Reliability and reproducibility of CBCT assessment of mandibular changes before and after treatment for Class III growing patients - an easy and quick way for evaluation.

The objective of this study was to evaluate intraobserver reliability and inter-observer reproducibility of a 3-dimensional (3D) assessment method for mandibular changes of growing patients after orthodontic treatment for Class III malocclusion.Methods Cone-beam computed tomography (CBCT) scans were performed before and after orthodontic treatment for 27 patients. During the scan, the patient was positioned such that his/her mandibular plane was parallel to floor. Three observers independently worked on the DICOM data, reconstructed the pre- and post-treatment 3D models in software, selected the stable anatomical structures (basal bone area from the lingual surface of the symphysis to the distal aspect of the first molars) to guide the automated superimposition process. Then, each observer registered 14 anatomical landmarks on the virtual models, for three times after suitable interval, to generate 3 sets of coordinates; the mean was taken as the coordinates for that particular landmark. The intraobserver reliability and inter-observer reproducibility of the method were analyzed.Results The ICCs was > 0.90 for 25 (92.6%) of the intraobserver assessments. The precision of the measurement method was < 0.3 mm in 24 (88.9%) cases. The interobserver reproducibility errors were < 0.3 mm in 21 of the 27 cases.Conclusions The intraobserver reliability and inter-observer reproducibility of 3D assessment of mandibular changes using the virtual models were excellent.

NR5A2 as a potential target for exercise to improve metabolic syndrome.

BACKGROUND: Metabolic syndrome is a syndrome of a variety of metabolic disorders. Exercise is beneficial to the human body. However, the association of NR5A2 and exercise with metabolic syndrome remains unclear. METHODS: Download the GSE10540 and GSE12385 from GEO database. Bioinformatics analysis was used to screen the hub molecular of the metabolic syndrome. Forty 3-week-old C57BL/6J male mice were used in this study. The mean body weight was (17.5 ± 2.1) g. After 10 days of adaptive feeding, they were randomly divided into 4 groups according to the random number table method: Model + Exercise (n = 10), Model (n = 10), Model/NR5A2-OE (n = 10), Model/NR5A2-KO (n = 10). Western Blotting was performed to detect the expression of hub genes and signaling pathway. RESULTS: There were 349 DEGs in GSE10540 and 49 DEGs in GSE12385. 10 core genes were obtained. GO showed that differentially expressed genes were mainly enriched in vascular morphogenesis, contractile fiber fraction, chemotaxis, and MAPK cascade regulation. KEGG showed that MAPK signaling pathway was a significant section in the metabolic syndrome. PIK3R2, STRA8, FLT1, DMRT1, FGF22, NR5A2, and FLT were up-regulated and PRDM14, POU5F1, and KDR were down-regulated in metabolic syndrome after exercise. CONCLUSION: The expression of NR5A2 is down-regulated in metabolic syndrome, and exercise can increase the expression level of NR5A2. NR5A2 might be used as a potential target for exercise to improve metabolic syndrome.

CGF Membrane Promotes Periodontal Tissue Regeneration Mediated by hUCMSCs through Upregulating TAZ and Osteogenic Differentiation Genes.

Concentrated growth factor (CGF) membranes are widely used in basic and clinical research of soft and hard tissue regeneration, but its effect on periodontal tissue regeneration is less studied. This study explored the role of CGF membranes in periodontal tissue regeneration mediated by human umbilical cord mesenchymal stem cells (hUCMSCs). HUCMSCs and human periodontal ligament fibroblasts (HPLFs) were extracted and identified by microscope and flow cytometry. The effects of the extracted CGF membrane on cell viability, osteogenic differentiation ability, osteopontin (OPN) expression, alkaline phosphatase (ALP) content, and osteogenic differentiation-related genes (Runt-related transcription factor 2 (RUNX2); osteocalcin (OCN); ALP), Tafazzin (TAZ) expression, and nuclear transfer were examined by MTT assay, alizarin red staining, immunofluorescence, enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qRT-PCR), and Western blot. Rescue experiments were performed to examine the effects of TAZ transfection and cell coculture. In the identified hUCMSCs (positive expressions of CD29, CD44, CD146, and CD105), overexpressed TAZ (pc-TAZ) enhanced the promotive effect of CGF membrane on cell viability, cell cycle, mineralization, ALP content and expressions of OPN, TAZ and osteogenic differentiation-related genes, and nuclear transfer. However, silencing TAZ showed opposite effects. The coculture of hUCMSCs and HPLFs further promoted the basic biological functions of HPLFs by upregulating osteogenic differentiation-related genes and COL-1 but downregulated MMP1 expression. Pc-TAZ could enhance the effect of CGF membrane on promoting periodontal tissue regeneration. CGF membrane promoted periodontal tissue regeneration through upregulating TAZ and osteogenic differentiation-related genes.

