Objective To investigate the effect of exogenous hydrogen sulfide (H 2 S) on H2 S concentration and cystathionine β-synthase (CBS) expression in hippocampus in a rat model of vascular dementia (VaD). Methods A rat model of VaD was induced by using the modified four -vessel occlusion. The rats were divided into sham operation, model, low -dose and high-dose NaHS groups using the random number table method. They were further redivided into one day, seven -day, and 30-day subgroups according to the time after modeling. After modeling respectively, NaHS 30 μmol/kg and 100 μmol/kg were injected intraperitoneally every day in the low -dose and high-dose NaHS groups. The normal saline was injected intraperitoneally every day in the sham operation group and the VaD model group. Morris water maze test was used to evaluate the learning and memory ability of the rats. The expression of CBS in hippocampus was detected by real-time fluorescent polymerase chain reaction. Western Blotting was used to detect expression of CBS protein in hippocampus. Results Morris water maze test showed that the escape latencies of the model group, low -dose and high-dose NaHS groups were prolonged significantly compared with the sham operation group (P <0.05); the times of crossing the platform were decreased significantly compared with the model group (P <0.05); and the escape latencies were shortened significantly in the low -dose and high-dose NaHS groups compared with the model group ( P <0.05). The H2 S content in hippocampus was decreased significantly in the model group, low -dose and high-dose NaHS groups compared with the sham operation group, but the low -dose and high-dose NaHS group was significantly higher than that in the model group (all P <0.05). The expression of CBS mRNA and protein in the model, low -dose and high-dose NaHS groups was significantly lower than that of the sham operation group (all P < 0.05), and there was no significant difference between the low -dose and high-dose NaHS groups and the model group. Conclusions Exogenous H2 S may improve the learning and memory ability of the VaD rats. It may be associated with the increased H2 S content in hippocampus. However, it has no effect on CBS expression.

Objective:The study focuses on the distinct distributions of γδT cells in various tissues and the changes after Sal-monella typhimurium infection,and attempts to explore the physiological significance of γδT cell distribution and the role of γδT cells in infectious diseases .Methods:Flow cytometry and PCR technique were used to detect the proportion of different γδ T cell subsets among thymus,spleen,lymph nodes,liver,skin,and intestinal intraepithelial lymphocytes .Flow cytometry was applied to detect the secretion of IFN-γand IL-17a.The changes of various γδT cell subsets in liver and intestinal intraepithelial lymphocytes were analyze after Slamonella typhimurium infection.Res ults: γδ T cells were rich in the intestinal epithelium , skin and liver, but poor in the thymus,spleen and lymph nodes .The distribution of different subsets was quite dissimilar .Vγ5+γδT cells chiefly existed in skin ,and Vγ1+,Vγ4+,Vγ7+γδ T cells largely existed in small intestine.γδ T cells in liver mainly secreted IL-17a;however,γδ T cells in intestinal intraepithelial secreted IFN-γ.After infection by Salmonella typhimurium , the proportion of γδ T cells in intestinal intraepithelial increased significantly ,particularly Vγ1 +γδT cells.In Liver,there was no significant change of total γδT cell ratio,but the ratio of Vγ1 +γδT cells reduced ,Vγ4 +γδT cells raised.Conclusi on:γδT cells are rich in the intestinal epithelium ,skin and liv-er.The distribution of different subgroups has specificity .There are large differences in the ability of cytokine secretion among various subgroups of γδT cells.The distribution of γδT cell subgroups in small intestine and liver changes during Salmonella typhimurium in-fection.

