中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
Chinese Journal of Experimental Surgery
2015年
9期
2096-2098
,共3页
谢仕刚%邱鹏%杨伟%张玉宝%徐同江%徐广明
謝仕剛%邱鵬%楊偉%張玉寶%徐同江%徐廣明
사사강%구붕%양위%장옥보%서동강%서엄명
垂体腺瘤%微小RNA Let-7b%细胞外基质金属蛋白酶诱导剂%基质金属蛋白酶-9
垂體腺瘤%微小RNA Let-7b%細胞外基質金屬蛋白酶誘導劑%基質金屬蛋白酶-9
수체선류%미소RNA Let-7b%세포외기질금속단백매유도제%기질금속단백매-9
Pituitary adenoma%MicroRNA Let-7b%Extracellular metalloproteinase inducer%Matrix metalloprotein-9
目的 探寻微小RNA Let-7b(Let-7b)在大鼠垂体腺瘤GH3细胞中的靶基因,并观察其对GH3细胞侵袭性生长的影响.方法 利用TargetScan、PicTar、MiRanda 3种不同的生物信息学方法对靶基因进行筛选;设计Let-7b模拟物(Let-7b mimic)及反义寡核苷酸链序列(AMO),采用慢病毒转染,分别外源性导入Let-7b mimic以增加Let-7b的表达量、AMO互补结合以阻碍Let-7b功能实现,转染不含有大鼠同源序列的空白病毒为对照组.实时荧光定量核酸扩增检测系统(FQ-PCR)检测各转染组Let-7b表达量的改变;Westen blot检测转染后各组GH3细胞中细胞外基质金属蛋白酶诱导剂(CD147)、基质金属蛋白酶-9(MMP-9)蛋白表达量的改变;Transwell细胞小室侵袭实验研究各组细胞侵袭能力的改变.结果 大鼠GH3细胞中,Let-7b与CD147存在直接的靶向关系,并对CD147、MMP-9蛋白表达及细胞的侵袭能力有明确的抑制作用.转染mimic组的GH3细胞Let-7b的表达量是空白对照组的(9.56±0.51)倍,CD147、MMP-9蛋白表达降低,细胞的侵袭性生长受到抑制,差异有统计学意义(P＜0.05);转染AMO组的GH3细胞Let-7b的表达为对照组的(0.31±0.01)倍,CD147、MMP-9蛋白表达量增高,细胞的侵袭能力增强,差异有统计学意义(P＜0.05);对照组与自然生长组Let-7b的表达量、CD147、MMP-9蛋白表达量及细胞侵袭能力差异均无统计学意义(P＞0.05).结论 Let-7b靶向CD147对GH3细胞的侵袭性有抑制作用.
目的 探尋微小RNA Let-7b(Let-7b)在大鼠垂體腺瘤GH3細胞中的靶基因,併觀察其對GH3細胞侵襲性生長的影響.方法 利用TargetScan、PicTar、MiRanda 3種不同的生物信息學方法對靶基因進行篩選;設計Let-7b模擬物(Let-7b mimic)及反義寡覈苷痠鏈序列(AMO),採用慢病毒轉染,分彆外源性導入Let-7b mimic以增加Let-7b的錶達量、AMO互補結閤以阻礙Let-7b功能實現,轉染不含有大鼠同源序列的空白病毒為對照組.實時熒光定量覈痠擴增檢測繫統(FQ-PCR)檢測各轉染組Let-7b錶達量的改變;Westen blot檢測轉染後各組GH3細胞中細胞外基質金屬蛋白酶誘導劑(CD147)、基質金屬蛋白酶-9(MMP-9)蛋白錶達量的改變;Transwell細胞小室侵襲實驗研究各組細胞侵襲能力的改變.結果 大鼠GH3細胞中,Let-7b與CD147存在直接的靶嚮關繫,併對CD147、MMP-9蛋白錶達及細胞的侵襲能力有明確的抑製作用.轉染mimic組的GH3細胞Let-7b的錶達量是空白對照組的(9.56±0.51)倍,CD147、MMP-9蛋白錶達降低,細胞的侵襲性生長受到抑製,差異有統計學意義(P＜0.05);轉染AMO組的GH3細胞Let-7b的錶達為對照組的(0.31±0.01)倍,CD147、MMP-9蛋白錶達量增高,細胞的侵襲能力增彊,差異有統計學意義(P＜0.05);對照組與自然生長組Let-7b的錶達量、CD147、MMP-9蛋白錶達量及細胞侵襲能力差異均無統計學意義(P＞0.05).結論 Let-7b靶嚮CD147對GH3細胞的侵襲性有抑製作用.
