Long-term exposure to microplastics induces intestinal function dysbiosis in rare minnow (Gobiocypris rarus).

Microplastics are ubiquitous in the natural environment, especially in waters, and their potential impact is also a key issue of concern. In this study, we used 1 µm, 1000 µg/L, polystyrene (PS-MPs) particles to analyze the effects after exposure for 14 and 28 days in rare minnow (Gobiocypris rarus). Results indicated that PS-MPs induce structural alterations in the intestinal tissue, including epithelial damage, villi damage and the inflammatory cell infiltration, while the changes were severer after exposure for 28 days. Polystyrene microplastics also significantly increased the activities of catalase (CAT, increased 142 % and 385 % in 14d and 28d), superoxide dismutase (SOD, increased 17.76 % and 23.43 % in the 14d and 28d) and the content of malondialdehyde (MDA, increased 14.5 % and 442 % in the 14d and 28d), glutathione (GSH, increased 146 % and 298 % in the 14d and 28d). The results not only showed the characterization of gut microbial communities in rare minnow, but also indicated that microbial diversity and composition were altered in gut of fish exposed to PS-MPs. In the control groups, Proteobacteria (31.36-54.54 %), Actinobacteriota (39.99-52.54 %), Fusobacteriota (1.43-1.78 %), Bacteriadota (0.31-0.57 %) were the four dominant bacterial phyla in the intestinal of rare minnow. After exposure to microplastics, In the gut microbiota, the proportion of Proteobacteria increased 9.27 % and 30 % with exposure time, while Actinobacteria decreased 37.89 % and significantly different after 28 days. In addition, metabolomic analysis suggested that exposure to PS-MPs induced alterations of metabolic profiles in rare minnow and differential metabolites were involved in energy metabolism, inflammatory responsible secretion, oxidative stress, nucleotide and its metabolomics. In conclusion, our findings suggest that long-term exposure to microplastics could induce intestinal inflammation, oxidative stress, microbiota dysbiosis and metabolic disorder in rare minnow, and the alterations and severity were exacerbated by prolonged exposure. This study has extended our cognition of the toxicity of polystyrene, and enriched theoretical data for exploring the toxicological mechanism of microplastics.

Microplastics (Polystyrene) Exposure Induces Metabolic Changes in the Liver of Rare Minnow (Gobiocypris rarus).

Microplastics are environmental contaminants and an emergent concern. Microplastics are abundant in freshwater and can cause biochemical stress in freshwater organisms. In the current study, rare minnows (Gobiocypris rarus) were exposed to 1µm polystyrene microplastics at 200 µg/L concentration. We observed various sublethal effects after four weeks of exposure but no mortality. Numerous cellular and tissue alterations were observed in the liver. Differential metabolites and differentially expressed genes between control and exposure groups were identified and mapped to pathways in the Kyoto Encyclopedia of Genes and Genomes. The combination of transcriptomic and metabolomic analyses revealed significantly varied metabolic pathways between the two groups. These pathways were involved in glucolipid, amino acid, and nucleotide metabolism. Results demonstrated that MP exposure induced immune reaction, oxidative stress, and disturbed glycolipid and energy metabolism. The current study provided novel insights into the molecular and metabolic mechanisms of microplastic ecotoxicology in rare minnow.

MiRNA-218 regulates osteoclast differentiation and inflammation response in periodontitis rats through Mmp9.

Periodontitis is a multiple infection and inflammatory disease featured by connective tissue homeostasis loss, periodontal inflammation, and alveolar bone resorption. MicroRNAs (miRNAs) are involved in the mediation of a large scale of pathological processes. Here, we show that miRNA-218 provides protective effect on periodontitis via regulation of matrix metalloproteinase-9 (Mmp9). This pathway is aberrant in periodontium from rats with periodontitis and human periodontal ligament progenitor cells stimulated by lipopolysaccharide, with downregulation of miR-218 and higher levels of Mmp9 compared with periodontium from healthy rats and cells without stimulation. Overexpression of miR-218 can suppress the degradation of Collagen Types I and IV and dentin sialoprotein (DSP), attenuate osteoclast formation, and inhibit the secretion of proinflammatory cytokines. On the other hand, overexpression of Mmp9 promotes the degradation of Collagen Types I and IV and DSP as well as RANKL-induced osteoclast formation and elevates inflammatory factors levels. Furthermore, the inhibitory effect of miR-218 was prevented by rescuing the Mmp9 expression. In addition, we also have showed that miR-218 was able to attenuate bone resorption and inflammation in a periodontitis rat model. Collectively, our findings therefore suggest that miR-218 acts as a protective role in periodontitis through the regulation of Mmp9.