Objective To observe the inhibitory effects of sub-MIC matrine alone and in combination with erythromycin on Staphylococcus epidermidis biofilms and their influences on morphological changes of the biofilms.Methods Minimum inhibitory concentrations (MIC) of matrine and erythromycin against Staphylococcus epidermidis were determined by the serial dilution method,antibacterial activity of matrine combined with erythromycin against planktonic S.epidermidis was evaluated by the checkerboard method.S.epidermidis biofilms were constructed in vitro,XTT reduction assay was used to evaluate influences of sub-MIC matrine alone and in combination with erythromycin on metabolism and adhesion of S.epidermidis biofilms,and scanning electronic microscope(SEM) was applied to observe the morphological and the structural changes of the biofilms.Results The MIC of erythromycin to S.epidermidis was 7.8125 μg/ml,while the MIC of matrine was greater than 1000 μg/ml,besides,a synergistic effect between erythronmycin and matrine on planktonic S.epidermidis was shown (FIC＜0.5).The sub-MIC matrine had no significant inhibitory effect on adhesion of S.epidermidis,and also the combination of the two agents was better than was used alone.However,the sub-MIC matrine had inhibitory effects on metabolism and morphology of S.epidermidis biofilms,and the combination of the two agents was weaker than was used alone.Conclusion Both the sub-MIC matrine and erythromycin had a significant inhibitory effect on S.epidermidis biofilm formation.Combination of the two agents showed synergistic effects on plankton and adhesion of S.epidermidis,but showed no synergistic effect on metabolism and morphology of the biofilms.

Objective:To remove murine embryonic stem cells(mESC)from the differentiating cell culture and purify the differentiated cells by Magnetic Activated Cell Sorting(MACS).Methods:Neural differentiation of mESC was induced by a 5-stage method.The specific cell surface marker,SSEA-1,was used to identify ES cells in the differentiating cells.The optimal dilutions of mouse anti mouse SSEA-1 IgM primary antibody and FITC conjugated goat anti mouse secondary antibody were determined before the flow cytometry test.The incubation time and incubation temperature of primary antibody were all optimized to make the cytometry test accurate.After the optimization,stage 4 cells were dissociated into single cell suspension,incubated with antibody of SSEA-1 and microbeads conjugated goat anti mouse IgM,and then sorted through the magnetic field.The rate of SSEA-1 positive cells in pre-and post-separation groups was assessed by flow cytometry,and the viability of cells was evaluated by trypan blue staining counting under light microscopy.Results:The proportion of SSEA-1 positive cells in the separated cells can be reduced from(7.19?1.36)% to(1.34?0.80)%.The survival rate of sorted cells was more than 92%,similar to that of pre-separation cells.Conclusions:The MACS system we used can effectively remove mESC from the differentiated cells.The sorted cells will be well provided for the subsequent studies about transplantation therapy.

OTX1 gene is one of the pivotal transcriptional factors involved in the neurogenesis.In order to overexpress the OTX1 gene in distinct cell types and find out its contribution to the proliferation and differentiation of stem cells in vitro,OTX1 cDNA was subcloned into lentiviral vectors.The resulting constructions pDUETOTX1,pDUETGFPOTX1 and pDUETGFP were packaged in 293 cells producing viral particles to transduce 293T cells,SY5Y cells,mouse embryonic stem cells and E15 neural stem cells.It was proved that the transferred OTX1 gene was located in the nuclei of the transduced cells in stead of plasma.Lentivirus is an ideal vector delivering gene to different cells.The overexpression of OTX1 in transduced 293T cells were validated by Western blot and immunofluorescence.

