Wnt3α and transforming growth factor-ß induce myofibroblast differentiation from periodontal ligament cells via different pathways.

Myofibroblasts are specialized cells that play a key role in connective tissue remodeling and reconstruction. Alpha-smooth muscle actin (α-SMA), vimentin and tenascin-C are myofibroblast phenotype, while α-SMA is the phenotypic marker. The observation that human periodontal ligament cells (hPDLCs) differentiate into myofibroblasts under orthodontic force has provided a new perspective for understanding of the biological and biomechanical mechanisms involved in orthodontic tooth movement. However, the cell-specific molecular mechanisms leading to myofibroblast differentiation in the periodontal ligament (PDL) remain unclear. In this study, we found that expression of Wnt3α, transforming growth factor-ß1 (TGF-ß1), α-SMA and tenascin-C increased in both tension and compression regions of the PDL under orthodontic load compared with unloaded control, suggesting that upregulated Wnt3α and TGF-ß1 signaling might have roles in myofibroblast differentiation in response to orthodontic force. We reveal in vitro that both Wnt3α and TGF-ß1 promote myofibroblast differentiation from hPDLCs. Dickkopf-1 (DKK1) impairs Wnt3α-induced myofibroblast differentiation in a ß-catenin-dependent manner. TGF-ß1 stimulates myofibroblast differentiation via a JNK-dependent mechanism. DKK1 has no significant effect on TGF-ß1-induced myofibroblastic phenotype.

Micro-flow cell washing technique combined with single-cell Raman spectroscopy for rapid and automatic antimicrobial susceptibility test of pathogen in urine.

Facing the rapid spread of antimicrobial resistance, methods based on single-cell Raman spectroscopy have proven their advances in reducing the turn-around time (TAT) of antimicrobial susceptibility tests (AST). However, the Raman-based methods are still hindered by the prolonged centrifugal cell washing procedure, which may require complex labor operation and induce high mechanical stress, resulting in a pretreatment time of over 1 h as well as a high cell-loss probability. In this study, we developed a micro-flow cell washing device and corresponding Raman-compatible washing chips, which were able to automatically remove the impurities in the samples, retain the bacterial cell and perform Raman spectra acquisition in situ. Results of washing the 5- and 10-µm polymethyl methacrylate (PMMA) microspheres showed that the novel technique achieved a successful removal of 99 % impurity and an 80 % particle retention rate after 6 to 10 cycles of washing. The micro-flow cell washing technique could complete the pretreatment for urine samples in a 96-well plate within 10 min, only taking 15 % of the handling time required by centrifugation. The AST profiles of urine sample spiked with E. coli 25922, E. faecalis 29212, and S. aureus 29213 obtained by the proposed Raman-based approach were found to be 100 % consistent with the results from broth micro-dilution while reducing the TAT to 3 h from several days which is required by the latter. Our study has demonstrated the micro-flow cell washing technique is a reliable, fast and compatible approach to replace centrifuge washing for sample pretreatment of Raman-AST and could be readily applied in clinical scenarios.

Catechol-Modified and MnO2-Nanozyme-Reinforced Hydrogel with Improved Antioxidant and Antibacterial Capacity for Periodontitis Treatment.