MicroRNA-21 promotes bone reconstruction in maxillary bone defects.

BACKGROUND: Bone reconstruction of the maxillary bone defects is an urgent issue due to its functional and aesthetic influence. MicroRNAs (miRNAs) are a class of non-coding RNAs that function in diverse biological and pathological processes. Recently, microRNA-21 (miR-21) was reported to play significant roles in bone formation, suggesting that miR-21 can be novel biomarker and therapeutic target for bone remodelling and skeletal diseases. However, the role of miR-21 in maxillary bone defects remains unclear. OBJECTIVE AND METHODS: This study aimed to investigate the effect of miR-21 on the bone reconstruction by inducing maxillary bone defects in wild-type (WT) and miR-21 knockout (miR-21-KO) mice and explore these mice as maxillary bone defect models. RESULTS: Micro-computed tomography (micro-CT) and histochemistry showed that the miR-21-KO mice had reduced bone reformation ability compared with the WT mice. The expression levels of alkaline phosphatase (ALP) and osteocalcin (OCN) were dramatically decreased in the miR-21-KO mice. In addition, injection of miR-21 agomir intra-peritoneally into miR-21-KO mice (miR-21-KO+ agomir) following the maxillary bone defects surgery displayed a significantly enhanced bone formation -promoting ability, which indicated that miR-21 agomir could ameliorate maxillary bone defects in miR-21-KO mice in vivo. Furthermore, immunohistochemistry suggested that ALP and OCN expressions were prominently increased in miR-21-KO+ agomir mice. CONCLUSION: These findings demonstrated that miR-21 deficiency impaired bone reformation and miR-21 contributed to the bone reconstruction of the maxillary bone defects. The evidence also supported the use of WT and miR-21-KO mice as maxillary bone defect models for further research.

The Contribution of PEG Molecular Weights in PEGylated Emulsions to the Various Phases in the Accelerated Blood Clearance (ABC) Phenomenon in Rats.

PEGylated preparations will be cleared rapidly from blood circulation when they are administrated twice in the same animal at a time interval, referred to as the "accelerated blood clearance" (ABC) phenomenon. Commonly, the study of the ABC phenomenon was investigated in two aspects: induction phase and effectuation phase. Herein, we report the influence of physicochemical properties (PEG molecular weights) in the induction phase and effectuation phase on the ABC phenomenon. In the experiment, on one hand, PEGylated emulsions with different molecular weights of PEG (refer to PEn, n = 400, 600, 800, 1000, 2000, and 5000) were injected for the first dose (induction phase) and induced PE2000 to produce ABC phenomenon. On the other hand, after PE2000 injected, PEn was injected for the second dose (effectuation phase). The results indicated that PE2000 and PE5000 induced an intense ABC phenomenon by their long-circulating characteristic. Interestingly, PE400, PE600, PE800, and PE1000 produced a consistent ABC phenomenon but different circulation time. Apparently, the induction of the ABC phenomenon is not only determined by the circulation time but also by the PEG molecular weights. When PEn is in the effectuation phase, the extent of the ABC phenomenon was not positively related to the molecular weights of PEG, increasing first and then weaken with the increase of molecular weights of PEG. These suggest that the number of -(CH2CH2O)n- repeat units of PE2000 was more conducive to interact with anti-PEG IgM. The results reported here clearly indicate that both the PEG molecular weights of prior dose and the subsequent dose of emulsion strongly influence the extent of the ABC phenomenon. Taken together, our observations in this study complete the effect of PEG molecular weights at a different phase of the ABC phenomenon. Furthermore, our findings have a significant impact on the choice of molecular weights for PEGylated formulations for use in cross-administration.