Icariin contributes to healing skull defects in rabbit model.

OBJECTIVE: To investigate the efficacy of icariin for healing skull defects in rabbit models. METHODS: Thirty-six 3-month-old female New Zealand white rabbits, weighing 1.8-2.0 kg each, were randomly divided into either the control or experimental group. Skull defect models were constructed in both groups, with the experimental group receiving oral icariin afterwards. At 4, 8 and 12 postoperative weeks, the rabbits were euthanized, and X-rays and samples were taken. Tissue sections were stained using hematoxylin and eosin, followed by additional processing using histological and bone metrological methods for observing the rate and quality of the bone formation. RESULTS: Histologically, additional mature lamellar bone formed in the defect area in the experimental group compared with that of the control group. Bone metrological methods showed that the bone mass, trabecular bone width and number of osteoblasts in the experimental group were significantly higher than those of the control group (P < 0.01). The number of osteoclasts did not significantly differ between the two groups (P > 0.05). CONCLUSION: Icariin increased the bone mass and improved the condition of the defect area in the rabbit skulls.

Knockdown of microRNA-584 promotes dental pulp stem cells proliferation by targeting TAZ.

Proliferation of dental pulp stem cells (DPSCs) is crucial in tooth development and damage repairing, also includes its therapy application for tissue engineering. MicroRNAs (miRNAs) are key players in biological processes of DPSCs, and transcriptional co-activator with PDZ-binding motif (TAZ) also plays important roles in cell proliferation and differentiation, however, the roles of miR-584 and TAZ in DPSCs are not known. We found up-regulated miR-584 expression and down-regulated TAZ expression levels in aging dental pulp tissue compare to those in young dental pulp tissue. In proliferating DPSCs we demonstrated the decreased miR-584 expression and increased TAZ expression. miR-584 mimics suppressed DPSCs proliferation and migration, and significantly reduced TAZ production, whereas miR-584 inhibition exerted the converse effects. Knocking down of the TAZ in DPSCs had a similar effect as overexpression of miR-584. Furthermore, luciferase reporter assay demonstrated that miR-584 could directly bind to the TAZ mRNA 3'UTR to repress its translation. Overexpression of TAZ can partly rescue miR-584 mimic-mediated the inhibition of proliferation. Additionally, miR-584 inhibited cell proliferation and downregulated expression of cell cycle proteins by AKT signaling pathway. Together, we identified that miR-584 may be a key regulator in the proliferation of DPSCs by regulating TAZ expression via AKT signaling pathway. It would be a promising biomarker and therapeutic target for pulp disease.

Concentrated Growth Factor Promotes Dental Pulp Cells Proliferation and Mineralization and Facilitates Recovery of Dental Pulp Tissue.

BACKGROUND Dental pulp cells (DPCs) play vital roles in the recovery of dental pulp tissue. Concentrated growth factor (CGF) can promote proliferation and mineralization of various cells. However, the functions of CGF on DPCs and dental pulp tissue are unclear. The object of our study was to identify the roles of CGF in DPCs proliferation and mineralization in vitro and to assess the effects of CGF on direct pulp capping in vivo. MATERIAL AND METHODS We performed CCK-8 and Transwell assay to detect proliferation and migration activity of DPCs. Alizarin Red staining was performed to examine mineralized nodules. Alkaline phosphatase activity test was used to measure the mineralization capacity of DPCs. We assessed the odontogenic differentiation gene expression level by Western blot and qPCR. The effect of CGF on direct pulp capping in vivo were evaluated by radiography and histopathology. RESULTS CGF increased the number of proliferative and migratory DPCs. CGF enhanced DPCs mineralized nodules and improved the gene expression levels of DSPP, DMP-1, BSP, and ALP. CGF upregulated the protein levels of ALP, BMP2, SMAD5, Runx2, and p-Smad, and the effect could be partially reversed by Noggin. CGF promoted pulp recovery and kept its vitality in directly pulp capping. CONCLUSIONS CGF promotes DPCs proliferation and mineralization. It regulates the mineralization of DPCs via the BMP2/SMAD5/Runx2 signaling pathway. CGF can be used as the effective graft for direct pulp capping.

