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Anti-HBV hairpin ribozyme-mediated cleavage of target RNA in vitro

World Journal of Gastroenterology 2002年第1期8卷 91-94页

作者： yu-husong Ju-ShengLin 等 InstituteofLiverDiseases TongjiHospitalTongjiMedicalCollegeHuazhongUniversityofScienceandTechologyWuhan430030HubeiProvinceChina InstituteofLiverDiseases TongjiHospitalTongjiMedicalCollegeHuazho

AIM: To study the preparation and cleavage activity of HpRzdirected against the transcript of HBV core gene in ***: HpRz gene designed by computer targeting thetranscript of HBV core gene was cloned into the vector p1 .5between 5'-cis-Rz and 3'-cis-Rz. 32p-labeled HpRz transcriptproved whether the vector fit for the preparation of hairpinribozyme in vitro. 32p-labeled pKC transcript containing HBVcore region as target-RNA was transcribed using T7 RNApolymerase and purified by denaturing PAGE. Cold HpRztranscript was incubated with 32p-labeled target-RNAs underdifferent conditions and radioautographed after denaturingpolyacrylamide gel ***: HpRz has the specific ability of cleavage of target-RNA at 37℃ and 12 mM MgCL2. Km = 26.31nmol/L, Kcat = 0.18/min. These results revealed that the design of HpRz ***: HpRz prepared in this study possessesspecific catalytic activity from the identification of cleavageactivity. These results indicate that hairpin ribozyme mayintracellularly inhibit the replication of HBV, therefore it maybecome a novel potent weapon for the treatment of hepatitisB.

关键词：乙型肝炎核酶 RNA分裂靶细胞抗病毒机制

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Preparation and identification of anti-transforming growth factor β1 U1 small nuclear RNA chimeric ribozyme in vitro

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World Journal of Gastroenterology 2003年第3期9卷 572-577页

作者： Ju-ShengLin yu-husong Xin-JuanKong BinLi Nan-ZhiLiu Xiao-LiWu You-XinJin InstituteofLiverDiseases TongjiHospitalTongjiMedicalCollegeHuazhongUniversityofScienceandTechnologyWuhan430030HubeiProvinceChina StateKeyLaboratoryofMolecularBiology InstituteofBiochemistryandCellBiologyShanghaiInstitutesforBiologicalSciencesChineseAcademyofSciencesShanghai200031China

AIM: To study the preparation and cleavage activity of antitransforming growth factor (TGF)β1 U1 small nuclear (sn)RNA chimeric hammerhead ribozymesin ***: TGFβ1 partial gene fragment was cloned into T-vector at the downstream of T7 promoter. 32p-labeled TGFβ1 partial transcripts as target RNA were transcribed in vitro and purified by denaturing polyacrylamide gel electrophoresis (PAGE). Anti-TGFβ1 ribozymes were designed by computer, then synthetic ribozyme fragments were cloned into the U1 ribozyme vector pZeoU1EcoSpe containing U1 snRNA promoter/enhancer and terminator.32p-labeled U1 snRNA chimeric ribozyme transcripts were gel-purified, incubated with target-RNAs at different conditions and autoradiographed after running denaturing ***: Active UlsnRNA chimeric ribozyme (U1Rz803)had the best cleavage activity at 50 °C; at 37 °C, it was active, Km=34.48 nmol/L, Kcat=0.14 min-1; while the point mutant ribozyme U1Rz803m had no cleavage activity, so these indicated the design of U1Rz803 was ***: U1Rz803 prepared in this study possessed the perfect specific catalytic cleavage activity. These results indicate U1 snRNA chimeric ribozyme U1Rz803 may suppress the expression of TGFβ1in vivo, therefore it may provide a new avenue for the treatment of liver fibrosis in the future.

