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OBJECTIVE: To investigate the role of N-methyl-D-aspartate receptor(NMDAR)/cyclic adenosine monophosphate(cAMP)/protein kinase A(PKA) signaling pathways in regulating 2-chloroethanol-induced aquaporin-4(AQP4) expression in astrocytes(AS). METHODS: i) AS in logarithmic growth phase were treated with 2-chloroethanol at the doses of 0.0, 7.5, 15.0 and 30.0 mmol/L for 12 hours, and the cells were collected for detection. ii) The AS in logarithmic growth phase were divided into blank control group, inhibitor control group, 2-chloroethanol group, and inhibitor intervention group. The inhibitor included dizocilpine(MK-801) and N-(2-[p-bromocinnamylamino-]ethyl)-5-isoquinolinesulfonamide(H89). The blank control group did not receive any treatment. The inhibitor control group was treated with a concentration of 10.0 μmmol/L MK-801 or 15.0 μmmol/L H89. The MK-801 intervention group was pretreated with MK-801 at a concentration of 10.0 μmmol/L for 30 minutes. The H89 intervention group was pretreated with H89 at a concentration of 15.0 μmmol/L for 1 hour. After the intervention, the AS in 2-chloroethanol group and MK-801, H89 intervention group were stimulated with 2-chloroethanol at a dose of 30.0 mmol/L for 12 hours. iii) The AS in each group were collected and used for Western blotting and real-time fluorescence quantitative polymerase chain reaction analysis to detect the protein and mRNA expression of AQP4, NMDAR receptor main subunit(NR1), NMDAR receptor 2 B subunit(NR2 B) and calmodulin dependent protein kinaseⅡ(CaMKⅡ). The Western blotting was adopted to detect the expression of phosphorylase-CaMKⅡ(p-CaMKⅡ) and PKA. Colorimetric method was used to detect the concentration of calcium(Ca~(2+)) in AS. The enzyme-linked adsorption test was used to measure adenylate cyclase(AC) activity and cAMP levels. RESULTS: i) The relative expression of protein and mRNA of AQP4, NR1 and NR2 B, PKA at protein level and CaMKⅡ at mRNA level, and the ratio of p-CaMKⅡ/CaMKⅡ protein, the concentration of Ca~(2+), AC activity and cAMP level in 30.0 mmol/L group were higher then those of 0.0 mmol/L group in AS(P<0.05). The relative protein expression of PKA and the concentration of Ca~(2+) increased with the increase of 2-chloroethanol(P<0.05). ii) The relative protein expression of AQP4 and the concentration of Ca~(2+) in the 2-chloroethanol group were higher than that of the blank control group and MK-801 control group(P<0.05). The relative protein expression of AQP4 and the concentration of Ca~(2+) in MK-801 intervention group were lower than that in 2-chloroethanol group(P<0.05). The relative protein expression of AQP4 and PKA in 2-chloroethanol group were higher than that of the blank control group and H89 control group(P<0.05). The relative protein expression of AQP4 and PKA in H89 intervention group was lower than that in 2-chloroethanol group(P<0.05). CONCLUSION: The 2-chloroethanol timulation induces the expression of AQP4 by activating NMDAR/cAMP/PKA signaling pathway in AS.
