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International Journal of Biological... Dec 2023Amelogenin and its derived peptides have exhibited excellent efficacy in promoting enamel biomimetic remineralization. However, little is known about their specific...
Amelogenin and its derived peptides have exhibited excellent efficacy in promoting enamel biomimetic remineralization. However, little is known about their specific action mechanisms. Herein, by combining experiments and computer simulation, the mechanism of an amelogenin-derived peptide QP5 in regulating enamel biomimetic remineralization is unveiled for the first time. In experiments, peptide QP5 was separated into (QPX)5 and C-tail domains, the interactions of peptide-minerals in nucleation solution and the regulation of peptide on enamel biomimetic remineralization were explored. QP5 exhibited an unordered conformation when mineral ions existed, and it could adsorb on minerals through its two domains, thereby inhibiting spontaneous nucleation. The remineralized enamel regulated by C-tail showed better mechanical properties and formed more biomimetic crystals than that of (QPX)5, indicating the C-tail domain of QP5 played an important role in forming enamel-like crystals. The simulation results showed that the conformation of QP5 changed greatly, mainly exhibiting β-bend, β-turn, and coil structures, and it eventually adsorbed on enamel through negatively charged residues of the C-tail domain, then captured Ca from solution to promote enamel remineralization. This study improved the evaluation methods of the mechanism of biomimetic peptides, and laid a theoretical basis for the amelioration and clinical transformation of peptide QP5.
Topics: Amelogenin; Biomimetics; Computer Simulation; Minerals; Peptides
PubMed: 37848117
DOI: 10.1016/j.ijbiomac.2023.127322 -
Journal of Molecular Histology Apr 2024Cytodifferentiation of odontogenic cells, a late stage event in odontogenesis is based on gene regulation. However, studies on the identification of the involved genes... (Review)
Review
Cytodifferentiation of odontogenic cells, a late stage event in odontogenesis is based on gene regulation. However, studies on the identification of the involved genes are scarce. The present study aimed to search for molecules for the cytodifferentiation of ameloblastic cells in rats. Differential display-PCR revealed a differentially expressed gene between cap/early bell stage and hard tissue formation stage in molars. This gene was identified as N-myc Downregulated Gene 1 (Ndrg1), which is the first report in tooth development. Real time PCR and western blotting confirmed that the mRNA level of Ndrg1 was higher during enamel formation than the cap stage. Ndrg1 expression was upregulated in the early bell, crown, and root stages in a time-dependent manner. These patterns of expression were similar in Ndrg2, but Ndrg3 and Ndrg4 levels did not change during the developmental stages. Immunofluorescence revealed that strong immunoreactivity against Ndrg1 were detected in differentiated ameloblasts only, not inner enamel epithelium, odontoblasts and ameloblastic cells in defected enamel regions. Alkaline phosphatase and alizarin red s stains along with real time PCR, revealed that Ndrg1 and Ndrg2 were involved in cytodifferentiation and enamel matrix mineralization by selectively regulating amelogenin and ameloblastin genes in SF2 ameloblastic cells. These results suggest that Ndrg may play a crucial functional role in the cytodifferentiation of ameloblasts for amelogenesis.
Topics: Animals; Rats; Ameloblasts; Amelogenesis; Molar; Muscle Proteins; Nerve Tissue Proteins; Odontogenesis; Proteins
PubMed: 38407765
DOI: 10.1007/s10735-024-10182-9 -
International Dental Journal Apr 2024The aim of this research was to compare the efficacy of the remineralising potential of self-assembling peptides (SAPs): Curodont Repair (P11-4), P26, and leucine-rich...
BACKGROUND
The aim of this research was to compare the efficacy of the remineralising potential of self-assembling peptides (SAPs): Curodont Repair (P11-4), P26, and leucine-rich amelogenin peptides (LRAP) with the standard 5% NaF varnish (Duraphat) on early enamel caries lesions (EECLs).
