The aim of this study is to investigate the influence of different posts on the fracture mechanics of endodontically-treated teeth with open apex. Forty-eight human maxillary anterior teeth were collected, and the root was transversely sectioned 12 mm under the cementoenamal junction (CEJ). These samples were then randomly divided into two groups, i.e., minor diameter open apex root (group A) and major diameter open apex root (group B), with mineral trioxide aggregate (MTA) placed into the apical 4 mm in the root canals. Subsequently, both groups were respectively further divided into three subgroups as follows: fiber-post (subgroup 1), metal post (subgroup 2) and non-post (subgroup 3) group. Teeth were restored with a composite resin crown and tested by using a universal testing machine at the rate of 1 mm/min cross-head. Values of the maximum fracture resistance and failure patterns were recorded and compared among all subgroups. In addition, the changes of MTA properties were carefully examined via X-ray photography. Our results indicate that (1) In group A, the mean value of fracture resistance for teeth restored with fiber posts were statistically higher than that with either metal post or non-post; (2) In group B, there was no statistically significant difference in the mean value of fracture resistance among three subgroups; (3) No statistical significance in the mean value of fracture resistance was found between group A and group B; (4) The failure modes of most samples (58%) were irreparable; (5) MTA in two teeth developed cracks after loading tests. In conclusion, endodontically-treated teeth restored with fiber posts are more resistant to fracture than those restored with either metal posts or non-post, and most of the fracture modes are catastrophic in nature.

BACKGROUND:Long non-coding RNAs (lncRNAs) have became the hot topic in current studies and play an important role in the tumorigenesis. However, lncRNAs involved in the osteoblast differentiation remain poorly reported.
OBJECTIVE:To investigate the role of human LncRNAs in osteogenic differentiation of C3H10T1/2 cells induced by bone morphogenetic protein-2 and explore action mechanism.
METHODS:The induction of bone morphogenetic protein-2 was validated by alkaline phosphatase staining and the expression of corresponding genes was detected. The lncRNA expression profile was analyzed using the Arraystar lncRNA array in C3H10T1/2 MSCs undergoing early osteoblast differentiation. The expression with or without bone morphogenetic protein-2 induction was compared with high-flux sequencing, and the down-regulated genes were screened. The effect of lncRNA overexpression on osteogenic differentiation of C3H10T1/2 cells induced by bone morphogenetic protein-2 was observed.
RESULTS AND CONCLUSION:The bone morphogenetic protein-2 induced C3H10T1/2 cells led to increased alkaline phosphatase activity. After 72 hours of bone morphogenetic protein-2 induction, alkaline phosphatase, Id1,osteocalcin, Runx2, sp7 expression were increased (P<0.05). There were 24 down-regulated lncRNAs identified between bone morphogenetic protein-2 treated and untreated groups, the decrease of expression was 1.5 folds, and among them, only AK035085 contained intron. Compared with control group with no AK03508 expression, over-expression lncRNA AK035085 decreased the expression of alkaline phosphatase, Id1, osteocalcin, Runx2, sp7 (P<0.05). Experimental findings indicate that bone morphogenetic protein-2 induces osteogenic differentiation of C3H10T1/2 cells and AK035085 inhibits the osteogenic differentiation.

Aim To investigate the effect of insulin glargine and human insulin on proliferation of a human breast cancer cell line MDA-MB-231 and the role of ERK in the process.Methods MDA-MB-231 cells were incubated with insulin glargine and human insulin at different concentrations and for different time courses.A specific ERK1/2 inhibitor,PD98059,was used either alone or in combination with insulin glargine or human insulin to test the involvement of ERK pathway in cell growth.Cell proliferation was evaluated using cell counting kit-8 reagents.Cell cycle distribution was analyzed by flow cytometry.Results Both insulin glargine and human insulin dose-dependently enhanced MDA-MB-231 cell proliferation at the concentrations from 1 to 100 IU?L-1 after treatment for 96 h.At the concentration of 10 IU?L-1,both drugs promoted cell growth at 48,72,and 96 h.The percentage of S+G2/M cells was significantly increased in both insulin glargine and human insulin treated groups as compared to untreated controls.No significant difference was observed between insulin glargine and human insulin in their effects on cell proliferation and cell cycle distribution.Cell proliferation was significantly inhibited by PD98059.However,in the presence of PD98059,both drugs still promoted cell proliferation significantly as compared to untreated controls.Conclusions Insulin galrgine and human insulin similarly promote proliferation of MDA-MB-231 cells independent of ERK activation.