Effects of Uncleavable and Cleavable PEG-Lipids with Different Molecular Weights on Accelerated Blood Clearance of PEGylated Emulsions in Beagle Dogs.

To investigate the effect of polyethylene glycol (PEG) molecular weights on circulation time of PEGylated emulsions and the second injection of injected PEGylated emulsions, we studied the effect of molecular weights on the pharmacokinetic behavior of PEG-DSPE (modified emulsions with 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-n-[methoxy (polyethyleneglycol)]) and PEG-CHMC (modified emulsions with poly(ethyleneglycol)-cholesteryl carbonate) emulsions in beagle dogs. The "accelerated blood clearance" (ABC) phenomenon was induced. Through this study, the contribution of PEG molecular weights on the ABC phenomenon was further clarified, and the results provided guidance for lessening or eliminating the ABC phenomenon. We injected different PEG-modified emulsions with 10% PEG-modified density into beagle dogs at 2 µmol phospholipids kg-1 and the blood samples were drawn after 1 min, 3 min, 5 min, 10 min, 15 min, 30 min, 60 min, 120 min, 240 min, 360 min, 600 min, and 24 h. Then, concentrations of the drug were assayed using high-performance liquid chromatography (HPLC). The results showed that the circulation times of PEG-DSPE-modified emulsions were significantly different because of the difference in molecular weights, whereas those of the PEG-CHMC modified emulsions were not. The spatial conformation of PEG with small molecular weights (PEG400, PEG600, and PEG800) was more likely to induce a strong ABC phenomenon. The results of our work suggest the interaction of circulation time and PEG molecular weights on the ABC phenomenon, implying that the spatial conformation of PEG has advantages that alleviate the ABC phenomenon. Importantly, the results have implications for the choice of molecular weights of PEG for PEGylated formulations.

Enhanced Opsonization-Independent Phagocytosis and High Response Ability to Opsonized Antigen-Antibody Complexes: A New Role of Kupffer Cells in the Accelerated Blood Clearance Phenomenon upon Repeated Injection of PEGylated Emulsions.

The accelerated blood clearance (ABC) phenomenon is an immune response against the first injection of PEGylated colloidal drug delivery systems (CDDSs), which causes the accelerated clearance of the second dose. The enhanced complement-mediated phagocytic activity of Kupffer cells is responsible for accelerated second-dose clearance. Nevertheless, few studies have focused on the role of Kupffer cells in the induction phase of the ABC phenomenon. In this study, the intrinsic phagocytic activity of Kupffer cells was significantly enhanced at 6 days after the initial injected PEGylated emulsions (PEs) using the carbon clearance test and single-pass liver perfusion experiment. Furthermore, PE could stimulate Kupffer cells activation, leading to enhanced cell viability of Kupffer cells and opsonization-independent cellular uptake. It was also found that the response ability of Kupffer cells to the antigen-antibody complexes was augmented by the first injection of PE. Conclusively, we proposed that, besides anti-PEG IgM and complement activation-mediated hepatic uptake, enhanced opsonization-independent phagocytosis of Kupffer cells and the high response ability to opsonized antigen-antibody complexes contribute to the accelerated clearance of the second administration. The results indicated that Kupffer cells play an indispensable role in the ABC phenomenon and provided novel insights into the current view on the mechanism of the ABC effect.

Expression and Function of Hypoxia Inducible Factor-1α and Vascular Endothelial Growth Factor in Pulp Tissue of Teeth under Orthodontic Movement.

