CAJ | 학술논문

目的探讨重组腺病毒介导的Ⅰ型人类免疫缺陷病毒蛋白R(Vpr)转染对胶质瘤U251细胞TGFβRⅢ表达及恶性生物学行为的影响.方法将U251细胞分为正常对照组,空载体组和实验组进行细胞培养,空载体组和实验组按感染复数(multiplicity of infection,MOI)=100分别进行空载腺病毒(Ad-GFP)和含有Vpr基因的重组腺病毒(Ad-Vpr)转染,转染后用Western blot检测Vpr、TGFβRⅢ、MMP-2和MMP-9表达水平,Transwell、划痕实验检测细胞迁移和侵袭能力.结果实验组U251细胞经Ad-Vpr转染后,可见Vpr蛋白表达,同时TGFβRⅢ蛋白表达升高,正常对照组、Ad-GFP组、Ad-Vpr组TGFβRⅢ与β-actin密度灰度比值分别为0.71±0.04,0.83±0.09和1.13±0.16,三组间差异有统计学意义(P＜0.05).MMP-2和MMP-9蛋白表达降低,Ad-Vpr组MMP-2和MMP-9与β-actin密度灰度比值分别为0.51±0.11和0.43±0.08,对照组为1.11±0.11和1.06±0.18,Ad-GFP组为1.23±0.05和1.16±0.12,与对照组和Ad-GFP组相比,Ad-Vpr组MMP-2和MMP-9表达明显降低(P＜0.05).Transwell体外侵袭实验结果显示,对照组、Ad-GFP组和Ad-Vpr组平均视野的穿膜细胞数分别为(123.12±10.82)个、(118.89±12.65)个和(80.64±7.72)个,与对照组和Ad-GFP组比较,Ad-Vpr转染组穿膜细胞数明显减少,差异具有统计学意义(P＜0.05).结论 Ad-Vpr可能通过上调TGFβRⅢ降低胶质瘤U251细胞的迁移和侵袭能力.
목적 탐토중조선병독개도적Ⅰ형인류면역결함병독단백R(Vpr)전염대효질류U251세포TGFβRⅢ표체급악성생물학행위적영향.방법 장U251세포분위정상대조조,공재체조화실험조진행세포배양,공재체조화실험조안감염복수(multiplicity of infection,MOI)=100분별진행공재선병독(Ad-GFP)화함유Vpr기인적중조선병독(Ad-Vpr)전염,전염후용Western blot검측Vpr、TGFβRⅢ、MMP-2화MMP-9표체수평,Transwell、화흔실험검측세포천이화침습능력.결과 실험조U251세포경Ad-Vpr전염후,가견Vpr단백표체,동시TGFβRⅢ단백표체승고,정상대조조、Ad-GFP조、Ad-Vpr조TGFβRⅢ여β-actin밀도회도비치분별위0.71±0.04,0.83±0.09화1.13±0.16,삼조간차이유통계학의의(P＜0.05).MMP-2화MMP-9단백표체강저,Ad-Vpr조MMP-2화MMP-9여β-actin밀도회도비치분별위0.51±0.11화0.43±0.08,대조조위1.11±0.11화1.06±0.18,Ad-GFP조위1.23±0.05화1.16±0.12,여대조조화Ad-GFP조상비,Ad-Vpr조MMP-2화MMP-9표체명현강저(P＜0.05).Transwell체외침습실험결과현시,대조조、Ad-GFP조화Ad-Vpr조평균시야적천막세포수분별위(123.12±10.82)개、(118.89±12.65)개화(80.64±7.72)개,여대조조화Ad-GFP조비교,Ad-Vpr전염조천막세포수명현감소,차이구유통계학의의(P＜0.05).결론 Ad-Vpr가능통과상조TGFβRⅢ강저효질류U251세포적천이화침습능력.
Objective To study the effect of recombinant adenovirus mediated transfection of human immunodeficiency virus type 1 viral protein R (Vpr) on TGFβR Ⅲ expression and malignant behavior of glioma cell U251.Methods U251 cells were divided into the control group,mock group and experiment group,the mock and experiment group were transfected with Ad-GFP and Ad-Vpr,respectively at a multiplicity of infection (MOI) of 100.The expression of Vpr,TGFβR Ⅲ,MMP-2 and MMP-9 were detected by western blot,the malignant biological behavior changes of U251 cells transfected by Ad-Vpr were evaluated by scratch assay and transwell assay.Results Western blot showed that the Vpr protein could be expressed after Ad-Vpr transfection,meanwhile,the expression of TGFβR Ⅲ was up regulated by Ad-Vpr transfection,Relative density values of TGFβR Ⅲ compared to β-actin in control,Ad-GFP and Ad-Vpr group were 0.71 ±0.04,0.83 ±0.09 and 1.13 ±0.16,respectively,there were significant different in three groups(P ＜ 0.05).While the expression of MMP-2 and MMP-2 were down regulated by Ad-Vpr transfection.Relative density values of MMP-2 and MMP-9 compared to β-actin were 0.51 ± 0.11 and 0.43 ±0.08 for Ad-Vpr group,1.11 ±0.11 and 1.06 ±0.18 for control group and 1.23 ±0.05 and 1.16 ± 0.12 for Ad-GFP group,respectively.The expression of MMP-2 and MMP-9 were significantly decreased comparing to control and Ad-GFP group (P ＜ 0.05).Transwell assay results showed that the average cells across the membrane were 123.12 ± 10.82,118.89 ± 12.65 and 80.64 ± 7.72 in control,Ad-GFP and Ad-Vpr group,respectively.Compared with control and the Ad-GFP group,the cells across the membrane in Ad-Vpr group were significantly reduced (P ＜ 0.05).Conclusions Vpr could decrease migratory and invasive behavior of glioma U251 cells probably by upregulating TGFβRⅢ expression.