Objective To investigate the effect of channel density on the gating properties of Anoctamin 1(Ano1,TMEM16A)Ca2+?activated chlo?ride channel. Methods Ano1 expression plasmids were transiently transfected into HEK293 cells. High density and low density of Ano1 was ob?tained after expressing the protein for 24 h and 6 h,respectively. Electrophysiological recordings were performed in the whole?cell patch clamp con?figuration. The activation kinetics of current traces was fitted by exponentials. Results The current density was significantly higher in cells express?ing Ano1 for 24 h than those expressing Ano1 for 6 h(P<0.05). The activation of Ano1 current in cells with low density was well fit by a single expo?nential withτslow of 292.71±38.11 ms. The activation of Ano1 current in cells with high density was well fit by two exponentials withτfast of 47.78±4.58 ms andτslow of 385.74±71.44 ms. ANO1 current in cells with high density has a rapid active component(τfast)more than low density. There was no significantly different of theτslow between cells with high density and low density of Ano1(P>0.05). Conclusion Our findings suggested that chan?nel density regulates the gating of Ano1. High channel density promotes activation of Ano1.

Objective:To explore the application and effect of team-based learning (TBL) combined with flipped classroom in the teaching of physiology.Methods:A total of 70 medical students were selected and randomly divided into two groups, experimental group ( n=34) and control group ( n=36). Both groups studied digestion physiology by different method. In the experimental group, the students were taught by TBL combined with flipped classroom. The control group was given traditional teaching. The students studied and discussed problems in a team, and shared the answers in the flipped classroom. The effects of teaching were evaluated by the final test scores and the self-made questionnaire. SPSS 17.0 was used for t test on data comparison between the two groups. Results:The test scores of digestion physiology in the experimental group were (5.47±1.02) points, which were significantly higher than those of the control group (4.42±1.63) points, with significant differences ( P=0.020). A total of 34 questionnaires were issued and 34 valid questionnaires were collected, with an effective rate of 100%. The questionnaire results showed that TBL combined with flipped classroom was accepted and approved by about 82% of students in the experimental group who agreed that the teaching model helps enhance students' initiative and interest in learning, develop their sense of cooperation and comprehensive application ability. Conclusion:TBL combined with flipped classroom is feasible and effective in the physiological teaching, and it can be popularized in medical courses.

ObjectiveTo investigate the effects of propofol on calcium/calmodulin-dependent protein kinase Ⅱ α ( CaMK Ⅱ α) activity in hippocampus in mentally depressed rats after electroconvulsive therapy (ECT).MethodsHealthy adult male SD rats aged 2-3 months weighing 180-220 g were used in this study.Mentally depressed model was induced by chronic unpredictable mild stress.Forty mentally depressed rats were randomly divided into 4 groups (n =10 each): mental depression group (group D),propofol group (group P),ECT group (group E),propofol + ECT group (group DPE).Groups D and P received intraperitoneal normal saline 8 ml/kg or propofol 80 mg/kg once a day for 7 consecutive days respectively.Group E received ECT once a day for 7 consecutive days.Group DPE received propofol 80 mg/kg + ECT once a day for 7 consecutive days.Sucrose preference test was performed at 1 d before and 1 d after treatment,and Morris water maze test was performed at 1 d before and 3 d after treatment.The rats were sacrificed after Morris water maze test,and hippocampi were removed for determination of CaMK Ⅱ α and phosphorylated CaMK Ⅱ α(pCaMK Ⅱ α )expression,and pCaMK Ⅱ α/CaMK Ⅱ α ratio was caculated.ResultsCompared with group D,the sucrose preference percentage was significantly increased in groups E and DPE,the escape latency prolonged and space exploration time shortened,and the expression of CaMK Ⅱ α and pCaMK Ⅱ α down-regulated,pCaMK Ⅱ α/CaMK Ⅱ α ratio decreased in group E,the escape latency was significantly shortened and space exploration time prolonged,and the expression of pCaMK Ⅱ α up-regulated in group DPE ( P ＜ 0.05).Compared with group E,the escape latency was significantly shortened,space exploration time prolonged,and the expression of CaMK Ⅱ α and pCaMK Ⅱ α up-regulated,and pCaMK Ⅱ α/CaMK Ⅱ α ratio increased in group DPE ( P ＜ 0.05).ConclusionPropofol can reduce the cognition impairment induced by ECT in mentally depressed rats through enhancing CaMK Ⅱ α activity in hippocampus.