Effects of Biomineralization on Osseointegration of Pure Titanium Implants in the Mandible of Beagles.

PURPOSE: The purpose of this study was to investigate the effect of a biologically active dental implant surface (treated with sandblasting and acid etching [SLA] followed by immersion in simulated body fluid [SBF]) on osseointegration. MATERIALS AND METHODS: We randomly divided 9 healthy adult male beagles (aged 8 months; body weight, 12 kg) into 3 groups: machined, SLA, and SLA-biomineralization (SLA-Bio). Six pure titanium implants (diameter of 3.5 mm and length of 8 mm) were used in the mandible of each dog after observation of the surface morphology, as well as analysis of the composition of the surface elements by scanning electron microscopy-energy dispersive x-ray spectroscopy. At 4, 8, and 12 weeks after implantation, animals were euthanized to collect the mandibles so that we could perform the removal torque test to evaluate the implant stability in bone and histomorphometry to analyze the implant-bone osseointegration. RESULTS: Scanning electron microscopy results showed that uniformly distributed sponge-like structures were found on the SLA-treated surface and an apatite layer was observed on the SLA-SBF-treated surface (SLA-Bio group). In the energy dispersive x-ray spectroscopy analysis, the elements titanium, oxygen, carbon, calcium, and phosphorus were found on the surfaces of the SLA-Bio group, whereas titanium was the only element found in the other groups. The removal torque test showed that the peak removal torque values of the 3 groups increased gradually with the passage of time, and the peak removal torque values of the SLA-Bio group were significantly higher than those of the other groups (P < .01) at 4, 8, and 12 weeks after implantation. Histomorphometric analysis showed that osseointegration was being enabled more rapidly in the SLA-Bio group, as well as that the mineral apposition rate and percentage of bone-to-implant contact of the SLA-Bio group were higher than those of the remaining groups at 4, 8, and 12 weeks after implantation (P < .01). CONCLUSIONS: Treating titanium implants with SLA-SBF can improve osseointegration as well as increase the interfacial shear strength.

Effect of Xylosyltransferase-I Silencing on Implanting Growth of Salivary Pleomorphic Adenoma.

BACKGROUND Salivary pleomorphic adenoma is one of the most common salivary gland tumors. It has a relatively high tendency to recur and a high risk of malignant transformation. The present study aimed to study the effect of XT-I gene silencing on the implanting growth of salivary pleomorphic adenoma. MATERIAL AND METHODS Primary cultures of SPA cells and fibroblasts from the same patient were assessed. The adenovirus vector Ad-shRNA-XT-I was constructed and transfected into SPA cells. The expression of XT-I gene and XT-I protein was detected by real-time PCR and Western blot. The contents of proteoglycans were detected. The SPA cells transfected with Ad-shRNA-XT-I (group SPA-XT-I) and Ad-shRNA-HK (group SPA-HK), as well as without transfection (group SPA), were implanted into ADM scaffold with fibroblasts and then transferred into 18 BALB/C-nu nude mice for 3 months. RESULTS Primary cultures showed SPA cells were positive for human CK and S-100 protein and the fibroblasts were positive for human vimentin. The expressions of XT-I gene and protein were decreased by 51% and 51.31%, respectively. The content of proteoglycans was reduced by 48.45%. The results of the implanting growth in vitro and in vivo of nude mice indicated that no tumors grew in the SPA-XT-I group, whereas SPA grew in groups SPA-HK and SPA positive for human a-SMA, S-100 protein, and calponin. CONCLUSIONS XT-I gene silencing effectively inhibited the implanting growth of SPA.

Mineralization Induction of Gingival Fibroblasts and Construction of a Sandwich Tissue-Engineered Complex for Repairing Periodontal Defects.

