Objective:The study aims to compare effect of inflammatory microenvironment on the regenerative capacitary of periodontal ligament stem cells (PDLSCs) andumbilical cord mesenchymal stem cells (UCMSCs). Methods：TNF-α was applied to mimic inflammatory microenvironment. Cell proliferation assay and proliferation marker genes CCNE1, CCND1 analysis were used to compare the proliferation capacity. AnnexinV/PI staining was used to detect cellular apoptosis. Alizarin red staining and osteogenesis marker genes RUNX2, OCN analysis were used to compare the capacity of osteogenic differentiation. Results：Treatment of TNF-α resulted in an obvious decrease in the proliferation and osteogenic differentiation capacity of PDLSCs, while an increase in apoptosis of PDLSCs. In contract, treatment of TNF-α leaded to a mild decrease in the capacity of proliferation and osteogenic differentiation in UCMSCs, and a slight increase in cellular apoptosis. Conclusions：Comparing with PDLSCs, UCMSCs are more resistant to the influence of inflammatory microenvironment.

Objective:?Establishing an animal model which can simulate the natural tooth developing environment in physiological condition for the researches of tooth regeneration, root development and tooth eruption. Methods:?The upper first molar of mouse at bell stage is used in this model for extraction and transplantation, comparing with the subrenal capsule transplantation model by radiological and histological observation postoperatively. Results:?The tooth germ of upper first molar by the postnatal 8th day has almost formed the crown and the root development is about to begin, which is a suitable subject of study in this model. Comparing with that in the subrenal capsule transplantation model, the root development of transplanted tooth germ in this model shows more similar to that of the natural tooth. Conclusions:?The extraction and orthotopic transplantation model of the upper first molar at bell stage in mice is a desired option for the observation of root development and tooth eruption.

Objective:?To explore the feasibility of using CFSE (carboxyfluorescein succinimidyl amino ester, CFSE) to sort senescent cells. Methods:?Senescent cells were induced by the CDK4/6 inhibitor LY2835219 on Cal27. CFSE staining was performed immediately after withdrawal of LY2835219, and then cells were cultured in the dark. The cells were stained with SA-β-gal on days 0, 2, 4, 6 and 8 after withdrawal. The cells with high fluorescence intensity of CFSE were sorted by a flow cytometer at a wavelength of 488 nm. Results:?Cells labeled with CFSE were consistent with SA-β-gal staining. The rate of positive result of SA-β-gal in the CFSE-high group sorted by CFSE on the 4th and 6th days after withdrawal was significantly higher than that in the unsorted group (P＜0.05). Conclusions:?Using CFSE to sort senescent cells is feasible, which could obtain live senescent cells.

Objective：To explore the role of miR-21 in inflammatory response and osteogenic differentiation ofperiodontal ligament stem cell (PDLSCs). Methods：Gingiva samples from periodontitis patients and healthy people were collected and real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to detect the expression of miR-21. 10 ng/mLtumor necrosis factor (TNF-α) was used to stimulate PDLSCs, mimicking inflammatory environment in vitro. QRT-PCR was performed to detect the expression of miR-21. MiR-21 mimics and inhibitors were utilized respectively to overexpress and knockdown miR-21 in PDLSCs. QRT-PCR and alkaline phosphatase (ALP) staining were performed to confirm the role of miR-21 on the osteogenic differentiation of PDLSCs. Results：The expression of miR-21 was upregulated in the gingiva tissues from periodontitis patients(P＜0.05). The miR-21 expression in PDLSCs was also upregulated under TNF-α stimulation (P＜0.01). Overexpression of miR-21 down-regulated the expression of cytokines IL-6, IL-1βinduced by TNF-α in PDLSCs(P＜0.05). Conversely, knocking down miR-21 promoted the expression of those genes(P＜0.05). During the osteogenesis induction of PDLSCs, the expression of miR-21 was significantly increased (P＜0.01). Overexpression of miR-21 enhanced and knocking down miR-21 alleviated the expression of osteogenic related genes RUNX2, OCNand the ALP staining (P＜0.05). In the inflammatory microenvironment, miR-21 overexpression could alleviate the inhibition effect of TNF-α stimulation on osteogenesis of PDLSCs (P＜0.05).Conclusions：MiR-21 alleviates the expression of cytokines induced by TNF-α in PDLSCs and promotes osteogenic differentiation of PDLSCs in the inflammatory microenvironment.

