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The study of human PDGF-B gene transferred to cat corneal endothelial cells

International Journal of Ophthalmology(English edition) 2012年第1期5卷 18-22页

作者： Wen-Juan Luo, chuan-fu wang Department of Ophthalmology the Affiliated Hospital of Medical College Qingdao University Qingdao 266003 Shandong Province China School of Pharmacy Jiangxi Science and Technology Normal University Nan Chang 330013 Jiangxi Province China Department of Pathology the Affiliated Hospital of Medical College Qingdao University Qingdao 266003 Shandong Province China

AIM: To demonstrate that human platelet-derived growth factor-B (PDGF-B) cDNA could be Expressed in primary cultured cat corneal endothelia cells by using gene transfer techniques; to explore a useful tool for the further studies of the molecular mechanisms of corneal endothelium failure and provide a potential effective genetic therapy for the blind patients. ' METHODS: Human PDGF-B cDNA was isolated from human placent by RT-PCR and inserted into pcDNA(4) vector to construct recombinant eukaryotic expression plasmid pcDNA(4)-PDGF-B. The full length was confirmed by the DNA sequencing analysis. By tearing endothelium technique we obtained pure single layer of cat corneal endothelial cells. The pcDNA(4)-PDGF-B eukaryotic Expression vector was transferred into cat corneal endothelial cells by Effectene (TM) lipofectine. The transfection efficiency of Effectene (TM) lipofectine in pcDNA(4)-B was detected with pcDNA(4)-GFP. 5 days later, RT-PCR was used to check the PDGF-B expression. Cell viability was tested by modified tertrozalium salt (MU) method. Cell morphology was observed under inverted phase contrast microscope. RESULTS: The human PDGF-B cDNA was isolated successfully from healthy parturien placent tissue and the sequence was confirmed by computer automatic sequence and PCR analysis. Pure single layer cat corneal endothelial cells were successfully cultured by tearing endothelium technique. Effectene (TM) lipofectine transfection technique could be effectively used to transfer pcDNA(4)-PDGF-B into cat corneal endothelial cells in vitro, the transfection efficiency was 30%. RT-PCR result showed that human PDGF-B gene was highly expressed in transfected cat corneal endothelial cells. The expressed PDGF-BB protein promoted the viability of cat corneal endothelial cells. CONCLUSION: Human platelet-derived growth factor-B (PDGF-B) cDNA could be highly Expressed in cultured cat corneal endothelial cells by gene transfection techniques. Expressed PDGF-BB protein sign

关键词： platelet-derived growth factor corneal endothelial cell viability gene transfection.

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Effect of basic fibroblast growth factor on cat corneal endothelial cell proliferation

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International Journal of Ophthalmology(English edition) 2011年第4期4卷 384-387页

作者： Wen-Juan Luo Yang Zhou Mei-Guang Liu and chuan-fu wang Department of Ophthalmology the Affiliated Hospital of Medical College Qingdao University Qingdao 266003 Shandong Province China

AIM: To investigate the function of basic fibroblast growth factor (bFGF) on cat corneal endothelial cells proliferation. METHODS: Cat corneal endothelial cells were primarily cultured, stimulated with bFGF for different period, the proliferation of cells was assayed by modified tertrozalium salt (MTT) method, and the morphologic changes were observed with inverted phase contrast microscope and transmission electron microscope. RESULTS: At 1, 3 and 5 days after bFGF was added to cat corneal endothelial cells, the result of MTT in 490nm showed significant difference than that in control group, and the difference was most significant in 10ng/mL group. CONCLUSION: bFGF can promote proliferation of cat corneal endothelial cells. 10ng/mL is the relatively most effective dose.