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Objective To investigate the impairment mechanism of learning and memory function induced by arsenite exposure through studying the effects of sodium arsenite on gliotransmitter release from astrocytes.Methods Primary cultured astrocytes were isolated from neonatal (0-3 days) Wistar rats and determined by glial fibrillary acidic protein (GFAP) immunofluorescence staining.The primary cultured astrocytes were randomly divided into four groups,in which astrocytes were exposed to 0.0,2.5,5.0,or 10.0 μmol/L sodium arsenite,respectively,for 24 h.Intracellular free Ca2+ concentration ([Ca2+]i) in astrocytes was measured by fluorescence dual wavelength spectrophotometer;,concentrations of glutamate,D-serine,glycine and γ-aminobutyric acid were measured by high performance liquid chromatography (HPLC).Results More than 95％ cells were positive for GFAP immunofluorescence staining.The difference of [Ca2+]i among groups treated with sodium arsenite was statistically significant (F =20.030,P ＜ 0.05).[Ca2+]i increased significantly in group treated with 10.0 μmol/L sodium arsenite [(263.27 ± 14.80)nmol/L] compared with those in groups treated with 0.0,2.5,5.0 μmol/L sodium arsenite [(204.24 ± 27.21),(214.49 ± 21.85),(232.74 ± 23.14)nmol/L,all P ＜ 0.05].The differences of the levels of D-serine,glycine and γ-aminobutyric acidamong groups treated with sodium arsenite were significant (F =26.599,33.539,5.599,all P ＜ 0.05).The levels of D-serine [(21.580 ± 1.313),(21.936 ± 1.539),(23.401 ± 1.648)μmol/L],glycine [(26.353 ± 2.449),(29.711 ± 1.530),(29.234 ± 2.057)μmol/L] and γ-aminobutyric acid [(27.277 ± 3.421),(30.213 ± 2.098),(29.364 ± 2.588)μmol/L] released by astrocytes increased significantly in groups treated with 2.5,5.0,10.0 μmol/L sodium arsenite compared with those in groups treated with 0.0 μmol/L sodium arsenite [(16.017 ± 1.046),(16.763 ± 3.007),(22.736 ± 4.139)μmol/L,all P ＜ 0.05].Conclusion Arsenite could affect gliotransmitter release from astrocytes,and further impair learning and memory function.
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Objective To observe and evaluate the application of case-based learning in occupational health teaching of clinical medicine students.Methods Seven-year undergraduates majored in clinical medicine were selected as study subjects,divided into two groups,and received case-based learning(55 persons,story case for theory course,field case for experiment course)and traditional teaching(56 persons,theoretical instruction for theory course,instructional video for experiment course),respectively.T test and x2 test were used to analyze and compare students' after-school evaluation results and the questionnaire survey results of both groups respectively.Results The total score of the after-school evaluation[(11.84 ± 2.86) vs.(9.80 ± 3.39),t=-3.419],basic cognitive ability score[(4.80 ± 1.08) vs.(4.05 ± 1.43),t=-3.097] and analyzing ability score[(7.04 ±2.21) vs.(5.75 ± 2.59),t=-2.813] of students in the case-based learning group were significantly higher than those of the students in the traditional teaching group(P＜0.05).Questionnaire survey showed that the number and proportion of case teaching group of students held a positive attitude towards the teaching methods used in improving students' ability to analyze problems(x2=4.757),increasing study interest (x2=4.371),promoting knowledge mastery (x2=4.276) is higher than traditional teaching group (P＜0.05).Conclusion Case-based learning can inspire students' learning initiative,improve their creative ability to use knowledge,analyze and solve practical problems,so as to promote the course scores.