Objective: To treat oral squamous cell carcinoma through via exosomes loaded with miRNA34a. Methods: First, exosomes ought to be extracted from the culture supernatants of HEK293 cells by means of ultracentrifugation and identified through integrating transmission electron microscopy, nanoparticle tracking analyzer (NTA) with Western blotting experiment, to load miRNA34a mimics modified by hydrophobicity into exosomes by utilizing the method of co-incubation, and to fabricate exo-miRNA34a for loading efficiency observation under an inverted fluorescence microscope. Second, exo-miRNA34a will be inoculated into HN6 cells, so that the up-taking ability of HN6 cells on exosomes will be observed via laser scanning confocal microscope, and the proliferation changes of HN6 cells will be also detected coupled with CCK-8 experiment. Results: Small vesicles of 40-150 nm in size will be observed under a transmission electron microscopy in culture supernatant fluid extract of HEK293 cells, the distribution maxima of particle size of the vesicles is 97.9 nm based on the observation results of NTA, and surface protein, TSG101 and CD63, of exosomes can be detected by the transmission electron microscopy. However, plasmosin- Calnexin turns out negative after detection, which verifies the successful extraction of exosomes. Furthermore, the observation results of fluorescence microscope show that miRNA34a mimics modified by hydrophobicity can be loaded into exosomes effectively during co-incubation, drug loading capacity is 47% approximately. The particle size of exosomes increases after drug loading, and its integrity is not affected. Besides, laser scanning confocal microscope displays that exo-miRNA34a enters into HN6 cells, CCK-8 detection finds that miRNA34a can inhibit the proliferation of HN6 cells obviously (P＜0.05). Conclusion: MiRNA34a mediated by exosomes can significantly inhibit the proliferation of HN6 cells in oral squamous cell carcinoma.

Objective:To prepare berberine (BBR)-loaded chitosan/sodium β-glycerophosphate (CS/β-GP) thermosensitive hydrogels and investigate its basic physical, chemical properties and biological characteristics, to evaluate the potential role in the treatment of chronic periodontitis.Methods:CS/β-GP thermosensitive hydrogel was prepared by physical cross-linking, and BBR was added by embedding. According to the concentration of BBR, 25, 50, 100 μg/mLBBR/CS/β-GP thermosensitive hydrogel was prepared, and CS/β-GP thermosensitive hydrogel was used as a control to characterize each gel gelation time and pH, rheological properties, micromorphology and drug release rate. CCK8 method was used to detect the effects of hydrogels on the proliferation of periodontal ligament cells. The inhibition zone experiment was used to analyze each group’s antibacterial effect of on periodontal pathogens Porphyromonas gingivalis (P.gingivalis) and Fusobacterium nucleatum (F. nucleatum).Results:Each group of gels has good injectability at 4℃, and phase transition can be completed around body temperature. The phase transition temperature of CS/β-GP is 37.26℃. The hydrogels of each group can be gelled within 3 minutes at 37℃. The pH is in the neutral range. The results of gel electron microscopy show a uniform interconnected scaffold structure. BBR/CS/β-GP hydrogel can continuously release BBR slowly invitro, and the cumulative release rate can reach 70%~85% in 12 hours; co-cultivation of gels and periodontal ligament cells in each group shows good biological properties for 1, 3 and 5 days; each group of hydrogels can inhibit the growth of P.gingivalis and F. nucleatum.The greater the drug concentration, the stronger the antibacterial effect.Conclusions:BBR/CS/β-GP has better physical and chemical properties and biological characteristics, and has a certain bacteriostatic effect on periodontal pathogens. It can be used as a topical treatment option for chronic periodontitis.

Objective： To investigate the effects of silencing RECQ1 gene on the proliferation, migration, and epithelial mesenchymal transition (EMT) of oral squamous cell carcinoma. Methods： Human normal oral keratinocytes HOK cells and oral squamous cell carcinoma Cal27 cells were cultured in vitro, and the expressions of RECQ1 mRNA and protein were detected by qRT-PCR and Western Blot. RNAi was used in targeted silence RECQ1 gene in oral squamous cell carcinoma Cal27 cells. The experiment was divided into five groups: si-RECQ1 group (si-RECQ1 transfected into Cal27 cells), si-RECQ1+transforming growth factorβ1 (TGF-β1) group (added TGF-β1 after RECQ1 silence), si-NC group, TGF-β1 group and blank control group.CCK-8 method was used to detect the inhibition rate of cell proliferation in each group. Transwell assay was used to detect the invasive ability of cells. Cell migration ability was measured by cell scratch test. The expressions of RECQ1 and EMT-related proteins, including E-cadherin, N-cadherin and Vimentin, were detected by qRT-PCR and Western Blot. Results： Compared with HOK cells, the expression of RECQ1 mRNA and protein in Cal27 cells increased significantly (P＜0.05); compared with the blank control group and the si-NC group, the expressions of RECQ1 mRNA and protein in Cal27 cells in the si-RECQ1 group decreased significantly (P＜0.05); compared with blank control group, si-NC group and si-RECQ1+TGF-β1 group, the proliferation inhibition rate, E-cadherin mRNA and protein expression levels of Cal27 cells in si-RECQ1 group were increased significantly, while TGF-β1 group decreased significantly (P＜0.05), and Cal27 cell invasion number, migration distance, Vimentin, N-cadherin mRNA and protein expression levels in si-RECQ1 group decreased significantly, but TGF-β1 group increased significantly (P< 0.05). Conclusions： RECQ1 silencing can significantly inhibit the proliferation, migration and invasion of oral squamous cell carcinoma, which may be achieved by inhibiting the EMT process.