关键词： basic fibroblast growth factor corneal endothelial cells modified tertrozalium salt assay

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Human β-NGF gene transferred to cat corneal endothelial cells

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International Journal of Ophthalmology(English edition) 2016年第7期9卷 937-942页

作者： Wen-Juan Luo Min Liu Gui-Qiu Zhao chuan-fu wang Li-Ting Hu Xiang-Ping Liu Department of Ophthalmology the Affiliated Hospital of Medical College Qingdao University Central Laboratory of the Affiliated Hospital of Medical College Qingdao University

AIM： To transfect the cat corneal endothelial cells （CECs） with recombinant human β-nerve growth factor gene adeno-associated virus （AAV-β-NGF） and to observe the effect of the expressed β-NGF protein on the proliferation activity of cat CECs. METHODS： The endothelium of cat cornea was torn under the microscope and rapidly cultivated in Dulbecco＇s modified Eagle＇s medium （DMEM） to form single layer CECs and the passage 2 endothelial cells were used in this experiment. The recombinant human AAV-β-NGF was constructed. The recombinant human AAV-β-NGF was transferred into cat CECs directly. Three groups were as following： normal CEC control group, CEC-AAV control group and recombinant CEC- AAV-β-NGF group. Forty-eight hours after transfection, the total RNA was extracted from the CEC by Trizol. The expression of the β-NGF target gene detected by fluorescence quantitative polymerase chain reaction; proliferation activity of the transfected CEC detected at 48h by MTT assay; the percentage of G1 cells among CECs after transfect was detected by flow cytometry method （FCM）; cell morphology was observed under inverted phase contrast microscope. RESULTS： The torn endothelium culture technique rapidly cultivated single layer cat corneal endothelial cells. The self-designed primers for the target gene and reference gene were efficient and special confirmed through electrophoresis analysis and DNA sequencing. Forty-eight hours after transfect, the human β-NGF gene mRNA detected by fluorescence quantitative polymerase chain reaction showed that there was no significant difference between normal CEC control group and CEC-AAV control group （P〉0.05）; there was significant difference between two control groups and recombinant CEC-AAV-β-NGF group （P〈0.05）. MTT assay showed that transfect of recombinant AAV-β-NGF promoted the proliferation activity of cat CEC, while there was no significant difference between normal CEC control group and CEC-AAV control group （P〉0.05）. FCM result show

关键词： nerve growth factor corneal endothelialcell transfect proliferation

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Cloning, expression and functional analyses of human platelet-derived growth factor-B chain peptide for wound repair of cat corneal endothelial cells

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Chinese Journal of Traumatology 2009年第1期12卷 -页

作者： Wen-juan LUO[a] Gui-qiu ZHAO[a] chuan-fu wang[a] Li-mei wang[b] Xiao-ji wang[c] Department of Ophthalmology Affiliated Hospital Medi-cal College Qingdao University Qingdao 266003 China Department of Ophthalmology Medicine and Drug College Ocean University of China Qingdao 266003 China Department of Ophthalmology School of Pharmacy Jiangxi Science and Technology Normal University Nanchang 330013 China

Objective: To investigate the biological function of platelet-derived growth factor B (PDGF-B) on the survival and proliferation of cat corneal endothelial cells so as to provide bases for further studies of its role in wound repair and its clinical ***: Total RNA was extracted from the placenta tissues of healthy pregnant women undergoing hysterotokotomy and PDGF eDNA was obtained with re-verse transcription-polymerase chain reaction (RT-PCR). The prokaryotic expression vector pET-PDGF-B was constructed and expressed the recombinant PDGF-B in Escherichia coli (***) BL21 (DE3). After purification and refolding on Ni2+-chelation affinity chromatography (NTA) column, it was used to culture cat corneal endothelial cells. Cell proliferation was tested by modified tertrazolium salt (MTT) and flow cytometer. And the morphologic change and the ultrastructure were ob-served under an inverted phase contrast microscope, a scan-ning electron microscope and a transmission electon microscope, ***: PDGF-B chain peptide (PDGF-BB) gene was successfully inserted into the prokaryotic expression vector, pET-28a(+). The purified recombined protein pET-PDGF-B showed a single band on sodium dodecyl sulfate polyacry-lamide gel electropheresis (SDS-PAGE) with the molecular weight of about 27 u, which was in agreement with the de-duced value. MTT and flow cytometry showed that PDGF-BB promoted the survival and proliferation of cat corneal en-dothelial ***: The construction of recombinant prokary-otic expression vector pET-PDGF-B and the preparation of PDGF-BB protein provide a foundation for further study of the function of PDGF-BB and producing biological PDGF-BB protein. The expressed PDGF-BB promotes the prolif-eration of cultured cat corneal endothelial cells.

