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Objective To evaluate the inductive effect of interferon-γ(IFN-γ) combined with tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) on programmed necrosis of the human immortalized keratinocyte cell line HaCaT,and to explore its mechanisms.Methods In vitro cultured HaCaT cells were divided into several groups:negative control group receiving no treatment,IFN-γ group treated with 50 μg/L IFN-γ,TRAIL group treated with 4 μg/L TRAIL,and cytokine combination group treated with 50 μg/L IFN-γ and 4 μg/L TRAIL or zVAD combination group pretreated with 40 μmo/L zVAD for 1 hour followed by the treatment with 50 μg/L IFN-γand 4 μg/L TRAIL.After 48-hour treatment,the morphology of HaCaT cells were observed under a light microscope,methyl-thiazolyl-tetrazolium assay was performed to evaluate the inhibitory effect of the treatment on the proliferation of HaCaT cells,and double staining flow cytometry to detect the necrosis of HaCaT cells.Meanwhile,real-time fluorescence-based quantitative PCR (qPCR) was conducted to determine the mRNA expression of receptor interaction protein kinase 3 (RIP3) and mixed lineage kinase domain-like protein (MLKL),Western blot analysis to determine the expression of RIP1,RIP3,MLKL proteins and their phosphorylated forms (pRIP1,pRIP3,pMLKL),immunofluorescent staining to observe the distribution of pRIP3 and pMLKL in HaCaT cells,and the level of reactive oxygen species (ROS) in HaCaT cells in the above groups was detected by the fluorescence probe DCFH-DA.Statistical analysis was carried out with SPSS 22 software by using one-way analysis of variance (ANOVA) for comparing indices among different groups,and least significant difference (LSD)-t test for multiple comparisons.Results After 48-hour treatment,HaCaT cells in the cytokine combination group and zVAD combination group showed necrosis-like morphologic features.Methyl-thiazolyl-tetrazoliumassay revealed significant differences in the survival rate of HaCaT cells among the IFN-γgroup,TRAIL group,cytokine combination group,zVAD combination group and negative control group (73.16％ ± 5.71％,81.46％ ± 4.68％,72.18％ ± 2.93％,69.67％ ± 3.24％ and 100％,respectively;F =24.34,P ＜ 0.001).The necrosis rate of HaCaT cells was notably higher in the cytokine combination group and zVAD combination group (9.86％ ± 1.31％,10.33％ ± 2.16％,respectively) than in the negative control group (5.26％ ± 0.91％,t =4.61,5.07,respectively,both P ＜ 0.05).qPCR revealed that the mRNA expression of RIP3 and MLKL significantly increased in the cytokine combination group and zVAD combination group compared with the negative control group (tRIP3 =0.99,1.84,tMLKL =1.51,2.17,respectively,all P ＜ 0.05).Western blot analysis suggested that the protein expression of RIP1,RIP3,MLKL,pRIP1,pRIP3 and pMLKL significantly increased in the cytokine combination group compared with the negative control group (all P ＜ 0.05),and the zVAD combination group showed significantly decreased caspase 8 expression and increased expression of the above proteins compared with the cytokine combination group.Fluorescence microscopy showed that enhanced green dot-like or clump-like fluorescent spots (representing pRIP3) could be observed in the cytoplasm,and red fluorescent spots (representing pMLKL) could be seen on the cell membrane in the cytokine combination group.The average fluorescence intensity of ROS was significantly higher in the cytokine combination group than in the negative control group (t =702.00,P ＜ 0.05).Conclusion IFN-γcombined with TRAIL can induce the programmed necrosis of HaCaT cells with increased level of ROS.