关键词： RNA化学构成酶微核抗转换生长因子β1 分离纯化肝纤维化

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Maxizyme-mediated specific inhibition on mutant-type p53 in vitro

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World Journal of Gastroenterology 2003年第7期9卷 1571-1575页

作者： Xin-JuanKong yu-husong Ju-ShengLin Huan-JunHuang Nan-XiaWang Nan-ZhiLiu BinLi You-XinJin InstituteofLiverDisease TongjiHospitalTongjiMedicalCollegeHuazhongUniversityofScienceandTechnologyWuhan430030HubeiProvinceChina LaboratoryofMolecularBiology shanghaiInstituteofBiochemistryChineseAcademyofSciencesShanghai200031China

AIM: To evaluate the specific inhibition of maxizyme directing against mutant-type p53 gene (mtp53) at codon 249 in exon 7 (AGG→AGT)in ***: Two different monomers of anti-mtp53maxizyme (maxizyme right MzR, maxizyme left MzL) and control mutant maxizyme (G5→A5) were designed by computer and cloned into vector pBSKU6 (pBSKU6MzR,pBSKU6MzL). After being sequenced, the restrictive endonuclease site in pBSKU6MzR was changed by PCR and then U6MzR was inserted into pBSKU6MzL, the recombinant vector was named pU6Mz and pU6asMz (mutant maxizyme).Mtp53 and wild-type p53 (wtp53) gene fragments were cloned into pGEN-T vector under the T7 promoter *** 32p-labeled mtp53 transcript was the target mRNA. Cold maxizyme transcripts were incubated with 32p-labeled target RNA in vitro and radioautographed after denaturing polyacrylamide gel ***: In cell-free systems, pU6Mz showed a specific cleavage activity against target mRNA at 37 ℃ and 25 mM MgCL2. The cleavage efficiency of pU6Mz was 42 %, while pU6asMz had no inhibitory effect. Wtp53 was not cleaved by pU6Mz ***: pU6Mz had a specific catalytic activity against mtp53 in cell-free system. These lay a good fundation for studying the effects of anti-mtp53 maxizyme in HCC cell lines. The results suggest that maxizyme may be a promising alternative approach for treating hepatocellular carcinoma containing mtp53.

关键词： p53基因基因突变 Maxizyme 肝细胞癌

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一道高考解析几何题的推广

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河北理科教学研究 2011年第2期 16-17页

作者：邹生书余护松湖北省阳新县高级中学

2010年高考全国卷Ⅰ第21题如下：已知抛物线C：y2=4x的焦点为F，过点K（-1，0）直线l与C相交于A，B两点，点A关于x轴的对称点为D．（1）证明：点F在直线BD上；（2）设FA·FB=8/9，求ΔBOD的内切圆M的方程．

关键词：解析几何题高考推广 2010年全国卷抛物线对称点内切圆

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小骨窗开颅经额入路脑室内血肿清除术治疗侧脑室出血并铸型的研究

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现代中西医结合杂志 2015年第35期24卷 3902-3904页

作者：何洪雨刘华新杨志强于帅匡虎松解放军第149医院江苏连云港222042

目的观察小骨窗开颅经额叶入路脑室内血肿清除术治疗侧脑室出血并铸型的效果。方法将90例侧脑室内出血并铸型的患者随机分为2组,观察组45例行小骨窗开颅经额入路脑室内血肿清除术,对照组45例行单纯侧脑室外引流术。术后随访3~6个月,按... 详细信息

目的观察小骨窗开颅经额叶入路脑室内血肿清除术治疗侧脑室出血并铸型的效果。方法将90例侧脑室内出血并铸型的患者随机分为2组,观察组45例行小骨窗开颅经额入路脑室内血肿清除术,对照组45例行单纯侧脑室外引流术。术后随访3~6个月,按照格拉斯哥预后评分（GOS）评价2组临床疗效。结果观察组脑室内引流管留置时间及住院时间均明显短于对照组（P均〈0.05）。观察组恢复良好率明显高于对照组（P均〈0.05）,病死率及颅内感染、脑脊液漏发生率明显低于对照组（P均〈0.05）。结论小骨窗开颅经额入路脑室内血肿清除术治疗脑室出血并铸型疗效明显优于单纯侧脑室外引流术。

关键词：小骨窗开颅脑出血脑室铸型脑室外引流

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