中华肿瘤杂志
中華腫瘤雜誌
중화종류잡지
CHINESE JOURNAL OF ONCOLOGY
2015年
8期
573-577
,共5页
王迪%韩毅%朱莉莉%邓立力%曲迪%崔凤%徐玉清
王迪%韓毅%硃莉莉%鄧立力%麯迪%崔鳳%徐玉清
왕적%한의%주리리%산립력%곡적%최봉%서옥청
肺肿瘤%细胞%贝伐单抗%细胞增殖%基质金属蛋白酶2%基质金属蛋白酶9%肝细胞生长因子受体
肺腫瘤%細胞%貝伐單抗%細胞增殖%基質金屬蛋白酶2%基質金屬蛋白酶9%肝細胞生長因子受體
폐종류%세포%패벌단항%세포증식%기질금속단백매2%기질금속단백매9%간세포생장인자수체
Lung neoplasms%Cells%Bevacizumab%Cell proliferation%Matrix metalloproteinase-2%Matrix metalloproteinase-9%Hepatocyte growth factor receptor
目的:体外观察血管内皮生长因子( VEGF)单克隆抗体贝伐单抗对人肺癌A549细胞增殖和侵袭力的影响,并初步探讨其可能的机制。方法以贝伐单抗处理A549细胞,采用细胞计数试剂盒8( CCK?8)和Transwell法检测A549细胞增殖活性及侵袭能力,实时荧光定量PCR和Western blot检测A549细胞中基质金属蛋白酶2( MMP?2)、基质金属蛋白酶9( MMP?9)和肝细胞生长因子受体( c?Met) mRNA和蛋白的表达水平。结果 CCK?8检测结果显示,贝伐单抗对A549细胞增殖活性的影响有双向作用。10μg/ml贝伐单抗处理A549细胞48 h和72 h的吸光度(A)值分别为0.79±0.10和0.90±0.18,对A549细胞增殖有抑制作用。100μg/ml贝伐单抗处理A549细胞48 h和72 h的A值分别为1.77±0.13和2.07±0.14,对A549细胞增殖有促进作用。侵袭实验显示,50μg/ml贝伐单抗组对A549细胞侵袭力的抑制作用最强,其穿过膜细胞数为(16406.19±5674.23)个,明显低于对照组的(36108.68±6263.83)个,差异有统计学意义(P<0.05)。实时荧光定量PCR检测显示,贝伐单抗处理A549细胞48 h对MMP?2、MMP?9和c?Met mRNA的表达均有双向调节作用。50μg/ml贝伐单抗对A549细胞中MMP?2和MMP?9 mRNA 表达的抑制作用最强,其表达水平分别为0.13±0.08和0.13±0.06。10μg/ml贝伐单抗对A549细胞中c?Met mRNA表达的抑制作用最强,其表达水平为0.18±0.04,明显低于对照组( P<0.01)。100μg/ml贝伐单抗对A549细胞中MMP?2、MMP?9和c?Met mRNA表达有促进作用,其表达水平分别为1.82±0.31、1.60±0.25和2.63±0.48,明显高于对照组( P<0.05)。10μg/ml 贝伐单抗对A549细胞中MMP?2和MMP?9 mRNA的抑制作用呈时间依赖性。 Western blot检测显示,贝伐单抗对A549细胞中MMP?2和c?Met蛋白表达有双向调节作用。100μg/ml贝伐单抗对A549细胞中MMP?2和c?Met蛋白表达有促进作用,其表达水平分别为1.07±0.09和0.98±0.06,明显高于对照组(P<0.05)。50μg/ml贝伐单抗对MMP?2蛋白表达的抑制作用最强,其表达量为0.11±0.04,明显低于对照组( P<0.01)。10μg/ml贝伐单抗对c?Met蛋白表达的抑制作用最强,其表达水平为0.38±0.01,明显低于对照组(P<0.01)。不同浓度贝伐单抗对A549细胞中MMP?9蛋白的表达影响均无统计学意义( P>0.05)。10μg/ml 贝伐单抗对A549细胞中MMP?2、MMP?9和c?Met蛋白表达的抑制作用随时间变化逐渐增强。结论一定浓度的贝伐单抗对A549细胞的增殖及侵袭力有明显的抑制作用,但较高浓度(100μg/ml)的贝伐单抗对A549细胞侵袭的抑制作用减弱,甚至对细胞增殖表现出促进作用,这可能与贝伐单抗对MMP?2和c?Met蛋白的非浓度依赖性抑制有关。
目的:體外觀察血管內皮生長因子( VEGF)單剋隆抗體貝伐單抗對人肺癌A549細胞增殖和侵襲力的影響,併初步探討其可能的機製。方法以貝伐單抗處理A549細胞,採用細胞計數試劑盒8( CCK?8)和Transwell法檢測A549細胞增殖活性及侵襲能力,實時熒光定量PCR和Western blot檢測A549細胞中基質金屬蛋白酶2( MMP?2)、基質金屬蛋白酶9( MMP?9)和肝細胞生長因子受體( c?Met) mRNA和蛋白的錶達水平。結果 CCK?8檢測結果顯示,貝伐單抗對A549細胞增殖活性的影響有雙嚮作用。10μg/ml貝伐單抗處理A549細胞48 h和72 h的吸光度(A)值分彆為0.79±0.10和0.90±0.18,對A549細胞增殖有抑製作用。100μg/ml貝伐單抗處理A549細胞48 h和72 h的A值分彆為1.77±0.13和2.07±0.14,對A549細胞增殖有促進作用。