Objective To investigate the effects of lentivirus-mediated heat shock protein 70 (HSP70) gene on calcium homeostasis and calcium channels of PC12 cells induced by ischemic and hypoxia and its mechanisms. Methods PC12 cells at logarithmic phase were collected, and they were divided into recombined lentiviral infection group [infected by lentivirus containing HSP70 and green fluorescent protein (GFP) fluorescin gene], lentivirus control group (infected by lentivirus containing GFP without HSP70 gene) and non-infection group. PC12 cells were subjected ischemia/hypoxia for 4, 8, 12, 24 hours, and the cell activity was determined by methylthiazolyl tetrazolium (MTT) assay test inorder to determine the best time for ischemia/hypoxia. The mRNA expressions of HSP70, muscle/endoplasmic reticulum Ca2+-ATP isoforms (SERCA2a, SERCA2b), ryanodine receptor 2 (RyR2), and inositol 1,4,5-triphosphate receptor 1 (IP3R1) were determined by real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR), and the protein expressions of HSP70, SERCA, and IP3R were determined by Western Blot at 8 hours after ischemic/hypoxia. Flow cytometry was used to determine the levels of intracellular reactive oxygen (ROS) and intracellular Ca2+ ([Ca2+]i). Results With the prolongation of time of ischemia/hypoxia, the cell viability in all groups showed an increase followed by a weakening, and peaked at 8 hours. The cell viability at 8 hours in lentiviral infection group was significantly higher than that of the non-infection group and lentivirus control group [A value (×10-2): 20.3±2.2 vs. 14.1±2.1, 15.0±1.6, both P < 0.01], the mRNA and protein expressions of HSP70 and SERCA in lentiviral infection group were significantly increased [HSP70 mRNA (2-ΔΔCt ): 0.785±0.018 vs. 0.428±0.019, 0.423±0.023; HSP70 protein (gray value): 2.72±0.20 vs. 1.56±0.36, 1.63±0.41; SERCA2a mRNA (2-ΔΔCt ): 0.971±0.037 vs. 0.367±0.014, 0.347±0.012; SERCA2b mRNA (2-ΔΔCt ): 8.869±0.162 vs. 3.015±0.091, 2.941±0.091; SERCA protein (gray value): 2.84±0.18 vs. 1.48±0.26, 1.52±0.29], and IP3R2 mRNA and protein expressions were significantly declined [IP3R2 mRNA (2-ΔΔCt ): 0.183±0.020 vs. 0.439±0.020, 0.433±0.040; IP3R2 protein (gray value): 1.15±0.12 vs. 1.91±0.20, 1.83±0.19], with statistically significant differences (all P < 0.01); no significant difference in RyR mRNA was found [2-ΔΔCt (×10-3): 1.97±0.63 vs. 2.02±0.22, 2.01±0.09, both P > 0.05]; the relative fluorescence intensity of ROS and [Ca2+]i in lentiviral infection group was significantly reduced (ROS: 30.54±1.23 vs. 58.03±1.97, 57.72±2.35; [Ca2+]i: 34.50±2.05 vs. 48.20±3.02, 46.80±2.75, all P < 0.01]. Conclusion Exogenous HSP70 can maintain calcium homeostasis in the endoplasmic reticulum of PC12 cells, affect the Ca2+ channel protein regulated by calcium channel IP3R and calcium pump SERCA, which may cause hypoxia/ischemia intracellular injury.

Objective To investigate the ultrasonic imaging features of thyroid follicular adenoma for conducting the correct diagnosis and differentiation diagnosis.Methods The clinical and imaging data in 64 cases of pathologically proven thyroid follicular adenoma were analyzed on the maximal diameter of tumor,nodularity number,high and low echogenicity,peripheral halo,echo hom-ogeneity,calcifications,and so on.The misdiagnosis causes were investigated.Results The mass was mainly solid or cystic-solid mixed echo.The ultrasonic imaging features of thyroid follicular adenoma were non-peripheral halo or thin wall halo,hyperecho or isoecho,internal macrocalcifications and peripheral calcifications,homogeneous echo structure.Conclusion The ultrasonographic examination can provide the better diagnosis and differentiation diagnosis on thyroid follicular carcinoma.

Objective To observe the effect of catechol-O-methyltransferase inhibitor (COMTI) tolcapone on levodopa-treated patients with Parkinson's disease (PD) and on the motor function fluctuation, and safety taking this drug.Methods50 PD patients treated with levodopa were divided into the trial group and control group with 25 cases in each group. The patients in the trial group were given tolcapone 100 mg three times per day. The cases in the control group were given placebo with the same dose. The hepatic function of the patients was examined every month after administration. The time variety of patients' motor function fluctuation was recorded by the diary. The time of clinical observation was 6 months.ResultsThe UPDRS scores of the trial group in the first and second months after taking tolcapone were not significantly different from that of the control group ( P>0.05), but scores of the third to sixth months were significantly different from that of the control group ( P<0.05～0.01). There was a significant difference between UPDRS scores of the trial group before and after treatment ( P<0.01～0.001). The Honhe-Yahr scores of the trial group in the first and second months after treatment were not significantly different from that before treatment ( P>0.05), but scores of the third to sixth months were significantly different from that before treatment ( P<0.05). The motor function fluctuation of the patients in the trial group improved significantly after treatment ( P<0.05). The numbers of the cases had dry mouth, nausea and astriction were 3 respectively; those had acratia, insomnia and diarrhea were 2 respectively; those had muscular soreness, abdominal distention, hidrosis and fidget were 1 respectively. All adverse effects had a little influence to administration. The hepatic function of all patients had no significant change.ConclusionTolcapone can increase the curative effect of PD patients treated with levodopa and improve the motor function fluctuation, and is safety after taken.