METHODS
A demineralising solution (DS) was used to create artificial EECLs in human dental enamel specimens, which were randomly allocated to treatment groups: P11-4; P26 solution; LRAP solution; 5% NaF varnish; and deionised water (DIW). Each specimen was subjected to 8 days of pH cycling. Specimens from each test group were subjected to microcomputed tomography (micro-CT) and nanomechanical testing to assess mineral density (MD), hardness (H), and elastic modulus (EM) properties of sound, demineralised, and treated enamel.
RESULTS
The mean MD percentage gain was highest in the P26 and P11-4 groups, followed by the LRAP, 5% NaF varnish, and DIW groups. There were statistically significant differences amongst groups. In the outer layer of EECLs, the EM and H were highest in P26 and P11-4 groups, followed by the LRAP and 5% NaF varnish. In the inner layer of EECLs, the EM and H were highest in P11-4 and P26 groups, indicative of enhanced penetration and remineralisation of the deeper parts of the artificial EECLs.
CONCLUSIONS
P26 and P11-4 SAPs are more effective than 5% NaF varnish in remineralising the depth of EECLs.
Topics: Humans; X-Ray Microtomography; Dental Enamel; Tooth Remineralization; Fluorides, Topical; Sodium Fluoride; Dental Caries; Peptides
PubMed: 37743135
DOI: 10.1016/j.identj.2023.07.003 -
Dental Materials : Official Publication... Feb 2024This study aims to synthesize novel chitosan nanoparticles loaded with an amelogenin-derived peptide QP5 (TMC-QP5/NPs), investigate their remineralization capability and...
OBJECTIVE
This study aims to synthesize novel chitosan nanoparticles loaded with an amelogenin-derived peptide QP5 (TMC-QP5/NPs), investigate their remineralization capability and inhibitory effects on endogenous matrix metalloproteinases (MMPs), and evaluate the dentin bonding properties of remineralized dentin regulated by TMC-QP5/NPs.
METHODS
TMC-QP5/NPs were prepared by ionic crosslinking method and characterized by dynamic light scattering method, scanning electron microscopy, transmission electron microscope, atomic force microscope, Fourier transform infrared spectroscopy, and differential scanning calorimetry. The encapsulation and loading efficiency of TMC-QP5/NPs and the release of QP5 were examined. To evaluate the remineralization capability of TMC-QP5/NPs, the mechanical properties, and the changes in structure and composition of differently conditioned dentin were characterized. The MMPs inhibitory effects of TMC-QP5/NPs were explored by MMP Activity Assay and in-situ zymography. The dentin bonding performance was detected by interfacial microleakage and microshear bond strength (μSBS).
RESULTS
TMC-QP5/NPs were successfully synthesized, with uniform size, good stability and biosafety. The encapsulation and loading efficiency of TMC-QP5/NPs was respectively 69.63 ± 2.22% and 13.21 ± 0.73%, with a sustained release of QP5. TMC-QP5/NPs could induce mineral deposits on demineralized collagen fibers and partial occlusion of dentin tubules, and recover the surface microhardness of dentin, showing better remineralization effects than QP5. Besides, TMC-QP5/NPs significantly inhibited the endogenous MMPs activity. The remineralized dentin induced by TMC-QP5/NPs exhibited less interfacial microleakage and higher μSBS, greatly improved dentin bonding.
SIGNIFICANCE
This novel peptide-loaded chitosan nanoparticles improved resin-dentin bonding by promoting dentin remineralization and inactivating MMPs, suggesting a promising strategy for optimizing dentin adhesive restorations.
Topics: Chitosan; Biomimetics; Nanoparticles; Peptides; Dentin; Matrix Metalloproteinases
PubMed: 37951748
DOI: 10.1016/j.dental.2023.11.003 -
BMC Oral Health Nov 2023Amelogenesis imperfecta (AI) is a developmental enamel defect affecting the structure of enamel, esthetic appearance, and the tooth masticatory function. Gene mutations...
BACKGROUND
Amelogenesis imperfecta (AI) is a developmental enamel defect affecting the structure of enamel, esthetic appearance, and the tooth masticatory function. Gene mutations are reported to be relevant to AI. However, the mechanism underlying AI caused by different mutations is still unclear. This study aimed to reveal the molecular pathogenesis in AI families with 2 novel pre-mRNA splicing mutations.
