To construct recombinant reporter plasmids containing different Alpha gene segments and Alpha1-TFEB fusion gene and to evaluate the promoter activity of the Alpha gene. Methods:Promoter regions of the Alpha gene were predicted using a software Primer 0.5. Five Alpha gene segments with different lengths and a normal TFEB gene promoter (pTFEB) were amplified via polymerase chain reaction, and recombinant reporter plasmids containing different Alpha gene segments and a normal TFEB gene pro-moter were constructed. Liposome transfection was used to transfect these vectors into the human embryo kidney 293T cells. The pro-moter activity of the Alpha gene was evaluated via luciferase assay. Meanwhile, the recombinant Alpha1-TFEB plasmid was construct-ed and transfected into the 293T cells. The TFEB expression of the recombinant Alpha1-TFEB plasmid was then detected via Western blot. Results: Recombinant reporter plasmids containing different Alpha gene segments and pTFEB were constructed successfully. Compared with the luciferase activity of pGL3-Basic, that of the groups with Alpha1, Alpha2, Alpha3, Alpha4 and Alpha5 significantly increased (P<0.01). The luciferase activity also increased significantly in the groups with Alpha1, Alpha2 and Alpha5 compared with that of the pTFEB group (P<0.01). The TFEB expression of the pGL3-Enhancer-Alpha1-TFEB was significantly higher compared with that of the pGL3-Enhancer group. Conclusion:In t(6;11) translocation RCC, the Alpha gene has a strong promoter activity and it en-hances TFEB expression. The strongest promoter activity region is in Alpha5 with a sequence from 643 bp to 693 bp.

Objective To study the curative effect of psychological intervention combined with drug treatment of children addicted to the network game.Methods One hundred and twelve children addicted to the network game were selected,who were admitted to the outpatient department of the Second Affiliated Hospital of Xinxiang Medical University from Feb.2009 to Oct.2013 were selected,according to the admission time sequence,they were randomly divided into drug treatment group,psychological intervention group(as control group) and psychological intervention combined with drug therapy group(test group).Each of the control groups had 38 cases of patients,while treatment group had 36 cases.Drug treatment group was treated with oral Fluoxetine Hydrochloride Capsules 10 mg,1 time/d; the psychological intervention group was given the general psychological conversation,until discharged from the hospital; the test group adopted the cognitive restructuring,imagine the debate,self suggestion,remind yourself of cognitive therapy and behavior contract,behavior reinforcement,behavior extinction,aversive stimulus,self-regulation of behavior therapy combined with drug therapy.Respectively,before treatment,2 weeks,4 weeks and 8 weeks after the end of the treatment,Chinese Internet Addiction Scale,Coping Style Questionnaire (CSQ) and symptom checklist 90 (SCL-90) of each factor were evaluated.Results By the end of the 2 weeks and 8 weeks after the treatment,CIAS scores in the treatment group were significantly lower than that in drug therapy and psychological intervention group,the differences were statistically significant (t =2.49,2.03,all P ＜ 0.05 ; t =2.72,2.72,all P ＜ 0.01) ; scores of 3 factors of fantasy,self accusation and retreat in CSQ group in coping style questiennaire in the coping style questionnaire in the treatment group were lower than those in control groups,the differences were statistically significant (t =3.99,3.78,2.15,all P ＜ 0.05; t=2.64,4.20,3.72,all P ＜ 0.01); the problem solving,help seeking factor in the test group were higher than those in the control groups,the differences were statistically significant (t =2.69,2.43,5.31,2.25; all P ＜ 0.01) ; variation test combined physicochemical factors scores at different stages had on difference comparison of those in the 2 control groups (all P ＞ 0.05).Comparison of the SCL-90 score between the test group and the 2 control groups,the differences were statistically significant (t =5.05,4.31,all P ＜ 0.01).There was no significant difference of the scores between the 2 control groups (all P ＞ 0.05).Conclusions Psycholoical intervention therapy including cognitive therapy,behavior therapy combined with drug treatment have good effect on children's addiction to Web games.

