Artesunate negatively controls expression of cyclin D1 and activity of AP-1 by inhibiting the activation of ERK1/2 protein in HSC-T6 cells

AIM: To investigate the effects of artesunate(Art) on the expression of ERK1/2, AP-1 and cyclin D1 in rat hepatic stellate cells (HSCs), and to elucidate the molecular mechanism of Art against hepatic fibrosis. METHODS: HSC-T6 cells were treated with platelet-derived growth factor BB(PDGF-BB) to induce cell proliferation. The cells were divided into control group, PDGF-BB group, PDGF-BB+Art groups (with 6.25 mg稬^-1, 25 mg稬^-1or 50 mg稬^-1 of Art) and PDGF-BB+PD98059 group. The level of collagen type I in the supernatant was detected by enzyme-linked immunosorbent assay (ELISA). The mRNA expression levels of ERK1/2 and cyclin D1 were measured by RT-PCR. The protein levels of p-ERK1/2 and cyclin D1 in HSC-T6 cells were detected by Western blotting. The activity of AP-1 was analyzed by electrophoretic mobility shift assay. RESULTS: The concentration of collagen type I was significantly higher in PDGF-BB group than that in control group (P<0.05), and decreased in PDGF-BB+Art group and PDGF-BB+PD98059 group in comparison with that in PDGF-BB group (P<0.05, P<0.01). The protein level of ERK1/2 in PDGF-BB+Art group (50 mg稬^-1) was lower than that in PDGF-BB group (P<0.05), and was even lower in PDGF-BB+PD98059 group (P<0.01). The mRNA expression of cyclin D1 in PDGF-BB+Art groups (25 mg稬^-1and 50 mg稬^-1) and PDGF-BB+PD98059 group were significantly lower than that in PDGF-BB group (P<0.05). The protein levels of p-ERK1/2 and cyclinD1 were the highest in PDGF-BB group, and significantly lower in PDGF-BB+Art groups (6.25 mg稬^-1, 25 mg稬^-1 and 50 mg稬^-1) and PDGF-BB+PD98059 group (P<0.05, P<0.01). The AP-1 binding activity in HSC-T6 cells was down-regulated by Art. CONCLUSION: Artesunate inhibits the proliferation of HSC-T6 cells in vitro by inhibiting the activation of ERK1/2, thus down-regulating the activity of AP-1 and expression of cyclin D1.     

Neuroprotective effect of L-serine on rats with permanent middle cerebral artery occlusion

AIM: To investigate the neuroprotective effect, therapeutic dosage and time window of L-serine against permanent cerebral injury in rats. METHODS: Permanent middle cerebral artery occlusion (pMCAO) was induced in the rats to determine the efficacy of L-serine (ip) by neurological evaluation, TTC staining and Nissl staining.L-serine was used at different doses (56 mg/kg, 168 mg/kg and 504 mg/kg) and for different time periods (1 h, 3 h, 6 h, 12 h and 24 h after pMCAO). Aminooxyacetic acid (AOAA), an inhibitor of serine racemase, was used to alter the efficacy of L-serine. Laser Doppler perfusion monitor was used to observe the regional cerebral blood flow (rCBF) in the ischemic cerebral cortex under the condition with or without L-serine treatment. RESULTS: Treatment with L-serine at doses of 168 mg/kg and 504 mg/kg at time point of 3 h after pMCAO greatly decreased the neurological deficit score and infarct volume,and attenuated the loss of hippocampal CA1 neuronal cells. In the observation of therapeutic time window, L-serine displayed a significant neuroprotective effect if used within 6 h after pMCAO, but did not exert any notable effect if used over 12 h after pMCAO. AOAA hardly changed the effect of L-serine. L-serine treatment notably raised rCBF in the area of ischemic cerebral cortex when it was injected 30 min after pMCAO. However, strychnine, an antagonist of strychnine-sensitive glycine receptor, did not alter this effect of L-serine. CONCLUSION: L-serine has neuroprotective effect on permanent ischemic brain injury in rats if administered early and sufficiently by augmentation of rCBF in the ischemic cerebral cortex.     