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<p><b>OBJECTIVE</b>To investigate the effect of lead-exposed astrocyte conditioned medium (ACM) on the synaptic formation of neurons and to provide reference for the mechanism of lead neurotoxicity.</p><p><b>METHODS</b>Astrocytes were cultured in the medium containing 50, 100, 200, 400, and 800 µmol/L lead acetate for 72 h. Alamar Blue was used to assess the cell viability of astrocytes, and then ACM was collected. Primarily cultured neurons were divided into six groups: pure culture group, non-glutamic acid (Glu)-induced ACM treatment group, Glu-induced lead-free ACM treatment group, and Glu-induced 50, 100, and 200 µmol/L lead acetate-exposed ACM treatment groups. Neurons were collected after being cultured in ACM for 24, 48, or 72 h. The content of synaptophysin (SYP) in neurons was determined by Western blot. The SYP expression in neurons was measured by immunofluorescence after being cultured in ACMfor 72 h.</p><p><b>RESULTS</b>In all lead-exposed groups, the cell viability of astrocytes declined with increasing concentration of lead (P < 0.05). The Western blot showed that compared with the pure culture group, the non-Glu-induced ACM treatment group and Glu-induced lead- free ACM treatment group had significantly increased content of SYP in neurons (P < 0.01); compared with the non-Glu-induced ACM treatment group, the Glu-induced ACM treatment groups had significantly reduced SYP expression in neurons (P < 0.05); compared with the Glu-induced lead-free ACM treatment group, all lead-exposed ACM treatment groups had the content of SYP in neurons significantly reduced with increasing concentration of lead after 72-h culture (P < 0.01), the 200 µmol/L lead-exposed ACM treatment group had significantly reduced content of SYP in neurons after 48-h culture (P < 0.01), and all lead-exposed ACM treatment groups showed no significant changes in the content of SYP in neurons after 24-h culture. Double-labeling immunofluorescence of SYP showed that all lead-exposed ACM treatment groups had a significant decrease in the number of SYP-fluorescent particles after 72-h culture (P < 0.05).</p><p><b>CONCLUSION</b>Astrocytes promote synaptic formation of neurons, which may be inhibited during lead exposure.</p>
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Astrocytes , Physiology , Cell Survival , Cells, Cultured , Culture Media, Conditioned , Metabolism , Glutamic Acid , Metabolism , Lead , Toxicity , Neurons , Synapses , PhysiologyABSTRACT
Objective To observe the validity and feasibility of PBL teaching method in occupational health and occupational medicine theory teaching.Methods Sixty-one five-year undergraduates majored in preventive medicine were selected and self-control was applied.Two teaching contents which had similar teaching requirements were selected,one applied PBL teaching method and the other applied traditional teaching method.Students' results and passing rate in examination were analyzed and compared.SPSS 16.0 software was used for statistical analysis and data were manifested as ((x-) ± s).T test was used to do comparison between average results and chi-square test was used to do comparison between rates.SAS 9.0 software was employed and Fisher's exact probability analysis was performed on percent of pass.P＜0.05 stands for statistically significant differences.Results Test scores of the students taught by PBL teaching method were significantly higher than those of students taught by traditional teaching method (P＜0.05).Both female and male students' test scores were improved (P＜0.05) and increase of female students' test scores were more obvious than male students'.Passing rate of female students taught by PBL teaching method was significantly higher than female students taught by traditional teaching method (P＜0.05).Conclusion PBL teaching method applied in occupational health and occupational medicine theory teaching can significandy improve the quality of teaching.
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Objective To observe the level of circulating CD31 +/CD42b-endothelial microparticles in patients with ST-elevated myocardial infarction (STEMI),and discuss the correlation between CD31 +/CD42b-and traditional myocardial injury index.Methods A total of 22 healthy subjects and 44 patients with angiographically confirmed coronary atherosclerotic lesions collected from January 2010 to December 2010 were studied prospectively.The patients were divided into SAP (stable angina pectoris,Canadian Cardiovascular Society,CCS Ⅱ to Ⅲ ) group (n =22) and STEMI group (n =22).The level of circulating CD31 +/CD42b-endothelial microparticles was detected by flow cytometric device after admission; creatine kinase (CK) and its isoenzymes (CK-MB) were detected by using biochemical analyzer; C-reactive protein was determined by a highly sensitive latex-enhanced turbidimetric immunoassay with a low detection limit of 0.25 mg/dl (IMMAGE,Beckman Coulter; Reagent from Orion Diagn Co.Ltd.,Vantaa,Finland).Cardiac troponin I was measured by chemiluminescence immunoassay (ACCESS2 from Beckman Coulter).In 22 STEMI patients,blood sample was taken not only after admission but subsequently at 4-hour intervals during the first 48 hours.Peak levels of myocardial enzymes after injury ( creatine kinase,CK; creatine kinase MB isoenzyme,CK-MB; c troponins I,cTnI) and high sensitive C reactive protein (hs-CRP) values were determined.The correlation between circulating level of CD31 +/CD42b(-) and peak levels of myocardial biomarkers after injury were analyzed.Results The endothelial micro-particles were significantly higher in STEMI patients than those in either SAP group or normal group ( P ＜ 0.01 ),whereas there was no difference between the latter two groups.In STEMI patients studied in these cross sectional study,circulating CD31 +/CD42b-microparticles were positively correlated with peak level of myocardial biomarkers after injury.Moreover,the correlations between myocardial biomarkers after injury ( CK,CK-MB,cTnl and hs-CRP) and circulating CD31 +/CD42b-microparticles were r =0.489,P =0.021; r =0.501,P=0.018; r=0.491,P=0.02; r=0.612,P=0.002.Conclusions The level of circulating blood CD31 +/CD42b-endothelial micro-particle was expected to become a predictive marker in STEMI.