Objective To explore the effects of under-expression of large tumor suppressor 1 (LATS1) on activation of Hippo signaling pathway and differentiation, proliferation, migration of bone marrow mesenchymal stem cells (mMSCs) of micein vitro.Methods mMSCs of C57BL/6 mice were divided into normal control (MSC) group, empty vector control (MSC-GFP) group, LATS1-over-expressing (MSC-LATS1) group, empty vector without LATS1 shRNA control (MSC-shControl) group and LATS1-under-expressing (MSC-shLATS1) group. Lentiviral vectors with activated,inactivated LATS1 (the key molecule of Hippo signaling pathway) modifications and empty vectors were constructed and were used to infect mMSCsin vitro. The transduction efficiencies mediated by the lentiviral vectors were evaluated by fluorescence microscopy and flow cytometry. The mRNA expression of LATS1 was quantified by quantitative real-time polymerase chain reaction (qRT-PCR), and the protein expressions of LATS1, YAP (p-YAP), 14-3-3 were quantified by Western Blot to evaluate the activation of Hippo signaling pathway. Osteogenic and adipogenic differentiation of mMSCs were evaluated through measurement of Runx2, OSX and C/EBPα, PPAR-γ mRNA by qRT-PCR, as well as Alizarin Red S and Oil red O staining. Proliferation of mMSCs was evaluated using methy thiazdyl tetrazolium (MTT) assay. The scratch test and Transwell chamber test were used to analyze the horizontal and vertical migration ability of mMSCs.Results The transduction efficiencies mediated by the lentiviral vectors were 94.74%-96.10%. Compared with MSC-GFP group, the activation of Hippo signaling pathway was promoted in MSC-LATS1 group [LATS1 mRNA (2-ΔΔCT): 4.37±0.21 vs. 1.20±0.04, LATS1 protein (gray value): 2.21±0.06 vs. 1.09±0.10, p-YAP/YAP protein (gray value): 1.51±0.13 vs. 0.98±0.05, 14-3-3 protein (gray value): 1.92±0.18 vs. 1.10±0.09, allP < 0.05], osteogenic and adipogenic differentiation of mMSCs were decreased in MSC-LATS1 group [mineralization (A value):0.13±0.02 vs. 0.40±0.03, Runx2 mRNA (2-ΔΔCT): 0.51±0.02 vs. 0.98±0.09, OSX mRNA (2-ΔΔCT): 0.41±0.04 vs. 1.04±0.09, lipid accumulation (A value): 0.10±0.02 vs. 0.25±0.03, C/EBPα mRNA (2-ΔΔCT): 0.33±0.03 vs. 1.11±0.09, PPAR-γ mRNA (2-ΔΔCT): 0.29±0.02 vs. 1.04±0.10, allP < 0.05], the proliferation rate of mMSCs at 4-7 days was decreased in MSC-LATS1 group and so were the horizontal and vertical migration of mMSCs [wound healing rate: (18.65±3.53)% vs. (40.29±1.87)%, migrated cells (cells/MP): 35.99±6.18 vs. 103.67±17.77, bothP <0.05]. Compared with MSC-shControl group, the activation of Hippo signaling pathway was inhibited in MSC-shLATS1 group [LATS1 mRNA (2-ΔΔCT): 0.16±0.01 vs. 0.98±0.03, LATS1 protein (gray value): 0.38±0.03 vs. 1.04±0.07, p-YAP/YAP protein (gray value): 0.58±0.04 vs. 1.05±0.06, 14-3-3 protein (gray value): 0.14±0.02 vs. 1.02±0.09, allP < 0.05], osteogenic and adipogenic differentiation of mMSCs were increased in MSC-shLATS1 group [mineralization (A value): 0.93±0.13 vs. 0.44±0.05, Runx2 mRNA (2-ΔΔCT): 1.44±0.12 vs. 0.95±0.04, OSX mRNA (2-ΔΔCT):1.67±0.06 vs. 1.10±0.11, lipid accumulation (A value): 0.47±0.06 vs. 0.28±0.04, C/EBPα mRNA (2-ΔΔCT):3.98±0.61 vs. 0.99±0.10, PPAR-γ mRNA (2-ΔΔCT): 3.05±0.36 vs. 0.98±0.14, allP < 0.05], the proliferation rate of mMSCs at 3-7 days was increased in MSC-shLATS1 group and so were the horizontal and vertical migration of mMSCs [wound healing rate: (80.18±6.98)% vs. (46.18±1.01)%, migrated cells (cells/MP): 212.69±41.21 vs. 115.87±35.15, bothP < 0.05].Conclusions Under-expression of LATS1 promotes the differentiation, proliferation, migration of mMSCs by inhibition of Hippo signaling pathwayin vitro.