METHODS
Two Chinese families with AI were recruited. Whole-exome sequencing and Sanger sequencing were performed to identify mutations in candidate genes. Minigene splicing assays were performed to analyze the mutation effects on mRNA splicing alteration. Furthermore, three-dimensional structures of mutant proteins were predicted by AlphaFold2 to evaluate the detrimental effect.
RESULTS
The affected enamel in family 1 was thin, rough, and stained, which was diagnosed as hypoplastic-hypomature AI. Genomic analysis revealed a novel splicing mutation (NM_001142.2: c.570 + 1G > A) in the intron 6 of amelogenin (AMELX) gene in family 1, resulting in a partial intron 6 retention effect. The proband in family 2 exhibited a typical hypoplastic AI, and the splicing mutation (NM_031889.2: c.123 + 4 A > G) in the intron 4 of enamelin (ENAM) gene was observed in the proband and her father. This mutation led to exon 4 skipping. The predicted structures showed that there were obvious differences in the mutation proteins compared with wild type, leading to impaired function of mutant proteins.
CONCLUSIONS
In this study, we identified two new splicing mutations in AMELX and ENAM genes, which cause hypoplastic-hypomature and hypoplastic AI, respectively. These results expand the spectrum of genes causing AI and broaden our understanding of molecular genetic pathology of enamel formation.
Topics: Humans; Female; Amelogenin; Amelogenesis Imperfecta; Dental Enamel Proteins; Mutation; Mutant Proteins; Extracellular Matrix Proteins
PubMed: 37985977
DOI: 10.1186/s12903-023-03508-8 -
Journal of Molecular Evolution Dec 2023Deletion/insertion polymorphism (DIP) is one of the more promising genetic markers in the field of forensic genetics for personal identification and biogeographic...
Deletion/insertion polymorphism (DIP) is one of the more promising genetic markers in the field of forensic genetics for personal identification and biogeographic ancestry inference. In this research, we used an in-house developed ancestry-informative marker-DIP system, including 56 autosomal diallelic DIPs, three Y-chromosomal DIPs, and an Amelogenin gene, to analyze the genetic polymorphism and ancestral composition of the Chinese Korean group, as well as to explore its genetic relationships with the 26 reference populations. The results showed that this novel panel exhibited high genetic polymorphism in the studied Korean group and could be effectively applied for forensic individual identification in the Korean group. In addition, the results of multiple population genetic analyses indicated that the ancestral component of the Korean group was dominated by northern East Asia. Moreover, the Korean group was more closely related to the East Asian populations, especially to the Japanese population in Tokyo. This study enriched the genetic data of the Korean ethnic group in China and provided information on the ancestry of the Korean group from the perspective of population genetics.
Topics: Humans; Ethnicity; Polymorphism, Genetic; Genetics, Population; China; Republic of Korea; Gene Frequency; Polymorphism, Single Nucleotide
PubMed: 38006428
DOI: 10.1007/s00239-023-10143-y -
Cureus Feb 2024The purpose of the study was to compare and histologically investigate pulpal response and dentin bridge formation after direct pulp capping using recombinant amelogenin...
The purpose of the study was to compare and histologically investigate pulpal response and dentin bridge formation after direct pulp capping using recombinant amelogenin and mineral trioxide aggregate (MTA). Recombinant amelogenin protein and MTA were used as pulp capping materials in 120 teeth from eight mongrel dogs. Dogs were sacrificed at two different evaluation times. Regenerative changes were evaluated histologically. At two weeks, in contrast to the MTA group, most of the amelogenin group showed moderately formed hard tissue formation and the pulp tissue was completely filling the entire pulp chamber. These results were statistically significant. At two months, all the samples of the amelogenin group showed complete dentin bridge formation and the pulp chamber was filled entirely with tissue-mimicking the authentic pulp in all the specimens of the amelogenin group. These results were statistically significant. In conclusion, direct pulp capping by recombinant amelogenin protein resulted in significantly better regeneration of the dentin-pulp complex than MTA.
