TAZ contributes to osteogenic differentiation of periodontal ligament cells under tensile stress.

BACKGROUND AND OBJECTIVE: Bone remodeling during orthodontic treatment is achieved by the osteogenesis of human periodontal ligament cells (PDLCs) subjected to mechanical loadings. Transcriptional co-activator with PDZ-binding motif (TAZ) mediates bone remodeling in response to extracellular mechanical signals. This study aims to investigate the role of TAZ in osteogenesis of PDLCs under tensile strain. MATERIALS AND METHODS: A uniaxial cyclic tensile stress (CTS) at 12% elongation and 6 cycles/min (5 s on and 5 s off) was applied to PDLCs. The osteogenic differentiation was determined by the protein and gene expressions of osteogenic markers using qRT-PCR and Western blot, respectively, and further by alkaline phosphatase (ALP) activity and Alizarin Red S staining. The interaction of TAZ with core-binding factor α1 (Cbfα1) was examined by co-immunoprecipitation. The immunofluorescence histochemistry was used to examine the nucleus aggregation of TAZ and the reorganization of actin filaments. Moreover, small interfering RNA-targeting TAZ (TAZsiRNA) was used for TAZ inhibition and Y-27632 was employed for Ras homologue-associated coiled-coil protein kinase (ROCK) signaling blockage. RESULTS: CTS clearly stimulated the nucleus accumulation of TAZ and its interaction with Cbfα1. CTS-induced osteogenesis in PDLCs was significantly abrogated by the infection with TAZsiRNA, as shown by the decreased stained nodules and protein expressions of Cbfα1, collagen type I, osterix, and osteocalcin, along with the inhibition of ß-catenin signaling. Moreover, ROCK inhibition by Y-27632 hindered TAZ nucleus aggregation and its binding with Cbfα1, which subsequently lead to the decreased osteoblastic differentiation of PDLCs. CONCLUSIONS: Taken together, we propose that TAZ nucleus localization and its interaction with Cbfα1 are essential for the CTS-induced osteogenic differentiation in PDLCs. And such TAZ activation by CTS could be mediated by ROCK signaling, indicating the pivot role of ROCK-TAZ pathway for PDLCs differentiation.

Enhanced adsorption of steroid estrogens by one-pot synthesized phenyl-modified mesoporous silica: Dependence on phenyl-organosilane precursors and pH condition.

In this study, the phenyl-modified mesoporous materials were successfully synthesized using phenyl-organosilanes (trimethoxyphenylsilane and triethoxyphenylsilanea) by one-pot co-condensation method for the removal of estrone (E1), 17ß-estradiol (E2), and 17α-ethinyl estradiol (EE2). Both the triethoxyphenylsilane-modified material (20%EtPh-MCM-41) and trimethoxyphenylsilane-modified material (20%MePh-MCM-41) could rapidly achieve equilibrium in 30 min at low adsorbent dosage of 0.025 g L-1. But the different hydrolysable groups of trimethoxyphenylsilane and triethoxyphenylsilane led to the discrepancies in physicochemical properties of the 20%EtPh-MCM-41 and 20%MePh-MCM-41, and thus affected adsorption performance. The 20%EtPh-MCM-41 exhibited the faster estrogen adsorption rates expressed in pseudo-second-order kinetic constant than the 20%MePh-MCM-41 due to the more hydrophobicity. Conversely, the 20%MePh-MCM-41 had much more estrogen adsorption capacities than the 20%EtPh-MCM-41 because of the more available adsorption sites. The addition of the phenyl-organosilane improved estrogen adsorption by π-π and hydrophobic interactions, and the Langmuir-model-based maximum adsorption amounts could reach 99.02, 83.47, and 53.60 mg g-1 for EE2, E2, and E1, respectively. But excessive concentration of phenyl-organosilane decreased adsorption capacities due to poor pore structure. Alkaline solution, which induced estrogen deprotonation and negative surface charge of absorbents, inhibited estrogen adsorption by electrostatic repulsion and the decreased hydrophobic interaction, but acidic and neutral solutions, ionic strength, and humic acid did not significantly affect estrogen removal. This work not only showed the high potential of trimethoxyphenylsilane-modified MCM-41 used in water purification for steroid estrogens, but also demonstrated the suitable selection of organosilane precursors was key in producing favorable materials with designed functionality.

N-propyl functionalized spherical mesoporous silica as a rapid and efficient adsorbent for steroid estrogen removal: Adsorption behaviour and effects of water chemistry.