Orthodontic force may lead to cell damage, circulatory disturbances, and vascular changes of the dental pulp, which make a hypoxic environment in pulp. In order to maintain the homeostasis of dental pulp, hypoxia will inevitably induce the defensive reaction. However, this is a complex process and is regulated by numerous factors. In this study, we established an experimental animal model of orthodontic tooth movement to investigate the effects of mechanical force on the expression of VEGF and HIF-1α in dental pulp. Histological analysis of dental pulp and expressions of HIF-1α and VEGF proteins in dental pulp were examined. The results showed that inflammation and vascular changes happened in dental pulp tissue in different periods. Additionally, there were significant changes in the expression of HIF-1α and VEGF proteins under orthodontic force. After application of mechanical load, expression of HIF-1α and VEGF was markedly positive in 1, 3, 7 d, and 2 w groups, and then it weakened in 4 w group. These findings suggested that the expression of HIF-1α and VEGF was enhanced by mechanical force. HIF-1α and VEGF may play an important role in retaining the homeostasis of dental pulp during orthodontic tooth movement.

A preliminary study of the innate immune memory of Kupffer cells induced by PEGylated nanoemulsions.

The accelerated blood clearance (ABC) phenomenon describes a dilemma of polyethylene glycol (PEG) applied in drug delivery system (DDS) caused by its immunogenicity, that results in the enhanced blood clearance rate and increased hepatic and splenic accumulation after secondary injection of PEGylated nanocarriers. However, the ABC index, as the judgement of ABC phenomenon, only describes the accelerated blood clearance rate, but ignores the enhanced hepatic and splenic accumulation. Therefore, we proposed the hepatic accumulation (HA) index and the splenic accumulation (SA) index as supplements for assessing the ABC phenomenon, to emphasize the contribution of liver and spleen, especially the liver, possessing the most population of tissue resident macrophages. By altering the first injection site from the tail vein to the liver portal vein, there was no impact on anti-PEG IgM production, and the secondary hepatic accumulation of PEGylated nanoemulsions (PE) was observed to be proportionate to the first PE stimulation strength on the liver. We also determined that Kupffer cells (KCs) were the main contributor to this enhancement. On this basis, we revealed a definite phenomenon that PE could induce innate immune memory in KCs, by enhancing the phagocytosis of KCs toward PE during the secondary stimulation. The PE-stimulated KCs could carry this memory to the naïve rats through adoptive transfer, resulting in increased hepatic accumulation in the recipient rats without antibody production. Studies examining the phagocytosis of KCs in vivo, ex vivo and in vitro revealed that the memory of KCs against PE triggered by first-stimulated PE could be maintained independently of other cells or components until 21 days after the first stimulation, and possessing specificity to PEG, which was invalid to long-circulating GE (GM1 modified nanoemulsions). The discovery of immune memory in KCs induced by PE highlights the importance of focusing on the relationship between the innate immune system and PEGylated nanocarriers during the development of DDS to improve medication safety in the clinic.

Compressive force stimulates the gene expression of IL-17s and their receptors in MC3T3-E1 cells.

During orthodontic tooth movement, cytokines released from periodontal ligament fibroblasts and alveolar bone osteoblasts can alter the process of bone remodeling. Recently, interleukin-17 (IL-17) was found to stimulate osteoclastic resorption through osteoblasts by inducing receptor activator of nuclear factor κB ligand (RANKL) expression. However, the relationship between mechanical stress and IL-17 production by osteoblasts is not clear. Therefore, we examined the effect of compressive force on the expressions of IL-17A, IL-17B, IL-17C, IL-17D, IL-17E, IL-17F, and their receptors (IL-17RA, IL-17RB, IL-17RC, IL-17RD, and IL-17RE) using MC3T3-E1 cells as osteoblast-like cells. We also examined the effect of IL-17A on the expression of IL-17Rs, RANKL, macrophage colony-stimulating factor (M-CSF), and osteoprotegerin (OPG). The cells were cultured with or without continuous compressive force (1.0 and 2.0 g/cm(2)) for up to 24 hr. The cells were also cultured with or without IL-17A (0.1, 1.0, or 10 ng/ml) for up to 72 hr. The mRNA expressions of IL-17s and their receptors were estimated by real-time polymerase chain reaction. The expression levels of IL-17s and their receptors increased depending on the compressive force. The addition of IL-17A increased the expression of IL-17RA, IL-17RB, IL-17RC, IL-17RE, RANKL, and M-CSF, whereas it decreased OPG expression. These results indicate that compressive force induces the expression of IL-17s and their receptors in osteoblast-like cells and that IL-17s and their receptors produced in response to compressive force may affect osteoclastogenesis through the expression of RANKL, M-CSF, and OPG.

