here we characterized normal Odaph expression by in situ hybridization, generated Odaph truncation mice using CRISPR/Cas9 to replace the TGC codon encoding Cys41 into a TGA translation termination codon, and characterized and compared molar and incisor tooth formation in Odaph+/+, Odaph+/C41*, and OdaphC41*/C41* mice. We also searched genomes to determine when Odaph first appeared phylogenetically. We determined that tooth development in Odaph+/+ and Odaph+/C41* mice was indistinguishable in all respects, so the condition in mice is inherited in a recessive pattern, as it is in humans. Odaph is specifically expressed by ameloblasts starting with the onset of post-secretory transition and continues until mid-maturation. Based upon histological and ultrastructural analyses, we determined that the secretory stage of amelogenesis is not affected in OdaphC41*/C41* mice. The enamel layer achieves a normal shape and contour, normal thickness, and normal rod decussation. The fundamental problem in OdaphC41*/C41* mice starts during post-secretory transition, which fails to generate maturation stage ameloblasts. At the onset of what should be enamel maturation, a cyst forms that separates flattened ameloblasts from the enamel surface. The maturation stage fails completely.
Assuntos
Ameloblastos/fisiologia , Amelogênese , Proteínas da Matriz Extracelular/metabolismo , Fosfoproteínas/metabolismo , Amelogênese Imperfeita/genética , Amelogênese Imperfeita/patologia , Animais , Esmalte Dentário/crescimento & desenvolvimento , Esmalte Dentário/ultraestrutura , Proteínas da Matriz Extracelular/química , Proteínas da Matriz Extracelular/genética , Técnicas de Introdução de Genes , Hibridização In Situ , Incisivo/anatomia & histologia , Camundongos , Dente Molar/anatomia & histologia , Odontogênese , Fosfoproteínas/química , Fosfoproteínas/genéticaRESUMO
Dental enamel forms extracellularly as thin ribbons of amorphous calcium phosphate (ACP) that initiate on dentin mineral in close proximity to the ameloblast distal membrane. Secreted proteins are critical for this process. Enam-/- and Ambn-/- mice fail to form enamel. We characterize enamel ribbon formation in wild-type (WT), Amelx-/- and Mmp20-/- mouse mandibular incisors using focused ion beam scanning electron microscopy (FIB-SEM) in inverted backscatter mode. In Amelx-/- mice, initial enamel mineral ribbons extending from dentin are similar in form to those of WT mice. As early enamel development progresses, the Amelx-/- mineral ribbons develop multiple branches, resembling the staves of a Japanese fan. These striking fan-shaped structures cease growing after attaining ~ 20 µm of enamel thickness (WT is ~ 120 µm). The initial enamel mineral ribbons in Mmp20-/- mice, like those of the Amelx-/- and WT, extend from the dentin surface to the ameloblast membrane, but appear to be fewer in number and coated on their sides with organic material. Remarkably, Mmp20-/- mineral ribbons also form fan-like structures that extend to ~ 20 µm from the dentin surface. However, these fans are subsequently capped with a hard, disorganized outer mineral layer. Amelogenin cleavage products are the only matrix components absent in both Amelx-/- and Mmp20-/- mice. We conclude that MMP20 and amelogenin are not critical for enamel mineral ribbon initiation, orientation, or initial shape. The pathological fan-like plates in these mice may form from the lack of amelogenin cleavage products, which appear necessary to form ordered hydroxyapatite.