To achieve an enhanced and selective adsorption of steroid estrogens, the n-propyl functionalization was applied to the mesoporous silica material (MCM-41) according to the physico-chemical property analysis of steroid estrogens. Adsorption behaviour and water chemistry effects were evaluated with the most concerned steroid estrogens: estrone (E1), 17ß-estradiol (E2) and 17α-ethinyl estradiol (EE2) based on the materials characterization. The results showed the uptakes of E1, E2, and EE2 onto the modified MCM-41 were enhanced and accelerated by the n-propyl functionalization, which was positively correlated with the hydrophobicity of the synthesized materials. Kinetic data fitted the pseudo-second-order model well. Based on the Langmuir model, the maximum adsorption capacities of the n-propyl modified MCM-41 were up to 119.87, 88.38, and 86.91 mg g-1 for EE2, E1, and E2, respectively. Importantly, both acid and neutral solutions were beneficial to estrogen removal, but ionic strength and humic acid did not affect the estrogen adsorption. The above results suggested that the n-propyl functionalized MCM-41 would be a promising adsorbent for the rapid and efficient removal of estrogens with the selectivity from natural organic matter like humic acid. Mechanism analysis showed the key role of hydrophobic interaction, and it also confirmed the contribution of the carbonylic lone pair electrons of E1, which helped the formation of stronger hydrogen bonds with silicon hydroxyls and enhanced the dipole-dipole interaction between E1 and the synthesized materials.

[Hypoxia regulates the expression of OPG/RANKL mRNA in rat bone marrow mesenchymal stem cells].

PURPOSE: To investigate the effects of hypoxia on the expression of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa B ligand (RANKL) mRNA in rat bone marrow mesenchymal stem cells (rBMSCs). METHODS: rBMSCs were isolated and cultured by whole bone marrow cell adherent method, and an optimal hypoxic preconditioning model was established with CoCl2 (cobalt chloride). rBMSCs were incubated in cell culture mediums with different concentrations of CoCl2 (final concentrations of CoCl2 were 0, 50, 100, 200, 400 µmol/L) and incubated for different times. MTT assay was applied to detect the effect of CoCl2 on cell proliferation. mRNA and protein expression of HIF-1α of rBMSCs was detected by real-time PCR and Western blot. After treated with 100 µmol/L CoCl2 for 0, 12, 24, 48, 72, 96 h, the expression of rBMSCs OPG/RANKL mRNA were detected by real-time PCR. The differences in distribution of each genotype were analyzed with SPSS 18.0 software package. RESULTS: Compared with the control group, 200, 400 µmol/L CoCl2 inhibited the proliferation of rBMSCs (P<0.05). However, 50, 100 µmol/L CoCl2 had no significant impact on the proliferation of rBMSCs (P>0.05). Real-time PCR and Western blot showed that HIF-1α expression in 50 µmol/L and 100 µmol/L CoCl2 groups was significantly higher than the control group; the effect of 100 µmol/L CoCl2 was significantly greater than 50 µmol/L CoCl2. After cultivated in hypoxia condition for 12 h, the expression of OPG and RANKL mRNA in rBMSCs didn't change significantly (P>0.05). After cultured hypoxia condition for 24, 48, 72, 96 h, the expression of OPG mRNA in rBMSCs increased while the RANKL decreased, thus the ratio of OPG/RANKL increased and the difference was significant (P<0.05). CONCLUSIONS: Hypoxia can regulate the mRNA expression of OPG and RANKL mRNA in rBMSCs and significantly promote osteogenic differentiation.

Baicalein enhances the osteogenic differentiation of human periodontal ligament cells by activating the Wnt/ß-catenin signaling pathway.

