【Objective】 To sequence P1 gene(184～1946 bp) includ ing SBR gene in expressional plasmid of transgenic plant. 【Methods】 pROP1 plas mid was extracted from E. coli DH5α, P1 gene was sequenced by PCR sequence kit and DNA autosequencer and compared with surface protein gene of Streptococc us mutans in GenBank. 【Results】 616 base pairs from 5′ end and 466 base pairs from 3′ end were sequenced, its accuracy is up to 98%. 【Conclusion】 The seq uence of P1 gene's 5′ end and 3′ end is consistent with DNA sequence of SBR re gion of Streptococcus mutans surface protein pac gene and may provide useful in formations to construct the transgenic plant anticaries vaccine.

Objective:To culture human alveolar osteoblasts and study its osteogenic characteristics in vitro.Methods:Human alveolar bone was obtained from human donor and cultured in explants.Cells migrating from explants were observed by inverted phase-contrast microscope and cell proliferation was detected by MTT assay.After cultured in conditional medium containing dexamethasone,?-glycerphosphoric sodium and ascorbic acid,cells were tested by ALP assay,Alizarin red assay and von Kossa assay,3H-proline labeling with collagenase digestion method.Results:The alveolar osteoblasts migrated from explants after 5 d culture.Cells could be passaged after 14-16 d culture.ALP values were obviously positive and cells showed positive reaction by Alizarin red assay and von Kossa assay in conditional medium.The cultured human alveolar osteoblasts secreted type I Collagen.Conclusion:The human alveolar osteoblasts cultured in this experiment grow well,and the morphological and biological characteristics of the culture cells are similar to those of osteoblasts.

Objective To investigate the the multi-directional differentiation potential between pluripotent of human gingival fibroblasts (HGFs) and human periodontal ligament cells (HPDLCs). Methods HPDLCs and HGFs were obtained from the primary culture. HPDLCs and HGFs at 3rd-4th passage were cultured in osteogenic, adipogenic or chondrogenic me-dium. Cells without differentiation were taken as control. Alizarin red, Alcian blue and oil red O staining were performed to detect osteogenic differentiation, chondrogenic and adipogenic differentiation in vitro, respectively. Reverse transcription polymerase chain reaction (RT-PCR) was applied to examine the expression of osteocalcin (OCN), runt-related transcription factor 2 (RUNX2) and collagen 1 (Col 1), peroxisome proliferator-activated receptor gamma 2 (PPARγ2) and collagen 10 (Col 10). Results HPDLCs and HGFs cultured in osteogenic medium showed massive calicium nodulus at day 28, but HP-DLCs formed more calicium nodulus than those of HGFs. The expressions of OCN, RUNX2 and Col 1 were significantly high-er in HPDLCs than those in HGFs (P<0.05). In chondrogenic medium both cells were found blue deposit at day 14, and the expression of Col 10 was significantly higher in HGFs than that of HPDLCs (P<0.01). Furthermore, in adipogenic medium HGFs showed more lipid-filled droplets stained with oil red O than HPDLCs at day 21. The expression of PPARγ2 was sig-nificantly higher in HGFs than that of HPDLCs (P<0.01). Conclusion HPDLCs has the better potency of osteogenic differ-etiation than HGFs, however, HGFs has the better potency of adipogenic and chondrogenic differentiation.

Objective To study the correlation between the expression of activator protein-1 (c-Fos/c-Jun) mRNA and gingival inflammation,so as to discuss the pathogenesis of periodontitis.Methods The gingival tissues were divided into three groups according to the gingival index (GI),including GI=0 group (control group,14 cases),GI=1 group (15 cases) and GI=2 group (11 cases).The total RNA in each gingival tissue was extracted,and cDNA was synthesized by reverse transcription synthesis.The expressions of c-Fos and c-Jun mRNA in healthy gingival tissue (GI=0 group) were detected by reverse transcription-polymerase chain reaction.The levels of c-Fos and c-Jun mRNA in all the groups were detected by real-time quantitative PCR.Results Both c-Fos and c-Jun mRNA was expressed in healthy gingival tissues.The levels of c-Fos and c-Jun mRNA in GI=1 group was 15.58±9.19 and 3.47± 1.77,respectively,which was significantly higher than 1.31±1.03 and 1.32±0.94 in GI=0 group,and the differences were statistically significant (all P＜0.05).The level of c-Fos mRNA in GI=2 group was 3.01±1.48,which was lower than that in GI=1 group (P＜0.05) and higher than that in GI=0 group (P＜0.05).The level of c-Jun mRNA in GI=2 group was 1.48±0.65,which was lower than that in GI=1 group,and had no significant difference with GI=0 group (P＞ 0.05).Conclusions Activator protein-1 (c-Fos/c-Jun) is associated with the degree of gingival inflammation,suggesting that it is involved in the occurrence and development of gingival inflammation.