Periodontitis is an inflammatory disease characterized by tooth loss and alveolar bone resorption. Bacteria are the original cause of periodontitis, and excess reactive oxygen species (ROS) encourage and intensify inflammation. In this study, a mussel-inspired and MnO2 NPs-reinforced adhesive hydrogel capable of alleviating periodontitis with improved antibacterial and antioxidant abilities was developed. The hydrogel was created by combining polyvinyl alcohol (PVA), 3,4-dihydroxy-d-phenylalanine (DOPA), and MnO2 nanoparticles (NPs) (named PDMO hydrogel). The hydrogel was demonstrated to be able to scavenge various free radicals (including total ROS─O2•- and OH•) and relieve the hypoxia in an inflammatory microenvironment by scavenging excess ROS and generating O2 due to its superoxide dismutase (SOD)/catalase (CAT)-like activity. Besides, under 808 nm near-infrared (NIR) light, the photothermal performance of the PDMO hydrogel displayed favorable antibacterial and antibiofilm effects toward Escherichia coli, Staphylococcus aureus, and Porphyromonas gingivalis (up to nearly 100% antibacterial rate). Furthermore, the PDMO hydrogel exhibited favorable therapeutic efficacy in alleviating gingivitis in Sprague-Dawley rats, even comparable to or better than the commercial PERIO. In addition, in the periodontitis models, the PDMO2 group showed the height of the residual alveolar bone and the smallest shadow area of low density among other groups, indicating the positive role of the PDMO2 hydrogel in bone regeneration. Finally, the biosafety of the PDMO hydrogel was comprehensively investigated, and the hydrogel was demonstrated to have good biocompatibility. Therefore, the developed PDMO hydrogel provided an effective solution to resolve biofilm recolonization and oxidative stress in periodontitis and could be a superior candidate for local drug delivery system in the clinical management of periodontitis with great potential for future clinical translation.

MiR-199a-5P promotes osteogenic differentiation of human stem cells from apical papilla via targeting IFIT2 in apical periodontitis.

Introduction: Periapical alveolar bone loss is the common consequence of apical periodontitis (AP) caused by persistent local inflammation around the apical area. Human stem cells from apical papilla (hSCAPs) play a crucial role in the restoration of bone lesions during AP. Studies have recently identified the critical role of microRNAs (miRNAs) involved in AP pathogenesis, but little is known about their function and potential molecular mechanism, especially in the osteogenesis of hSCAPs during AP. Here, we investigated the role of clinical sample-based specific miRNAs in the osteogenesis of hSCAPs. Methods: Differential expression of miRNAs were detected in the periapical tissues of normal and patients with AP via transcriptomic analysis, and the expression of miR-199a-5p was confirmed by qRT-PCR. Treatment of hSCAPs with miR-199a-5p mimics while loaded onto beta-tricalcium phosphate (ß-TCP) ceramic particle scaffold to explore its effect on osteogenesis in vivo. RNA binding protein immunoprecipitation (RIP) and Luciferase reporter assay were conducted to identify the target gene of miR-199a-5p. Results: The expression of miR-199a-5p was decreased in the periapical tissues of AP patients, and miR-199a-5p mimics markedly enhanced cell proliferation and osteogenic differentiation of hSCAPs, while miR-199a-5p antagomir dramatically attenuated hSCAPs osteogenesis. Moreover, we identified and confirmed Interferon Induced Protein with Tetratricopeptide Repeats 2 (IFIT2) as a specific target of miR-199a-5p, and silencing endogenous IFIT2 expression alleviated the inhibitory effect of miR-199a-5p antagomir on the osteogenic differentiation of hSCAPs. Furthermore, miR-199a-5p mimics transfected hSCAPs loaded onto beta-tricalcium phosphate (ß-TCP) scaffolds induced robust subcutaneous ectopic bone formation in vivo. Discussion: These results strengthen our understanding of predictors and facilitators of the key AP miRNAs (miR-199a-5p) in bone lesion repair under periapical inflammatory conditions. And the regulatory networks will be instrumental in exploring the underlying mechanisms of AP and lay the foundation for future regenerative medicine based on dental mesenchymal stem cells.

Preparation of magnetic molecularly imprinted polymers for separating rutin from Chinese medicinal plants.

The preparation of magnetic molecularly imprinted polymers (MMIPs) which can be used for the separation and purification of rutin from Chinese medicinal plants has been proposed. By applying the improved co-precipitation method, magnetic Fe(3)O(4) particles were easily prepared, followed by the modification of TEOS and functionalization with -CH=CH(2). Using functionalized Fe(3)O(4) particles as the magnetic cores, rutin as the template, and acrylamide as the functional monomer, MMIPs were synthesized by surface-imprinted polymerization under the protection of nitrogen gas and successive mechanical stirring at 60 °C for 24 h. Magnetic non-molecularly imprinted polymers (MNIPs) were also prepared with the same synthesis procedure as with MMIPs only without the presence of rutin. Magnetic particles were characterized by FT-IR, XRD, and TG analysis. And the selectivity of MMIPs was also investigated in detail. In addition, the performance of the MMIPs for the adsorption of rutin in the analysis of Chinese medicinal plants was assessed. The mean recoveries were 84.33% (RSD: 3.22%, n = 3) for Saururus chinensis (Lour.) Bail and 85.20% (RSD: 3.58%, n = 3) for Flos Sophorae, respectively, which showed that the prepared MMIPs with many advantages possess the value of practical application.

