上海交通大学医学院2012级研究生-文献导读如何全面把握科学文献的内容.ppt

1、How to Master the Topic from Scientific Paper 如何完整把握学术论文的主题内容,陈 万 涛上海交通大学医学院 附属第九人民医院口腔颌面外科 上海市口腔医学重点实验室 上海市重点学科（口腔基础医学）,Kinds of reading skill Intensive reading skill （精读） Extensive reading skill（泛读） Skimming reading skill（略读） Scanning reading skill（掠读）,Intensive reading 精读、细读 完全理解每词、每句和每段意思,Extens

2、ive reading 泛读、快读 理解段落的大体意思 理解关键字、关键句,Skimming reading 略读、快读 寻找文章的关键词、主题句 了解文章大体意思 查找、选取精读内容,Scanning reading 掠读、快速阅读 查找文章中特定信息,Reading fast & mastering the topic? 把握主题和快速阅读的可能技巧？,Title Abstract Results: Figures and Tibles Conclusions,Intensive reading skill,Results Introduction,Extensive reading sk

3、ill,Material and Methods Reference,Scanning reading skill,Discussion Reference,Skimming reading skill,Shanghai Museum,Selection and validation of differentially expressed genes in head and neck cancer,Paper 1,Selection and validation of differentially expressed genes in head and neck cancer,Title,Ab

4、stract We applied a robust combinatorial (multi-test) approach to microarray data to identify genes consistently up- or down-regulated in head and neck squamous cell carcinoma (HNSCC). RNA was extracted from 22 paired samples of HNSCC and normal tissue from the same donors and hybridized to the Affy

5、metrix U95A chip.,Abstract (continued) Forty-two differentially expressed probe sets (representing 38 genes and one expressed sequence tag) satisfied all statistical tests of significance and were selected for further validation. Selected probe sets were validated by hierarchical clustering,multiple

6、 probe set concordance, and target-subunit agreement. In addition, real-time PCR analysis of 8 representative genes performed on both microarray tested and independently obtained samples correlated well with the microarray data.,Abstract (continued),The genes identified and validated by this method

7、were in comparatively good agreement with other rigorous HNSCC microarray studies. We conclude that combinatorial analysis of microarray data is a promising technique for identifying differentially expressed genes with few false positives.,Abstract (continued),Differentially expressed genes in OSSC

8、detected by microarray,step I,Testable Hypothesis:Differentially expressed genescan be detected by microarray analysis,Normal tissue,OSSC tissue,GeneChip Human Genome U95A2 12,626 full-length genes. One sample One chip Total 22 pairs,Selection of differentially expressed genes between Tumor tissues

9、and Normal tissues,step II,a t-test b Wilcoxon rank-sum c paired t-test d SAM e PPV f MDMR g WEPO,Group A,Group B,Group C,七种统计学方法筛选差异表达基因,Combinatorial approach for analysis and selection,Step III Validation of Tier I genes,Hierarchical clustering All genes vs. Tier I genes RT-PCR Chip tested sample

10、s Independent (chip un-tested) samples,Hierarchical Clustering based on all (12642) probes,Hierarchical Clustering based on Tier I (42) probes,Selected genes may be the the useful target for molecular diagnosis of OSSC,Results of Real Time-PCR,Ratio grids showing selected and unselected probe sets f

11、or multi-probe set genes.,Tier I Genes Annotation Up-regulated Down-regulated,Structural: Col-1 (1, 2), Col-4 (1, 2), Col-5 (2), FN*, MFAP2 Enzymes: FAPA*, MMP1*, PLAU*, SERPINH2* TAA: OPN (x2)* Other: C5ORF13*, ITAG6, PTHLH*, Est,Structural: KRT*(4, 13), PPL*, SCEL* Enzymes: Enzyme: ACPP* (x2), CYP

12、3A5, DUSP5, GPX3*, HPGD, PP11, SPINK5*, TGM3* TAA: CEACAM* (1, 5), LAGY*, EMP1* Other: BLNK, IL1RA*, KIAA0790 MAL, NMU, PITX1*, ZNF185,Discussion,Study design and overall strategy The method is with fewer positive false, but some putative genes were missed. Rationale of the approach to selection of

13、differentially expressed genes. Validation of selected genes. Annotation and relevance of selected genes to malignancy.,conclusion,Combinatorial analysis of microarray data from 22 paired tumor/normal samples from HNSCC patients yielded 42 probe sets (representing 38 genes plus one EST) which satisf

14、y highly stringent criteria for significant over- or under-expression in HNSCC tissue. A subset of selected genes was validated by in silico analysis and real-time PCR of both chip-tested and independently obtained sample pairs to confirm their differential expression in HNSCC versus normal tissues.