Expression of ERK5 in colorectal cancer tissues and its correlation with distant metastasis in colorectal cancer patients

AIM: To investigate the expression of extracellular signal-regulated kinase 5 (ERK5) in primary colorectal cancer (CRC) and adjacent normal mucosa, and to analyze the relationship between ERK5 expression and clinicopathological parameters for exploring the functions of ERK5 in the occurrence and development of CRC. METHODS: The expression of ERK5 in carcinoma tissues and normal mucosa was examined by a set of tissue microarrays and the method of immunohistochemistry. The potential relationship between ERK5 expression and clinicopathological features was also analyzed. RESULTS: ERK5 expression was significantly higher in CRC tissues (134/338, 39.6%) than that in normal tissues (21/80, 26.2%; P<0.05). Overexpression of ERK5 in CRC tissues was significantly correlated with distant metastasis (P<0.05). However, no correlation between ERK5 expression and age at surgery, sex, tumor location, the depth of invasion, lymph node metastasis, TNM staging or differentiation grade was found (P>0.05). According to the Kaplan-Meier analysis, there is no significant difference in 5-year overall survival between the patients with ERK5 expression at high level and at low level. CONCLUSION: ERK5 protein is highly expressed in CRC with distant metastasis. This may be a promotive factor in the process of distant metastasis.     

北京鸭FABP2基因多态性与体尺和屠体性状的相关性研究

本研究以12个全同胞家系共224只G2代北京鸭资源群体为试验材料,分别利用DNA序列测定和PCR-RFLP方法检测G1和G2代个体基因型。结果在北京鸭小肠脂肪酸结合蛋白(FABP2)基因中共发现4处单核苷酸多态性(SNPs),外显子3中1处SNP(92bp)可导致氨基酸的改变(Gly→Ser),G2代中该SNP产生3种基因型:AA、AB和BB,其频率分别为0.103、0.629和0.268,群体中A基因频率为0.417,B基因频率为0.583。最小二乘分析表明,BB基因型个体的体斜长、胫围和颈长显著高于AB基因型;BB基因型个体胫蹼重、肌胃重和翅重显著高于AA基因型和AB基因型;BB基因型个体头重显著高于AA基因型(P<0.05)。综上所述,FABP2基因外显子3中(92bp)的SNP与北京鸭的部分体尺和屠体性状显著相关,可作为北京鸭体尺和屠体性状标记辅助选择的分子标记。     

北京市朝阳区景观连接度距离阈值研究

景观连接度方法是一种进行景观生态安全格局定量化评价的较好方法。此方法的应用中,景观连接度适宜距离阈值是一个关键因素,以北京市朝阳区为例对其展开研究。选取景观组分数(NC)、整体连接度指数(ⅡC)以及可能连接度指数(PC)来表征景观连接度以及计算斑块重要性值(dI);预设9个距离阈值,探讨不同距离阈值下景观连接度和斑块重要性值的变化规律。结果表明:北京市朝阳区景观连接度的距离阈值以400～600 m为宜。在此距离阈值的约束下分析表明,小型生态斑块对提高区域景观连接度有重要作用。     

参与式乡村评估在青海海东地区CDM造林再造林项目中的应用

基于实地踏查、问卷调查、社区会议及关键人物访谈等参与式乡村评估工具对青海海东地区清洁发展机制(CDM)造林再造林项目进行评估,结果表明:拟议项目能很好地反映当地社区生计发展需求,不仅将对缓解气候变化做出贡献,还将为社区经济发展和环境保护做出贡献。农户、社区和政府等相关利益群体对项目实施提出进行更多的技术和业务培训,合理选择资金安排,采用混交种植方式等有益建议。     