PubMed: 38516479
DOI: 10.7759/cureus.54560 -
Forensic Science International. Genetics Jul 2024The Precision ID NGS System from Thermo Fisher Scientific is a mainstream next-generation sequencing (NGS) platform used in forensic laboratories to detect almost all...
The Precision ID NGS System from Thermo Fisher Scientific is a mainstream next-generation sequencing (NGS) platform used in forensic laboratories to detect almost all commonly used forensic markers, except for Y-chromosomal short tandem repeats (Y-STRs). This study aimed to: 1) develop a Y-STR panel compatible with the automatic workflow of the NGS system using Ion AmpliSeq Technology, 2) evaluate the panel performance following the SWGDAM guidelines, and 3) explore the possibility of using a combination workflow to detect autosomal STRs and Y-STRs (AY-STR NGS workflow). The GrandFiler Y-STR Panel was successfully designed using the 'separating' and 'merging' strategies, including 102 Y-STRs and Amelogenin with an average amplicon length of 133 bp. It is a mega Y-STR multiplex system in which up to 16 samples can be sequenced simultaneously on an Ion 530 ™ Chip. Developmental validation studies of the performance of the NGS platform, species specificity, reproducibility, concordance, sensitivity, degraded samples, case-type samples, and mixtures were conducted to unequivocally determine whether the GrandFiler Y-STR Panel is suitable for real scenarios. The newly developed Y-STR panel showed compelling run metrics and NGS performance, including 92.47% bases with ≥ Q20, 91.80% effective reads, 2106 × depth of coverage (DoC), and 97.09% inter-locus balance. Additionally, it showed high specificity for human males and 99.40% methodological and bioinformatical concordance, generated complete profiles at ≥ 0.1 ng input DNA, and recovered more genetic information from severely degraded and diverse case samples. Although the outcome when used on mixtures was not as expected, more genetic information was obtained compared to that from capillary electrophoresis (CE) methods. The AY-STR NGS workflow was established by combining the GrandFiler Y-STR Panel with the Precision ID GlobalFiler ™ NGS STR Panel v2 at a 2:1 concentration ratio. The combination workflow on NGS performance, reproducibility, concordance, and sensitivity was as stable as the single Y-STR NGS workflow, providing more options for forensic scientists when dealing with different case scenarios. Overall, the GrandFiler Y-STR Panel was confirmed as the first to effectively detect a large number of Y-STR markers on the Precision ID NGS System, which is compatible with 51 Y-STRs in commercial CE kits and 51 Y-STRs in commercial NGS kits and the STRBase. The panel is as robust, reliable, and sensitive as current CE/NGS kits, and is suitable for solving real cases, especially for severely degraded samples (degradation index > 10).
Topics: Humans; Chromosomes, Human, Y; Microsatellite Repeats; High-Throughput Nucleotide Sequencing; Male; DNA Fingerprinting; Reproducibility of Results; Sequence Analysis, DNA; Species Specificity; Animals; Amelogenin; Polymerase Chain Reaction
PubMed: 38749212
DOI: 10.1016/j.fsigen.2024.103059 -
BMC Genomics Apr 2024Previously, a novel multiplex system of 64 loci was constructed based on capillary electrophoresis platform, including 59 autosomal insertion/deletions (A-InDels), two...
Joint application of A-InDels and miniSTRs for forensic personal, full and half sibling identifications, and genetic differentiation analyses in two populations from China.
BACKGROUND
Previously, a novel multiplex system of 64 loci was constructed based on capillary electrophoresis platform, including 59 autosomal insertion/deletions (A-InDels), two Y-chromosome InDels, two mini short tandem repeats (miniSTRs), and an Amelogenin gene. The aim of this study is to evaluate the efficiencies of this multiplex system for individual identification, paternity testing and biogeographic ancestry inference in Chinese Hezhou Han (CHH) and Hubei Tujia (CTH) groups, providing valuable insights for forensic anthropology and population genetics research.