Molecularly imprinted polymer for solid-phase extraction of rutin in complicated traditional Chinese medicines.

In the present work, an improved and direct approach for the preparation of molecularly imprinted polymers (MIPs) was proposed. The MIPs were prepared based on bulk polymerization by water-bath heating and ultrasonic elution of the template, using rutin as the template, acrylamide (AM) as the functional monomer and 2,2'-azobisisobutyronitrile (AIBN) as the cross linker. Molecularly imprinted polymers prepared by other elution methods, including microwave-assisted extraction and conventional Soxhlet extraction, were used for comparison and the results showed that the ultrasonic elution method is the best. The synthesized MIPs were characterized by Fourier transform infrared (FT-IR) spectroscopy and scanning electron microscopy (SEM). High performance liquid chromatography (HPLC) was used to evaluate the adsorption properties and recognition mechanism of the MIPs. Structurally similar compounds including quercetin and genistein were utilized for verifying the molecular selectivity and characterizing the recognition capability of the MIPs. The MIPs were used as a sorbent for the solid phase extraction of rutin, and the resultant cartridge showed a good extraction performance. Thus, a molecularly imprinted solid-phase extraction (MISPE) procedure for selective pre-concentration of rutin from complicated traditional Chinese medicine (TCM) samples was proposed. Various elution parameters that affect the adsorption capacity of the polymer were evaluated to optimize the selective pre-concentration of rutin. The characteristics of the MISPE method were validated by HPLC. The recoveries ranged from 85% to 91% for TCMs, which demonstrated that this MISPE-HPLC method could be applied to pre-concentrate and determinate rutin directly from complicated TCM samples in the presence of other interfering substances.

[Maxillary dental and palatal morphological characteristics in 62 children with impacted maxillary canine in mixed dentition].

PURPOSE: The aim of this study was to compare the difference of dental arch and palatal morphology between children with impacted maxillary canine and without impacted maxillary canine, to provide reference for early diagnosis and treatment of impacted maxillary canine. METHODS: Sixty-two children, 8-11 years of age (9.8±1.04 years), were divided into experimental group and control group, with 31 children in each group. Digital casts were obtained using 3D laser scanner from maxillary models. Dental arch width, length, basal bone width, palatal width, height, surface area and volume were measured. Paired t test was performed with SPSS 24.0 software package. RESULTS: Dental arch width and palatal width were significantly narrower in experimental group than in control group(P<0.05), but there was on significant difference in dental arch length, basal bone width and palatal height between the two groups(P>0.05).Additionally, palatal surface area and volume were significantly larger in control group than in experimental group(P<0.05). CONCLUSIONS: Children with impacted maxillary canine have smaller maxillary dental arch width, palatal width, palatal surface area and volume compared to control group. Dental arch width, palatal width, palatal surface area and volume should be taken into consideration when early identifying impacted maxillary canine.

[Three-dimensional morphological analysis of the palate of mouth-breathing children in mixed dentition].

OBJECTIVE: To study the effects of mouth-breathing on maxillary arch development by comparing the palatal morphology of mouth- and nose-breathing children in mixed dentition. METHODS: Children in mixed dentition were enrolled and categorized into mouth-breathing (test group) and nose-breathing groups (control group) according to their breathing patterns. Children's plaster models were scanned with 3D laser scanner, and the 3D data were reconstructed and measured using Minics 15.0 and Geomagic 12.0 software. Measurement data (inter-molar width, palatal height, palatal volume, and palatal surface area) of the two groups were compared, and the correlation among the four measurement items was analyzed. RESULTS: The participants were 73 children (37 in test group and 36 in control group) with a mean age of (8.63±0.78) years old. The test group had significantly smaller inter-molar width, palatal volume, and palatal surface area but significantly higher palatal height than the control group (P<0.05). Inter-molar width and palatal volume were positively correlated with the palatal surface area in the test group (P<0.05). Inter-molar width and palatal height were positively correlated with the palatal surface area in the control group (P<0.01). CONCLUSIONS: Mouth-breathing children have significantly reduced inter-molar width, palatal volume, and surface, and substantially increased palatal height, leading to different developmental patterns of the palatal morphology.