Objective To observe the effects of basic fibroblast growth factor (bFGF) on osteogenic differentiation abili?ty and cell proliferation of human gingival fibroblasts (HGFs), and to explore the role of bFGF on the process of osteogenic differencitiaion in vitro. Methods HGFs were cultured in vitro until the 3rd passage when they were divided into four groups:normal medium as group 1, normal medium with 10μg/L bFGF as group 2, osteogenic medium as group 3 and osteo?genic medium with 10μg/L bFGF as group 4. MTT assay was used to evaluate the proliferation of HGFs. Alkaline phospha?tase (ALP) staining and Alizarin red staining were applied to investigate osteogenic potential of HGFs under different culture conditions. Results bFGF at concentration of 10 μg/L could increase HGFs proliferation in both normal and osteogenic medium (P<0.01). HGFs could be induced towards osteogenic differentiation and form mineralized nodule in osteogenic me?dium. However, 10μg/L bFGF had no effects on ALP activity and mineralized nodule formation of HGFs during osteogenic differentiation. Conclusion bFGF could promote the proliferation of HGFs but show no effects on osteogenic differentiation of HGFs at concentration of 10μg/L.

Objective To study the HPLC fingerprints of isoflavones from different medicinal plants of G.max and establish a sensitive and specific method for controlling the quality of the soybean isoflavone.Methods The gradient elution mode was applied in chromatographic separation.A C_(18) column(250 mm?4.6 mm,5 ?m) was used with the mobile phase: ice acetic acid water solution(pH value 3.2)-methanol,flow rate: 0.6 mL/min,detecting wavelength: 261 nm,and the column temperature: room temperature.All 25 samples collected from different species were determined.The clustering analysis and the software of similarity analysis were applied for datum analysis.Results This method had a good repeatability and reproducibility.The ratio of peaks′ area from distinct samples were different.Conclusion The method can show the difference of chemical compositions among species completely and can be used as a quality control method for soybean.

Objective To investigate the pluripotency of human gingival fibroblasts (hGFs), and provide a novel cell source for tissue engineering. Methods With informed consent from volunteers, fresh and healthy gingiva were collected. The hGFs were obtained from the gingiva by tissue culture. The third passage of hGFs was cultured in osteogenic medium, chondrogenic medium and adipogenic medium. Cells without differentiation were taken as control. Cells were examined by al?kaline phosphatase (ALP) staining, Alizarin red staining, Alcian blue staining and oil red O staining for detecting of the abili?ty of differentiation pluripotency. Real-time polymerase chain reaction was applied to examine the expression of osteogenic marker genes ALP, runt-related transcript factor 2 (Runx2), chondrogenic marker aggrecan (AGR) and adipogenic marker peroxisome proliferator-activated receptor gamma 2 (PPARγ2). Results The hGFs cultured in osteogenic medium showed massive violet deposit at day 7 and calcium nodulus at day 28, meanwhile, the expressions of ALP and Runx2 were higher than those of control (P<0.01). In chondrogenic group cells were found blue deposit at day 14. In adipogenic group lipid-filled droplets stained with oil red O were found in cells at day 14. However, hGFs in control group had no any positive stain?ing. Furthermore, expressions of AGR and PPARγ2 were significantly higher than those of control (P<0.01). Conclusion Human gingival fibroblasts have the pluripotency of osteogenic, adipogenic and chondrogenic differentiation.

OBJECTIVETo investigate the influence of high glucose on Porphyromonasgingivalis (Pg) lipopolysaccharide (LPS) stimulating human gingival fibroblasts (HGF) to secret the cytokines.

METHODSHGF were obtained from the primary culture of the tissue explants. Cells were divided into four groups, low glucose (5.5 mmol/L) + 1 mg/L Pg LPS (group A);low glucose + 10 mg/L Pg LPS (group B); high glucose (25 mmol/L) +1 mg/L Pg LPS(group C); high glucose+10 mg/L Pg LPS (group D). The levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in cell supernatants were detected by enzyme- linked immunosorbent assay at 6 h and 12 h. The expressions of toll-like receptor 2, 4 (TLR-2, 4) were examined by real-time polymerase chain reaction. After pretreatment with anti-TLR2 and anti-TLR4 monoclonal antibody in HGF, TNF-α and L-1β levels were detected.

