2003 Fiscal Year Final Research Report Summary
Analysis of gene alteration in volved in endometrial carcinogensis using in vitro model of human normal endometrial glandular cells
| Project/Area Number |
14370527
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Obstetrics and gynecology
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| Research Institution | Shinshu University School of Medicine |
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Principal Investigator |
SHIOZAWA Tanri Shinshu University School of Medicine, assistant professor, 医学部附属病院, 講師 (20235493)
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| Co-Investigator(Kenkyū-buntansha) |
KONISHI Ikuo Shinshu University School of Medicine, Professor, 医学部, 教授 (90192062)
NIKAIDO Toshio Shinshu University Graduate School of Medicine, associate professor, 大学院・医学研究科, 助教授 (50180568)
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| Project Period (FY) |
2002 – 2003
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| Keywords | endometrium / endometrial carcinoma / immortalization / large T antigen / telomerase / subtraction / microarray |
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| Research Abstract |
The molecular mechanisms of endometrial carcinogenesis have not fully been understood. To analyze the pathogenesis of endometrial carcinoma in vitro, we planned to establish immortalized endometrial glandular cell lines. We first established an in vitro culture system of the normal endometrial glandular cells and we then transfected a large T antigen expression plasmid to the cultured cells. The transfected cells successfully survived until 20th passage of culture. We are now in preparation to transfect the large T antigen using viral vectors for better transfection efficiency to obtain longer viable passages.
To discover new gene alterations involved in endometrial carcinogenesis, we isolated mRNA from frozen tissue sections of the normal, hyperplastic and malignant endometrial glands of the identical patients using laser-captured microdissection. The isolated mRNA was then amplified using T7-RNA polymerase-based amplification, and the amplified mRNA were subjected to cDNA subtraction and microarray to detect genes whose expression differ among the normal, hyperplastic and malignant tissues. Subsequently, we cloned approximately 30 new genes which are overexpressed in neoplastic tissues compared to the normal counterparts, and approximately 20 new genes which are down-regulated in the neoplastic lesions. The tissue specificity of most of these gene were confirmed by semi-quantitative RT-PCR. We are currently investigating the further tissue specificity of the expression of these genes using in situ hybridization.
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