Aim To explore the effects and its mechanisms of ginsenoside-Rg1 on level of t-PA and PAI-1.Methods Type 1 plasminogen activator inhibitor(PAI-1) and tissue type plasminogen activator(t-PA) activity in plasma were assayed using chromogenic substrate.Results The results showed that ginsenoside-Rg1 in vitro or in vivo significantly inhibited PAI-1activity,while increased t-PA activity.These effects were concentration-dependent.Intravenous Panax notoginsenoside Rg1 at 30,60,120 and 240 mg?kg~(-1) markedly suppressed PAI-1 level in plasma as well as platelet-released substances stimulated by thrombin,while increased plasma t-PA activity.And release level of PAI-1 owing to blood platelet was greatly decreased by ginsenoside-Rg1.Conclusion Ginsenoside-Rg1 showed potent antithrombosis due to the inhibition of PAI-1 and increase of t-PA.It might also be a advantagous mechanism to its antithrombsis.

This study is to investigate the effect of the major chemical composition in rhizome of Pterocypsela elata, lactuside B, on expression of bcl-2, bax mRNA and their protein in rats' cerebral cortex after cerebral ischemia-reperfusion injury. First, middle cerebral artery ischemia-reperfusion injury model was established, and each group was treated with the corresponding medicines. Animals were separately sacrificed at 24 h and 72 h. The brain infarct volumes were detected by TTC dye, bcl-2 and bax mRNA expression was checked by RT-PCR, and the proteins of bcl-2 and bax were explored by two-step immunohistochemistry in cerebral cortex of rats. Lactuside B can reduce brain infarct volume of cerebral cortex of rats, increase the expression of bcl-2 mRNA and decrease that of bax mRNA. Moreover, the ratio of bcl-2 to bax mRNA is higher in 12.5 and 25 mg kg(-1) dose group, respectively, which is significantly different from that of model group (P < 0.05 or P < 0.01). Generally, either 12.5 or 25 mg kg(-1) dose group is better than positive control medicine nimodipine (P < 0.05 or P < 0.01). In addition, the expression of bcl-2 and bax protein is consistent with their gene expression. Infarct volume and the ratio of bcl-2 to bax mRNA expression are significantly different (P < 0.05 or P < 0.01) between 72 h and 24 h group. The results demonstrated that lactuside B could play a good role in resisting cerebral ischemia by upregulating the expression of bcl-2 mRNA and protein and downregulating that of bax mRNA and protein.

BACKGROUND: Retinitis pigmentosa (RP) is a non-inflammatory, bilaterally progressive, retinal degeneration characterized by loss of photoreceptor cells via an apoptotic mechanism, and it eventually leads to blindness.Research shows that the traditional Chinese medicines of Astragalus has great prospect on blocking the progression of RP disease.OBJECTIVE: To observe the protective effect of Astragalus on N-methylN-nitrosourea (MNU)-induced retinal damage in Sprague-Dawley (SD) rats and provide the optimal treatment for RP in humans.DESIGN: Randomized controlled experiment.SETTING: School of Pharmaceutical Sciences, Xinxiang Medical College.MATERIALS :The experiment was completed in Pharmacological Laboratory of Zhongshan Ophthalmic Centre, Sun Yat-sen University between March to December 2004. Totally 114 female SD rats were purchased from the Animal Center of Zhongshan Medical College, Sun Yat-sen University.MNU was purchased from Sigma Company of America. Astragalus injection was purchased from Chengdu Diao Jiuhong Pharmaceutical Factory (Batch No. Z99060535, 2 mL/ampoule, main ingredient: Astragalus).METHODS: Among 114rats, 30 were for morphometric analysis of retinal layers, 30 were for detection of apoptosis and 54 were for detection of NF-κB p65 activity. All of them were randomly divided into different groups and each group had 6 rats. Astragalus at doses of 2.5, 5 and 10 g/kg were injected intraperitoneally into 47-day rats once a day. Meanwhile, a single intraperitoneal injection of 60 mg/kg MNU was given to 50-day rats in model and Astragalus groups. At different intervals after MNU treatment,the animals were sacrificed. Retinal damage was evaluated based on retinal thickness, the apoptotic index of the photoreceptor cells was detected by TUNEL labeling and the DNA-binding activity of NF-κB p65 was analyzed according to transcription factor assay kit.MAIN OUTCOME MEASURES: Comparison of retinal thickness, apoptotic index and the activity of nuclear NF-κB p65.RESULTS: Totally 114 rats entered the result analysis. Pretreatment with Astragalus could dose-dependently suppress MNU-induced photoreceptor cell loss and decreased the apoptotic index. Astragalus at dose of 10 g/kg also time-dependently up-regulated the activity of nuclear NF-κB p65.However, protective effect of Astragalus on MNU-induced central retinal damage was not found.CONCLUSION: Astragalus partially protects against MNU-induced retinal damage by up-modulating the activity of nuclear NF-κB p65 to inhibit apoptosis of photoreceptor cells in a dose-dependent manner.