15、,Conclusion(continued),Several genes identified in the present study were also found to be concordant with genes identified in other HNSCC microarray analyses of similar rigor. The strength of validation of these targets suggests that the intersection of mathematically distinct statistical methods c

16、an yield a reliable list of differentially expressed genes containing fewer false positives than a single method. This approach may have broad utility in the analysis of gene expression in HNSCC and other malignancies.,MA kuriakose,Fangan Chen,Zhiyuan Zhang,Ping Zhang,Wantao Chen,Identification of g

17、enes associated with cisplatin resistance in human oral squamous cell carcinoma cell line,Paper 2,Identification of genes associated with cisplatin resistance in human oral squamous cell carcinoma cell line,Title,Abstract,Tca induced by cisplatin,Established a drug-resistant cell line to cisplatin,

18、named Tca/cisplatin. Its IC50 was 6.5 times as that of parent cells,Step I,Tca cells,2 months,2 weeks,cisplatin,Growth of Cisplatin resistant variant cells when exposed in cisplatin,Tca,Tca/cisplatin,DNA staining by AO and EB Viable: green, dead: red,Viable: attached, dead: disattached,Platinum/DNA

19、(pg/ g),Platinum accumulation on DNA in Tca/cisplatin and Tca8113.,Effects of 10M cisplatin on Tca/cisplatin and Tca cell cycle distribution,Gene expression profile,Affymetrix HG-U95Av2,Tca,Tca/cisplatin,Step II,Differential expression gene analysis,DataMining Tool,Suite 5.0,25 decrease,38 increase,

20、12626 probe sets,Signal 500 Ratio2 P0.05,RT-PCR RT-PCR Western blot Western blot,CCND1 CCND3 Pgp GST-Pi,口腔鳞癌耐药相关基因的验证,Step III,Validation of gene expression profile,Cyclin D1,Cyclin D3,GST-Pi,P-gp,Tca/cisplatin,Tca,Tca/cisplatin,Tca,RT-PCR,Western blot,细胞周期调控基因CCND1和CCND3的表达,4 proteins examined by I

21、HC in clinical OSSC samples (n:50),The Tca/cisplatin and Tca8113 cell lines are useful models for identifying candidate genes responsible for the mechanism of cisplatin-resistance.,Conclusion,Sixty-three genes related to cisplatin-resistance were identified. Among these, decreases in cell cycle arre

22、st genes and increase in oncogenes, cell cycle regulation gene and genes involved in metabolism and synthesis led to the cell cycle acceleration, increased proliferation rate and resistance to cisplatin-induced apoptosis in Tca/cisplatin cells.,CCND1 and CCND3 seemed to be closely involved in cispla

23、tin resistance.,Conclusion(continued),The data from this study provide useful clues to screen candidate targets for early diagnosis and intervention in cisplatin-resistance.,Inhibition of cyclin D1 expression by cyclin D1 shRNAs in human oral squamous cell carcinoma cells is associated with increase

24、d cisplatin chemosensitivity,Paper 3,Inhibition of cyclin D1 expression by cyclin D1 shRNAs in human oral squamous cell carcinoma cells is associated with increased cisplatin chemosensitivity,Title,Cycling D1 is a well-known cell cycle regulator. Recently,its prosurvival function has been revealed i

25、n several tumors.Because increasing expression of cyclin D1 is a common event in OSCC and has been correlated with cisplatin resistance, we investigated if cyclin D1 inhibition could increase cisplatin chemosensitivity of OSCC.Five cyclin D1 shRNAs were prepared and 3 were selected for subsequent ex

26、periments. IC50 values for cisplatin were determined by an MTT assay. Cisplatin-induced apoptosis and cell cycle block were investigated.,Abstract,A tumor transplantation model was generated to examinethe cisplatin sensitivity of Tca/cisplatin after in vivo cyclin D1silencing.The most effective shRN

27、A resulted in 84.51% knockdown of the cyclin D1 protein level.After the transfection with the 2 most effective shRNAs, the cisplatin IC50 decreased from 5.88 ug/ml to 1.36 ug/ml and 2.47 ug/ml, overexpression of cyclin D1 rendered OSCC cells more resistant to cisplatin treatment (IC50 increased 1 ti

28、me),Abstract(continued),This decreasing IC50 was correlated with the down-regulationof cisplatin-induced NF-kB activity in cyclin D1 knockdowncells, and was independent of CDK4 function. In vivo, tumor transplantation models also confirmed a cisplatin-sensitizing effect of cyclin D1 shRNA in OSCC. A

29、 TUNEL assay validated the increase in apoptosis as induced by cisplatin in cyclin D1 knockdown cells. Cyclin D1 may be an important target for future therapy in patients with OSCC.,Abstract(continued),FIGURE 1 Cisplatin IC50 and cyclin D1 expression of OSCC cell lines.,FIGURE 2 Cyclin D1 shRNA decreased the expression