运用噬菌体展示随机肽库筛选与内毒素结合的高亲和性多肽

目的 从噬菌体展示随机肽库中筛选与内毒素结合的多肽序列,并进行鉴定.方法 以内毒素脂多糖(lipopolysaccharide,LPS)为靶分子对噬菌体展示随机十二肽库进行4轮亲和筛选,获得与LPS结合的噬菌体克隆,应用结合实验和克隆斑抑制实验进一步确证.挑选结合力强的克隆进行DNA测序,推导出呈现的多肽序列,应用生物信息学软件进行多肽序列分析和同源性分析.结果 经4轮亲和筛选从噬菌体展示随机十二肽库中筛选获得了86个克隆,挑选12个结合力强的克隆进行DNA序列测序及生物信息学分析,结合本项目组的噬菌体展示随机七肽库筛选结果推导出呈现的多肽序列为HWQWPHWSPPP(命名为P11肽).检索相关数据库发现此肽序列未被申请专利,体内约908种蛋白与其结构相匹配,其中包含有与LPS相互作用的位点.结论 通过对噬菌体展示随机肽库的淘选,获得与LPS结合的高亲和性多肽,为进一步以这些多肽为先导物进行定向进化研究提供了实验依据和结构基础.     

HCCR-2反义核酸对肝癌HepG2细胞的作用

Objective To investigate the effects of antisense recombinant euraryotic expression vector of HCCR-2 on the proliferation and apoptosis of HepG2. Methods The antisense recombinant eukaryotic expression vector of HCCR-2 was constructed. The vector was stably transfected to the HepG2 cells, and positive clones were selected by G418 (antiseuse vector group), pIRES2-EGFP vector was transfected into the HepG2 cells in the same way (pIRES2-EGFP group). The conditions of the nontransfected HepG2 cells were used as control (HepG2 group). Changes in cell growth curve, cell cycle, cell apoptosis and morphology of HepG2 cells after the transfec-tion were detected by MTT method, flow cytometry and transmission electron microscopy, respectively. All the data were analyzed by one-way ANOVA and chi-square test. Results The expression level of HCCR-2 mRNA was down-regulated to 0.39±0.04 in antisense vector group, and the expression level of HCCR-2 mRNA in pIRES2-EGFP group and HepG2 group were 0.62±0.06 and 0.72±0.03, respectively, with significant difference among the 3 groups (F=43.701, P<0.05). The apoptotic rate of HepG2 cells in antisense vector group, pIRES2-EGFP grop and HepG2 group were 13.30%, 2.51% and 2.07%, respectively, with significant difference among the 3 group (χ2=6.793, 8.721, P<0.05). The growth of HepG2 cells in antisense vector group was retarded, and was blocked in G0/G1 stage. Conclusions The HCCR-2 antisense recombinant eukaryotic expression vector can inhibit the mRNA expression of HCCR-2 and promote the apoptosis of cells. HCCR-2 may be involved in cell regulation and the proliferation of hepatocellular carcinoma cells.     

HCCR-2反义核酸对肝癌HepG2细胞的作用

Objective To investigate the effects of antisense recombinant euraryotic expression vector of HCCR-2 on the proliferation and apoptosis of HepG2. Methods The antisense recombinant eukaryotic expression vector of HCCR-2 was constructed. The vector was stably transfected to the HepG2 cells, and positive clones were selected by G418 (antiseuse vector group), pIRES2-EGFP vector was transfected into the HepG2 cells in the same way (pIRES2-EGFP group). The conditions of the nontransfected HepG2 cells were used as control (HepG2 group). Changes in cell growth curve, cell cycle, cell apoptosis and morphology of HepG2 cells after the transfec-tion were detected by MTT method, flow cytometry and transmission electron microscopy, respectively. All the data were analyzed by one-way ANOVA and chi-square test. Results The expression level of HCCR-2 mRNA was down-regulated to 0.39±0.04 in antisense vector group, and the expression level of HCCR-2 mRNA in pIRES2-EGFP group and HepG2 group were 0.62±0.06 and 0.72±0.03, respectively, with significant difference among the 3 groups (F=43.701, P<0.05). The apoptotic rate of HepG2 cells in antisense vector group, pIRES2-EGFP grop and HepG2 group were 13.30%, 2.51% and 2.07%, respectively, with significant difference among the 3 group (χ2=6.793, 8.721, P<0.05). The growth of HepG2 cells in antisense vector group was retarded, and was blocked in G0/G1 stage. Conclusions The HCCR-2 antisense recombinant eukaryotic expression vector can inhibit the mRNA expression of HCCR-2 and promote the apoptosis of cells. HCCR-2 may be involved in cell regulation and the proliferation of hepatocellular carcinoma cells.     

人子宫颈癌基因-2干扰抑制HepG2肝癌细胞mRNA的表达

目的利用构建的特异性针对HCCR-2的干扰质粒RNAi-H1,研究HCCR-2干扰对HepG2肝癌细胞mRNA的影响.方法将干扰质粒用脂质体法转染HepG2细胞,经G418持续压力选择和有限稀释法获得稳定转染的细胞系.Real-timePCR检测HCCR-2mRNA表达水平.结果Real-time PCR结果表明HepG2-H1mRNA表达明显减少.结论HCCR-2干扰可以抑制HepG2肝癌细胞mRNA的表达.     