BACKGROUND The ideal healing technique for periodontal tissue defects would involve the functional regeneration of the alveolar bone, cementum, and periodontal ligament, with new periodontal attachment formation. In this study, gingival fibroblasts were induced and a "sandwich" tissue-engineered complex (a tissue-engineered periodontal membrane between 2 tissue-engineered mineralized membranes) was constructed to repair periodontal defects. We evaluated the effects of gingival fibroblasts used as seed cells on the repair of periodontal defects and periodontal regeneration. MATERIAL AND METHODS Primitively cultured gingival fibroblasts were seeded bilaterally on Bio-Gide collagen membrane (a tissue-engineered periodontal membrane) or unilaterally on small intestinal submucosa segments, and their mineralization was induced. A tissue-engineered sandwich was constructed, comprising the tissue-engineered periodontal membrane flanked by 2 mineralized membranes. Periodontal defects in premolar regions of Beagles were repaired using the tissue-engineered sandwich or periodontal membranes. Periodontal reconstruction was compared to normal and trauma controls 10 or 20 days postoperatively. RESULTS Periodontal defects were completely repaired by the sandwich tissue-engineered complex, with intact new alveolar bone and cementum, and a new periodontal ligament, 10 days postoperatively. CONCLUSIONS The sandwich tissue-engineered complex can achieve ideal periodontal reconstruction rapidly.

Treatment of obstructive sleep apnoea-hypopnea syndrome by mandible advanced device reduced neuron apoptosis in frontal cortex of rabbits.

Objective: To investigate effects of mandible advanced device (MAD) therapy for obstructive sleep apnoea-hypopnea syndrome (OSAHS) on the neuron apoptosis and acetylcholine esterase activity in frontal cortex. Materials and methods: Thirty male New Zealand white rabbits were randomly divided into three groups (n = 10 in each group): group OSAHS, group MAD, and control group. Hydrophilic polyacrylamide gel was injected into soft palate of the animals to induce OSAHS in group OSAHS and group MAD. The group MAD animals wore MAD to relief the obstructiveness. The control group was not given any treatment. Computed tomography (CT) examination of the upper airway and polysomnography (PSG) recordings were performed in supine position. All rabbits were induced to sleep in a supine position for 4 to 6 hours every day and were observed for consecutive 8 weeks. The frontal cortices of three groups were dissected and the neuron apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and flow cytometry. Acetylcholine esterase (AchE) activity in the frontal cortex was measured by spectrophotometry. Results: The group OSAHS exhibited high neuron apoptosis rate and low AchE activity than those of group MAD and control group. The blood oxygen saturation was negatively correlated with neuronal apoptosis rate and positively correlated with AchE activity. Applying MAD in OSAHS animals significantly improve the neuronal damage and function deficits by apnoea-hypoxia caused by narrowed upper airway. Conclusion: This study provided evidence that MAD therapy for OSAHS can significantly decrease neuronal apoptosis and increase AchE activity in the frontal cortex.

Cytocompatibility and biologic characteristics of synthetic scaffold materials of rabbit acellular vascular matrix combining with human-like collagen I.

Scaffold material provides a three-dimensional growing environment for seed cells in the research field of tissue engineering. In the present study, rabbit arterial blood vessel cells were chemically removed with trypsin and Triton X-100 to prepare rabbit acellular vascular matrix scaffold material. Observation by He&Masson staining revealed that no cellular components or nuclei existed in the vascular intima and media after decellularization. Human-like collagen I was combined with acellular vascular matrix by freeze-drying to prepare an acellular vascular matrix-0.25% human-like collagen I scaffold to compensate for the extracellular matrix loss during the decellularization process. We next performed a series of experiments to test the water absorbing quality, biomechanics, pressure resistance, cytotoxicity, and ultra-micro structure of the acellular vascular matrix composite material and natural rabbit artery and found that the acellular vascular matrix-0.25% human-like collagen I material behaved similarly to natural rabbit artery. In conclusion, the acellular vascular matrix-0.25% human-like collagen I composite material provides a new approach and lays the foundation for novel scaffold material research into tissue engineering of blood vessels.

Study of composite vascular scaffold combining with differentiated VSMC- and VEC-like cells in vitro and in vivo.