Accuracy of surgical positioning of orthodontic miniscrews with a computer-aided design and manufacturing template.

INTRODUCTION: Our objective was to enable accurate miniscrew placement after preoperative simulation. We developed a new template for miniscrew placement and evaluated its accuracy. METHODS: Eleven patients who had bimaxillary protrusion were scanned with computed tomography. The 3-dimensional computed tomography data were used to produce, with stereolithography apparatus, a template for accurate miniscrew placement. The interradicular space available for miniscrew placement was calculated in the 3-dimensional images. Postoperative computed tomography images were matched with preoperative images to calculate the deviations between the planned and actual placements. RESULTS: The distance for placement of a miniscrew between 2 roots was 4.12 mm (SD, 0.25 mm; range, 3.7-4.5 mm). The placed miniscrews showed an average angular deviation of 1.2 degrees (SD, 0.43 degrees ; range, 0.6 degrees -2.41 degrees ) compared with the plan, whereas the mean linear distomesial deviation was 0.42 mm (SD, 0.13 mm; range, 0.15-0.6 mm) at the tip. CONCLUSIONS: The proposed template has high accuracy and will be especially useful for patients who require precise miniscrew placement.

A myxobacterial LPMO10 has oxidizing cellulose activity for promoting biomass enzymatic saccharification of agricultural crop straws.

Myxobacteria are soil microorganisms with the ability to break down biological macromolecules due to the secretion of a large number of extracellular enzymes, but there has been no research report on myxobacterial lytic polysaccharide monooxygenases (LPMOs). In this study, two LPMO10s, ViLPMO10A and ViLPMO10B, from myxobacterium Vitiosangium sp. GDMCC 1.1324 were characterized. Of which, ViLPMO10B is a C1-oxidizing cellulose-active LPMO. Moreover, ViLPMO10B could decrease the degree of polymerization of crop straws cellulose and synergize with commercial cellulase to promote the saccharification. When the weight ratio of commercial cellulase to ViLPMO10B was 9:1, the conversion efficiency of corn stalk, sugarcane bagasse, and rice straw into reducing sugar was improved by 17%, 16%, and 22%, respectively, compared with commercial cellulase without ViLPMO10B. These results indicate that ViLPMO10B has the potential to be a component of a high-efficient cellulase cocktail and has application value in the saccharification of agricultural residual biomasses.

Biologic response of rapid tooth movement with periodontal ligament distraction.

INTRODUCTION: Length of treatment is a complaint of many orthodontic patients. The purpose of this study was to evaluate the security and feasibility of rapid tooth movement with periodontal ligament distraction. METHODS: Eight male beagles, aged 13 to 16 months, were used in this study. Extraction of the mandibular second premolar and alveolar surgery to reduce the osteal resistance on the mesial side of the extraction socket were performed on the experimental side. Then a device was placed to distract the first premolars distally on the experimental side; on the control side, the first premolars were distalized with nickel-titanium coil springs. The beagles were killed in the first, second, fourth, and eighth weeks after orthodontic force application. RESULTS: The first premolar on the experimental side moved more rapidly than that on the control side (P <0.05). Histologic data indicated that more new bone was deposited on tension area of the experimental side than on the control side. Active and extensive bone resorption in the compressive area and bone deposition in the tension area were observed on the experimental side. CONCLUSIONS: These results suggest that the periodontal ligament can be rapidly distracted without complications. The rapid orthodontic tooth movement by distracting the periodontal ligament cannot be emulated by current conventional orthodontic concepts and methods.

Nanosilver/poly (dl-lactic-co-glycolic acid) on titanium implant surfaces for the enhancement of antibacterial properties and osteoinductivity.