RESULTSTNF-α concentration increased obviously in high glucose 6 h and 12 h after Pg LPS stimulation (P < 0.01). IL-1β secretion increased (P < 0.01). Meanwhile, TLR2, 4 mRNA expression increased, especially in high glucose+10 mg/L Pg LPS (P < 0.01). After inhibition of the TLR2, 4 in high glucose + 10 mg/L Pg LPS respectively, TNF-α level [(297.16±11.49), (390.01±12.81) ng/L] decreased (F = 166.02, P < 0.01), and IL-1β level [(49.90±4.08), (99.35±5.01) ng/L] also decreased (F = 153.51, P < 0.01).

CONCLUSIONSHigh glucose may promote Pg LPS to stimulate the secretion of TNF-α and IL-1β through regulating TLR2, 4 expression, which suggests that the elevating blood glucose precipitate in aggravating the process of periodontal disease.

Antibodies, Monoclonal ; Drug Synergism ; Fibroblasts ; drug effects ; metabolism ; Glucose ; administration & dosage ; pharmacology ; Humans ; Interleukin-1beta ; metabolism ; Periodontal Diseases ; etiology ; Polysaccharides, Bacterial ; toxicity ; Porphyromonas gingivalis ; chemistry ; Time Factors ; Toll-Like Receptor 2 ; immunology ; metabolism ; Toll-Like Receptor 4 ; immunology ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

Objective: To detect and analyze the differential expression profile of long non-coding RNA (lncRNA) in aggressive periodontitis (AgP) and healthy gingival tissues, in order to explore the role of lncRNA in AgP. Methods: After the informed consents were obtained, gingival tissues from AgP patients (n=40) and healthy volunteers (n=40) were collected in Department of Periodontology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine (from Mar. 2012 to Aug. 2012) and Department of Periodontology, Hospital of Stomatology, Tianjin Medical University (from Oct. 2016 to Apr. 2017). The differential expression of lncRNA of tissues from AgP patients (n=20) and healthy volunteers (n=20) were examined via microarray assay. Bioinformatics was applied to analyze the expression data of lncRNA and correlative mRNA. Two lncRNAs (lncRNA-TNFRSF13C and lncRNA-API5) were chosen to verify the microarray results by using real time quantitative polymerase chain reaction (PCR) in the other gingival tissues. Results: Compared with the result of healthy gingival tissues, totally 8 632 lncRNAs were differentially expressed in tissues from AgP patients. From these data, 1 986 lncRNAs were significantly upregulated while 6 646 lncRNAs were downregulated, amongst which 48 lncRNAs were upregulated (>10 times) (P<0.05), 14 lncRNAs were downregulated (>10 times) (P<0.05). Furthermore, totally 5 519 correlative mRNAs were differentially expressed, amongst which 1 676 mRNAs were upregulated (≥2 times, P<0.05) and 3 843 mRNAs were downregulated≤0.5 (P<0.05). The selected lncRNA-TNFRSF13C and lncRNA-API5 were up-regulated in AgP (P<0.05), which confirmed the results of microarray. From bioinformatics, differential expression lncRNAs were in association with many signal pathways including toll-like receptor signaling pathway, cell cycle and apoptosis pathway, and tumor necrosis factor receptor superfamily pathway. Conclusions LncRNA may be involved in the pathogenesis of AgP through various pathways, which need to be further explored.

Polymeric hydrogels have been widely researched as drug delivery systems, wound dressings and tissue engineering scaffolds due to their unique properties such as good biocompatibility, shaping ability and similar properties to extracellular matrix. However, further development of conventional hydrogels for biomedical applications is still limited by their poor mechanical properties and self-healing properties. Currently, nanocomposite hydrogels with excellent properties and customized functions can be obtained by introducing nanoparticles into their network, and different types of nanoparticles, including carbon-based, polymer-based, inorganic-based and metal-based nanoparticle, are commonly used. Nanocomposite hydrogels incorporated with polymeric micelles can not only enhance the mechanical properties, self-healing properties and chemical properties of hydrogels, but also improve the
Biocompatible Materials ; Hydrogels ; Micelles ; Nanocomposites ; Polymers