Although research into the tissue engineering of vessels has proceeded at a tremendous pace, many deficiencies still need to be resolved. A well-adopted constructed vessel requires both functional and structural properties to stimulate the native vessel and resist stress and tension in vivo. In the present study, we developed a novel three-layer composite vascular scaffold consisting of differentiated vascular smooth muscle cell-, vascular endothelial cell-like cells, and a rabbit acellular vascular matrix (ACVM)-0.25% HLC-I scaffold. HE staining, immunohistochemical assays, immunofluorescence assays (IFAs), and scanning electron microscopy were performed to monitor the growth status of cells on the scaffold material in vitro. After the vascular endothelial cell -vascular smooth muscle cell-scaffold was implanted into nude mice for three, six, and nine weeks, samples were harvested from the implanted mice and observed visually or by HE staining and IFAs for cell viability and morphology. Additionally, burst pressure resistance experiments were used to assess the maximal pressure that the engineered vessel could resist. We found that the engineered vascular endothelial cell-vascular smooth muscle cell-scaffold vessel possessed favorable biocompatibility and considerable strength, matching native vessels in vivo and in vitro, and may be significant in the future clinical implantation of tissue-engineered vasculature.

Effects of TAZ on human dental pulp stem cell proliferation and migration.

Transcriptional coactivator with PDZ­binding motif (TAZ) acts as the key downstream regulatory target in the Hippo signaling pathway. TAZ overexpression has been reported to promote cellular proliferation and induce epithelial­mesenchymal transition in human mammary epithelial cells. However, the effects of TAZ in the regulation of human dental pulp stem cell (hDPSC) proliferation and migration, as well as the molecular mechanisms underlying its actions, remain to be elucidated. The present study demonstrated that TAZ was expressed in hDPSCs. TAZ silencing, following hDPSC transfection with TAZ­specific small interfering (si)RNA (siTAZ), inhibited cellular proliferation and migration in vitro. These effects appeared to be associated with the downregulation of connecting tissue growth factor (CTGF) and cysteine­rich angiogenic inducer (Cyr) 61 expression. Further investigation of the mechanisms underlying the actions of TAZ in hDPSCs revealed that TAZ silencing suppressed CTGF and Cyr61 expression by interfering with transforming growth factor (TGF)­ß signaling pathways. The present results suggested that TAZ may be implicated in the proliferation and migration of hDPSCs, through the modulation of CTGF and Cyr61 expression via a TGF­ß­dependent signaling pathway.

Morphological Observation on Critical-Sized Cranial Defect Repaired by Icariin and Autologous Concentrate Growth Factors in Rabbits.

BACKGROUND Morphological changes repaired by icariin and autologous concentrate growth factors (ACGF) in critical-sized cranial defect were observed and their promoting effects were investigated. MATERIAL AND METHODS Seventy-two New Zealand white rabbits weighing 1.8~2.0kg were used to build a critical-sized cranial defect model and were randomly divided into 3 groups. X-ray, HE staining, general and histological observation, and immunohistochemistry were used to describe the changes caused by normal saline, icariin, and ACGF. RESULTS Cranial defects were covered with newly formed bone tissue at the 12th week in icariin and ACGF groups, with red color, hard surface, and no obvious boundary. Densities were the same in 2 groups at 4 timepoints. HE staining showed defects filled with a large amount of fibrous connective tissue, thick collagen fibers, and abundant osteoclasts. No new bone matrix appeared in any of the 3 groups. Trabecular area, trabeculae width, and osteoblast number in 2 groups were more than that of the control group, and osteoclast number was lower. However, osteoclast number among the 3 groups at the 12th week had no significant difference, which was the same with 4 indicators between the icariin and ACGF groups. From the 4th to 12th week, regenerated cartilage was formed and showed positive reaction with BMP-2 and TGFß1 from primary bone, which also was demonstrated by granulation tissue and uniform dyeing. CONCLUSIONS ACGF and icariin both can increase new bone quantity and improve bone quality, which can also promote healing. The effects and mechanisms of icariin and ACGF on the expression of gene are not exactly the same.

Fundamental research on the action mechanism of the 800 nm semiconductor laser on skin blackheads and coarse pores.

