Novel Protein Tyrosine Phosphatase 1B Inhibitor-Geranylated Flavonoid from Mulberry Leaves Ameliorates Insulin Resistance.

Mulberry leaf is a common vegetable with a variety of beneficial effects, such as hypoglycemic activity. However, the underlying mechanism of its hypoglycemic effect have not been fully revealed. In this study, two flavonoid derivatives were isolated from mulberry leaves, a new geranylated flavonoid compound (1) and its structural analogue (2). The structures of compounds 1 and 2 were elucidated using spectroscopic analysis. To study the potential hypoglycemic properties of these compounds, their regulatory effects on protein tyrosine phosphatase 1B (PTP1B) were investigated. In comparison to oleanolic acid, compounds 1 and 2 showed significant inhibitory activities (IC50 = 4.53 ± 0.31 and 10.53 ± 1.76 µM) against PTP1B, the positive control (IC50 = 7.94 ± 0.76 µM). Molecular docking predicted the binding sites of compound 1 to PTP1B. In insulin-resistance HepG2 cell, compound 1 promoted glucose consumption in a dose-dependent manner. Furthermore, western blot and polymerase chain reaction analyses indicated that compound 1 might regulate glucose consumption through the PTP1B/IRS/PI3K/AKT pathway. In conclusion, geranylated flavonoids in mulberry leaves inhibite PTP1B and increase the glucose consumption in insulin-resistant cells. These findings provide an important basis for the use of mulberry leaf flavonoids as a dietary supplement to regulate glucose metabolism.

[Ammonia Oxidation in a Neutral Purple Soil Measured by the 13C-DNA-SIP Method].

Increasing evidence suggests that ammonia oxidation in acidic soils is primarily catalyzed by ammonia-oxidizing archaea (AOA), while ammonia-oxidizing bacteria (AOB) drive ammonia oxidation in neutral and alkaline soils in which AOA overwhelmingly outnumber AOB. Therefore, neutral purple soil with a pH of 7.2 was selected to study the composition of the active ammoxidation microbial community with a stable isotope nucleic acid probe technique combined with cloning sequencing. Results showed that the nitrification rate was 9.68 mg·(kg·d)-1, and AOA and AOB were abundant in neutral purple soils. By using DNA-based stable isotope probing (SIP), we gathered strong evidence of archaeal ammonia oxidation by AOA and AOB. Phylogenetic analysis indicated that the Nitrosospira Cluster 3a.1 AOB was dominant in terms of quantity at 0 days, and the Nitrosospira Cluster 3a.2 only accounted for a small part. After 56 days of cultivation, the Nitrosospira Cluster 3a.2 replaced the Nitrosospira Cluster 3a.1 as the active AOB that dominated ammonia oxidation. The AOA that predominated quantitatively at day 0 was Nitrososphaera Subcluster 9, but after cultivation this became Nitrososphaera Subcluster 3.2/3.3. Thus, the community structure of AOA and AOB changed. Active autotrophic nitrification was found in this neutral purple soil. Sequencing analysis of the 13C-labeled DNA provided robust evidence that both archaea and bacteria played important roles in the nitrification and not all ammonia oxidizers in native soil were active in the nitrification. Phylogenetic analysis clearly showed that the dominant active archaea and bacteria during the incubation were affiliated with Nitrososphaera Subcluster 3.2/3.3 within the soil group 1.1b lineage and Nitrosospira Cluster 3a.2, respectively, which were different from the dominant ammonia oxidizers at the beginning of the incubation. These results suggest that the community structure of ammonia oxidizers can shift quickly upon changes in the substrate availability in soils.

[Effect of Naringin on proliferation and osteogenic differentiation of bone marrow stromal cells in vitro].

PURPOSE: To investigate the differentiation and osteogenic activity of Naringin-induced bone marrow stromal cells (BMSCs) in dogs. METHODS: BMSCs were separated and cultured in vitro and identified by FCM. Then different concentration of Naringin (1×10⁻5, 1×10⁻6, 1×10⁻7, 1×10⁻8 and 1×10⁻9 mol/L) were added to cell culture media to induce BMSCs. The effect of Naringin on BMSCs was evaluated respectively by CCK-8 method and measuring the activity of alkaline phosphatase (ALP). The formation of nodules of calcium was detected by von Kossa staining. The data was analyzed with SPSS20.0 software package. RESULTS: The result of cell-surface marker displayed that the expression of CD34 and CD45 were negative while the expression of CD90 was positive. The values were 0.126%, 0.075% and 95.4%, respectively. Naringin could obviously promote cell proliferation, which exhibited the best effect on proliferation and osteogenic differentiation at concentration of 10-6 mol/L. Calcium nodule (von Kossa) staining was positive. CONCLUSIONS: Naringin-induced bone marrow stromal cells can be differentiated into osteoblasts. Naringin at the concentration of 10-6 mol/L can enhance the proliferation and osteogenic differentiation of BMSCs.