Prevalence and care index of early childhood caries in mainland China: evidence from epidemiological surveys during 1987-2013.

Early childhood caries (ECC) is the most common chronic disease in young children. Its reported prevalence varies greatly across China. This systematic review aimed to explore the epidemiological characteristics of ECC in mainland China from 1987 to 2013. In total, 102 articles were included. The pooled national prevalence and care index (ft/dmft%) for ECC were 65.5% and 3.6%, respectively. The overall ECC prevalence declined from 77.9% during 1987-1994 to 56.4% during 2010-2013. The pooled ECC prevalence for children aged 1-6 years was 0.3%, 17.3%, 40.2%, 54.4%, 66.1%, and 70.7%, respectively. There was no significant difference in prevalence between boys (59.1%) and girls (58.9%); and the care index was also similar (8.1% versus 7.7%). Slightly higher ECC prevalence was observed in rural areas (63.5%) compared with urban areas (59.5%) (RR = 1.08, 95% CI: 1.02-1.14); but a much higher care index was reported in urban children (6.0%) than their rural counterparts (1.6%) (RR = 3.68, 95% CI: 2.54-5.35). The 2006-2013 map of ECC prevalence among 5-year-olds showed wide geographic variations across China. Four adjacent provinces, including Sichuan, Chongqing, Hubei, and Shaanxi, constituted the areas with the lowest ECC prevalence in mainland China.

Sclerostin is essential for alveolar bone loss in occlusal hypofunction.

Bone loss is caused by occlusal hypofunction and is a serious health concern. This is particularly true of tooth loss, which is common in the elderly. However, the cellular and molecular mechanisms underlying bone loss have yet to be fully elucidated. Sclerostin and Wnt/ß-catenin signaling have previously been reported to serve important roles in regulating bone remodeling. Therefore, the present study aimed to investigate the involvement of sclerostin and Wnt/ß-catenin signaling in occlusal hypofunction-induced alveolar bone remodeling. The unilateral maxillary molars of 14 male Sprague-Dawley rats were extracted in order to establish a model of occlusal hypofunction. For each rat, the non-extraction side was treated as the control group for comparisons with the extraction side. At 8 weeks after tooth extraction, the rats were sacrificed and alveolar bone specimens were harvested for X-ray radiography, micro-computed tomography (CT) and histological and immunohistochemical examinations. Bone loss and architecture deterioration were observed at the occlusal hypofunction side. The bone mineral density was markedly decreased and the ratio of bone volume to total volume was significantly decreased at the hypofunction side, as compared with the control side (P<0.001). In addition, the number of osteoclasts at the hypofunction side were significantly increased compared with that in the control side (P<0.001), as demonstrated using tartrate-resistant acid phosphatase staining. Furthermore, the protein expression levels of sclerostin and receptor activator of nuclear factor-κB ligand were increased, whereas those of ß-catenin were decreased, at the hypofunction side when compared with the control side. In conclusion, the results of the present study suggested that occlusal hypofunction-induced bone loss may be associated with upregulated expression of sclerostin, which, in turn, may inhibit the activity of the Wnt/ß-catenin signaling pathway.

Morphological characteristics influencing the orthodontic extraction strategies for Angle's class II division 1 malocclusions.