The aim of the study was to determine the mechanism of action of the 800 nm semiconductor laser on skin blackheads and coarse pores. A total of 24 healthy purebred short-haired male guinea pigs, weighing 350-400 g, were selected and smeared with 0.5 ml coal tar suspension evenly by injector once daily. Treatment was continued for 14 days to form an experimental area of 8×3 cm on the back of the guinea pigs. The animals were divided into the following groups: Normal control group (NC), low-dose laser treatment group (L-LS), high-dose laser treatment group (H-LS), and Q-switched Nd:YAG treatment group (QC). Samples were extracted 1, 7 and 14 days after surgery and hematoxylin and eosin staining was used to identify the following: Epidermis, dermis, sebaceous gland change and hair follicle damage; the expression of proliferating cell nuclear antigen (PCNA) of sebaceous gland cells using immunohistochemistry; sebaceous gland cell apoptosis using TUNEL; and the protein expression of caspase-3, Bax and Bcl-2 using western blot analysis. With the extension of time, we observed inflammatory cell infiltration, an increase in hair follicle distortion and necrosis of the surrounding hair follicles. The expression levels of PCNA of the L-LS, H-LS and QC groups decreased with time. Regarding the respective time points, the NC group was highest, the L-LS and H-LS groups were next highest and the H-LS group was lowest. The difference was statistically significant (P<0.05). The apoptotic rate of the L-LS, H-LS and QC groups increased with time. With regard to the respective time points, the NC group was lowest, the L-LS and QC groups were next lowest and the H-LS group was highest. The difference was statistically significant (P<0.05). The protein expression of caspase-3, Bax and Bcl-2 of the L-LS, H-LS and QC groups increased with time. Regarding the respective time points, caspase-3 and Bax protein expression of the NC group was lowest, the L-LS and QC groups were next lowest and the H-LS group was highest. Bcl-2 protein expression of the NC group was highest, protein expression of the NC group was next highest and the H-LS group was lowest. The difference was statistically significant (P<0.05). In conclusion, the low-dose 800 nm semiconductor laser is an effective treatment on skin blackheads and coarse pores, and promotes hair follicle cell apoptosis without reducing the expression of PCNA.

The clinicopathologic relevance of RECK gene polymorphisms in ameloblastoma.

OBJECTIVE: To investigate the relationship between RECK gene polymorphisms and the clinicopathologic features of ameloblastoma. DESIGN: Normal gingival mucosa specimens were obtained from 10 healthy volunteers. Ameloblastomas were surgically removed from 30 patients and part of the tumor specimens were used to detect RECK gene polymorphisms by using PCR-single-strand conformation polymorphism (PCR-SSCP) and DNA sequencing analysis. Expression of RECK and MMP-9 protein was measured using western blot. RESULTS: The overall SNP rate was 46.7% (14/30). Four polymorphisms were detected in exon 9, 11, 13, 15 of the RECK gene: two synonymous (P520P and R625R) and two missense SNPs (V275I and I395V). RECK protein expression in specimens with minor RECK SNPs was lower than that in specimens without RECK SNPs (P<0.05), and, RECK protein expression in specimens with and without RECK SNPs was lower than that in the normal gingiva specimens (P<0.05). MMP-9 protein expression in specimens with minor RECK SNPs was higher than that in specimens without RECK SNPs (P<0.05), and MMP-9 protein expression in specimens with and without RECK SNPs was higher than that in normal gingiva specimens (P<0.05). RECK gene polymorphisms were closely associated with active proliferation, capsular invasion, and clinical recurrence of ameloblastoma. CONCLUSION: The rs16932912(G/A) SNP in the RECK gene was closely associated with active proliferation, capsular invasion, and clinical recurrence of ameloblastoma. RECK protein expression was closely associated with the presence of the rs16932912(G/A) SNP.

Induced differentiation of human gingival fibroblasts into VSMC-like cells.