목적 탐심미소RNA Let-7b(Let-7b)재대서수체선류GH3세포중적파기인,병관찰기대GH3세포침습성생장적영향.방법 이용TargetScan、PicTar、MiRanda 3충불동적생물신식학방법대파기인진행사선;설계Let-7b모의물(Let-7b mimic)급반의과핵감산련서렬(AMO),채용만병독전염,분별외원성도입Let-7b mimic이증가Let-7b적표체량、AMO호보결합이조애Let-7b공능실현,전염불함유대서동원서렬적공백병독위대조조.실시형광정량핵산확증검측계통(FQ-PCR)검측각전염조Let-7b표체량적개변;Westen blot검측전염후각조GH3세포중세포외기질금속단백매유도제(CD147)、기질금속단백매-9(MMP-9)단백표체량적개변;Transwell세포소실침습실험연구각조세포침습능력적개변.결과 대서GH3세포중,Let-7b여CD147존재직접적파향관계,병대CD147、MMP-9단백표체급세포적침습능력유명학적억제작용.전염mimic조적GH3세포Let-7b적표체량시공백대조조적(9.56±0.51)배,CD147、MMP-9단백표체강저,세포적침습성생장수도억제,차이유통계학의의(P＜0.05);전염AMO조적GH3세포Let-7b적표체위대조조적(0.31±0.01)배,CD147、MMP-9단백표체량증고,세포적침습능력증강,차이유통계학의의(P＜0.05);대조조여자연생장조Let-7b적표체량、CD147、MMP-9단백표체량급세포침습능력차이균무통계학의의(P＞0.05).결론 Let-7b파향CD147대GH3세포적침습성유억제작용.
Objective To investigate target genes of microRNA Let-7b in GH3 cells of rat,and study the influence of Let-7b on invasive growth of GH3 cells.Methods Three different bioinformatic ways including TargetScan,PicTar and MiRanda were used to identify the target genes.The Let-7 b mimic and anti-miRNA oligonuclotide (AMO) were designed and transfected into GH3 cells via lentivirus,and the GH3 cells which transfected with vehicle were used as the control group.Real-time quantitative PCR detecting system (FQ-PCR) was used to detect the expression of Let-7b after different transfection.Westen blotting was used to detect the expression of extracellular metalloproteinase inducer (CD147) and matrix metalloprotein-9 (MMP-9).Transwell experiments were used to detect the invasive ability.Results Let-7b directly targeting CD147 as an inhibitor,suppressed the protein expression of CD147 and MMP-9,and inhibited invasive ability of GH3 cells.After transfection with Let-7b mimic,the mRNA level of Let-7b was 9.56 ± 0.51 times higher than that in the control group (P ＜ 0.05),the protein expression of CD147 and MMP-9 were reduced,the invasive growth of GH3 cells were inhibited.Inhibiting the Let-7b expression by transfection with AMO,the mRNA level of Let-7b was 0.31 ±0.01 times lower than that in the control group (P ＜ 0.05),the protein expression of CD147 and MMP-9 were increased,the ability of invasive growth was enhanced.There were no significant differences in the expression of Let-7b,the protein expression of CD147 and MMP-9,and invasive ability of GH3 cells between the control group and the natural growth group.Conclusion Let-7b targeting CD147 inhibits the invasive growth of GH3 cells,it may be used as drug target for gene treatment and clinical prognostic indices.