BACKGROUND: Extraction has now been accepted widely in various malocclusions including Angle's class II division 1. However, the levels of scientific evidence in orthodontic treatment planning have been weak, and it is unlikely to systematically provide a rationale and consistent basis in decisions of extraction. This study was retrospectively designed to investigate the initial morphologic characteristics of class II division 1 subjects involving four different extraction strategies, to determine the relevant influential factors when choosing extraction strategies with the most commonly used mechanics and the principle of simplicity in orthodontic treatment based on cases diagnosed and treated by an experienced orthodontist. METHODS: One hundred and ten samples of Angle's class II division 1 malocclusion with good facial and occlusal outcomes after orthodontic treatment were selected and divided into four groups according to different extraction patterns. For each case, pretreatment models and the lateral radiographs were analyzed. Significant variables of models and craniofacial structures of each group were identified by comparing the measurements using one-way analysis of variance (ANOVA) at a significance level of P < 0.05. Then, binary logistic regression analysis was used and a regression equation was established to quantify the correlations among the significant variables and their contributions to the extraction decisions. RESULTS: Molar relationship, lower anterior crowding, anterior Bolton index, and anterior overjet measured from models, as well as ANS-Xi-Pm, NBa-PtGn, Li-NsPog', U1-NPog and L1-NPog measured from lateral radiographs were found to be statistically significant. Binary logistic regression analysis revealed that lower anterior crowding, molar relationship, and growth pattern were the three most relevant influential factors with a declining impact contributing to the extraction decisions for Angle's class II division 1 malocclusions. CONCLUSIONS: Angle's class II division 1 malocclusions exhibit various morphological characteristics. Orthodontists should comprehensively consider the reciprocal impact of multiple factors when choosing different extraction strategies for Angle's class II division 1 malocclusions.

Preparation of molecular imprinted polymers using bi-functional monomer and bi-crosslinker for solid-phase extraction of rutin.

Molecular imprinted polymers (MIPs) were prepared using rutin as the template, different reagents as the functional monomer and different reagents as the cross-linker by solution polymerization. Several parameters that would influence the performance of MIPs were investigated including the type of functional monomer (single or double) and cross-linker (single or double), and the molar ratio of the template, the functional monomer and the cross-linker. The optimum synthesis conditions of MIPs were found to be bi-monomers (acrylamide-co-2-vinyl pyridine, 3:1) and bi-crosslinker (ethylene glycol dimethacrylate-co-divinylbenzene, 3:1). The ratio of the template, the functional monomer and the cross-linker was found to be 1:6:20. MIPs synthesized under these conditions were filled into the cartridges as the adsorbents of solid-phase extraction (SPE). A competition test was conducted to authenticate the selectivity and the specificity of molecularly imprinted solid-phase extraction (MISPE) for rutin using the mixture solution of standard rutin and its structural analogs including quercetin, naringenin and kaempferol. Compared with purchased SPE including C(18), silica and PCX, MISPE showed better selectivity and enrichment property for rutin in the extracted solutions of Chinese medicinal plants than any others. The mean recoveries were 85.93% (RSD: 3.04%, n=3) for Saururus chinensis (Lour.) Bail and 88.61% (RSD: 3.36%, n=3) for Flos Sophorae, respectively, which indicated that the optimized rutin-MIPs possess the value of practical application.

Development and characterization of molecularly imprinted polymers for the selective enrichment of podophyllotoxin from traditional Chinese medicines.

In the present work, microwave heating initiated precipitation polymerization was developed to prepare podophyllotoxin (PPT) molecularly imprinted polymers (MIPs), resulting in much shorter polymerization time and better particle morphology. Prior to the polymerization, ultraviolet and FTIR spectroscopy were used to study the interactions between PPT and the functional monomers. The synthesized parameters were respectively optimized and the optimal conditions for the efficient adsorption property were template: PPT, 1 mmol; functional monomer: acrylamide, 6 mmol; bi-crosslinker: ethylene glycol dimethacrylate, 20 mmol and divinylbenzene, 20 mmol; porogen: acetonitrile, 40 mL; initiator: azobisisobutyronitrile, 0.01mol L⁻¹; polymerization temperature: 60°C. FTIR spectroscopy, SEM and thermal analysis were used to characterize the MIPs. The results of the equilibrium rebinding experiments and the competitive adsorption experiments showed that these imprinted polymers exhibited good adsorption ability for the PPT. Scatchard analysis illustrated that two and one types of binding sites were generated in the MIPs and non-imprinted polymers (NIPs), respectively. Using the prepared MIPs as the solid phase extraction (SPE) sorbent, PPT was extracted selectively and efficiently from Dysosma versipellis, Sinopodophyllum hexandrum and Diphylleia sinensis. The regression equation was y=5.873×106x+17075.659 with the correlation coefficient of 0.9994 in the concentration range of 0.005-0.4 mg mL⁻¹. After washing and eluting the SPE column with methanol and MeOH/acetic acid solution (v/v, 9:1), the limits of detection were 0.12-0.18 µg mL⁻¹ and their recoveries were in the range of 89.5-91.1% with all RSDs lower than 3.7.