Vascular smooth muscle cells (VSMCs) are major component of the vascular wall, and they play an essential role in maintaining the basic physiological function and stable structure of the vascular wall. In the present study, human gingival fibroblasts (HGFs) were cultured and induced into VSMC-like cells in vitro to confirm that HGFs with properties of stem cells have the potential for differentiation. The epithelium isolated from patients was extracted from normal human gingiva consisting of epithelium and connective tissue. HGFs were first identified by morphological examination, as well as specific gene and protein expression, and then induced by 10ng/mL PDGF-BB combined with 2ng/mL of TGF-ß1 for 28 days. After induction, ICS data indicated that induced VSMC-like cells were positive for α-SMA and SM-MHC, and IFA data showed that induced cells were positive for SM22α and Cnn1. RT-PCR results demonstrated that α-SMA and SM-MHC mRNA were specifically expressed, and myofilament-like structures also appeared in induced cells. In conclusion, the data indicated that HGFs could differentiate to VSMC-like cells with typical VSMC morphologic, ultrastructural, and immunological characteristics via induction with PDGF-BB and TGF-ß1.

ClC-3 Promotes Osteogenic Differentiation in MC3T3-E1 Cell After Dynamic Compression.

ClC-3 chloride channel has been proved to have a relationship with the expression of osteogenic markers during osteogenesis, persistent static compression can upregulate the expression of ClC-3 and regulate osteodifferentiation in osteoblasts. However, there was no study about the relationship between the expression of ClC-3 and osteodifferentiation after dynamic compression. In this study, we applied dynamic compression on MC3T3-E1 cells to detect the expression of ClC-3, runt-related transcription factor 2 (Runx2), bone morphogenic protein-2 (BMP-2), osteopontin (OPN), nuclear-associated antigen Ki67 (Ki67), and proliferating cell nuclear antigen (PCNA) in biopress system, then we investigated the expression of these genes after dynamic compression with Chlorotoxin (specific ClC-3 chloride channel inhibitor) added. Under transmission electron microscopy, there were more cell surface protrusions, rough surfaced endoplasmic reticulum, mitochondria, Golgi apparatus, abundant glycogen, and lysosomes scattered in the cytoplasm in MC3T3-E1 cells after dynamic compression. The nucleolus was more obvious. We found that ClC-3 was significantly up-regulated after dynamic compression. The compressive force also up-regulated Runx2, BMP-2, and OPN after dynamic compression for 2, 4 and 8 h. The proliferation gene Ki67 and PCNA did not show significantly change after dynamic compression for 8 h. Chlorotoxin did not change the expression of ClC-3 but reduced the expression of Runx2, BMP-2, and OPN after dynamic compression compared with the group without Cltx added. The data from the current study suggested that ClC-3 may promotes osteogenic differentiation in MC3T3-E1 cell after dynamic compression. J. Cell. Biochem. 118: 1606-1613, 2017. © 2016 Wiley Periodicals, Inc.

The relationship between proteoglycan inhibition via xylosyltransferase II silencing and the implantation of salivary pleomorphic adenoma.

OBJECTIVE: To study the relationship between proteoglycan inhibition and the implantation of salivary pleomorphic adenoma (SPA). METHODS: SPA fresh tissue was primitively cultured and identified. The Ad-shRNA-XT-II adenovirus vector was constructed and transfected into SPA cells to silence the XT-II gene. The expression of the XT-II gene and protein was detected using real-time PCR and Western blotting, respectively. The proteoglycan content of the cells was determined 48 h after transfection. The invasion and migration of SPA cells were observed using Matrigel invasion and wound-healing assays. Fibroblasts from the tumour capsule of the same patient were obtained, cultured and seeded onto an acellular dermal matrix (ADM) scaffold. Tumour cells were seeded onto the scaffold with the fibroblasts and then transferred to BALB/C-nu nude mice and allowed to grow in vivo for 3 months. RESULTS: The SPA cells cultures were positive for human calponin, S-100 protein, α-SMA and CK. XT-II gene and protein expression was decreased by 42.72% and 34%, respectively. The proteoglycan content was downregulated by 41.15%. XT-II gene silencing decreased the invasion and migration abilities of SPA cells. The assessment of SPA growth in nude mice indicated an absence of tumour growth in the SPA-XT-II group (in which the XT-II gene was silenced), whereas SPA growth was observed in the other two groups (in which the XT-II gene was not silenced), and the tumour tissue was positive for the human S-100 protein, α-SMA and CK8&18. CONCLUSION: Proteoglycan inhibition induced via XT-II gene silencing inhibited the implantation of SPA.