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Effect of 5-Aza-2’-Deoxycytidine on Growth of Renal Carcinoma Line OS-RC-2 and Hypermrthylation of γ-Catenin Gene

Author: TaoMeiMan
Tutor: SunHao
School: Jiangsu University
Course: Surgery
Keywords: 5 - aza -2'- deoxycytidine OS-RC-2 renal cancer cell lines γ-catenin gene Apoptosis Immunohistochemical Western blot Methylation
CLC: R737.11
Type: Master's thesis
Year: 2009
Downloads: 60
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Abstract


Objective: To study 5 - aza-2'-deoxycytidine (5-Aza-CdR), the proliferation of human renal cell carcinoma OS-RC-2 cell lines, apoptosis, and further analysis of 5-Aza-CdR treatment of γ- catenin gene methylation status, and explore the feasibility of kidney cancer pathogenesis and clinical treatment. Methods: (1) using different concentrations (10 -7 10 -6 10 -5 , 10 -4 mol / L) of the specific DNA methyltransferase inhibitors 5-Aza-CdR treatment OS-RC-2 renal cancer cell lines. (2) inverted microscope and transmission electron microscope observation of morphological changes in cells before and after 5-Aza-CdR treatment OS-RC-2 renal cancer cell lines; (3) four methyl thiazolyl tetrazolium (MTT) assay 5-Aza-CdR OS-RC-2 inhibition rate of renal cancer cell line; (4), flow cytometry (FCM) detection of 5-Aza-CdR OS-RC-2 renal cancer cell line apoptosis; (5) cells immunochemical detection of 5-Aza-CdR treatment OS-RC-2 renal cancer cell lines after γ-catenin expression changes; (6) Western blot assay (Western blot) 5-Aza-CdR treatment OS-RC-2 kidney cancer changes in γ-catenin protein expression in the cell lines; (7) methylation-specific PCR (methylation-specific PCR, MSP) before and after the treatment of the detection cell γ-catenin gene methylation status. Results: (1) morphological observation: the cancer volume reduced under an inverted microscope, nuclear pyknosis, chromatin condensation into blocks; transmission electron microscopy organelle swelling mitochondria cavitation lysosomes nucleoplasm uneven. (2) MTT detected :5-Aza-CdR significantly inhibited OS-RC-2 renal cancer cell proliferation, and the drug concentration and duration of action within a certain range into a positive correlation (P <0.05). (3) Flow cytometry showed: 10 -7 10 -6 10 -5 mol/L5-Aza-CdR OS-RC-2 renal carcinoma cell line after treatment, the increased rate of apoptosis (3.74 ± 0.34)% and (7.85 ± 0.59)%, (12.93 ± 1.32)% vs (0.86 ± 0.08)%, in a dose dependent (F = 189.7, P <0.01). (4) cells by immunohistochemistry and Western blot show drug treatment OS-RC-2 renal cancer cell lines after γ-catenin protein to restore expression, and γ-catenin protein expression levels of 5-Aza-CdR concentration was positively correlated (P < 0.05). (5) without 5-Aza-CdR treatment of OS-RC-2 renal cancer cells, γ-catenin gene promoter region hypermethylation, 10 -5 mol/L5-Aza-CdR for 72h, γ-catenin gene promoter region hypermethylation is reversed. Conclusion :5-Aza-CdR effective reversal of renal cell carcinoma OS-RC-2 cells γ-catenin gene abnormal methylation restore gamma-catenin protein expression to induce tumor cell apoptosis and inhibit the growth of kidney cancer cells.

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CLC: > Medicine, health > Oncology > Genitourinary tumors > Urinary tumors > Kidney,renal pelvis tumor
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