Preparation of codeine-resinate and chlorpheniramine-resinate sustained-release suspension and its pharmacokinetic evaluation in beagle dogs.

Using ion exchange resins (IERs) as carriers, a dual-drug sustained release suspension containing codeine, and chlorpheniramine had been prepared to elevate drug safety, effectiveness and conformance. The codeine resinate and chlorpheniramine resinate beads were prepared by a batch process and then impregnated with Polyethylene glycol 4000 (PEG 4000), respectively. The PEG impregnated drug resinate beads were coated with ethylcellulose as the coating polymer and di-n-butyl-phthalate as plasticizer in ethanol and methylene chloride mixture by the Wurster process. The coated PEG impregnated drug resinate beads were dispersed in an aqueous suspending vehicle containing 0.5% w/w xanthan gum and 0.5% w/w of hydroxypropylmethylcellulose of nominal viscosity of 4000 cps, obtaining codeine resinate and chlorpheniramine resinate sustained-release suspension (CCSS). Codeine phosphate and chlorpheniramine maleate were respectively loaded onto AMBERLITE IRP 69, and PEG 4000 was used to impregnate drug resinate beads to maintain their geometry. Ethylcellulose with di-n-butyl-phthalate in ethanol and methylene chloride mixture for the coating of drug resinate beads was performed in Glatt fluidized bed coater, where the coating solution flow rate was 8-12 g/min, the inlet air temperature was 50-60 degrees C, the outlet air temperature was 32-38 degrees C, the atomizing air pressure was 2.0 bar and the fluidized air pressure was adjusted as required. Few significant agglomeration of circulating drug resinate beads was observed during the operation. The film weight gained 20% w/w and 15% w/w were suitable for the PEG impregnated codeine resinate and chlorpheniramine resinate beads, respectively. Residual solvent content increased with coating level, but inprocess drying could reduce residual solvent content. In the present study, the rates of drug release from both drug resinate beads were measured in 0.05 M and 0.5M KCl solutions. The increased ionic strength generally accelerated the release rate of both drugs. But the release of codeine from its resinate beads was much more rapid than chlorpheniramine released from its resinate beads in the same ionic strength release medium. The drug release specification of the CCSS, where release mediums were 0.05 M KCl solution for codeine and 0.5 M KCl solution for chlorpheniramine, was established to be in conformance with in vivo performance. Relative bioavailability and pharmacokinetics evaluation of the CCSS, using commercial immediate-release tablets as the reference preparation, were performed following a randomized two-way crossover design in beagle dogs. The drug concentrations in plasma were measured by a validated LC-MS/MS method to determine the pharmacokinetic parameters of CCSS. This LC-MS/MS method demonstrated high accuracy and precision for bioanalysis, and was proved quick and reliable for the pharmacokinetic studies. The results showed that the CCSS had the longer value of Tmax and the lower value of Cmax, which meant an obviously sustained release effect, and its relative bioavailability of codeine and chlorpheniramine were (103.6 +/- 14.6)% and (98.1 +/- 10.3)%, respectively, compared with the reference preparation. These findings indicated that a novel liquid sustained release suspension made by using IERs as carriers and subsequent fluidized bed coating might provide a constant plasma level of the active pharmaceutical ingredient being highly beneficial for various therapeutic reasons.