Objective To study the antidepressant-like effect of Butanol Extract from Stevia (BEFS) and its possible corresponding mechanisms.Methods Forty-five mice were randomly divided into control group,model group,model + Fluoxetine group,model + BEFS-3 g/kg group and model + BEFS-6 g/kg group.The chronic unpredictable mild stress(CUMS) for 3 weeks was used to make up depressive animal model.It was based on body weight,crossing numbers,rearing numbers and sucrose preference to assess whether the model was a success.After continuous lavage giving BEFS and fluoxetine for 2 weeks and subsequent behavioral test,the mRNA transcription of Single immunoglobulin interleukin-1 receptor-related molecule (SIGIRR) and nuclear factor kappa-B (NF-κB) in liver tissue of mice were detected by using quantitative real time-PCR.Results Three weeks after CUMS,behavior parameters were significantly decreased in mice exposed to CUMS compared with control group[(17.28 ±0.49) g vs (22.47 ± 0.82) g,P =0.000;(43.35 ± 1.68) scores vs (125.25 ± 3.07) scores,P =0.000;(3.92 ± 0.18) scores vs (15.92 ± 1.78) scores,P =0.000;(49.39 ± 2.05) ％ vs (67.18 ± 4.46) ％,P =0.001].However,2 weeks after dosing,body weight,crossing numbers,rearing numbers and sucrose preference were (19.03 ± 0.44) g,(70.50 ± 6.56) scores,(3.08 ± 0.77) scores and (41.27 ± 7.59) ％,respectively in model group.Compared to model group,behavior parameters of BEFS (3 and 6 g/kg BEFS) groups were significantly increased [BEFS-6 g/kg group:(24.42 ± 1.46) g,(99.75 ± 5.07) scores,(13.08 ± 1.60) scores,(64.31 ± 1.92) ％,all P ＜ 0.05;BEFS-3 g/kg group:(23.82 ± 1.72) g,(91.67 ± 5.69) scores,(6.92 ± 1.05) scores,(59.56 ± 5.31) ％,respectively,all P ＜ 0.05].Correspondingly,BEFS could reverse the mRNA expression of SIGIRR lowering and NF-κB rising in the liver of mice exposed to CUMS(BEFS-6 g/kg group:1.208 1 ± 0.102 2,1.437 7 ± 0.352 9;BEFS-3 g/kg group:0.768 0 ± 0.108 6,2.186 2 ± 0.203 8,all P ＜ 0.05).Conclusions BEFS can remarkably improve depression-like behaviors in CUMS mice and its antidepressant activity is mediated,at least in part,by upregulating SIGIRR and downregulating NF-κB in the liver.

Objective To investigate the major chemical composition in rhizome of Pterocypsela elata and explore the activity of lactuside B against brain ischemia.Methods The compounds were isolated and purified by silica gel column chromatography and elucidated by spectroscopic experiments;The model of partial brain ischemia was used to detect water content,MDA and SOD levels in brain tissue in order to observe the activity of lactuside B against brain ischemia.Results Ten compounds were obtained and established as lactuside B(1),11?,13-dihydrolactucin acetate(2),?-sitosterol(3),daucosterol(4),(24R)-5?-stigrnast-7,22(E)-dien-3?-ol(5),3,3',4-trimethoxylellagic acid(6),?-amyrin(7),oleanolic acid(8),n-hexacosanic acid(9),and stearic acid(10).Lactuside B was a key component,and it's yield was 0.15%.Contents of water and MDA level in the brain tissue were significantly decreased,and the SOD content notably increased in all groups of lactuside B.Conclusion Ten compounds are all isolated from this plant for the first time.Compound 1 is a key component which possesses obvious activity against brain ischemia.