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ObjectiveTo investigate the effect of immunoglobulin G (IgG) extravasated from blood circulation on the expression of toll-like receptor 4 (TLR4) induced by peripheral lipopolysaccharide (LPS) in rat brain. Methods The rats were divided into four groups in random, 5 rats in each. Group one received LPS 100μg/kg by intraperitoneal administration, normal saline was given by intravenous injection 6 hours later; group two was injected with adrenalin (AD) 15μg/kg intravenously; group three was treated with LPS intraperitoneally, AD was injected 6 hours later; group four was injected normal saline intravenously as control. For all groups, the animals were sacrificed 30 min after the last injection, and the brains were taken for investigation of the TLR4 expressions by immunofluorescence staining and RT-PCR. Result Immunofluorescence staining showed that IgG immunoreactive product was patch-like, distributed in the brain parenchyma in all the animals that received AD. In the LPS+normal saline group, IgG was found merely around the blood vessels. Meanwhile, in LPS+AD animals, TLR4 immunoreactive product coexisted with microglia marker Iba-1 within the IgG extravasated area. The double-labeled cells dispersed in the brain parenchyma and near to the cerebral vessels. In the LPS+saline group, TLR4 positive cells were endothelial-like. RT-PCR results indicated that the expression level of TLR4 in the LPS+AD group were significantly higher than that in the LPS+saline group or AD group or the saline control (P<0.01). Conclusion Extravasated circulating IgG may enhance the TLR4 expression in the rat brain induced by peripheral LPS.
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Objective To study the influence of the measurement of the platelet-monocyte aggregates (PMAs) by using of flow cytometry (FCM).Methods Anticoagulated peripheral venous bloods from nine healthy donors were incubated with a PE-CD14 MAb (monocyte marker) and a FITC-CD42a MAb (platelet marker) for 20 min and the formations of PMAs were measured by use of FCM.The factors such as fixative, anticoagulant, storage time and temperature were analyzed.Results The PMAs of citrated whole blood increased with the time elapsed in 6 h after blood drawing when they were stored in the room temperature.The PMAs of each time point showed significant difference (P
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<p><b>OBJECTIVE</b>To explore whether there is difference in arsenicals-induced DNA damage of human lymphocyte.</p><p><b>METHODS</b>Lymphocyte were sterilely collected from healthy donor and exposed to sodium arsenite (AsIII), sodium arsenate(AsV) and methyl sodium arsenate(MAsv) at 1,5,10,20 and 50 mumol/L. After incubation of 24 hours, cells were collected by centrifugation and DNA damage was detected by single cell gel electrophoresis (SCGE).</p><p><b>RESULTS</b>The comet frequency distribution of all groups except 1 mumol/L group of MAsV were significantly different from that of control. The comet length of all groups except 1 mumol/L group of AsV and 1.5 mumol/L groups of MAsV were significantly higher than that of control. There were correlations between the doses of arsenicals and the ratios of comet cell or length of comet(rAsIII = 0.8134, rAsV = 0.8734, rMAsV = 0.8994).</p><p><b>CONCLUSION</b>DNA damage in human lymphocyte were induced by all the three arsenicals. A dose-effect relationship was observed between exposure doses of the same arsenical and DNA damage. With different arsenicals but the same exposure dose, the DNA damage level was as follow: AsIII > AsV > MAsV.</p>
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Humans , Arsenates , Toxicity , Arsenites , Toxicity , Comet Assay , DNA Damage , Dose-Response Relationship, Drug , LymphocytesABSTRACT