Assuntos
Ameloblastos/fisiologia , Amelogênese , Amelogenina/fisiologia , Esmalte Dentário/metabolismo , Metaloproteinase 20 da Matriz/fisiologia , Ameloblastos/ultraestrutura , Animais , Esmalte Dentário/ultraestrutura , Proteínas do Esmalte Dentário/metabolismo , Incisivo/ultraestrutura , CamundongosRESUMO
Non-syndromic inherited defects of tooth dentin are caused by two classes of dominant negative/gain-of-function mutations in dentin sialophosphoprotein (DSPP): 5' mutations affecting an N-terminal targeting sequence and 3' mutations that shift translation into the - 1 reading frame. DSPP defects cause an overlapping spectrum of phenotypes classified as dentin dysplasia type II and dentinogenesis imperfecta types II and III. Using CRISPR/Cas9, we generated a Dspp-1fs mouse model by introducing a FLAG-tag followed by a single nucleotide deletion that translated 493 extraneous amino acids before termination. Developing incisors and/or molars from this mouse and a DsppP19L mouse were characterized by morphological assessment, bSEM, nanohardness testing, histological analysis, in situ hybridization and immunohistochemistry. DsppP19L dentin contained dentinal tubules but grew slowly and was softer and less mineralized than the wild-type. DsppP19L incisor enamel was softer than normal, while molar enamel showed reduced rod/interrod definition. Dspp-1fs dentin formation was analogous to reparative dentin: it lacked dentinal tubules, contained cellular debris, and was significantly softer and thinner than Dspp+/+ and DsppP19L dentin. The Dspp-1fs incisor enamel appeared normal and was comparable to the wild-type in hardness. We conclude that 5' and 3' Dspp mutations cause dental malformations through different pathological mechanisms and can be regarded as distinct disorders.
Assuntos
Dentinogênese Imperfeita/genética , Proteínas da Matriz Extracelular/genética , Fosfoproteínas/genética , Sialoglicoproteínas/genética , Animais , Esmalte Dentário/metabolismo , Dentina/metabolismo , Dentinogênese Imperfeita/metabolismo , Dentinogênese Imperfeita/fisiopatologia , Modelos Animais de Doenças , Proteínas da Matriz Extracelular/metabolismo , Feminino , Mutação da Fase de Leitura/genética , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Fenótipo , Fosfoproteínas/metabolismo , Sialoglicoproteínas/metabolismo , Dente/metabolismoRESUMO
Montmorillonite grafted polyacrylic acid composite (GNM) was prepared by using ultraviolet radiation grafting method in this work. The synthesized materials were characterized by XRF, SEM, FTIR, XRD, TG, and XPS. The experimental equilibrium data indicates that the adsorbent is suitable for the Langmuir model and belongs to the pseudo-second-order kinetic model. The entire adsorption process is spontaneous, endothermic, and chaotically enhanced by thermodynamic analysis. The maximum adsorption capacity of La(III) by GNM was 280.54 mg/g at 313.15 K. In addition, the regeneration experiment shows that the adsorbent has good reusability and stable desorption efficiency. This study demonstrates that GNM has high adsorption performance and La(III) adsorption and regeneration capabilities to solve the water pollution caused by rare earth ions and regeneration capabilities for La(III).
Assuntos
Resinas Acrílicas/química , Bentonita/química , Lantânio/química , Poluentes Químicos da Água/química , Adsorção , Cinética , Termodinâmica , Raios Ultravioleta , Poluentes Químicos da Água/análise , Purificação da ÁguaRESUMO
BACKGROUND: Ameloblastin (AMBN) is a secreted matrix protein that is critical for the formation of dental enamel and is enamel-specific with respect to its essential functions. Biallelic AMBN defects cause non-syndromic autosomal recessive amelogenesis imperfecta. Homozygous Ambn mutant mice expressing an internally truncated AMBN protein deposit only a soft mineral crust on the surface of dentin. METHODS: We characterized a family with hypoplastic amelogenesis imperfecta caused by AMBN compound heterozygous mutations (c.1061T>C; p.Leu354Pro/ c.1340C>T; p.Pro447Leu). We generated and characterized Ambn knockout/NLS-lacZ (AmbnlacZ/lacZ ) knockin mice. RESULTS: No AMBN protein was detected using immunohistochemistry in null mice. ß-galactosidase activity was specific for ameloblasts in incisors and molars, and islands of cells along developing molar roots. AmbnlacZ/lacZ 7-week incisors and unerupted (D14) first molars showed extreme enamel surface roughness. No abnormalities were observed in dentin mineralization or in nondental tissues. Ameloblasts in the AmbnlacZ/lacZ mice were unable to initiate appositional growth and started to degenerate and deposit ectopic mineral. No layer of initial enamel ribbons formed in the AmbnlacZ/lacZ mice, but pockets of amelogenin accumulated on the dentin surface along the ameloblast distal membrane and within the enamel organ epithelia (EOE). NLS-lacZ signal was positive in the epididymis and nasal epithelium, but negative in ovary, oviduct, uterus, prostate, seminal vesicles, testis, submandibular salivary gland, kidney, liver, bladder, and bone, even after 15 hr of incubation with X-gal. CONCLUSIONS: Ameloblastin is critical for the initiation of enamel ribbon formation, and its absence results in pathological mineralization within the enamel organ epithelia.