Background: Despite titanium (Ti) implants have been commonly used in the medical device field due to their superior biocompatibility, implant-associated bacterial infection remains a major clinical complication. Nanosilver, an effective antibacterial agent against a wide spectrum of bacterial strains, with a low-resistance potential, has attracted much interest too. Incorporation of nanosilver on Ti implants may be a promising approach to prevent biofilm formation. Purpose: The objective of the study was to investigate the antibacterial effects and osteoinductive properties of nanosilver/poly (dl-lactic-co-glycolic acid)-coated titanium (NSPTi). Methods: Gram-positive methicillin-resistant Staphylococcus aureus (MRSA) and the Gram-negative opportunistic pathogen Pseudomonas aeruginosa (PAO-1) were used to evaluate the antibacterial activity of NSPTi implants through the analysis of bacterial colonization in vitro and in vivo. Furthermore, we examined the osteoinductive potential of NSPTi implants by investigating the proliferation and differentiation of MC3T3-E1 preosteoblast cells. In vivo, the osteoinductive properties of NSPTi implants were assessed by radiographic evaluation, H&E staining, and Masson's trichrome staining. Results: In vitro, bacterial adhesion to the 2% NSPTi was significantly inhibited and <1% of adhered bacteria survived after 24 hours. In vitro, the average colony-forming units (CFU)/g ratios in the 2% NSPTi with 103 CFU MRSA and PAO-1 were 1.50±0.68 and 1.75±0.6, respectively. In the uncoated Ti groups, the ratios were 1.03±0.82×103 and 0.94±0.49×103, respectively. These results demonstrated that NSPTi implants had prominent antibacterial properties. Proliferation of MC3T3-E1 cells on the 2% NSPTi sample was 1.51, 1.78, and 2.22 times that on the uncoated Ti control after 3, 5, and 7 days' incubation, respectively. Furthermore, NSPTi implants promoted the maturation and differentiation of MC3T3-E1 cells. In vivo, NSPTi accelerated the formation of new bone while suppressing bacterial survival. Conclusion: NSPTi implants have simultaneous antibacterial and osteoinductive activities and therefore have the potential in clinical applications.

Down-regulation of long non-coding RNA MEG3 suppresses osteogenic differentiation of periodontal ligament stem cells (PDLSCs) through miR-27a-3p/IGF1 axis in periodontitis.

OBJECTIVE: This study aimed to investigate the roles of long noncoding RNA (lncRNA) maternally expressed gene 3 (MEG3) in osteogenic differentiation of periodontal ligament stem cells (PDLSCs) in periodontitis. METHODS: Differentially expressed lncRNAs and mRNAs between periodontitis periodontal ligament tissues and healthy periodontal ligament tissues were selected out using R project. PDLSCs were identified using flow cytometry. Western blot was employed to detect pathway relative proteins. Besides, targeted relationships between lncRNA and miRNA, as well as miRNA and mRNA were verified by dual luciferase reporter gene assay. Osteogenic differentiation of PDLSCs was assessed by alkaline phosphatase (ALP) staining and Alizarin Red Staining (ARS). Markers for osteoblast (Runx2, Osterix, Osteocalcin, Colla1) were detected using western blot. RESULTS: LncRNA MEG3 and IGF1 were both down-regulated, while miR-27a-3p was up-regulated in periodontitis samples compared with healthy samples. Overexpression of MEG3 promoted osteogenic differentiation by enhancing expression of IGF1 yet suppressing expression of miRNA-27a-3p. Meanwhile, the results of ALP and ARS staining indicated that up-regulation of lncRNA MEG3 or IGF1 promoted osteogenic differentiation in PDLSCs, which could be reversed with up-regulation of miRNA-27a-3p. CONCLUSION: Down-regulation of MEG3 suppressed osteogenic differentiation of PDLSCs through miR-27a-3p/IGF1 axis in periodontitis.

Expression of Osterix in mechanical stress-induced osteogenic differentiation of periodontal ligament cells in vitro.

