人体系统与功能整合课程是以器官系统为中心对基础医学课程进行整合而成的.本课题基于网络教学平台构建了人体系统与功能整合课程的网络课程,将混合式教学设计应用于该整合课程的教学实践中,并对该教学模式进行了综合评价,发现学生对该教学模式普遍接受,学习主动性有所提高,且可取得比单纯传统的教学模式更好的教学效果.

设计性实验是机能学实验中最重要的一个阶段,是一种模拟科研过程,由学生自行选题、大胆设计及探索研究的实验教学项目。以病理生理学为切入点,在2006级临床本科尝试开出的机能学设计性实验,结果表明通过设计性实验的开展有助于学生综合素质的培养和创新能力的提高,是未来机能学实验教学改革的重点。

Objective To evaluate tumor-targeting of transgenic bifidobacterium infants to melanoma in mice.Methods After bolus administration of transgenic bifidobacterium infantis,the values of radioactivity in tumor and organs were examined.Anaerobic culture and histological observation of tumor and normal tissues were taken for examination of tumor-targeting characteristics of transgenic bifidobacterium infantis.Results The radioactivity in melanoma tissue increased progressively,while the radioactivity in normal organs became attenuated with time.The anaerobic culture showed an obvious proliferation of bifidobacterium infantis in tumor tissue.A lame part of area was Gram in the tumor tissue section,whereas the normal tissue was Gram negative.Conclusion Bifidobacterium infantis has good tumor-targeting to melanoma in mice,and recombined gene can be transported technically to solid tumors.

Objective To construct CD/5-FC tumor gene therapy system delivered by bifidobacterium infantis, and to investigate the killing effect on melanoma B16-F10 cell of mice.Methods Recombined plasmid pGEX-CD was constructed and transferred into bifidobacterium infants by electroporation. Positive recombined bacteria was selected and identified, and then was incubated with 5-FC in anaerobic condition. The supernatant of bacteria was collected and added to the melanoma B16-F10 cells. The cell growth status was evaluated by MTT assay, and the morphologic changes of cells were observed. Results The CD/5-FC tumor gene therapy system delivered by bifidobacterium infantis has been constructed successfully. The survival rate of cells was significantly lower in CD group than those in control group and in pGEX group,and tumor cells of CD group were remarkably damaged morphologically. Conclusion The CD/5-FC system delivered by bifidobacterium infantis has the significant killing effect on melanoma B16-F10 cell of mice in vitro.

Designing experimentation is the most important phase in function experimentation teaching,and a simulation process of science research.Students choose the topic freely,design schema and practice by themselves.Based on pathophysilolgy,the article has practiced functional designing experimentation among the students of clinical medicine in 2006 grade.Results indicated that the practice of designing experimentation is helpful for raising general diathesis and innovative ability of students,and it will be the focus of function experimentation innovation in future.

Objective To construct CD/5-FC tumor gene therapy system delivered by Bifidobacterium Infants,and to investigate the killing effect on melanoma B16-F10 cell of mice.Methods Recombined plasmid pGEX-CD was constructed and transferred into bifidobacterium infants by electroporation.Positive recombined bacteria was selected and identified,and then was incubated with 5-FC in anaerobic condition.The supernatant of bacteria was collected and added to the melanoma B16-F10 cells.The cell growth status was evaluated by MTT assay,and the morphologic changes of cells were observed.Results The CD/5-FC tumor gene therapy system delivered by Bifidobacterium Infantis has been constructed successfully.The survival rate of cells was significantly lower in CD group than in control group and in pGEX group,and tumor cells of CD group were remarkably damaged morphologically.Conclusion The CD/5-FC system delivered by Bifidobacterium Infantis has the significant killing effect on melanoma B16-F10 cell of mice in vitro.