AIM:To investigate the production and activation of caspase-3 in primary rat renal proximal tubule cells in response to tumor necrosis factor-α(TNF-α) and the implication of nuclear factor-κB (NF-κB) in the process. METHODS:Isolated rat renal proximal tubule cells (PTCs) from male adult Sprague Dawley rats were treated with TNF-α according to the indicated time courses. A specific NF-κB inhibitor,Bay11-7082,was used alone or as a pretreatment for 1 h followed by exposure to TNF-α for 24 h.The protein levels of cleaved caspase-3,caspase-3,I-κBα,phosphorylated I-κBα,and GAPDH were detected by Western blotting using specific antibodies. RESULTS:The protein level of cleaved caspase-3 relative to caspase-3 was significantly increased in the presence of TNF-α for 6 h,12 h,and 24 h. Protein levels of caspase-3 were significantly decreased by 12 h and returned to baseline by 24 h in the presence of TNF-α. Treatment with Bay11-7082 for 25 h alone or pretreatment with Bay11-7082 for 1 h followed by addition of TNF-α for 24 h caused a remarkable reduction in both cleaved caspase-3 and caspase-3 as compared to control and TNF-α treated groups. An increase in phosphorylated I-κBα was observed from 15 min to 60 min after treatment with TNF-α at a dose of 10 μg/L in PTCs. CONCLUSION:NF-κB is not only associated with the activation of caspase-3 but also the production of caspase-3 in primary rat renal proximal tubule cells in response to TNF-α.

BACKGROUND: The preparation and antibacterial activity of metal ionic zirconium phosphates has been systemically investigated now, but the applications are limited owing to the discoloration or the low antibacterial activity. Here we prepared new antibacterial agents of quaternized zirconium phosphates by introducing quaternary ammonium salt bactericidal agent with high-effective, broad-spectrum and low-toxic into sodium zirconium phosphate through an ion-exchange method.OBJECTIVE: To explore the component structure and antibacterial activity of quaternized zirconium phosphates.DESIGN, TIME AND SETTING: An in vitro observational experiment was performed at Research Laboratory of Department of Chemistry, Jinan University from June to August 2009.MATERIALS: Quaternized zirconium phosphates were prepared by introducing dodecyl dimethyl benzyl ammonium chloride into sodium zirconium phosphate through an ion-exchange method.METHODS: The mol ratios of quaternary ammonium cations to cation exchange capacity of sodium zirconium phosphate in reaction solutions were 0.25: 1,0.5: 1, 1.0: 1, and 1.5 : 1, respectively, and four kinds of quaternized zirconium phosphates containing different contents of quaternary ammonium cations (QZrP-1, QZrP-2, QZrP-3, QZrP-4) were prepared through an ion-exchange method.MAIN OUTCOME MEASURES: The component structure and heat resistance of samples were measured by using an IR spectrometer, an elemental analyzer and a thermal analyzer, respectively. The minimum inhibitory concentrations (MICs) and minimal bactericidal concentrations (MICs) of the samples against Escherichia coli (E. co/i) and Staphylococci aureus (S. aureus) were estimated by a tube broth method.RESULTS: Quaternized zirconium phosphates were prepared, and the quaternary ammonium cation content increased with increasing the concentration of quaternary ammonium cations in reaction solution. The mass fraction of quaternary ammonium cations of QZrP-1, QZrP-2, QZrP-3, and QZrP-4 was 3.70%, 5.00%, 6.96%, and 10.01%, respectively. The onset temperatures of the decomposition for quaternary ammonium cations in quaternized zirconium phosphates were all higher than 345 °C, and they were preferable thermal stability. The antibacterial activity was higher when the quaternary ammonium cation content of quaternized zirconium phosphates increased. For quaternized zirconium phosphates QZrP-3 containing 6.96% mass fraction of quaternary ammonium cations, showed excellent antibacterial activity against E. coli and S. aureus.CONCLUSION: Quaternized zirconium phosphates QZrP-3 containing 6.96% mass fraction of quaternary ammonium cations,exhibited excellent thermal stability and antibacterial activity.