Osterix (Osx) is an osteoblast-specific transcription factor required for the differentiation of pre-osteoblasts into functional osteoblasts. This study sought to examine the changes of Osx expression in periodontal ligament cells (PDLC) subjected to mechanical force, and to investigate whether Osx is involved in the mechanical stress-induced differentiation of PDLC. Human PDLC were exposed to centrifugal force for 1-12 h. Real-time polymerase chain reaction (PCR), western blot, and immunofluorescence assays were used to examine the mRNA and protein expression of Osx and its subcellular localization. Furthermore, PDLC were transfected with the expression vector pcDNA3.1 flag-Osx and subjected to mechanical force for 6 h. The changes in alkaline phosphatase (ALP) activity and in the expression of core-binding factor alpha1 (Cbfa1), ALP, osteopontin, bone sialoprotein, osteocalcin, and collagen I were measured. After the application of mechanical force, Osx was upregulated in a time-dependent manner at both mRNA and protein levels, and Osx protein was translocated from the cytosol into the cell nuclei. Overexpression of Osx did not affect the expression of Cbfa1, but it significantly enhanced the ALP activity and the mRNA expression of all the aforementioned osteogenic marker genes, all of which increased further under mechanical stress. These results suggest that Osx might play an important role in the mechanical stress-induced osteogenic differentiation of PDLC and therefore be involved in alveolar bone remodeling during orthodontic therapy.

Long noncoding RNA TUG1 facilitates osteogenic differentiation of periodontal ligament stem cells via interacting with Lin28A.

Periodontal ligament stem cells (PDLSCs) are mesenchymal stem cells derived from dental tissues with multidirectional differentiation potential and excellent self-renewing ability. Recently, long noncoding RNAs (lncRNAs) have been shown to play important roles in MSC osteogenic differentiation. In this study, we found that taurine upregulated gene 1 (TUG1), an evolutionarily conserved and widely present lncRNA was significantly upregulated in osteogenically induced PDLSCs compared to their undifferentiated counterparts. Further investigation demonstrated that the expression of TUG1 was positively correlated with the osteogenic differentiation of PDLSCs following the induction, as evidenced by the increase in cellular alkaline phosphatase (ALP) level, formation of calcium nodules, and the upregulation of several osteogenic-related gene markers such as ALP, osteocalcin (OCN), and runt-related transcription factor 2 (Runx2). Conversely, TUG1 knockdown was demonstrated to inhibit the potential of PDLSCs for osteogenic differentiation. Using bioinformatics analysis, we identified lin-28 homolog A (Lin28A) as a potential target of TUG1 during osteogenic differentiation of PDLSCs. Lin28A was found to be significantly downregulated in TUG1-repressed PDLSCs and contained multiple binding sites for lncRNA TUG1. Moreover, suppression of Lin28A was shown to be able to inhibit osteogenic differentiation and decreased the expression of several osteogenic genes. Taken together, these results could help researchers better understand the mechanism that governs the osteogenic differentiation of PDLSCs, and also serve as a stepping stone for the development of novel therapeutic strategies that can be used to regenerate dental tissues.

Profiling of subgingival plaque biofilm microbiota in adolescents after completion of orthodontic therapy.

BACKGROUND: Fixed orthodontic treatment is the most common method for malocclusion but has the potential risk of periodontal complication with unclear outcomes of whether microbiologic and clinical changes could be reversible in adolescents after orthodontic therapy. METHODS: Twenty adolescents with orthodontic treatment were enrolled in the study as the case group at end of the therapy, while 19 periodontally healthy adolescents were involved in the control group. At baseline (T0), clinical parameters including gingival index, probing depth and sulcus bleeding index were tested, and subgingival plaque samples were collected from the lower incisors. The counts of A. actinomycetemcomitans, P. gingivalis, P. intermedia, T. forsythia and total bacteria were determined by real-time PCR. All parameters were reassessed after 1 month (T1) and 3 months (T2) in the case group and compared with that of the controls. RESULTS: At baseline (T0), clinical parameters (including GI, PD, SBI) of the test sites in the case group were significantly higher than that of the control group (P<0.05 or P<0.01). At 3 months (T2), no differences were noticed in GI and SBI between two groups. The prevalence and counts of periodontopathogens tend to be normal (P>0.05), while PD and the amount of P.intermedia were still significantly higher compared with that of the control group (P<0.05 or P<0.01). CONCLUSION: After removal of appliances, the periodontal changes induced by orthodontic therapy are only partially reversible at 3 months after removal.