Assuntos
Ameloblastos , Amelogênese Imperfeita , Proteínas do Esmalte Dentário , Mutação , Ameloblastos/metabolismo , Ameloblastos/patologia , Amelogênese Imperfeita/genética , Amelogênese Imperfeita/metabolismo , Amelogênese Imperfeita/patologia , Animais , Proteínas do Esmalte Dentário/genética , Proteínas do Esmalte Dentário/metabolismo , Dentina/metabolismo , Dentina/patologia , Técnicas de Introdução de Genes , Humanos , Camundongos , Camundongos Transgênicos , Especificidade de ÓrgãosRESUMO
Mutations in the dentin sialophosphoprotein (DSPP) gene cause dentinogenesis imperfecta. After synthesis, DSPP is proteolytically processed into NH2- and COOH-terminal fragments. The NH2-terminal fragment of DSPP is highly glycosylated but not phosphorylated, whereas the COOH-terminal fragment (named "dentin phosphoprotein" or "DPP") is highly phosphorylated but not glycosylated. These two fragments are believed to perform distinct roles in dentin formation. To analyze the functions of DPP in dentinogenesis, we created "Dspp-/-;DPP Tg mice", which expressed transgenic DPP driven by a Type I collagen promoter but lacked the endogenous Dspp gene. We characterized the dentin of the Dspp-/-;DPP Tg mice using X-ray radiography, histology, scanning electron microscopy, double fluorochrome labeling, immunohistochemistry and in situ hybridization. Micro-computed tomography analyses revealed that at postnatal 6 months, the transgenic expression of DPP increased the dentin thickness of the Dspp-null mice by 97.1% and restored the dentin material density by 29.5%. Histological analyses showed that the Dspp-null mice manifested an abnormal widening of the predentin while the predentin in Dspp-/-;DPP Tg mice was narrower than in the Dspp-null mice. Scanning electron microscopy analyses showed that the dentinal tubules in the Dspp-/-;DPP Tg mice were better organized than in the Dspp-null mice. The double fluorochrome labeling analyses demonstrated that the dentin mineral deposition rate in the Dspp-/-;DPP Tg mice was significantly improved compared to that in the Dspp-null mice. These findings indicate that the transgenic expression of DPP partially rescued the dentin defects of the DSPP-null mice, suggesting that DPP may promote dentin formation and that the coordinated actions between DPP and the NH2-terminal fragment of DSPP may be necessary for dentinogenesis.