关键词： Platelet derived growth factor Gene expression Corneal endothelium Proliferation

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Conventional interferon-α plus ribavirin therapy yields 98% sustained virologic response in children aged 1-5 years with unsafe injection-acquired chronic hepatitis C

Conventional interferon-α plus ribavirin therapy yields 98%...

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中华医学会第十三次全国感染病学术会议

作者： ShiShu Zhu Qing-Lei Zeng Yi Dong Zhi-Qiang Xu Li-Min wang Da-Wei Chen Jin Han Yu Gan fu-chuan wang Jian-Guo Yan Li-Li Cao Pu wang Xue-Xiu Zhang Hong-Fei Zhang fu-Sheng wang Treatment and Research Center for Childrens Liver Disease Beijing 302 Hospital The Institute of Translational Hepatology Beijing 302 HospitalPeking University Research Center for Biological Therapy Beijing 302 Hospital

Backgroud During the winter of 2011,a public health emergency occurred,wherein hundreds of children contracted hepatitis C virus(HCV)infection caused by unsafe injection praetiees in a rural clinic at the border between the Henan and Anhui provinces in ***,epidemiologieal,clinical,and antiviral efficaey data for children aged i-5 years is very *** We collected detailed data of the epidemiological and clinical eharacteristies of 256 children aged 1-5 years with HCV infection after *** therapy with conventional interferon-αplus ribavirin was administered to 162 children with HCV RNA-positive ehronie

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矿床成矿系列——五论矿床的成矿系列问题

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地球学报 2016年第5期37卷 519-527页

作者：陈毓川裴荣富王登红黄凡中国地质科学院北京100037 中国地质科学院矿产资源研究所国土资源部成矿作用与资源评价重点实验室北京100037

矿床的成矿系列(简称成矿系列)是矿床学领域的一个理论性概念,由五级序次组成。矿床成矿系列是成矿系列的第二序次,是成矿系列的核心部分。矿床成矿系列的划分,以岩浆、沉积、变质、表生和流体(非岩浆-非变质成因流体)5种成矿作用为基... 详细信息

矿床的成矿系列(简称成矿系列)是矿床学领域的一个理论性概念,由五级序次组成。矿床成矿系列是成矿系列的第二序次,是成矿系列的核心部分。矿床成矿系列的划分,以岩浆、沉积、变质、表生和流体(非岩浆-非变质成因流体)5种成矿作用为基本原则,以成矿地质环境为基础,结合成矿的时段与形成的矿床组合进行划分。本文对矿床成矿系列时空范围、时空组成结构、矿化强度与演化、成矿区带内不同矿床成矿系列之间的演化、叠加和复合作用及对指导找矿的意义进行了论述。

关键词：矿床成矿系列概念划分原则时空圈定组成与演化找矿意义

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真丝织物乳液型环氧化合物抗皱整理

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丝绸 2001年第9期38卷 7-9页

作者：蒋国川王福爱蔡再生东华大学上海200051 上海海关化验中心上海200120

通过单因素试验和正交试验 ,讨论了抗皱整理过程中乳液型含硅环氧交联剂EPSIB浓度、添加剂浓度、焙烘温度、焙烘时间等因素对整理绸弹性折皱回复角和表面摩擦特性及刚柔性的影响。经EPSIB整理后的真丝绸干弹性折皱回复角提高 6 0°左右 ... 详细信息

通过单因素试验和正交试验 ,讨论了抗皱整理过程中乳液型含硅环氧交联剂EPSIB浓度、添加剂浓度、焙烘温度、焙烘时间等因素对整理绸弹性折皱回复角和表面摩擦特性及刚柔性的影响。经EPSIB整理后的真丝绸干弹性折皱回复角提高 6 0°左右 ,湿弹性折皱回复角提高 80°左右 ,表面平滑性。

关键词：真丝绸抗皱整理环氧交联剂丝织物

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Local recurrence after intended curative excision of presacral lesions:causes and preventions

Local recurrence after intended curative excision of presacr...