[Clinical application of modified casting post with zirconia crown in restoration of anterior teeth].

PURPOSE: To evaluate the clinical effect of modified casting post with zirconia crown in restoration of anterior teeth. METHODS: A total of 45 anterior teeth (32 patients) were treated with modified gold-alloy post-cores with zirconia crowns. All roots were greatly tilted to the labial side. Marginal integrity, anatomic form, surface, color and the gingival situation after restorations were assessed at 6 and 12 months with the California Dental Association (CDA) quality assessment system. RESULTS: The clinical outcome of 44 zirconia crowns achieved rank A (97.8%), while the gingival situation in one patient (2.2%) reached rank B. CONCLUSIONS: Modified gold-alloy post-core with zirconia crown contributes to the aesthetic effect of the anterior teeth which need to change the angle.

Catalytic enantioselective addition of terminal 1,3-diynes to N-sulfonyl aldimines: access to chiral diynylated carbinamines.

An efficient method for the asymmetric synthesis of chiral diynylated carbinamines is described. The direct catalytic enantioselective addition of terminal 1,3-diynes to N-sulfonyl aldimines proceeded smoothly under mild reaction conditions to produce diynylated carbinamines in up to 98% yield and 99% ee.

Catalytic enantioselective addition of terminal 1,3-diynes to aromatic ketones: facile access to chiral nonracemic tertiary alcohols.

An efficient, catalytic, and enantioselective 1,2-addition of terminal 1,3-diynes to aromatic ketones was realized in the presence of 10 mol% of a Cu(OTf)(2)-hydroxycamphor-sulfonamide complex, affording chiral tertiary alcohols in up to 94% yield and 90% ee.

[Localization of enamelin in developing rat incisor by immunofluorescence].

PURPOSE: To investigate the expression of enamelin in developing rat incisor. METHODS: Immunofluorescence staining for the specimens of developing rat incisor was used to localize the expression of enamelin. RESULTS: Enamelin was not detected at the proliferation phase, and early differentiation phase in preameloblasts. At the advanced differentiation phase, enamelin expressed weakly in ameloblasts and odotoblasts. At the early maturation phase enamelin was detected in ameloblasts and odotoblasts. Enamelin expressed strongly in ameloblasts and enamel matrix but strongly in odotoblasts at the intermediate maturation phase and weakly at the advanced maturation phase. Enamelin was not found in the other stages and position. CONCLUSION: Enamelin,involved in enamel and dentin matrix formation, may play an important role in signal transduction of matrix formation. Supported by Project of the "Tenth-Five Year" Science Support Plans and Project "211" of Tongji University (Grant No.1504102011).

[Experimental study of subcutaneous osteogenesis by bone marrow stromal cells with A-PCPC in Beagle dogs].

PURPOSE: To study the osteogenic capability of the construct combined dog's bone stromal cells with active porous calcium phosphate cement (A-PCPC) in nude mice in vivo. METHODS: Isolated bone marrow stromal cells (BMSCs) were expanded and osteogenically induced in vitro. Their osteogenic phenotype was evaluated by cytochemistry. The tissue engineering complex was constructed with BMSCs/A-PCPC in vitro. After SEM scanning, the complex of BMSCs/A-PCPC was implanted into the subcutaneous tissue of the nude mice as experimental group, and A-PCPC as control group. The engineered bone was harvested 2,4,8weeks post-implantation and processed for HE staining, then evaluated by histology and histomorphometry. RESULTS: Cytochemistry showed alkaline phosphate activity, Von Kossa staining proved the formation of mineralization nodules. Scanning electron microscopy showed the cells adhered to the inner surface of the A-PCPC. HE staining showed a small group of woven bone formation 2 weeks later in the experimental group, while the formation of bone less in the control group. Woven bone turned into trabecular bone gradually at 4 weeks in the experimental group, while the control group showed a large number of bone-like tissue. Histomorphometry showed more mature bone in the experimental group than the control group at 8 weeks. CONCLUSIONS: The A-PCPC/BMSCs composites show good osteogenetic activity and could promote mineralization of the immature bone. It can be used as the bone tissue engineering scaffolds. Supported by Innovation Fund for Science and Technology Development of Pudong New District(Grant No. PKJ2009-Y19).