侵襲實驗顯示,50μg/ml貝伐單抗組對A549細胞侵襲力的抑製作用最彊,其穿過膜細胞數為(16406.19±5674.23)箇,明顯低于對照組的(36108.68±6263.83)箇,差異有統計學意義(P<0.05)。實時熒光定量PCR檢測顯示,貝伐單抗處理A549細胞48 h對MMP?2、MMP?9和c?Met mRNA的錶達均有雙嚮調節作用。50μg/ml貝伐單抗對A549細胞中MMP?2和MMP?9 mRNA 錶達的抑製作用最彊,其錶達水平分彆為0.13±0.08和0.13±0.06。10μg/ml貝伐單抗對A549細胞中c?Met mRNA錶達的抑製作用最彊,其錶達水平為0.18±0.04,明顯低于對照組( P<0.01)。100μg/ml貝伐單抗對A549細胞中MMP?2、MMP?9和c?Met mRNA錶達有促進作用,其錶達水平分彆為1.82±0.31、1.60±0.25和2.63±0.48,明顯高于對照組( P<0.05)。10μg/ml 貝伐單抗對A549細胞中MMP?2和MMP?9 mRNA的抑製作用呈時間依賴性。 Western blot檢測顯示,貝伐單抗對A549細胞中MMP?2和c?Met蛋白錶達有雙嚮調節作用。100μg/ml貝伐單抗對A549細胞中MMP?2和c?Met蛋白錶達有促進作用,其錶達水平分彆為1.07±0.09和0.98±0.06,明顯高于對照組(P<0.05)。50μg/ml貝伐單抗對MMP?2蛋白錶達的抑製作用最彊,其錶達量為0.11±0.04,明顯低于對照組( P<0.01)。10μg/ml貝伐單抗對c?Met蛋白錶達的抑製作用最彊,其錶達水平為0.38±0.01,明顯低于對照組(P<0.01)。不同濃度貝伐單抗對A549細胞中MMP?9蛋白的錶達影響均無統計學意義( P>0.05)。10μg/ml 貝伐單抗對A549細胞中MMP?2、MMP?9和c?Met蛋白錶達的抑製作用隨時間變化逐漸增彊。結論一定濃度的貝伐單抗對A549細胞的增殖及侵襲力有明顯的抑製作用,但較高濃度(100μg/ml)的貝伐單抗對A549細胞侵襲的抑製作用減弱,甚至對細胞增殖錶現齣促進作用,這可能與貝伐單抗對MMP?2和c?Met蛋白的非濃度依賴性抑製有關。
목적:체외관찰혈관내피생장인자( VEGF)단극륭항체패벌단항대인폐암A549세포증식화침습력적영향,병초보탐토기가능적궤제。방법이패벌단항처리A549세포,채용세포계수시제합8( CCK?8)화Transwell법검측A549세포증식활성급침습능력,실시형광정량PCR화Western blot검측A549세포중기질금속단백매2( MMP?2)、기질금속단백매9( MMP?9)화간세포생장인자수체( c?Met) mRNA화단백적표체수평。결과 CCK?8검측결과현시,패벌단항대A549세포증식활성적영향유쌍향작용。10μg/ml패벌단항처리A549세포48 h화72 h적흡광도(A)치분별위0.79±0.10화0.90±0.18,대A549세포증식유억제작용。100μg/ml패벌단항처리A549세포48 h화72 h적A치분별위1.77±0.13화2.07±0.14,대A549세포증식유촉진작용。침습실험현시,50μg/ml패벌단항조대A549세포침습력적억제작용최강,기천과막세포수위(16406.19±5674.23)개,명현저우대조조적(36108.68±6263.83)개,차이유통계학의의(P<0.05)。실시형광정량PCR검측현시,패벌단항처리A549세포48 h대MMP?2、MMP?9화c?Met mRNA적표체균유쌍향조절작용。50μg/ml패벌단항대A549세포중MMP?2화MMP?9 mRNA 표체적억제작용최강,기표체수평분별위0.13±0.08화0.13±0.06。10μg/ml패벌단항대A549세포중c?Met mRNA표체적억제작용최강,기표체수평위0.18±0.04,명현저우대조조( P<0.01)。100μg/ml패벌단항대A549세포중MMP?2、MMP?9화c?Met mRNA표체유촉진작용,기표체수평분별위1.82±0.31、1.60±0.25화2.63±0.48,명현고우대조조( P<0.05)。10μg/ml 패벌단항대A549세포중MMP?2화MMP?9 mRNA적억제작용정시간의뢰성。 Western blot검측현시,패벌단항대A549세포중MMP?2화c?Met단백표체유쌍향조절작용。100μg/ml패벌단항대A549세포중MMP?2화c?Met단백표체유촉진작용,기표체수평분별위1.07±0.09화0.98±0.06,명현고우대조조(P<0.05)。50μg/ml패벌단항대MMP?2단백표체적억제작용최강,기표체량위0.11±0.04,명현저우대조조( P<0.01)。10μg/ml패벌단항대c?Met단백표체적억제작용최강,기표체수평위0.38±0.01,명현저우대조조(P<0.01)。불동농도패벌단항대A549세포중MMP?9단백적표체영향균무통계학의의( P>0.05)。10μg/ml 패벌단항대A549세포중MMP?2、MMP?9화c?Met단백표체적억제작용수시간변화축점증강。결론일정농도적패벌단항대A549세포적증식급침습력유명현적억제작용,단교고농도(100μg/ml)적패벌단항대A549세포침습적억제작용감약,심지대세포증식표현출촉진작용,저가능여패벌단항대MMP?2화c?Met단백적비농도의뢰성억제유관。