In this study ,a wild type Salmonella typhimurium (S .typhimurium) strain was isolated and identified in Hong Kong (S129) ,then the asdA gene was knocked out and replaced with kanamycin resistant gene in a Salmonella typhi-murium strain S129 using the λ RED-mediated recombination method .The constructed mutant asdAΔS129 was validated by culturing in the presence or absence of 2 ,6-diaminopimelic acid (DAP) growth in vitro and evaluating its virulence in BALB/c mice challenge assay .Therefore ,this study has demonstrated that an asdA mutant Salmonella typhimurium has been success-fully constructed .

BACKGROUND: Exercise has been proved to accelerate the proliferation of intervertebral disc cells and extracellular matrix production in healthy rats. For the degenerative intervertebral disc, whether exercise also has positive effects on its cell proliferation, extracellular matrix production or pain relief remains unclear. OBJECTIVE: To investigate the effect of exercise on the extracellular matrix production in a rat model of intervertebral disc degeneration.METHODS: A rat model of intervertebral disc degeneration was prepared by Freund's complete adjuvant injection into the intervertebral disc at L5-6 levels. Then, the model rats were allowed to have a rest for 2 weeks. All rats were then randomly divided into exercise and control groups. Rats in the exercise group were forced to run every day, while the controls allowed free activities in the cage. The behavioral tests were performed at 7, 14, 28, 42, 56 and 70 days after modeling; meanwhile, the intervertebral disc samples were collected used for alcian blue staining and immunohistochemical staining to detect the levels of proteoglycan, aggrecan and collagen type Ⅱ in the intervertebral disc cells, respectively.RESULTS AND CONCLUSION: Vocalization threshold on the rat back of punctured disc was significantly decreased, while grooming and wet-dog shaking were significantly increased at 7 days after modeling compared with the baseline (P < 0.05), suggesting that Freund's complete adjuvant injection successfully induces disc degeneration, hyperalgesia and abnormal behaviors. Further, the vocalization threshold and wet-dog shaking in the exercise group showed significant improvement compared with the control group after 14 days of exercise (P < 0.05), while the grooming was significantly reduced until the 28th day (P < 0.01), indicating that exercise can alleviate pain caused by disc degeneration in model rats. At 21 days after modeling, the levels of proteoglycan, aggrecan and collagen type Ⅱ in the nucleus pulposus and annulus fibrosus were significantly decreased compared with the baseline (P < 0.01), indicating the occurrence of disc degeneration. After 14 days of training, the levels of proteoglycan, aggrecan, and collagen type Ⅱ in the nucleus pulposus and annulus fibrosus in the exercise group were significantly increased compared with the control group (P < 0.01). Moreover, after 8-week exercise, the level of proteoglycan in the nucleus pulposus and annulus fibrosus in the exercise group was increased by 4-5 times compared with the control group, and levels of aggrecan and collagen type Ⅱ in the nucleus pulposus in the exercise group also was increased by 3-4 times compared with the control group. To conclude, exercise can promote extracellular matrix increased by production by increasing the levels of proteoglycan, aggrecan, and collagen type II in the degenerative intervertebral disc.

OBJECTIVEConditioned taste preference (CTP) is a taste learning reflex by which an animal learns to prefer a substance which tastes not well and has been studied with much interest in recent years. However, the neural substrates of CTP are less known. This study aimed to determine the possible neural path- ways of CTP and whether serum leptin level and the leptin receptor (OB-Rb) in the hind brain are involved following CTP formation.