OBJECTIVETo investigate the effects of gallic acid against Candida albicans biofilms in vitro.

METHODXTT reduction assay was performed to determine the effect of gallic acid on C. albicans biofilms and its adherence, and microscopic examination was conducted to assess the effect of gallic acid on morphogenesis of C. albicans biofilms; and cytotoxic assay was used to measure the adverse effects of gallic acid.

RESULTSMIC50, SMIC50 of gallic acid against C. albicans biofilms were 500, 1000 mg x L(-1), respectively; 100 mg x L(-1) and 1000 mg x L(-1) of gallic acid could inhibit the initial adherence and filamentous growth, and the agent showed poor cytotoxic activity.

CONCLUSIONgallic acid displayed potent activity against C. albicans biofilm.

Biofilms ; drug effects ; Candida albicans ; cytology ; drug effects ; physiology ; Cell Adhesion ; drug effects ; Dose-Response Relationship, Drug ; Gallic Acid ; pharmacology

OBJECTIVETo evaluate the adhesion, proliferation and differentiation of osteoblast-like cells on the ultraviolet (UV)-treated titanium in different storing environment, and to find a way to enhance the bioactivity of titanium and to prevent its age-related degradation.

METHODSAcid-etched titanium disks stored under ambient conditions for 4 weeks and treated with UV light for 48 h.Then disks were divided into three groups and placed in a sealed container for 0 h (no-stored,NO group) , 4 weeks (air-stored, AS group) or in a sealed container filled with nitrogen for 4 weeks (nitrogen-stored,NS group) respectively. A group of UV-untreated titanium served as negative control (NC group).The surface morphology was evaluated using scanning electron microscopy (SEM), and hydrophilicity of disks were measured using contact angle measuring device. Cell counting kit-8 was used to measure the cell adhesion and proliferation. Cell differentiation was evaluated by testing alkaline phosphatase (ALP) activity using ALP reagent kit.

RESULTSThere was no difference in surface topography among groups.Contact angels in NS group [(67.70 ± 3.59)°] and NO group [(0.70 ± 0.28)°] were smaller than the others (P < 0.05). Cell adhesion in NS group at 2 h and 4 h point was (0.237 ± 0.006) and (0.578 ± 0.039), respectively, and proliferation at 3 d and 5 d point was (0.743 ± 0.026) and (1.548 ± 0.046) respectively, which were significantly higher than those in AS group [(0.158 ± 0.036), (0.400 ± 0.010), (0.499 ± 0.019) and (1.174 ± 0.062)] and in NC group [(0.161 ± 0.024), (0.390 ± 0.011), (0.508 ± 0.015) and (1.209 ± 0.025)] at the same time point (P < 0.05). How ever the results mention above in the NS group were lower than those in the NO group (P < 0.05). No difference were found between data from the AS group and NS group (P > 0.05). Osteoblast-like cells had an abundant spread on NS and NO group during 2 h incubation, but did not exactly spread on AS and NC group after incubation for 4 h. No difference were found in ALP among groups.

CONCLUSIONSUV treatment can enhance bioactivity of titanium, and nitrogen storage can slow down its biological aging.

Alkaline Phosphatase ; metabolism ; Animals ; Biocompatible Materials ; chemistry ; Cell Adhesion ; radiation effects ; Cell Differentiation ; radiation effects ; Cell Proliferation ; radiation effects ; Cells, Cultured ; Mice ; Microscopy, Electron, Scanning ; Nitrogen ; chemistry ; Osteoblasts ; cytology ; metabolism ; Surface Properties ; Titanium ; chemistry ; radiation effects ; Ultraviolet Rays