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中华医学会肿瘤学分会第七届全国中青年肿瘤学术会议——中华医学会肿瘤学分会“中华肿瘤明日之星”大型评选活动暨中青年委员全国遴选

作者： Xian Hua Gao,Wei Zhang,chuan Gang fu,Hao wang,En Da Yu,Rong Gui Meng,Lian Jie Liu Department of Colorectal Surgery of Changhai Hospital,Second Military Medical University, Shanghai 200433,China.

Objective To explore causes for local recurrence of presacral lesions after intended curative surgery and discuss prevention strategies. Methods Medical data of presacral lesions in our hospital from January 2001 to September 2009 were retrospectively studied,including preoperative examinations,intraoperative findings and postoperative histopathologies.

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前部完全包裹式羟基磷灰石义眼台植入术的效果

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国际眼科杂志 2009年第1期9卷 89-91页

作者：姜涛姜靖杨珊珊赵桂秋林静高岩王传富青岛大学医学院附属医院眼科中国山东省青岛市266003 青岛大学医学院附属医院内科中国山东省青岛市266003

目的：探讨前部完全包裹式羟基磷灰石（hydroxyapatite，HA）义眼台植入术的效果。方法：对52例患者施行前部完全包裹式羟基磷灰石义眼台植入术，术后观察外观、义眼台活动度情况及并发症的出现情况。结果：术后随访6—24mo。52例HA义... 详细信息

目的：探讨前部完全包裹式羟基磷灰石（hydroxyapatite，HA）义眼台植入术的效果。方法：对52例患者施行前部完全包裹式羟基磷灰石义眼台植入术，术后观察外观、义眼台活动度情况及并发症的出现情况。结果：术后随访6—24mo。52例HA义眼台居中，眼窝凹陷消失，义眼与健眼对称，外观满意。HA义眼台活动度优者42例（81％），良者10例（19％）。无HA义眼台脱出。出现结膜切口裂开5例（10％），结膜肉芽肿1例（2％），结膜囊狭窄1例（2％）。结论：HA义眼台植入术加强了义眼台的前部包裹物，降低了结膜切口裂开及义眼台暴露的发生率。

关键词：羟基磷灰石义眼台前部完全包裹式

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流动VLBI站氢原子钟的研制与性能测试(英文)

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计量学报 2002年第1期23卷 44-47页

作者：张为群林传富王关忠张一平于双林张健杨佩红中国科学院上海天文台上海200030

氢脉泽是至今为止除极短时间测量间隔之外最稳定的频率标准 ,它是射电天文所必需的频率标准。本文讨论了为流动VLBI站研制的新一代氢脉泽的物理结构特点 ,包括对腔泡结构、磁屏蔽、原子束光学系统等采用的新技术。性能测试表明 ,在 80s... 详细信息

氢脉泽是至今为止除极短时间测量间隔之外最稳定的频率标准 ,它是射电天文所必需的频率标准。本文讨论了为流动VLBI站研制的新一代氢脉泽的物理结构特点 ,包括对腔泡结构、磁屏蔽、原子束光学系统等采用的新技术。性能测试表明 ,在 80s到 1d之间其频率稳定度优于 1× 10 - 1 4 ,有了较大的提高。

关键词：氢脉泽频率稳定度腔调谐环境灵敏度相位锁定电子学频率标准流动VLBI站氢原子钟性能测试

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