Objective To observe the influence of nursing intervention on postpartum depression and breastfeeding compliance in primipara.Methods From August 2013 to April 2015,146 women received cesarean section in North Jiaochang Branch of Hanzhong Central Hospital were randomly divided into intervention group and control group with 73 cases in each group.The control group was treated with routine nursing measures,and the intervention group with both routine care and nursing intervention.Self-Rating Depression Scale (SDS) and Self-Rating Anxiety Scale (SAS) were used to evaluate the negative emotions of maternal postpartum.Breastfeeding Self-Efficacy Scale (BSES) were used to evaluate the confidence of maternal breast feeding.The breast feeding compliance between the two groups was compared.The breast feeding rates of the two groups were compared in 1 week and 1 month after discharge.Results Compared with the control group,the SDS and SAS scores of the intervention group were significantly lower (P＜0.05).The breastfeeding confidence of the intervention group was significantly better than the control group and the difference was statistically significant (P＜0.05).The compliance of breast feeding of the intervention group was 97.26％ significantly higher than that of the control group (78.08％)with a statistically significant difference (P＜0.05).After one-week and one-month follow-up,the rate of breastfeeding of the intervention group was significantly higher than that of the control group (95.89％ VS 83.56％;91.78％ VS 72.60％,P＜0.05).Concltsion The nursing intervention measures for primipara after cesarean section can significantly break bad mood,enhance maternal breastfeeding confidence,increase the rate of breastfeeding compliance and are worthy of promotion.

BACKGROUND:Long non-coding RNA (lncRNA) regulates a series of physiological processes and it is considered to play important roles in the gene regulation of development, differentiation and metabolism. MC3T3-E1, C2C12 and C3H10T1/2 cells are able to differentiate into different celllineages, such as bone cells and muscle cells, and they can be used in the study of musculoskeletal diseases.
OBJECTIVE:To study the role of lncRNA in osteogenic differentiation induced by bone morphogenetic protein 2.
METHODS:Osteogenic differentiation of MC3T3-E1, C2C12 and C3H10T1/2 cells was induced by bone morphogenetic protein 2, and microarray expression profiling of lncRNA was undertaken in osteogenic differentiation. LncRNA simultaneous changes in three cells were found out. The siRNA interference of the lncRNA was used to study its effects on the osteogenic differentiation induced by bone morphogenetic protein 2. Real-time PCR and alkaline phosphatase staining were applied to detect osteogenesis related indicators.
RESULTS AND CONCLUSION:In the process of osteogenic differentiation induced by bone morphogenetic protein 2, osteogenic differentiation indicators were increased, while myogenic differentiation indicator myogenin was reduced. LncRNA AK007000 was screened out to play a role in osteogenic differentiation induced by bone morphogenetic protein 2. Knockdown of lncRNA AK007000 decreased the expression of osteogenic differentiation indicators, while increased the expression of myogenin. Therefore, AK007000 may play a role in promoting osteogenic differentiation and inhibiting myogenic differentiation.

