CAJ | 학술논문

目的通过构建靶向于肾上腺髓质素（AM）基因的shRNA并转染卵巢癌细胞，观察抑制AM基因表达对卵巢癌细胞活性及化疗敏感性的影响。方法体外合成针对AM基因的shRNA序列，亚克隆至载体pRNA- U6.1/neo中，构建重组质粒pRNA- U6.1/neo- AM shRNA，转染至卵巢癌细胞中；通过RT- PCR检测转染后细胞内AM mRNA转录水平；MTT法检测细胞增殖活性及对顺铂敏感性变化；Western blot检测细胞外调节蛋白激酶（ERK）活性以及金属基质蛋白酶-9（MMP-9）表达。结果 RT- PCR表明，转染后细胞内AM mRNA水平明显下降，与阴性对照组相比，有统计学差异（P＜0.05）；MTT法结果表明，下调AM的表达后细胞增殖受到抑制，用顺铂处理后，AM shRNA组细胞存活率下降最为显著，与阴性对照组相比，有统计学差异（P＜0.05）；Western blot结果表明，下调卵巢癌细胞中AM的表达，可以抑制ERK的活性及MMP-9蛋白的表达。结论利用RNAi及基因转染技术下调卵巢癌细胞中AM的表达可以明显抑制卵巢癌细胞的增殖、转移能力并提高其对化疗药物顺铂的敏感性。
목적통과구건파향우신상선수질소（AM）기인적shRNA병전염란소암세포，관찰억제AM기인표체대란소암세포활성급화료민감성적영향。방법체외합성침대AM기인적shRNA서렬，아극륭지재체pRNA- U6.1/neo중，구건중조질립pRNA- U6.1/neo- AM shRNA，전염지란소암세포중；통과RT- PCR검측전염후세포내AM mRNA전록수평；MTT법검측세포증식활성급대순박민감성변화；Western blot검측세포외조절단백격매（ERK）활성이급금속기질단백매-9（MMP-9）표체。결과 RT- PCR표명，전염후세포내AM mRNA수평명현하강，여음성대조조상비，유통계학차이（P＜0.05）；MTT법결과표명，하조AM적표체후세포증식수도억제，용순박처리후，AM shRNA조세포존활솔하강최위현저，여음성대조조상비，유통계학차이（P＜0.05）；Western blot결과표명，하조란소암세포중AM적표체，가이억제ERK적활성급MMP-9단백적표체。결론이용RNAi급기인전염기술하조란소암세포중AM적표체가이명현억제란소암세포적증식、전이능력병제고기대화료약물순박적민감성。
Objective To investigate the effect of short hairpin RNA (shRNA) targeting adrenomedul in (AM) gene on chemosensitivity of ovarian cancer cel s. Methods Smal interfering RNA(siRNA) targeting AM gene was constructed and trans-fected to ovarian cancer HO8910 cel s. The expression of AM was detected by RT- PCR, Cel proliferation was examined by MTT assay and the expression of MMP- 9 and ERK was evaluated by Western blot. Results After transfection with siRNA targeting AM, the expression of AM gene in HO8910 cel s was inhibited and the expression of MMP- 9 and p- ERK was down- regulated. Meanwhile, the chemosensitivity of HO8910 cel s was increased. Conclusion Silencing of AM gene in ovarian cancer HO8910 cel s increases the chemosensitivity of the cel s, which is associated with the down- regulation of MMP- 9 and p- ERK proteins.