AIM:To investigate the effect of rhein on bleomycin-induced pulmonary fibrosis and the expression of microRNA-21 (miR-21) and transforming growth factor-β1 (TGF-β1)/Smad signaling molecules in rats.METHODS:A single dose of bleomycin was intratracheal injected into the SD rats to induce pulmonary fibrosis .After injection of bleo-mycin, the rats were randomly divided into low-, medium-and high-dose rhein treatment groups and model group .The rats that were instilled with normal saline intratracheally served as control group .After the treatment for 28 d, the pulmonary pathologic changes were observed under microscope with hematoxylin-eosin staining .The lung coefficient and hydroxypro-line content were also measured .The expression of miR-21 and the mRNA levels of TGF-β1 and Smad7 in the lung tissues were detected by real-time PCR.The protein levels of TGF-β1 and Smad7 were determined by Western blot .RESULTS:Rhein significantly attenuated the experimental alveolitis , pulmonary fibrosis , lung coefficient and hydroxyproline contents in the rats.Rhein obviously decreased the expression of miR-21,and the mRNA and protein levels of TGF-β1, but signifi-cantly increased the mRNA and protein levels of Smad 7 in the lung tissues .CONCLUSION: Rhein effectively prevents bleomycin-induced pulmonary fibrosis by inhibiting the expression of miR-21 and promoting the expression of Smad 7, thus regulating the TGF/Smad signaling pathway to decrease extracellular matrix deposition .

Objective To investigate the effects of intervention with the fluoxetine and the enriched environment on chronic stress induced depression behavior of rats,and the changes of myelin basic protein in hippocampus and prefrontal regions.Methods 50 adult male SD rats were randomly divided into control group,fluoxetine group,model group,enriched environment (EE) group and EE plus fluoxetine group.Fluoxetine group,model group,EE group and EE plus fluoxetine group underwent chronic unpredictable stress stimulus in the first to third week,and fluoxetine group,EE group,EE plus fluoxetine group underwent the intervention with EE and (or) fluoxetine in the fourth to sixth week.The changes of behavior in rats were evaluated by sucrose water consumption,open field test and weight changes.The content of MBP in each subregion of hippocampus and prefrontal regions of rats was measured with immunocytochemical methods.Results At the third weekend,the assessed behaviors of stressed rats decreased significantly compared with control group (P＜0.05);and at the sixth weekend,the behaviors of stressed rats restored after treated with EE and (or) fluoxetine.The content of MBP in the rat hippocampus CA1,DG area and prefrontal area of model group declined clearly compared with control group (mean density of model group orderly:0.199±0.024,0.204±0.021,0.225±0.028;control group orderly:0.279±0.034,0.288±0.043,0.308±0.053,P＜0.05).The content of MBP in the rat of fluoxetine group,EE group and EE plus fluoxetine group increased obviously compared with model group (fluoxetine group orderly:0.259± 0.047,0.266± 0.052,0.284 ± 0.031;EE group orderly:0.257±0.038,0.258±0.042,0.286±0.037;EE plus fluoxetine group orderly:0.271± 0.046,0.279±0.040,0.289±0.041,P＜0.05).Conclusion The depression-like behavior of rats induced by chronic unpredictable stress is associated with the change of the content of MBP in hippocampal CA1,DG area and prefrontal area;and the depression-like behavior and the content of MBP decrease are reversed after the intervention with fluoxetine and EE.

Objective To explore the effects of notoginsenoside-Rg_1 on brain-derived neurotrophic factor(BDNF) protein in cerebrum cortex of MCAO/R injury,as well as to investigate whether notoginsenoside-Rg_1 can up-regulate the protein content of BDNF of the positive neurons or the amount of the po-sitive neurons.Methods Adult male SD rats(60) were randomly divided into model group,notoginsenoside-Rg_1 high,middle,and low dose(200,100,50 mg/kg) groups,and the positive control(Nimodipine,1 mg/kg) group.All drugs were given once a day by ip till the mouse was killed.The focal cerebral(ischemia-)reperfusion model was made with thread-occluded method.Their frozen brain tissue were sliced(into) section of 12 ?m thickness.The four rats were randomly taken from each groups to be treated as specimens after surgical handle in 1,3,and 7 d.The slices were according to the immunohistochemical ABC techniques.The protein content of BDNF of the positive neurons and the amount of the positive neurons in cerebrum cortex of rat were observed and counted by HPIAS—1000 analytic system,and the nervous deficit symptoms after the cerebral ischemia were observed.Results Comparing with the model group,all notoginsenoside-Rg_1 treated groups obviously improved some nervous deficit symptoms of cerebral ischemia-reperfusion rats,and increased the protein content of BDNF and the amount of the positive neurons in the cerebrum cortex of model rats(P