<p><b>OBJECTIVE</b>To investigate the change in the antibiotic resistant spectrum of Pseudomonas aeruginosa (PA) isolated from burn wounds and the production of inducible beta-lactamase.</p><p><b>METHODS</b>VITEK-AMS system (total automatic bacterial identification and drug sensitivity system) and E-test concentration gradient were employed to perform bacterial identification and antibiotic sensitivity tests. K-B method was applied to detect inducible enzyme.</p><p><b>RESULTS</b>The resistance of PA to Cephalosporin and Imipenem was increased in the past 4 years from June of 1996 to June of 2000. Whereas the resistance to Cefoperazone/Sulbactam was least. There was an obvious difference of the resistance of PA to antibiotics during the 4 years (P < 0.05). The resistant rate to Imipenem ranged from 20% to 40%. PA was able to produce inducible enzymes among 120 strains of wild type of PA occupying 72.5% with Imipenam as the inducing agent.</p><p><b>CONCLUSION</b>The analysis of the antibiotic resistance of PA and the detection of inducible enzymes could be monitored from time to time and helpful in the correction of the use of antibiotics. Constant monitoring of antibiotic resistance might be beneficial to the prevention of outbreak of epidemics of PA infection in a burn unit.</p>
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Humans , Burns , Microbiology , Cephalosporins , Pharmacology , Drug Resistance, Bacterial , Enzyme Induction , Imipenem , Pharmacology , Microbial Sensitivity Tests , Pseudomonas aeruginosaABSTRACT
AIM: To investigate whether celecoxib,a cyclooxygenase-2(COX-2) inhibitor,potentiates the anti-leukemia activity of STI571 in K562 cells.METHODS: K562 cells were treated with STI571,celecoxib or combination of both at different concentrations in suspension culture.Cell proliferation was documented by MTT assay,and cell apoptosis was determined by flow cytometry and morphology.Meanwhile,RT-PCR was applied to analyze the probable mechanism underlying the effects of the drugs.RESULTS: The combination of STI571 and celecoxib dramatically suppressed the proliferation of K562 cells,in which 0.25 ?mol/L STI571 and 40.0 ?mol/L celecoxib enhanced the inhibiting rate to 76.1%?1.6%.Furthermore,the combining administration of drugs significantly promoted the apoptosis induced by STI571,which showed characteristic changes of morphologic features and increase in sub-G_1 cells.By using RT-PCR technique,the expression of COX-2 had no decline by single administration of celecoxib or STI571.However,a progressive down-regulation was caused by coadministration of two drugs.In contrast with COX-2,the expression of VEGF had no changes at any time.CONCLUSION: The administration of celecoxib alone only inhibits the proliferation of K562 cells.Combination treatment with STI571 and celecoxib promotes the apoptosis induced by STI571.
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AIM To establish an animal experimental model for study on prevention of cisplatin toxicity and explore the possible mechanisms of the toxicity induced by cisplatin administration. METHODS Cisplatin was administered i.p consecutively for five days to male mice weighted from 28 to 30 g. The toxic effects induced by different doses of cisplatin on hearing, liver and kidney were determined. RESULTS Dose dependent decrease of body weight, abnormality of kidney and liver coefficients, levels of BUN and activities of ALT in serum were induced by cisplatin administration. Furthermore, Levels of GSH, activities of GSH Px and SOD increased significantly in kidney. Reversely, levels of GSH, activities of GSH Px and SOD in liver decreased and levels of LPO increased significantly in animals given cisplatin compared with those in control animals. CONCLUSION Obvious damage on hearing, liver and kidney of mice could be induced after consecutively 5 days administration of cisplatin with doses range from 3 0 to 4 0 mg?kg -1 (body weight). Oxidative damage is one of the mechanisms of these toxic effects on liver and kidney induced by cisplatin. But for different organs or at different stages of cisplatin administration, the main mechanism may be different.