CAJ | 학술논문

目的分离并鉴定胆囊癌和胆囊良性组织的差异表达蛋白质,以发现可能用于早期诊断胆囊癌的肿瘤标志物.观察RNA干扰沉默膜联蛋白A3(AnnexinA3)基因对胆囊癌细胞增殖及周期的影响.方法提取人胆囊癌和胆囊良性组织的总蛋白质,用双向电泳分离蛋白并进行比较.选择差异表达超过2倍的蛋白点进行MALDI-TOF/TOF质谱和生物学分析.miRNA干扰胆囊癌细胞株中AnnexinA3的表达后,通过噻唑蓝(MTT)实验及流式细胞仪观察癌细胞增殖能力及细胞周期的变化.结果筛选出在胆囊癌组织中明显差异表达的46个蛋白点,共有17个蛋白质被成功鉴定,其中在胆囊癌组织中高表达的为9个,低表达的为8个,包括AnnexinA3、TTR蛋白等.RNA干扰胆囊癌细胞株AnnexinA3蛋白的表达后,MTT显示干扰后细胞增殖明显下降效率为44.14%(P<0.05).流式细胞仪观察显示干扰后G1期增加(P<0.05),S期减少(P<0.05).结论胆囊癌组织相对于胆囊良性组织蛋白存在明显差异.干扰AnnexinA3蛋白的表达可以改善胆囊癌细胞的某些恶性生物学行为.
목적 분리병감정담낭암화담낭량성조직적차이표체단백질,이발현가능용우조기진단담낭암적종류표지물.관찰RNA간우침묵막련단백A3(AnnexinA3)기인대담낭암세포증식급주기적영향.방법 제취인담낭암화담낭량성조직적총단백질,용쌍향전영분리단백병진행비교.선택차이표체초과2배적단백점진행MALDI-TOF/TOF질보화생물학분석.miRNA간우담낭암세포주중AnnexinA3적표체후,통과새서람(MTT)실험급류식세포의관찰암세포증식능력급세포주기적변화.결과 사선출재담낭암조직중명현차이표체적46개단백점,공유17개단백질피성공감정,기중재담낭암조직중고표체적위9개,저표체적위8개,포괄AnnexinA3、TTR단백등.RNA간우담낭암세포주AnnexinA3단백적표체후,MTT현시간우후세포증식명현하강효솔위44.14%(P<0.05).류식세포의관찰현시간우후G1기증가(P<0.05),S기감소(P<0.05).결론 담낭암조직상대우담낭량성조직단백존재명현차이.간우AnnexinA3단백적표체가이개선담낭암세포적모사악성생물학행위.
Objective To find potential molecular targets for gallbladder cancer diagnostics by analyzing and comparing the proteomes expressed in human gallbladder carcinoma and benign gallbladder tissues, and investigate whether annexinA3 down-regulation by miRNA leads to inhibition of proliferation and arrest of cell cycle in human gallbladder cancer cell line SGC-996. Methods The proteins expressed in these tissues were analyzed by the two-dimensional gel electrophoresis. Proteins expressed differently of a 2-fold change were cut and analyzed by MALDI-TOF/TOF mass spectrometry. After miRNA expression plasmid targeting to annexinA3 gene was constructed and transfected into cancer cells, the methyl thiazolyl tetrazolium ( MTT) assay was performed for measuring cell proliferation and flow cytometic analysis for cell cycle. Results Protein extracts of individual sample in each type of tissues were separated on two-dimensional gels. There were 46 differentially expressed proteins in the tissues of gallbladder cancer. Seventeen proteins were successfully identified by MS, in which 9 proteins were overexpressed in tumors and the rest 8 proteins were underexpressed, including Annexi-nA3, TTR protein, etc. After interfering annexinA3 in gallbladder cancer SGC-996 cell line, the cell proliferation was suppressed. Flow cytometry revealed that after interfering annexinA3 in gallbladder cancer SGC-996 cell line, the cells in G1 phase were increased, while those in S phase were declined as compared with control group (P<0.05). Conclusion Significant discrepancies in protein expression exist between gallbladder cancer tissues and benign gallbladder tissues. Some of the differentially expressed proteins found by the proteomic approach may be potential molecular targets for early diagnosis of gallbladder cancer. miRNA interfering to annexi-nA3 effectively reverses some of the malignant biological characters of gallbladder cancer cells.