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Objective To investigate the expressions of miR-17 and miR-20a in CD4+ T cells from patients with systemic lupus erythematosus (SLE) and their correlations with disease activity.Methods CD4+ T cells were isolated from the venous blood of 30 patients with SLE and 18 healthy human controls.RNA was extracted from the CD4+T cells,and real-time fluorescence-based quantitative PCR (qPCR) was performed to determine the expression levels of miR-17 and miR-20a.The intergroup comparison of miR-17 and miR-20a expression levels was done using t test and Mann-Whitney test,and the correlations of miR-17 and miR-20a expressions with clinical parameters were assessed using Pearson or Spearman correlation coefficients.Results The patients with SLE showed increased expression levels (2-△c~) of miR-17 and miR-20a compared with the healthy controls (0.24 ± 0.08 vs.0.15 ± 0.06 for miR-17,0.19 ± 0.10 vs.0.12 ± 0.09 for miR-20a,both P ＜ 0.05).The miR-17 and miR-20a expression levels were also significantly higher in patients with active SLE than in those with inactive SLE and the healthy controls (0.27 ± 0.07 vs.0.20 ± 0.05 and 0.16 ± 0.08 for miR-17,0.22 ± 0.10 vs.0.15-0.07 and 0.13 ± 0.09 for miR-20a,all P ＜ 0.05),but similar between the patients with inactive SLE and healthy controls (both P ＞ 0.05).The expression levels of both miR-17 and miR-20a were positively correlated with SLE disease activity index (SLEDAI)(r =0.45,0.38,respectively,both P ＜ 0.05) and anti-dsDNA antibody level (r =0.54,0.46,respectively,both P ＜0.05),but negatively correlated with serum C3 levels (r =-0.43,-0.42,respectively,both P ＜ 0.05).Condusions The expressions of miR-17 and miR-20a are increased in CD4+ T cells from patients with SLE,and correlated with the disease activity in SLE.
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Several problems were existed in present military hygiene teaching.In recent years,integrated reforms were done about the improving undergraduate and teacher's activities,strictly choosing teaching contents,doing well extra curriculum,and serious examination in nursing teaching.Furthermore,the new presumptions were also discussed.
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BACKGROUND: Decreasing of absorption of zinc from small intestine can induce dermatitis, alopecia, growth and developmental disorder and so on. It is not very clear that how to keep homeostasis when there is low zinc concentration. The discoveries of zinc transporters and relative researches provide new study direction.OBJECTIVE: To study the effect of different zinc concentration on the expressions of divalent metal transporter 1 (DMT1) and human zinc-regulated transporter (ZRT), iron-regulated transporter (IRT)-like protein (ZIP) 4mRNA, and analyze the potential pathway of absorption of zinc from small intestine in low zinc concentration.DESIGN: Blank-controlled experiment.SETTING: Department of Military Hygiene, Navy Faculty, Second Military Medical University of Chinese PLA.MATERIALS: The experiment was accomplished in Department of Military Hygiene, Navy Faculty of Second Military Medical University of Chinese PLA between October 2004 and May 2005. Materials were human colon adenocarcinoma cells Caco2 that were purchased from Shanghai Institute of Cell of Chinese Academy of Sciences.Time-dependent effect: The expression of DMT1 and ZIP4 mRNA was detected with reverse transcription-polymerase chain reaction (RT-PCR) at 0,dependent effect: The expression of DMT1 and ZIP4 mRNA were measured after the 0, 2.5, 5, 7.5 and 10 μmol/L TPEN exposure, respectively, by RT-PCR.MAIN OUTCOME MEASURES: Time-and dose-dependent effect of zinc on the expression of DMT1 and ZIP4 mRNA in Caco2 cells.RESULTS: ①Time-dependent effect: Compared with 0 hour, the expression of DMT1 mRNA obviously increased at 6, 8 and 10 hours (P ＜ 0.05 ), but there was no significant change at 2 and 4 hours. The expression of ZIP4 mRNA markedly increased in all timing, and ZIP4 mRNA level at the 6th hour reached the peak with the prolongation of low zinc duration,which was 2.1 times ofthat at 0 hour. ②Dose-dependent effect: The expression of DMT1 mRNA distinctly increased at the concentration of 7.5 and 10 μmol/L TPEN exposure as compared with that of the control group (P ＜ 0.05), but there was no significant change at 2.5 and 5 μmol/L. The ZIP4 mRNA expression increased with the increasing of the concentration of TPEN, the expression of ZIP4 mRNA was 2.7 times of that at 0 μmol/L. CONCLUSION: Zinc can regulate the expression of DMT1 and ZIP4 mRNA in Caco2 cells, and there is time-and dose-dependent effect. But the mRNA expression of ZIP4 is more sensitive and prompt than DMT1. Cells can upregulate the expression of DMT1 and ZIP4 mRNA to keep the homeostasis in low zinc condition.