飞行员服用西沙必利致心律失常1例

1 临床资料 患者,男,24岁,歼击机飞行员,飞行时间1600h。该飞行员患慢性浅表性胃炎伴胆汁返流,但不影响飞行。2000年1月患者听说西沙必利治胃病效果好,遂自购西沙必利2盒,每次口服10 mg,每日3次,自觉效果非常显著。服药数日后,在一次打篮球中曾出现轻度头晕、乏力。航医检查其脉搏为61次/min,血压100/70 mmHg(该飞行员平时安静状态下脉搏77次/min,血压115/75 mmHg),认为是疲劳所致,未在意,建议其适当休息。此后断续服用西沙必利,用药     

老年多器官功能不全综合征患者免疫功能探讨

目的:检测老年多器官功能不全综合征(MODSE)患者的细胞免疫、体液免疫指标,了解其免疫水平及其与疾病的关系。方法:将老年患者根据诊断分为MODSE组(46例)和对照组(40例),用双/三色直接免疫荧光标记全血溶血法检测细胞免疫CD3~+、CD3~+CD4~+、CD3~+CD8~+、CD4~+/CD8~+、NK、CIK指标;用免疫比浊法检测体液免疫C3,C4,CH50,IgG,IgA,IgM指标。结果:两组患者CD3~+,NK,IgA差异有统计学意义(P<0.05)。两组免疫功能比较有明显差异:对照组CD3~+高,MODSE组NK、IgA高。结论:MODSE组患者存在免疫功能低下。     

龙血竭乙醇液与云南白药气雾剂治疗急性软组织损伤的临床对照研究

目的研究龙血竭乙醇液外用治疗急性软组织损伤的临床疗效及不良反应。方法将急性软组织损伤患者148例随机分为两组,治疗组75例用龙血竭粉与75%乙醇液以1：2比例稀释涂敷患处,对照组73例用云南白药气雾剂外喷患处.两组疗程均为14d。结果两组治疗方法对急性软组织损伤均有良好的治疗作用,治疗组疗效优于对照组。结论龙血竭乙醇液外用治疗急性软组织损伤具有快速祛瘀、消肿、镇痛和加速受损组织修复作用,值得推广。     

内毒素结合肽模拟肽P11的体外活性研究

目的 对从噬菌体展示随机肽库筛选获得的内毒素结合肽模拟肽进行体外拮抗内毒素活性鉴定.方法 采用FMOC固相合成法化学合成内毒素结合肽模拟肽P11,并进行拮抗内毒素活性和细胞毒性测定.结果 亲和ELISA检测显示P11与LPS有较高的亲和力,通过生长曲线和流式细胞学分析细胞周期显示P11对人U937细胞生长无明显影响.流式细胞检测显示P11呈剂量依赖性抑制FITC-LPS与人外周血单核细胞(PBMC)结合.细胞因子生成抑制实验显示10 μg/ml P11可显著抑制LPS诱导PBMC和U937细胞TNF-α mRNA转录和蛋白表达.结论 体外活性鉴定结果表明化学合成的模拟肽P11可抑制LPS诱导的炎性反应.     

飞行员非阵发性房室交界性心动过速1例

1 临床资料 患者男性,31岁,歼-6飞行员,飞行时间1000h。无明显诱因突发心悸,伴头晕、恶心、持续约3min来诊。就诊时症状已缓解。查体:生命体征平稳。心电图示:心率88次/min,律齐。Ⅱ、Ⅲ、aVF导联P波侄倒置,Ⅱ、Ⅲ导联还可见房性融合波。其余导联为窦性P波,P-R间期0.10s,QRs     

HCCR-2反义核酸对肝癌HepG2细胞的作用

Objective To investigate the effects of antisense recombinant euraryotic expression vector of HCCR-2 on the proliferation and apoptosis of HepG2. Methods The antisense recombinant eukaryotic expression vector of HCCR-2 was constructed. The vector was stably transfected to the HepG2 cells, and positive clones were selected by G418 (antiseuse vector group), pIRES2-EGFP vector was transfected into the HepG2 cells in the same way (pIRES2-EGFP group). The conditions of the nontransfected HepG2 cells were used as control (HepG2 group). Changes in cell growth curve, cell cycle, cell apoptosis and morphology of HepG2 cells after the transfec-tion were detected by MTT method, flow cytometry and transmission electron microscopy, respectively. All the data were analyzed by one-way ANOVA and chi-square test. Results The expression level of HCCR-2 mRNA was down-regulated to 0.39±0.04 in antisense vector group, and the expression level of HCCR-2 mRNA in pIRES2-EGFP group and HepG2 group were 0.62±0.06 and 0.72±0.03, respectively, with significant difference among the 3 groups (F=43.701, P<0.05). The apoptotic rate of HepG2 cells in antisense vector group, pIRES2-EGFP grop and HepG2 group were 13.30%, 2.51% and 2.07%, respectively, with significant difference among the 3 group (χ2=6.793, 8.721, P<0.05). The growth of HepG2 cells in antisense vector group was retarded, and was blocked in G0/G1 stage. Conclusions The HCCR-2 antisense recombinant eukaryotic expression vector can inhibit the mRNA expression of HCCR-2 and promote the apoptosis of cells. HCCR-2 may be involved in cell regulation and the proliferation of hepatocellular carcinoma cells.