OBJECTIVE: Periodontium regeneration is one of the most important processes for periodontitis therapy. Human periodontal ligament cells (hPDLCs) play a vital role in the repair and regeneration of periodontal tissues. Our study aimed to investigated the mechanisms underlying the promotion of hPLDCs osteogenic differentiation by baicalein. DESIGN: hPDLCs were obtained from periodontal ligament (PDL) tissues by primary culture. The MTT assay was used to determine the growth curves of hPDLCs treated with different concentrations of baicalein (1.25, 2.5, 5, or 10µM). Alkaline phosphatase (ALP) staining and Alizarin red S staining were performed to assess osteogenic differentiation of hPDLCs administered baicalein. Osteogenic differentiation-related gene and protein expression levels and Wnt/ß-catenin pathway signal changes were assessed by qRT-PCR and Western blotting analysis. RESULTS: The results showed that baicalein decreased the growth of hPDLCs slightly and increased ALP activity and calcium deposition in a dose-dependent manner. The expression of runt-related transcription factor 2 (RUNX2), bone morphogenetic protein 2 (BMP2), Osterix (OSX) and osteocalcin (OCN) were elevated after baicalein administration. Moreover, baicalein strongly activated the Wnt/ß-catenin pathway and up-regulated the expression of ß-catenin, lymphoid enhancer factor 1 (LEF1) and Cyclin D1. Dickkopf-related protein 1 (DKK-1) significantly reversed the effects of baicalein on hPDLCs. CONCLUSIONS: Our findings indicated that baicalein enhanced the osteogenic differentiation of hPDLCs via the activation of the Wnt/ß-catenin signaling pathway, which may represent a potential candidate for periodontitis therapy.

[Nitrogen Removal Performance and Microbial Community Analysis of Activated Sludge Immobilization].

Immobilization of activated sludge was used to further remove nitrogen from secondary effluent. Intermittent sequencing batch reactor experiments were conducted to measure nitrogen removal in synthetic wastewater with initial total nitrogen concentrations (TN) of 10-45 mg·L-1 and C/N ratio of 1.78-10, and microbial community characteristic of embedding beads was investigated. When the packing ratio of embedding beads was 10%, and the temperature of wastewater, dissolved oxygen (DO), initial concentration of chemical oxygen demand (COD) were maintained at 10-15℃, 2-4 mg·L-1, and 80-100 mg·L-1, respectively, the results showed that the maximum total nitrogen removal loads ranged from 7.78 to 23.18 mg·(L·h)-1during the stable phase. SEM observations showed that the embedding beads were highly porous and microorganisms adhered to the interior and external surface of embedding beads, demonstrating that embedding beads acted as an ideal support material. Based on high-throughput sequencing analysis, the structure of microbial communities in the beads'interior and exterior changed significantly compared with embedding activated sludge. The advantage of denitrifying bacteria in embedding beads was obvious and the microbial diversity was good. Some microorganisms which can conduct both heterotrophic nitrification and aerobic denitrification, were identified. These processes may facilitate pathways for untraditional biological denitrification in the beads'interior.

Extraction techniques using isopropanol and Tenax to characterize polycyclic aromatic hydrocarbons bioavailability in sediment.

Polycyclic aromatic hydrocarbon (PAH)-degrading bacterium strain J1-q (Sphingomonas pseudosanguinis strain J1-q) was isolated from Yangtze River surface sediment in the downtown area of Chongqing in a previous study. Isopropanol and Tenax extraction techniques were used to characterize the bioavailability of target PAH compounds. Phenanthrene (Phe) and fluoranthene (Fluo) were the target PAHs due to their significant background concentrations in surface sediment samples. Isopropanol solutions at concentrations of 50-100% and residual Phe and Fluo concentrations in sediment were correlated, with R2 values of 0.9846 and 0.9649, respectively. The quantities of the Phe and Fluo fractions extracted for 3days with isopropanol from sediment were closely related with the corresponding quantities of PAHs degraded by bacterial strain J1-q when the extracting concentrations were 55% and 80%, respectively. The quantity of Phe extracted by Tenax agreed with the total quantity biodegraded when the Tenax: sediment mass ratio was 0.25 and the target PAHs were degraded for 30d, whereas the extracted quantity of Fluo accounted for 93.30% of the total quantity biodegraded by the bacterium. The triphasic model was appropriate to simulate the consecutive Phe and Fluo extraction process using Tenax at various Tenax: sediment ratios, and all simulated correlation coefficients were >0.9151. A 24-h extraction period was adequate to estimate the rapidly desorbing fractions when they were extracted with Tenax. Isopropanol extraction was preferable to characterize Phe and Fluo bioavailability under the experimental conditions, whereas Tenax extraction was useful to predict bioavailability of the two target PAHs with particular selectivity.

Combined effects of proinflammatory cytokines and intermittent cyclic mechanical strain in inhibiting osteogenicity in human periodontal ligament cells.