Assuntos
Proteínas da Matriz Extracelular/genética , Expressão Gênica , Fosfoproteínas/genética , Sialoglicoproteínas/genética , Transgenes , Animais , Colágeno Tipo I/metabolismo , Cadeia alfa 1 do Colágeno Tipo I , Proteínas da Matriz Extracelular/metabolismo , Genótipo , Imuno-Histoquímica , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Odontoblastos/metabolismo , Fosfoproteínas/metabolismo , Radiografia , Sialoglicoproteínas/metabolismo , Dente/diagnóstico por imagem , Dente/metabolismo , Dente/patologia , Microtomografia por Raio-XRESUMO
Bone morphogenetic protein 1 (BMP1) and tolloid-like 1 (TLL1) belong to the BMP1/tolloid-like proteinase family, which cleaves secretory proteins. The constitutive deletion of the Bmp1 or Tll1 genes causes perinatal or embryonic lethality in mice. In this study, we first studied the ß-galactosidase activity in mice in which an IRES-lacZ-Neo cassette was inserted in the intron of either the Bmp1 or the Tll1 gene; the ß-galactosidase activities were used to reflect the expression of endogenous Bmp1 and Tll1, respectively. Our X-gal staining results showed that the odontoblasts in the tooth and cells in the periodontal ligament express both Bmp1 and Tll1. We then created Bmp1 flox/flox and Tll1 flox/flox mice by removing the IRES-lacZ-Neo cassette. By breeding 2.3 kb Col1a1-Cre mice with the Bmp1 flox/flox and Tll1 flox/flox mice, we further generated Col1a1-Cre;Bmp1 flox/flox ;Tll1 flox/flox mice in which both Bmp1 and Tll1 were inactivated in the Type I collagen-expressing cells. We employed X-ray radiography, histology and immunohistochemistry approaches to characterize the Col1a1-Cre;Bmp1 flox/flox ;Tll1 flox/flox mice. Our results showed that the molars of the Col1a1-Cre;Bmp1 flox/flox ;Tll1 flox/flox mice had wider predentin, thinner dentin and larger pulp chambers than those of the normal controls. The dentinal tubules of the molars in the Col1a1-Cre;Bmp1 flox/flox ;Tll1 flox/flox mice appeared disorganized. The level of dentin sialophosphoprotein in the molars of the 6-week-old Col1a1-Cre;Bmp1 flox/flox ;Tll1 flox/flox mice was lower than in the normal controls. The periodontal ligaments of the Col1a1-Cre;Bmp1 flox/flox ;Tll1 flox/flox mice were disorganized and had less fibrillin-1. Our findings indicate that the proteinases encoded by Bmp1 and Tll1 genes play essential roles in the development and maintenance of mouse dentin and periodontal ligaments.
Assuntos
Proteína Morfogenética Óssea 1/metabolismo , Colágeno Tipo I/metabolismo , Dentina/patologia , Ligamento Periodontal/patologia , Metaloproteases Semelhantes a Toloide/deficiência , Animais , Proteína Morfogenética Óssea 1/deficiência , Proteínas da Matriz Extracelular/metabolismo , Camundongos , Dente Molar/patologia , Odontoblastos/metabolismo , Fosfoproteínas/metabolismo , Sialoglicoproteínas/metabolismoRESUMO
Young patients are expected to place increased demands on total hip arthroplasty (THA) because they are more active and have a longer life expectancy. The long-term outcome of the Charnley low-friction arthroplasty in young, active patients has been shown to be associated with a high degree of polyethylene wear and osteolysis. However, cementless acetabular components have been shown to be successful in young patients.We evaluated the clinical and radiographic results of 77 cementless THAs in 81 young, active patients at a mean follow-up of 7.5 years. Mean preoperative Harris Hip Score improved from 46.24 to 96.5 points at 6 years. One acetabular component showed aseptic loosening 10 years postoperatively. Three patients (3 hips) underwent bone grafts and liner renewal for severe osteolysis around the acetabular component. The rate of survival at 6 years for loosening was 98.8%, and for revision of the liner was 95.5% (95% confidence interval, 93%-98%). Mean liner wear rate was 0.125 mm/year (range, 0.0-0.39 mm/year). Acetabular osteolysis was found in 14% (9 hips) of the 67 hips and was related to polyethylene wear (P=.0024). Although there was only 1 cup loosening in this study, there was a high rate of linear wear of the polyethylene liner and acetabular osteolysis in young, active patients. Further follow-up is therefore needed, and osteolysis and polyethylene wear should continue to be observed in young, active patients.