[Effect of fluorosis on the expression of basic fibroblast growth factor in rat incisors].

OBJECTIVE: To investigate the effects of overdose fluoride on the expression of basic fibroblast growth factor (bFGF) in rat incisors. METHODS: Twenty Wistar rats were randomly divided into 2 groups: group I (control group) distilled water was given; group II (experimental group) 100 mg/L NaF was given. The rats were killed at the end of 8 th week. Immunohistochemical staining was used to study the expression of bFGF in rat incisors. RESULTS: Immunohistochemical results demonstrated the presence of bFGF in ameloblasts, odontoblasts of rat incisors. The expression of bFGF was reduced in group II (P < 0.01). CONCLUSIONS: Overdose fluoride inhibits the expression of bFGF and affects the interaction between dental epithelium and dental mesenchyme, which leads to the enamel demineralization.

[Effect of overdose fluoride on the expression of enamelin in rat mandibular incisor].

OBJECTIVE: To observe the effect of overdose fluoride on the expression of enamelin in rat mandibular incisor. METHODS: Twenty Wistar rats were divided randomly into two groups. Animals were maintained in standard environment with free access to food and distilled water (control group) or water added with 100 mg/L F-(experimental group). The rats were killed in the eighth week. HE staining was used to observe the morphology of ameloblasts. Immunohistochemical staining was adopted to study the expressions of enamelin in rat incisor. RESULTS: The ameloblasts of the treated rat were arranged in multi-layer. The ameloblasts in group II were thinner than those in group I. The structure of enamel matrix was in disorder. The expressions of enamelin in ameloblasts and odontoblasts were obviously inhibited in group II (P < 0.01). CONCLUSION: The overdose fluoride inhibits the secretion of enamelin and leads to the abnormal development of enamel matrix.

[Effects of overdosed fluoride on rat's incisor expression of matrixmetalloproteinase-20 and tissue inhibitors of metalloproteinase-2].

OBJECTIVE: To investigate the effects of overdosed fluoride on the expression of MMP-20 and TIMP-2 in rat incisor. METHODS: 20 Wistar rats were randomly divided into experiment group and control group. The distilled water was given in control group. 100 mg/L fluoride- was given in experiment group. After 8 weeks treatment, the rats were killed. Immunohistochemical staining was used to observe the expression of MMP-20 and TIMP-2 in rat incisors. RESULTS: Immunohistochemical results demonstrated the presence of MMP-20 and TIMP-2 protein in ameloblasts, odontoblasts, stratum intermedium and the stellate reticulum of rat incisor. The imagination analysis results showed that the expression of MMP-20 was reduesed in experiment group (P<0.01), and the expression of TIMP-2 had no significant difference (P>0.05). CONCLUSION: The overdosed fluoride inhibits the secretion of MMP-20 and leads to the disturbed balance between MMP-20, TIMP-2 in rat incisor, which leads to the delay of the amelogenin removal and the enamel demineralization.

[Effect of overdose fluoride on expression of bone sialoprotein in developing dental tissues of rats].

PURPOSE: To investigate the changes of bone sialoprotein (BSP) in developing dental tissues of rats exposed to fluoride. METHODS: Twenty rats were randomly divided into two groups, one was with distilled water (control group), the other was with distilled water treated by fluoride (experimental group). When the fluorosis model was established, the changes of the expression of BSP were investigated and compared between the two groups. HE staining was used to observe the morphology of the cell, and immunohistochemisty assay was used to determine the expression of BSP in rat incisor. Student's t test was used for statistical analysis. RESULTS: The ameloblasts had normal morphology and arranged orderly. Immunoreactivitis of BSP was present in matured ameloblasts, dentinoblasts, cementoblasts, and the matrix in the control group. But in the experimental group the ameloblasts arranged in multiple layers, the enamel matrix was confused and the expression of BSP was significantly lower than that of the control group. Statistical analysis showed significant differences between the two groups (P<0.01). CONCLUSION: Fluoride can inhibit the expression of BSP in developing dental tissues of rats, and then inhibit differentiation of the tooth epithelial cells and secretion of matrix. This is a probable intracellular mechanism of dental fluorosis.