Mechanical strain plays an important role in bone formation and resorption during orthodontic tooth movement. The mechanism has not been fully studied, and the process becomes complex with increased amounts of periodontal patients seeking orthodontic care. Our aims were to elucidate the combined effects of proinflammatory cytokines and intermittent cyclic strain (ICS) on the osteogenic capacity of human periodontal ligament cells. Cultured human periodontal ligament cells were exposed to proinflammatory cytokines (interleukin-1ß 5 ng/mL and tumor necrosis factor-α 10 ng/mL) for 1 and 5 days, and ICS (0.5 Hz, 12% elongation) was applied for 4 h per day. The autocrine of inflammatory cytokines was measured by enzyme-linked immunosorbent assay. The expression of osteoblast markers runt-related transcription factor 2 and rabbit collagen type I was determined using real-time polymerase chain reaction and Western blot. The osteogenic capacity was also detected by alkaline phosphatase (ALP) staining, ALP activity, and alizarin red staining. We demonstrated that ICS impaired the osteogenic capacity of human periodontal ligament cells when incubated with proinflammatory cytokines, as evidenced by the low expression of ALP staining, low ALP activity, reduced alizarin red staining, and reduced osteoblast markers. These data, for the first time, suggest that ICS has a negative effect on the inductive inhibition of osteogenicity in human PDL cells mediated by proinflammatory cytokines.

Biodegradation potential of polycyclic aromatic hydrocarbons by bacteria strains enriched from Yangtze River sediments.

Microbial degradation is an effective method for the removal of polycyclic aromatic hydrocarbons (PAHs) compounds from polluted sediments. Surface sediments collected from Yangtze River in the downtown area of Chongqing were found to contain PAH concentrations to various different degrees. Two bacteria strains (termed PJ1 and PJ2) isolated from the sediment samples could use phenanthrene (Phe) and fluoranthene (Flu) as carbon sources for growth thereby degrading these two PAH compounds. Using 16S rDNA gene sequencing, the isolates were identified as Sphingomonas sp. and Klebsiella sp., respectively. Biodegradation assays showed that the PJ1 presented an efficient degradation capability compared to PJ2 in cultures with the initial Phe and Flu concentrations ranging from 20 to 200 mg/L. The highest rates of Phe and Flu biodegradation by PJ1 reached 74.32% and 58.18% after incubation for 15 and 30 days, respectively. This is the first report on the biodegradation potential of the bacterial from surface sediments of an industrial area upstream of the Gorge Reservoir.

[MRI diagnosis of osteochondral lesions in the talus and the dynamic follow up analysis after osteochondral transplantation].

OBJECTIVE: To investigate MRI findings of osteochondral lesions in the talus;to evaluate the value of MRI in diagnosing and determining the stage of osteochondral lesions;to analyze the follow up clinical value of MRI in osteochondral transplantation of autologous bone. METHODS: A total of 79 patients from February 2013 to March 2015 had been retrospectively analyzed. All the patients were treated in our hospital. The ankle arthroscopy results were used as the reference standard, and the accuracy of MRI in diagnosis and Hepple staging had been investigated. Fifteen patients with cartilage transplantation of autologous bone were followed up with MRI examination and evaluation of cartilage repair score(MOCART) after one year. The values of MRI in the postoperative follow up were analyzed. RESULTS: Hepple staging of 79 patients was shown as follows:7 cases of stage I, 12 cases of stage II, 24 cases of stage III, 16 cases of stage IV, and 20 cases of stage V. Ankle arthroscopy grading of 59 patients in this group(in addition of 20 cases of stage V):2 cases of grade A, 2 cases of grade B, 4 cases of grade C, 14 cases of grade D, 22 cases of grade E, and 15 cases of grade F. The accuracy rate of MRI in determining Hepple V was set at 100%, and Hepple stage I corresponds to the arthroscopic A, B, C stage, stage II corresponds to D stage, stage III corresponds to E stage, stage IV corresponds to F stage. The accuracy rate of MRI in determining Hepple stage I to IV was 87.5%, 85.7%, 95.4% and 93.3% respectively. After cartilage transplantation of autologous bone, MRI of 15 patients showed cartilage surface in transplanted area was smooth, bone healed well, and the surrounding edema disappeared. The MOCART was 30 to 80 scores with an average score 59.0±15.6;9 cases of these 15 patients were(9/15, 60%) higher than 60 score. CONCLUSIONS: MRI plays a significant role in clinical diagnosis and staging of the talus osteochondral injury. As a method of long term follow up after cartilage transplantation, MRI can well evaluate the rapair of the postoperative osteochondral injury.