Objective To investigate protection of ischemic precondition(IPC)and postcondition(IPO)on renal ischemia reperfusion(I/R)injury in rabbits.Methods Firstly renal I/R injury models were used in this experiment.Forty male rabbits were randomly divided into 4 groups(n=10 each):groupⅠ:sham operation,groupⅡ:I/R injury,groupⅢ:IPC,groupⅣ:IPO.After reperfusion 60min,blood MDA content and SOD activity were measured.The changes of renal tissue were detected by microscope.Results SOD activity was significantly lower whereas the serum MDA content was significantly higher in I/R group、IPC group and IPO group than in sham group(P0.05).The serum SOD activity was significantly higher whereas MDA content was significantly lower in IPC group and IPO group than in I/R group.Microscopic examination showed that the renal I/R injury was significantly attenuated in IPC group and IPO group.Conclusion IPC and IPO attenuate the renal injury induced by I/R in rabbits,and the protective effect is associated with the enhancement of the body antioxidation.

Objective To evaluate the UPRT-enhanced antitumor effect of CD/5-FC suicide gene system.Methods Recombined plasmid pGEX-UPRT was constructed and transferred into bifidobacterium infants by electroporation.Positive recombined bacteria was selected and identified,and then was incubated with 5-FC in anaerobic condition.The supernatant of bacteria was collected and added to the melanoma B16-F10 cells.The cell growth status was evaluated by MTT assay,and the morphologic changes of cells were observed.Results Recombinant bifidobacterium infants could express UPRT correctly.The survival rate of cells was significantly lower in CD group、UPRT group and CD/UPRT group than in control group;and the survival rate of cells was significantly lower in CD/UPRT group than in CD group and UPRT group.The cells of CD/UPRT group were remarkably damaged morphologically,and the cell growth was significantly inhibited as compared with that of other groups.Conclusion Recombinant bifidobacterium infants with UPRT gene can significantly enhance antitumor effect of CD/5-FC suicide gene system on mice melanoma B16-F10 cells.

OBJECTIVE This study was conducted by using Bifidobacterium Infantis as a delivery system to transport suicide gene CD and UPRT to tumors in an attempt to assess the UPRT-enhanced antitumor effect of CD/5-FC. METHODS The recombinant plasmid pGEX-UPRT was constructed and transfered into Bifidobacterium Infantis by electroporation and then identified. The synergistic antitumor effect of coexpression CD and UPRT was determined by MTT method. And the morphologic changes of B16-F10 cells were observed. RESULTS Recombinant Bifidobacterium Infantis could express UPRT correctly. The suvival rate of cells administrated CD+ UPRT and 5-FC was significantly lower than that of control (P<0.01), and the 5-FC sensitivity (IC50 = 0.015 micromol/mL) exhibited a 8. 5-fold increase when compared with that of cells administrated CD alone (IC50 = 0.127 micromol/mL). The cells treated with CD+UPRT were remarkably damaged morphologically, and the growth of cells was significantly inhibited as compared with that of other groups. CONCLUSION Recombinant Bifidobacterium Infantis with UPRT gene can significantly enhance the killing effect of CD/5-FC suicide gene system on melanoma B16-F10 cells of mice.

甲状腺癌是一类常见的发生在内分泌腺的恶性肿瘤,也是头颈部最常见癌瘤之一.近年来,伴随着其他恶性肿瘤基础和临床研究的发展,甲状腺癌的外科临床和基础研究亦有长足进步,这对于帮助了解甲状腺癌及其诊治开辟了广阔的前景.甲状腺癌的发生、发展与其他恶性肿瘤一样,与多种癌基因、抑癌基因、转移相关基因关系密切[1],现将国内外甲状腺癌的分子病理学研究现状及动态进展介绍如下.