Objective:?To determine the metabolic differences between oral squamous cell carcinoma (OSCC) tissue and the paired adjacent noncancerous tissue by liquid chromatography-mass spectrometry. Methods:Fifteen pairs of OSCC tissues and matched normal tissues were analyzed by liquid chromatography-mass spectrometry. The differential metabolites were screened and determined by principal component analysis and partial least squares discriminant analysis.?Using MetaboAnalyst database to do the metabolic set enrichment analysis and metabolic pathway topological analysis of metabolites for determining abnormal metabolic pathways. Results:? Sixteen different metabolites can we got in OSCC tissues and adjacent tissues after analysis. By metabolic pathway analysis, there may be abnormal metabolic pathways such as sphingolipid metabolism, arginine and proline metabolism, pyrimidine metabolism and glycerophospholipid metabolism in OSCC patients. Conclusions:?There are obvious metabolic abnormalities in the tissues of OSCC and the paired adjacent noncancerous tissue, and abnormal metabolites with diagnostic value can be used as early diagnostic markers for OSCC patients.

Objective: To evaluate the effect of 2. 26 mg/mL fluoride varnish(Duraphat) applications on prevention for enamel demineralization during orthodontic treatment with fixed appliance．Methods: The study was based on a random clinical trial. In accordance with inclusion criteria，60 orthodontic patients were randomly selected into a test group and a control group. The patients in test group were used topical application of fluoride varnish at every 6 months during 24 months of orthodontic treatment and follow-up．Results: The enamel demineralization was significantly lower in test group than in control group during orthodontic treatment with fixed appliance(P＜0. 05).Conclusion: The effect of 2. 26 mg/mL fluoride varnish used for prevention of enamel demineralization during orthodontic treatment was remarkable．

Objective The primary approach to treating large cystic lesions is controversial. The aims of the present study were to assess the effects of decompression before enucleation for the treatment of large cystic lesions and to compare the rate of decrease of the lesions every three months so as to provide some guidance for clinicians on when to do the enucleations. Patients and Methods The study included 46 patients with large mandibular lesions (>3 cm). Decompression was used to release intraluminal pressure and decrease the volume of the lesion. Cone Bean computed tomography was applied to all patients every three months after decompression. Volumetric analysis was performed using software designed for 3-dimensional measurement of volumes. Other variables, such as age, gender, and rate of decrease, were recorded. Results: There were important differences in rates of decrease as time went by. Statistical analyses showed signi?cant differences in initial volume and duration of the lesions, while showed no signi?cant differences among groups for gender, and histologic lesion type (P > .05). Conclusion: Decompression of large cystic lesions could be useful for surgical interventions without complications.

Objective：To investigate whether follistatin-like protein 1 (FSTL1) can affect the osteogenic performance of mouse bone marrow mesenchymal stem cells (mBMSCs) in an inflammatory environment.?Methods:?mBMSCs were transfected with lentivirus to change the expression of FSTL1, and the proliferation activity of these cells was detected by CCK-8experiment. The mBMSCs transfected with lentivirus were stimulated with 10?ng/mL TNF-α, and then induced into osteoblasts.?Alkaline phosphatase (ALP) and alizarin red staining experiments were used to detect the changes of osteogenic ability of mBMSCs; ?the expression of collagen type Ⅰ collagen (Col1), osteopontin (Opn), osteocalcin (Ocn) was detected by real-time quantitative PCR.?Results:?Lentivirus transfection could effectively change the expression of FSTL1 gene in mBMSCs, and there was no significant difference in the proliferation activity of these cells (P＞0.05). In the inflammatory environment containing 10?ng/mL TNF-α, compared with the control group, mBMSCs overexpressing FSTL1 had lighter ALP and alizarin red staining results after osteogenic induction, and the expression of Col1, Opn, and Opn genes decreased (P＜0.05)；mBMSCs knockeddown FSTL1 had darker ALP and alizarin red staining results after osteogenic induction, and the expression of Col1, Opn, and Opn genes increased (P＜0.05). ?Conclusions:?FSTL1 promotes the inhibitory effect of inflammation on the osteogenic performance of mBMSCs, and inhibiting FSTL1 can effectively reduce the effect of inflammation on BMSCs.

Mesenchymal stem cells (MSCs) have been one of the most important cell sources in regenerative medicine on account of some outstanding advantages, e.g. low immunogenicity, strong anti - inflammatory and immune regulation. Nevertheless, the clinical development and application of MSCs are exceedingly restricted due to the characteristic of easy-aging in vitro culture. The issue on how to alleviate the senescence of MSCs is drawing much attention of both domestic and overseas researchers. This review summarizes MSCs aging signaling pathways and the process of related researches, to provide new insights to effective clinical applications of MSCs.