METHODSWe established CTP of quinine in rats with a 2-bottle preference test. The serum leptin concentrations were detected, the expression of c-fos in the rat brain was tested to determine the nuclei in relation with establishment of CTR Finally, the OB-Rb mRNA expression was examined by RT-qPCR assay in parabrachial nucleus (PBN) and the nucleus of the solitary tract (NST) of the hind brain.

RESULTSCompared with control group, the level of serum leptin was higher in the CTP group (4.58 ± 0.52 vs 1.67 ± 0.25 µg/L, P < 0.01); increased c-fos positive cells were found in the anterior hypothalamus (AH, 221.75 ± 4.96 vs. 178.50 ± 6.63 cells/mm², P < 0.05), the basal lateral amygdala (BLA, 70.75 ± 6.17 vs 56.50 ± 3.62 cells/ mm², P < 0.05) and the nucleus of the solitary tract (NST, 41.25 ± 1.32 vs 32.50 ± 1.02 cells/mm², P < 0.05). But in ventromedial nucleus of the hypothalamus (VMH, 20.75 ± 2.73 vs 38.5 ± 1.54 per 1 mm², P < 005), PBN (21.50 ± 2.24 vs 36.25 ± 1.49 cells/mm², P < 0.05) and the central nucleus of the amygdala (CeA, 22.25 ± 1.53 vs 35.50 ± 2.11 cells/mm², P < 0.05), the number of c-fos positive cells was decreased in the CTP group. In addition, we found OB-Rb mRNA expression in PBN of CTP group rats was higher than that of control group (0.95 ± 0.055 vs 0.57 ± 0.034, P < 0.05), while there was no significant difference of OB-Rb mRNA expression in NST between the two groups.

CONCLUSIONNuclei AH, BLA, NST, VMH, PBN and CeA participate in the formation of CTP. Leptin and its receptor in PBN may be involved in the formation and maintenance of CTP.

Animals ; Conditioning (Psychology) ; Leptin ; blood ; Rats ; Receptors, Leptin ; physiology ; Rhombencephalon ; physiology ; Taste ; physiology

Objective To study hemodynamics of cerebral blood flow in newborns with asphyxia.Method Bilateral cerebral electrical admittance plethysmogram(BCEAP) was used to explore characteristics of cerebral blood flow in 20 healthy newborns and 20 newborns with asphyxia respectively from first to fourth day after birth.Results Both the ratio of Hs to b-S(Hs/ b-S) and index of admittance differential loop(ADL) Ⅰ+Ⅱ decreased significantly in newborns with asphyxia compared to normal control from first to fourth day after birth(P