Objective: To construct Bifidobacterium Infantis/CD targeting gene therapy system. Methods:CD gene was amplified from E. Coli K12λ using PCR method. pGEX-1LamdaT plasmid and CD gene weredigested with dual restriction endonucleas of EcoR I and BamH I and two segments of 4.9 kb and 1.3 kb wereobtained. T4 DNA ligase was added to these two segments to make a recombinant CD/pGEX-1LamdaTplasmid. Then the recombinant plasmid was transfected into Bifidobacterium Infantis by electroporation.The recombinant plasmid was extracted from the positively transfected Bifidobacterium Infantis and digestedwith dual restriction endonucleases. Then the size of digested fragments was detected and sequencingof the gene segment inserted in extracted recombinant plasmid was performed according to the methodof Sanger dideoxynucleotide triphosphate chain termination. Results: 6.2 kb recombinant plasmid wasobtained from the positively transfected bacterial colony of Bifidobacterium Infantis. After being digestedwith dual restriction endonucleases, two segments of approximate 4.9 kb and 1.3 kb were gained from theextracted recombinant plasmid, which were equal to the size of pGEX-1LamdaT plasmid and CD gene,respectively. The full length and sequence of nucleotide acid of the inserted gene in extracted recombinantplasmid was completely identical to the CD gene. Conclusion: The foreign gene, CD gene was correctlyinserted into pGEX-1LambdaT plasmid and transferred into Bifidobacterium Infantis. BifidobacteriumInfantis/CD targeting gene therapy system was successfully constructed.

Aim: To construct a Bifidobacterium infantis/CD targeting gene therapy system and observe the antitumor effect of cytosine deaminase/5-fluorocytosine (CD/5-FC) suicide gene therapy system mediated by Bifidobacterium infantis on melanoma in vitro and in vivo. Methods: A recombinant CD/pGEX-1 LamdaT plasmid was transfected into Bifidobacterium infantis by electroporation. Bifidobacterium infantis transfected by recombinant CD/pGEX-1LamdaT plasmid was incubated with 5-FC anaerobically. Then the supernatant fluid was collected and added to melanoma B 16-F10 cells to observe the killing effect for B 16-F10 cells.Mice were inoculated with melanoma B 16-F10 cells to establish animal models.The mice were then injected with 5-FC and Bifidobacterium infantis transfected by recombinant CD/pGEX-1LamdaT plasmid. Results:Two segments of approximate 4.9kb and 1.3 kb were extracted from the 6.2 kb recombinant plasmid, which were equal to the size of the pGEX-1LamdaT plasmid and CD gene, respectively.Sequencing results showed that the full length and sequence of nucleotide acid of the inserted gene in extracted recombinant plasmid was completely identical to the CD gene. In vitro, B 16-F10 cells treated by supernatant fluid were remarkably damaged morphologically, and the cell growth was significantly inhibited. Experiments on the mice melanoma model showed that after treatment with a combination of transfected Bifidobacterium infantis and 5-FC, the tumor volume was significantly inhibited compared with controls. Conclusion: The foreign gene,CD gene, was correctly inserted into pGEX-1LambdaT plasmid and transferred into Bifidobacterium infantis. CD/5-FC suicide gene therapy system mediated by Bifidobacterium infantis demonstrated a good antitumor effect on melanoma in vitro and in vivo.

Aim: To construct a Bifidobacterium infantis /CD targeting gene therapy system and observe the antitumor effect of cytosine deaminase/5-fluorocytosine (CD/5-FC) suicide gene therapy system mediated by Bifidobacterium infantis on melanoma in vitro and in vivo . Methods: A recombinant CD/pGEX-1LamdaT plasmid was transfected into Bifidobacterium infantis by electroporation. Bifidobacterium infantis transfected by recombinant CD/pGEX-1LamdaT plasmid was incubated with 5-FC anaerobically. Then the supernatant fluid was collected and added to melanoma B16-F10 cells to observe the killing effect for B16-F10 cells. Mice were inoculated with melanoma B16-F10 cells to establish animal models. The mice were then injected with 5-FC and Bifidobacterium infantis transfected by recombinant CD/pGEX-1LamdaT plasmid. Results: Two segments of approximate 4.9 kb and 1.3 kb were extracted from the 6.2 kb recombinant plasmid, which were equal to the size of the pGEX-1LamdaT plasmid and CD gene, respectively. Sequencing results showed that the full length and sequence of nucleotide acid of the inserted gene in extracted recombinant plasmid was completely identical to the CD gene. In vitro , B16-F10 cells treated by supernatant fluid were remarkably damaged morphologically, and the cell growth was significantly inhibited. Experiments on the mice melanoma model showed that after treatment with a combination of transfected Bifidobacterium infantis and 5-FC, the tumor volume was significantly inhibited compared with controls. Conclusion: The foreign gene, CD gene, was correctly inserted into pGEX-1LambdaT plasmid and transferred into Bifidobacterium infantis . CD/5-FC suicide gene therapy system mediated by Bifidobacterium infantis demonstrated a good antitumor effect on melanoma in vitro and in vivo .