乙型脑炎病毒NS1蛋白的可溶性表达与抗原性分析

为获得可溶性表达的乙型脑炎病毒(JEV)NS1蛋白,将编码JEV SA14-14-2株NS1蛋白基因克隆至原核表达载体pMAL-c5X中,构建重组融合表达质粒pc5X-NS1;用重组质粒转化宿主菌ER2523后,经IPTG诱导得到可溶性的融合蛋白MBP-NS1;优化诱导表达条件,于28℃、0.3 mmol/L IPTG诱导4 h;最后融合蛋白经直链淀粉树脂柱纯化。结果表明:重组融合蛋白MBP-NS1具有良好的抗原性和特异性。     

斯氏艾美耳球虫ADF基因部分序列的克隆与分析

为了获得我国斯氏艾美耳球虫长春株ADF基因并分析其与柔嫩艾美耳球虫长春株相应序列的同源性,试验根据GenBank中公布的柔嫩艾美耳球虫ADF基因序列设计引物,采用RT-PCR方法获得我国斯氏艾美耳球虫长春株ADF基因部分序列,然后将其克隆到pMD18-T载体中,应用DNAStar软件对测得序列进行序列分析。结果表明:斯氏艾美耳球虫长春株ADF基因大小为357 bp;经DNAStar分析,我国斯氏艾美耳球虫长春株与柔嫩艾美耳球虫长春株ADF核苷酸序列和氨基酸序列同源性均为99.2%。说明ADF基因在进化过程中高度保守。     

人员因素对猪活体测定结果的影响

为检验人员因素在猪的活体测定中对数据的影响程度,采用随机试验的方法将测定人员分为A组和B组,在测定方法相同的情况下,对各组人员进行独立多次测量,并对测量结果进行方差分析。结果表明:两组人员因素对猪活体膘厚的测量结果均具有极显著差异(P<0.01);对于猪眼肌高度,A组人员因素差异不显著(P>0.05),B组人员因素则差异显著(P<0.05)。说明人员因素对活体测定结果有显著的影响。笔者建议使用B型超声波测定仪用于活体测定时,在数据读取方法上需进一步规范和统一,并提升人员技能,不断提高测定准确性,促进我国种猪遗传评估效率的逐步提升。     

肉兔机体组织中脂肪酸检测方法的选择与组成分析

为了更好地研究肉兔肌肉组织、皮下结缔组织和肠系膜组织中的脂肪酸组成,先用索氏提取法和氯仿-甲醇法从肉兔脂肪组织中提取油脂,继而进行酸法和碱法甲酯化,经气相色谱技术,对其脂肪酸组成进行了分析。结果表明:采用氯仿-甲醇法提取,碱法甲酯化法处理的效果最好。以塞北兔为研究对象,采用这种方案对油脂进行处理,利用气相色谱技术和气质联用仪对脂肪酸组成进行定性和定量分析。结果表明:肉兔肌肉间脂肪由12种脂肪酸组成,其中4种饱和脂肪酸中C16:0含量最高,为27.30%;4种单不饱和脂肪酸中C18:1含量最高,为24.02%;4种多不饱和脂肪酸中9,12-C18:2含量最高,为26.06%;P:S值为0.88。皮下结缔组织脂肪由13种脂肪酸组成,其中4种饱和脂肪酸中C16:0含量最高,为26.75%;5种单不饱和脂肪酸中C18:1含量最高,为29.07%;4种多不饱和脂肪酸中9,12-C18:2含量最高,为21.99%;P:S值为0.75。肠系膜脂肪由21种脂肪酸组成,其中7种饱和脂肪酸中C18:0含量最高,为25.47%,7种单不饱和脂肪酸中C18:1含量最高,为26.25%,7种多不饱和脂肪酸中9,12-C18:2含量最高,为27.52;P:S值为0.99。肌肉间、皮下和肠系膜上脂肪酸组成种类和含量有差异。肠系膜内脂肪酸种类最多,肌肉间较少。P:S比值肠系膜组织最高,皮下结缔组织最低。