BACKGROUND:A series of studies indicate that autophagy is closely linked with differentiation. Bone morphogenetic protein 2 (BMP-2) is the classical pathway for C2C12 and MC3T3-E1 osteoblast differentiation, and the ideal model to study osteogenic differentiation process.
OBJECTIVE:To observe the relationship between autophagy and BMP-2-induced celllines C2C12, MC3T3-E1 osteoblast differentiation.
METHODS:Real-Time PCR was applied to detect osteogenic differentiation and autophagy related index after C2C12 and MC3T3-E1 were induced with BMP-2 (100μg/L) for 72 hours. The osteogenic index alkaline phosphatase in BMP-2-induced MC3T3-E1 and C2C12 cultured with different concentrations of 3-methyladenine (0, 1, 5, 10 mmol/L) was determined with alkaline phosphatase staining. Western blot analysis was applied to detect LC3-I/II expression levels in C2C12 and MC3T3-E1 induced with BMP-2 for different time points (0, 12, 24, 48, 72, 96 hours).
RESULTS AND CONCLUSION:The autophagy-related mRNA and protein expression showed an increasing tendency and autophagy-related protein LC3 levels was increased, which was associated with the time, during the BMP-2-induced celllines C2C12, MC3T3-E1 osteoblast differentiation. Meanwhile, alkaline phosphatase expression levels were inhibited by autophagy in the process of osteogenic differentiation. Therefore, there is a close relationship between autophagy and the BMP-2-induced celllines C2C12, MC3T3-E1 osteoblast differentiation.

Background Subretinal transplantation of retinal pigment epithelium (RPE) cells for the treatment of age-related macular degeneration (AMD) have accelerated the drive to develop xeno-free cultivation system that support the rapid differentiation of human embryonic stem cells (hESCs) into ES-RPE cells.Objective This study was to report a modified xeno-free culture system and method for accelerating derivation of hESCs to differentiate into RPE cells.Methods This study was approved by Ethic Committee of Zhongshan Ophthalmic Center.HESC H1 line was cloned and cuhured in Vitronectin XFTM-coated 6-well dish with xenogenetic-free medium.Cells were cultured in 50 ng/ml noggin,10 ng/ml DKK-1 and 10 ng/ml insulin like growth factor-1 (IGF-1) medium for 2 days,and then the concentration of noggin was decreased to 10 ng/ml and 5 ng/ml basic fibroblast growth factor (bFGF) and cultured for the following 2 days.Sequentially,noggin and bFGF were removed and cultured for 2 days.Finally,1 μmol/L CHIR99021 was added in medium for 6 days.Morphological changes in the progress of ESCs differentiation into RPE were observed by Living Cell Imaging System.The expression of Mitf and RPE65,RPE cellsspecific markers,in the cells were detected by immunofluorescence technique,and the relative expression levels of RPE cells-specific marker mRNA were assayed using real time fluorescent quantitation PCR.Results Polygonalshape monolayer cells which contained pigments were initially observed at day 14 after cultured with the cobblestonelike arrangement.Mitf and RPE65 were strongly expressed in the hES-derived RPE cells 35 days after induced,showing red fluorescence,and the cells presented hexagonal shape at cultured day 60 with numerous pigment granules in cytoplasm.Compared with before differentiation,the expression levels of Mitf mRNA in hES-RPE cells increased by (3.43±2.77) folds and (8.91 ± 2.83) folds,and the expression levels of RPE65 mRNA increased by (14.60 ± 3.94) folds and (87.16 ±9.32) folds at day 7 and day 14 after differentiation,respectively (all at P＜0.05).Conclusions A defined xeno-free culture system is successfully established by adding niacinamide,DKK-l,noggin,IGF-1 and CHIR99021 in xeno-free medium,and this system can accelerate the derivation and differentiation of hESCs into RPE-like cells.