BACKGROUND To construct the Bifidobacterium Infantis/CD tumor targeting gene therapy system. METHODS PCR expanded CD gene and pGEX-1LambdaT plasmid was transferred into E.Coli. JM109. with TSS solution. Then a dual restriction enzyme (EcoR I and BamH I) digestion was carried out to cut CD gene and pGEX-1LambdaT plasmid. Two segments (4.9 kb and 1.3 kb) were selected and extracted from the gel. T4 DNA Ligase was added to these two segments' mixture. Finally, reconstruction fragment was transferred into Bifidobacterium Infantis by electroporation. The plasmid was extracted from the positive clone selected and testified by an agarose eletrophoresis after enzymed. RESULTS A positive transferred Bifidobacterium Infantis with 6.2 kb long reconstruction plasmid fragment with CD gene was established which was in accordance with theoretical calculation. CONCLUSIONS The Bifidobacterium Infantis/CD tumor targeting gene therapy system could be successfully constructed.

Hypoxia-inducible factor is a nuclear transcriptio na l factor that exists all over the body.It has α and β subunits.The HIF-1 is a nucleoprotein that produced by cells undergone hypoxia.Its binding to the targe t genes will regulate many genes’ expression such as endothelin、VEGF and iNOS and therefore lead to the systematic responses to hypoxia.The activation of HIF -1 is of very importance to the development of the heat and to the compensation of the myocardium in hypoxia and ischemia diseases.Investigation to the relatio nship between HIF-1 and cardiac adaptation to hypoxia and ischemia will give a new way to the therapy of cardiac diseases.

目的将携带胞嘧啶脱氨酶(CD)基因的重组婴儿双歧杆菌应用于肿瘤的基因导向酶前药物疗法,利用厌氧菌趋低氧代谢的特性构建一种全新的肿瘤靶向基因治疗系统.方法PCR扩增CD目的基因、酶切后连接目的基因与质粒片段,电穿孔法将重组体转染婴儿双歧杆菌,筛选阳性克隆抽提质粒,酶切后琼脂糖凝胶电泳鉴定,并体外检测CD基因的表达.结果携带自杀基因CD的重组载体成功地被转染进婴儿双歧杆菌,并能表达CD酶活性,当加入CD酶底物5-FC后,显示出对小鼠黑色素瘤B16-F10细胞明显的杀伤效应.结论成功构建了携带自杀基因CD的重组婴儿双歧杆菌,重组菌在体外显示出对B16-F10肿瘤细胞有明显的抑制效应.

基因导向性酶前体药物疗法(GDEPT)为肿瘤生物治疗最引人注目的研究领域,是继手术、放疗和化疗等传统疗法之后,一种具有良好应用前景的肿瘤治疗新措施,主要包括HSV-tk-GCV系统疗法和CD-5-FC系统疗法.为克服传统GDEPT疗法的不足,目前多主张联合基因治疗肿瘤,有人还提出一种全新的GDEPT疗法.现综述近几年来GDEPT在肿瘤基因治疗中的研究进展.