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Effects and Potential Mechanism of Notch1 Signal on Adhesion and Migration of Human Glioma U251 Cells

Author: WangXinYang
Tutor: ZhengZhiFu
School: Fujian Medical
Course: Biochemistry and Molecular Biology
Keywords: Glioma Notch signaling pathway Adhere to Migrate cadherin / catenin complex
CLC: R735.34
Type: Master's thesis
Year: 2011
Downloads: 51
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Abstract


Objective: To observe the Notch1 gene on human glioma U251 cell adhesion and migration ability; 2 to explore the mechanism of Notch1 gene affect U251 cell adhesion and migration. Methods: 1, the use the Notch1-shRNA and pNL-NICD/EGFP of slow virus infection of human glioma U251 cells, RT-PCR and Western Blotting method screening and identification of Notch1 downregulation or Notch1 intracellular domain (Notch1 intracellular domain, NICD) expression increase in U251 cells. 2, fibronectin (fibronectin, Fn) cell adhesion test, scratch test observation of adhesion and migration of cells in each group. 3, the cells in each group N-cadherin, αN-catenin and beta-catenin gene expression by RT-PCR and Western blotting assay, analysis of Notch1 three. 4, using co-immunoprecipitation method to detect N-cadherin, αN-catenin and beta-catenin relationship between the three. Results: 1, Notch1-shRNA lentiviral vector can effectively cut U251 cells expression of Notch1, pNL-NICD/EGFP lentiviral vector can be significantly up-regulated expression of U251 cells NICD. 2, Fn test, scratch test observed, of Notch1 down significantly enhanced cell adhesion, migration markedly weakened; decreased NICD increase cell adhesion, migration significantly enhanced. 3 of Notch1 down and NICD increases in cell N-cadherin, beta-catenin mRNA and protein expression with the control group showed no significant change. Notch1 downregulation αN-catenin mRNA and protein expression compared with the control group increased significantly, in the NICD raised cells αN-catenin mRNA and protein expression with the control group were significantly decreased compared. 4, the use of beta-catenin antibody immune co-precipitation experiments, be prepared to detect N-cadherin and αN-catenin, αN-catenin antibody co-immunoprecipitation, and also be prepared to detect N-cadherin and beta-catenin. Conclusion: 1, the of Notch1 gene with human glioma U251 cell adhesion, migration are closely related. Notch1 knockdown adhesion significantly enhanced migration capability significantly weakened; weakened NICD upregulation of cell adhesion, migration ability. In glioma U251 cells, N-cadherin, αN-catenin and beta-catenin between the three protein interaction. Notch1 signaling may increase the level of αN-catenin expression, affecting the cadherin / catenin complex thus changing the U251 cell adhesion and migration.

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CLC: > Medicine, health > Oncology > Gastrointestinal Cancer > Intestinal neoplasms > Colorectal tumors
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