Prevalence of fimA genotypes of Porphyromonas gingivalis in adolescent orthodontic patients.

BACKGROUND: The placement of fixed orthodontic appliances may alter the composition of oral microbiota and has the potential risk of periodontal complication. Porphyromonas gingivalis fimbriae play a critical role in colonization of P. gingivalis in subgingival regions. In this study, we investigated the association between the prevalence of P. gingivalis-specific fimA genotypes and periodontal health status in adolescent orthodontic patients, to identify the pathogencity of P. gingivalis during orthodontic therapy. METHODS: Sixty-one adolescent orthodontic patients were enrolled in the case group, while the control group consisted of 56 periodontally healthy adolescents. At baseline (T0), clinical parameter (gingival index) was tested, and subgingival plaque samples were obtained from the lower incisors. The incidences of P. gingivalis and fimA genotypes were detected by polymerase chain reaction. All parameters were reassessed after 1 month (T1), 2 months (T2), 3 months (T3), and 6 months (T4) in the case group and then compared with those of the controls. RESULTS: Both microbiological and clinical parameters from orthodontic patients started to increase after placement of fixed appliances. Maximum values were reached at 3 months after placement and followed by their decreases at six months. However, the microbiological and clinical parameters in the case group were significantly higher than those of the control group. The GI of fimA II, IV-positive samples was significantly higher than that of negative samples. CONCLUSION: P. gingivalis carrying fimA II or IV was closely related to orthodontic gingivitis. In addition, proper oral hygiene control could lead to little increase in dental plaque accumulation, and exert a beneficial effect to periodontal tissues.

PERK-eIF2α-ATF4 pathway mediated by endoplasmic reticulum stress response is involved in osteodifferentiation of human periodontal ligament cells under cyclic mechanical force.

To prevent excess accumulation of unfolded proteins in endoplasmic reticulum (ER), eukaryotic cells have signaling pathways from the ER to the cytosol or nucleus. These processes are known as the endoplasmic reticulum stress (ERS) response. Protein kinase R like endoplasmic reticulum kinase (PERK) is a major transducer of the ERS response and it directly phosphorylate α-subunit of eukaryotic initiation factor 2 (eIF2α), resulting in translational attenuation. Phosphorylated eIF2α specifically promoted the translation of the activating transcription factor 4 (ATF4). ATF4 is a known important transcription factor which plays a pivotal role in osteoblast differentiation and bone formation. Furthermore, ATF4 is a downstream target of PERK. Studies have shown that PERK-eIF2α-ATF4 signal pathway mediated by ERS was involved in osteoblastic differentiation of osteoblasts. We have known that orthodontic tooth movement is a process of periodontal ligament cells (PDLCs) osteodifferentiation and alveolar bone remodeling under mechanical force. However, the involvement of PERK-eIF2α-ATF4 signal pathway mediated by ERS in osteogenic differentiation of PDLCs under mechanical force has not been unclear. In our study, we applied the cyclic mechanical force at 10% elongation with 0.5Hz to mimic occlusal force, and explored whether PERK-eIF2α-ATF4 signaling pathway mediated by ERS involved in osteogenic differentiation of PDLCs under mechanical force. Firstly, cyclic mechanical force will induce ERS and intensify several osteoblast marker genes (ATF4, OCN, and BSP). Next, we found that PERK overexpression increased eIF2α phosphorylation and expression of ATF4, furthermore induced BSP, OCN expression, thus it will promote osteodifferentiation of hPDLCs; mechanical force could promote this effect. However, PERK(-/-) cells showed the opposite changes, which will inhibit osteodifferentiation of hPDLCs. Taken together, our study proved that PERK-eIF2α-ATF4 signaling pathway mediated by ERS involved in osteoblast differentiation of PDLCs under cyclic mechanical force.