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The Influence of Cell Growth under the Regulation Mechanism of Let-7 in Lung Cancer

Author: PanLin
Tutor: GongChaoHui
School: Ningbo University
Course: Biochemistry and Molecular Biology
Keywords: Small cell lung cancer Small RNA (microRNA) Lin-28 let-7
CLC: R734.2
Type: Master's thesis
Year: 2011
Downloads: 89
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Abstract


Background significant lung cancer is the most common malignancy in the world, and participate in the development of a multi-factor, multi-gene changes and the development of a multi-stage process complex lesions. Small RNA (microRNA, miRNA) expression abnormalities and is closely related to the development of lung cancer and the prognosis of patients with lung cancer. Many tumor suppressor miRNA, let-7 expression levels in a variety of lung tissue and cells down, leading many downstream oncogenes such as Ras gene expression levels raised. Tumor levels of let-7 primary transcripts (pri-let-7) did not change significantly, but significantly decreased the expression levels of mature let-7. Latest studies show, some embryonic cells or tumor cells, RNA combined with protein Lin-28 can be specific with pri-let-7 and the let-7 before the body (pre-let-7) combined, affect RNase Ⅲ Drosha and Dicer of let-7 cutting. Meanwhile, Lin-28 also synergies with other proteins in the pre-let-7 3 'end formation of poly-uracil (Poly U), the formation of unstable intermediates, pre-let-7 degradation in the cytoplasm. Currently, very little let-7 in lung maturity studies on the regulation, therefore, to explore the regulatory mechanism of let-7 and its impact of the growth of lung cancer cells, lung cancer and development is important to reveal. The purpose of this study is to explore whether lung cancer cells NCI-H446 Lin-28 protein on the regulation of let-7 and the regulation of cell growth and the impact of important experimental evidence for a reasonable evaluation of lung cancer occurrence and development. Lin-28 siRNA, chemically synthesized siRNA sequences alternate research methods (1) is designed according to the principles of siRNA design; (2) as a negative control non-specific siRNA liposome lin-28 siNRA sequence and negative control sequence transfected cultures NCI-H446 cells; (3) by Real time RT-PCR and Western blot analysis of Lin-28 gene and protein expression levels change before and after transfection, judge lin-28 siRNA inhibition efficiency, while application Real time RT-PCR analysis of let-7a -1 and changes in expression levels of let-7g evaluation Lin-28 (4) using MTT and flow cytometry (FACS) analysis of lin-28 siRNA transfection of let-7a-1 and let-7g expression levels; After NCI-H446 cell proliferation and cell cycle changes, evaluation of the let-7 expression level changes affect the growth of lung cancer cells. Real time RT-PCR analysis of cell cycle-related protein gene expression levels to determine the reason for the change of the cell cycle. Results (1) in the human small cell lung cancer cell line NCI-H446, lin-28 expression levels raised pri-let-7a-1 and pri-let-7g expression levels lowered mature let-7a-1 expression levels significant changes, the expression levels of mature let-7g down. (2) lin-28 siRNA transfection, lin-28 mRNA levels and protein water dropped by an average of mature let-7g expression level increases at the same time; (3) lin-28 siRNA transfection, NCI-H446 cell growth slower changes in cell cycle and cell cycle arrest in G1 phase, Real time RT-PCR results showed that the cdc25A gene expression level down. Conclusion This study demonstrated that in human small cell lung cancer cell line NCI-H446, lin-28 gene expression levels, and inhibiting the expression of let-7g. If the lin-28 gene expression is suppressed, can significantly increase the level of expression of let-7g, to change the cell cycle, inhibition of cell growth.

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CLC: > Medicine, health > Oncology > Respiratory system tumors > Lung tumors
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