Objective To determine if there is any difference in neuronal and glial(astrocytic and microglial)activation in the spinal cord in three rat models of neuropathic pain.Methods Twenty-four SD rats weighing 150-200 g were randomly divided into 4 groups(n=6 each):Ⅰ control group;Ⅱ chronic constrictive injury group(CCI);Ⅲ spinal nerve ligation group(SNL)and Ⅳ spared nerve injury(SNI).No operation was performed in control group.In CCI group left sciatic nerve was exposed and loosely ligated with catgut.In SNL group the L_5 spinal nerve was exposed and ligated with silk suture and cut.In SNI group tibial nerve and common fibular nerve were ligated and cut.Pain threshold was measured using plantar tactile stimulator(Ugo,Basile Co. Italy)every other day from 3 days before until 15 days after operation.50% paw withdrawal threshold was measured using up-and-down sequential mechanical stimulation of different intensity(0.45,0.70,1.20,2.00, 3.63,5.50,8.50,15.10 g)applied to the plantar surface of the injured paw.On the 15~(th) day after operation after pain threshold was measured the animals were anesthetized with intraperitoneal 10% chloral hydrate 400 mg? kg~(-1).The L_(5,6) segment of the spinal cord was isolated.Neuronal,astrocytic and microglial activation was determined by immuno-histochemistry with antibodies of c-Fos(a proto-oncogene protein),GFAP(an astrocyte marker)and OX-42(a microglial marker).Results The 50% paw withdrawal threshold reached the lowest level on the 7~(th) day after operation.The lowest level was maintained until the 15~(th) day after operation in group CCI,SNL and SNI.The 50 % paw withdrawal threshold was(14.1+1.5)g in control group,(2.5+0.5)g in CCI group, (1.5?0.6)g in group SNL and(0.8?0.4)g in group SNI.The number of c-Fos positive neurons in laminae Ⅳ-Ⅵ of dorsal horn was significantly greater in group CCI,SNL and SNI than in control group,but there was no significant difference among the 3 peripheral nerve injury groups.The activation of astrocytes and microglias in laminae Ⅰ-Ⅳ of dorsal horn was significantly increased in group CCI,SNL and SNI than in control group but there was no significant difference among the 3 peripheral nerve injury groups.Condusion There is no significant difference in activation of neurons and astrocytes and microglias in the ipsilateral dorsal horn among the 3 pain models.

Relapse after allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains a main question on treatment failure.Current strategies for management that usually include salvage chemotherapy,donor lymphocytic infusion and second transplantation.Our study assessed the efficacy of decitabine (DAC) for treating patients with acute lymphoblastic leukemia (ALL) who relapsed after allogeneic hematopoietic stem cell transplantation (allo-HSCT).We retrospectively analyzed the outcomes of 12 patients with relapsed ALL after allo-HSCT who received DAC therapy.Nine patients received DAC combined with chemotherapy and donor stem cell infusion,and 3 patients received single-agent DAC.Ten of the 12 patients achieved complete remission (CR),1 achieved a partial remission (PR),and 1 had no response (NR) after treatment at the latest follow-up (LFU),the median survival was 11.2 months (range,3.8-34,7 months).The 1-and 2-year overall survival (OS) rates were 50％ (6/12) and 25％ (3/12),respectively.Five patients were still alive;4 had maintained CR and 1 was alive with disease.Patients with Philadelphia chromosome-positive ALL had higher survival rate than patients with Philadelphia chromosome-negative ALL (57.1％ vs.20％).No aggravated flares of graft-versus-host disease (GVHD) were observed during DAC treatment.Therefore,DAC may be a promising therapeutic agent for ALL recurrence after allo-HSCT.

OBJECTIVETo detect and compare the transcriptional activities of prostate-specific membrane antigen (PSMA) promoter and enhancer and survivin promoter in different human prostate cancer cell lines, and to search for some evidence for the targeting gene therapy of human prostate cancer.

METHODSThe fragments of the PSMA promoter and enhancer and survivin promoter were amplified by PCR and inserted into pGL3-Basic. The recombinant plasmids were transiently transfected into human prostate cancer cell lines and normal Chang liver cells, and, their transcriptional activities in various cells were determined by measuring the expression of luciferase.

RESULTSThe survivin promoter exhibited a higher transcriptional activity than PSMA promoter and enhancer in tumor cell lines, and the S2pro promoter showed the highest activity, reaching one third of that of the CMV promoter.

CONCLUSIONThe survivin promoter is highly activated in prostate cancer cell lines and may serve as a new tool for the transcriptional targeting gene therapy of prostate cancer.

Antigens, Surface ; genetics ; Cell Line, Tumor ; Glutamate Carboxypeptidase II ; genetics ; Humans ; Inhibitor of Apoptosis Proteins ; genetics ; Male ; Plasmids ; Promoter Regions, Genetic ; Prostatic Neoplasms ; genetics ; therapy ; Transcription Initiation Site ; Transcriptional Activation ; Transfection