Performance linked to residence time distribution by a novel wool-based bioreactor for tertiary sewage treatment.

Laboratory-scale experiments were carried out using up-flow 7 L Submerged Aerated Filter reactors packed with wool fibre or commercial plastic pall rings, Kaldnes, (70% by volume) support media for the tertiary treatment of sewage. The performance of the wool bioreactor was more consistent than that with Kaldnes medium, for both TOC removal (93%) and SS removal (90%). Both plastic and wool-packed bioreactors achieved complete nitrification at the load of about 0.4 kgCOD/m(3)/day. The sludge yield of the wool bioreactor was almost half that of the bioreactor with Kaldnes suggesting that wool could retain residual organics and particulates. The wool however was degraded and it was concluded that wool would have to be considered as additional sacrificial adsorption capacity rather than an alternative medium. The performance was linked to the residence time distribution studies and these changes in the wool structure. Biomass growth increased the retention of the tracer in the wool reactor by, it was suggested, exposing a greater surface area. Results from the plastic media on the other hand showed increased mixing possibly by increasing the mobility of the plastic. Aeration increased the mixing in both reactors, and patterns were in all cases predominantly well-mixed.

Wound repair and proliferation of bronchial epithelial cells regulated by CTNNAL1.

Adhesion molecules play vital roles in airway hyperresponsiveness (AHR) or airway inflammation. Our previous study indicated that adhesion molecule catenin alpha-like 1 (CTNNAL1) is relevant closely to asthma susceptibility, but its biological function or significance is still unclear. In the present study, we observed the temporal and spatial distribution of CTNNAL1 expression in mouse lung tissue with the OVA-sensitized asthma model and found that the level of CTNNAL1 mRNA showed a prominent negative correlation with pulmonary resistance (R(L)). To study the function of CTNNAL1 in airway, effects of CTNNAL1 on proliferation and wound repair activity of human bronchial epithelial cells (HBEC) was investigated with antisense oligonucleotide (ASO) technique. The results showed that: (1) CTNNAL1 ASO could decelerate the repairing velocity and proliferation of HBEC; (2) CTNNAL1 expression was increased on the edge cells of mechanic wounded area in culture; (3) extracellular matrix component fibronectin (Fn) obviously promoted wound repair activity and proliferation of HBEC, which could be blocked by CTNNAL1 ASO; (4) Western blot showed that Fn could promote FAK phosphorylation, which also be inhibited by CTNNAL1 ASO. In conclusion, the level of CTNNAL1 mRNA expression is highly correlated to airway resistance; CTNNAL1 may contribute to the wound repair and proliferation of HBEC. Furthermore, it may serve to Fn mediated cell-extracellular adhesion and its signal transduction.

[Determination of main flavone glycosides in Flos Chrysanthemi and observation of factors influenced contents].

OBJECTIVE: To determine and compare the content of luteolin-7-O-beta-D-glucoside and apigenin-7-O-beta-D-glucoside in Flos Chrysanthemi from different collection time, sources, grades and processes. METHOD: The contents were determined by RP-HPLC. Zorbax SB C18 column (4.6 mm x 250 mm, 5 microm) was used as analysis column, the mobile phase was acetonitrile-pH 2.0 phosphate buffer solution with gradient elution, the detector was set at 338 nm. RESULT: The contents of two components changed at some degree in Flos Chrysanthemi from different collection time, different plant sites or with different grades, while the contents varied obviously among Flos Chrysanthemi from different source and different sorts. No obvious difference was found in Flos Chrysanthemi from different year. CONCLUSION: The contents of two components were influenced by process, plane site, source and sorts, especially by source and sorts.

[Determination of luteolin and luteolin-7-beta-D-glucoside in Chrysanthemum morfolium Ramat. from different collection time by RP-HPLC].

OBJECTIVE: To observe the content variation of luteolin and luteolin-7-beta-D-glucoside in Chrysanthemum morifolium Ramat. (CMR) from different collection time. METHODS: RP-HPLC was used to analyze these two components in CMR collected in 2001 and 2002. RESULT: The content of luteolin was significantly lower than that of luteolin-7-beta-D-glucoside. Furthermore, the former showed no marked changes during collection, while the latter did not varied markedly in early collection but decreased significantly in later collection. CONCLUSION: The content of luteolin-7-beta-D-glucoside reflects the quality of Chrysanthemum morifolium Ramat. more viably than that of luteolin.