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In teaching the course of Environmental Hygiene to the senior major of the four-year undergraduates of the Department of Public Administration,we have tried to reform the teaching program for the course of Public Place Hygiene.The related theories are taught right on the spot of public places instead of in classrooms.Students are organized to carry out such activities on their own as studying subject matters,consulting related documents and data,working out and implementing programs for on-the-spot monitoring and inspection,and then exchanging and discussing the results of monitoring and inspection,so that their interest in the courses is increased and their understanding of what they have learned is deepened.In this way they are able to apply their knowledge to practice,improve their practical capabilities,and enhance their overall capacity for analyzing and finding solutions to problems they are faced with.
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The new course standard of nursing specialty in military hygiene was established against the main problems in military hygiene teaching and considering the peacetime and wartime need of nursing post.In this paper,guiding ideology,content choosing and the request of operational skill of this standard were discussed in detail.
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Objective:To observe the growth and development of the weaned mice fed with different levels of dietary zinc and to explore the expression of zinc transporter 3(ZnT3) mRNA induced by different dietary zinc intakes. Methods: Twenty male weaned mice (postnatal day 21) were divided into 4 groups: zinc deficient (ZD), zinc adequate(ZA), zinc supplemental (ZS) and pair fed(PF). Mice were fed with different levels of dietary zinc for 3 weeks (from postnatal day 21 to postnatal day 42) ;the zinc contents of ZD, ZA, ZS and PF group were 1 mg/kg, 30 mg/kg, 180 mg/kg and 180 mg/kg respectively. From postnatal day 21 to postnatal day 42, the diet intakes and weight of the mice were measured everyday. On postnatal day 42, the mice were sacrificed and tissues were immediately isolated and frozen lor RNA extraction. The serum zinc concentrations were measured by AAS and the expression of ZnT3 mRNA was determined by semiquantitative RT-PCR. Results: The dietary intakes and weight of ZD mice were much lower than that of other groups(P3
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Objective:To study the concentration response and time course of zinc transporter 1 (ZnT 1) and metallothionein (MT1/MT2) mRNA,as well as the cell viability to zinc exposure in primary cultured new born rats hippocampal neuron s. Methods: The cell viability were determined by trypan blue s taining at various concentrations of Zn 2+ (0, 50, 100, 125, 150, 175, 200 ?mol/L). The expression of ZnT 1, MT1 and MT2 mRNA to various concentrations of zinc exposure(0, 50, 75, 100, 125, 150 ?mol/L) and to 100 ?mol/L zinc exposure for different times (0, 2, 4, 6, 8 h) were determined b y real time fluorescent quantitative PCR. Results: The viabilit y of the neurons decreased significantly after 48 h once the concentration of Zn 2+ exceeded 125 ?mol/L,and the expression of ZnT 1 mRNA was in proportion to the increment of zinc.The expression of MT1/MT2 mRNA reached a plateau when the zinc concentration exceeding 75 ?mol/L. The expression of ZnT 1 mRNA peaked on about 2 h.However, the expression of MT1/MT2 mRNA reached its maximal around 6 h at the concentration of 100 ?mol/L. Conclusion: These results imply that although both MTs and ZnT 1 can attenuate the zinc toxicity, they may play different roles at different phases. ZnT 1 enhance the efflux of zinc prior to the sequesteration by MTs. Th e action of ZnT 1 may be durable, but the role of MTs may be satiable.