Objective To study the effect and mechanism of bevacizumab on proliferation and invasion of human lung cancer A549 cells. Methods A549 cells were treated with bevacizumab. Proliferation and invasion of the bevacizumab?treated A549 cells were detected using cell counting kit CCK?8 and Transwell assay, respectively. The expression of the mRNA and protein of MMP?2, MMP?9 and c?Met Results Proliferation activity was inhibited at the concentration of 10 μg/ml and promoted at the concentration of 100 μg/ml bevacizumab. Bevacizumab in the concentration of 50 μg/ml had a stronger inhibitory effect on the invasion of A549 cells (16 406.19±5 674.23 penetrated cells ) than that of control group (36 108.68±6 263.83, P<0.05). The real?time PCR showed that bevacizumab had a stronger inhibitory effect on the expression of MMP?2 and MMP?9 mRNA at the concentration of 50 μg/ml and on the expression c?Met mRNA at the concentration of 10 μg/ml bevacizumabin the A549 cells. However bevacizumab at the concentration of 100μg/ml showed a promoting effect on the expression of MMP?2, MMP?9 and c?Met mRNA ( 1. 82 ± 0. 31, 1. 60 ± 0. 25, 2. 63 ± 0.48), significantly higher than that of the control group (1.00±0.19, 1.00±0.23, 1.00±0.22, P<0.05). The expression of MMP?2, MMP?9 and c?Met mRNA and protein was inhibited by 10μg/ml bevacizumab in a time?dependent manner. The Western blot assay showed that bevacizumab had a bi?directional effect on the expression of MMP?2 and c?Met proteins in the A549 cells:a promoting effect at 100 μg/ml and inhibitory effect on the expression of MMP?2 at 50μg/ml bevacizumab, and inhibitory effect on the expression of c?Met protein at 10 μg/ml bevacizumab. Conclusions Our findings indicate that in a certain range of concentrations, bevacizumab has prominent inhibitory effect on the proliferation and invasion of A549 cells. However,over the concentration of 100 μg/ml, bevacizumab shows a weakening anti?invasion effect, even has a promoting effect on cell proliferation. This phenomenon may be related to the inhibiting effect on the expression of MMP?2 and c?Met proteins in a non?concentration?dependent manner by bevacizumab.