RESULTS
The cumulative values of power of discrimination (CDP) and probability of exclusion (CPE) for the 59 A-InDels and two miniSTRs were 0.99999999999999999999999999754, 0.99999905; and 0.99999999999999999999999999998, 0.99999898 in CTH and CHH groups, respectively. When the likelihood ratio thresholds were set to 1 or 10, more than 95% of the full sibling pairs could be identified from unrelated individual pairs, and the false positive rates were less than 1.2% in both CTH and CHH groups. Biogeographic ancestry inference models based on 35 populations were constructed with three algorithms: random forest, adaptive boosting and extreme gradient boosting, and then 10-fold cross-validation analyses were applied to test these three models with the average accuracies of 86.59%, 84.22% and 87.80%, respectively. In addition, we also investigated the genetic relationships between the two studied groups with 33 reference populations using population statistical methods of F, D, phylogenetic tree, PCA, STRUCTURE and TreeMix analyses. The present results showed that compared to other continental populations, the CTH and CHH groups had closer genetic affinities to East Asian populations.
CONCLUSIONS
This novel multiplex system has high CDP and CPE in CTH and CHH groups, which can be used as a powerful tool for individual identification and paternity testing. According to various genetic analysis methods, the genetic structures of CTH and CHH groups are relatively similar to the reference East Asian populations.
Topics: Humans; Phylogeny; Siblings; Genetics, Population; China; INDEL Mutation; Microsatellite Repeats; Forensic Genetics; Gene Frequency
PubMed: 38566035
DOI: 10.1186/s12864-024-10187-4 -
Journal of Periodontal Research Jun 2024In order to evaluate the effect of methacrylated hyaluronic acid (HAMA) hydrogels containing the recombinant human amelogenin (rhAm) in vitro and in vivo.
OBJECTIVES
In order to evaluate the effect of methacrylated hyaluronic acid (HAMA) hydrogels containing the recombinant human amelogenin (rhAm) in vitro and in vivo.
BACKGROUND
The ultimate goal in treating periodontal disease is to control inflammation and achieve regeneration of periodontal tissues. In recent years, methacrylated hyaluronic acid (HAMA) containing recombinant human amyloid protein (rhAm) has been widely used as a new type of biomaterial in tissue engineering and regenerative medicine. However, there is a lack of comprehensive research on the periodontal regeneration effects of this hydrogel. This experiment aims to explore the application of photoresponsive recombinant human amelogenin-loaded hyaluronic acid hydrogel for periodontal tissue regeneration and provide valuable insights into its potential use in this field.
MATERIALS AND METHODS
The effects of rhAm-HAMA hydrogel on the proliferation of human periodontal ligament cells (hPDLCs) were assessed using the CCK-8 kit. The osteogenic differentiation of hPDLCs was evaluated through ALP staining and real-time PCR. Calvarial parietal defects were created in 4-week-old Sprague Dawley rats and implanted with deproteinized bovine bone matrix in different treatment groups. The animals were euthanized after 4 and 8 weeks of healing. The bone volume of the defect was observed by micro-CT and histological analysis.
RESULTS
Stimulating hPDLCs with rhAm-HAMA hydrogel did not significantly affect their proliferation (p > .05). ALP staining and real-time PCR results demonstrated that the rhAm-HAMA group exhibited a significant upregulation of osteoclastic gene expression (p < .05). Micro-CT results revealed a significant increase in mineralized tissue volume fraction (MTV/TV%), trabecular bone number (Tb.N), and mineralized tissue density (MTD) of the bone defect area in the rhAm-HAMA group compared to the other groups (p < .05). The results of hematoxylin and eosin staining and Masson staining at 8 weeks post-surgery further supported the results of the micro-CT.
CONCLUSIONS
The results of this study indicate that rhAm-HAMA hydrogel could effectively promote the osteogenic differentiation of hPDLCs and stabilize bone substitutes in the defects that enhance the bone regeneration in vivo.
Topics: Hyaluronic Acid; Animals; Bone Regeneration; Amelogenin; Humans; Rats, Sprague-Dawley; Periodontal Ligament; Rats; Hydrogels; Cell Proliferation; Cell Differentiation; Recombinant Proteins; Osteogenesis; Male; X-Ray Microtomography; Cells, Cultured; Methacrylates; Biocompatible Materials
PubMed: 38481308
DOI: 10.1111/jre.13235