CAJ | 학술논문

目的以单纯疱疹病毒1-胸苷激酶(HSV1-tk)为报告基因,以131I标记的氟-碘阿糖基脲嘧啶(FIAU)为报告探针,对心肌内转染HSV1-tk的家兔行心脏报告基因单光子发射计算机断层(SPECT)显像,以探讨该报告系统应用于SPECT心脏报告基因显像的可行性,为SPECT报告基因心脏显像最佳转染滴度、显像时间选择提供依据.方法制作腺病毒(Ad)为载体携带HSV1-tk基因的重组体Ad-tk,家兔心肌内注射Ad-tk.实验分2部分:(1)转染报告基因后不同时间埘显像的影响:家兔心肌注射不同滴度的Ad-tk(1×109、5×108、1×108、5×107、1×107 pfu),2 d后耳缘静脉注射131IFIAU 600 μCi行6、24、48和72 h心肌SPECT显像,注射1×109 pfu Ad-tk的家兔继续行96和120 h显像.(2)不同Ad-tk滴度对显像的影响:家兔心肌注射不同滴度的Ad-tk(1×109、5×108、1×108、5×107、1×107pfu),2 d后耳缘静脉注射131I-FIAU 600 μCi行6、24和48 h心肌SPECT显像,显像结束后处死家兔,取出心脏,测量γ计数值,行逆转录-聚合酶链反应(RT-PCR)研究,并对SPECT图像和RT-PCR图像行半定量分析.结果 SPECT图像可见Ad-tk在注射局部表达,而非注射部位未见表达,对照组心肌除6 h可隐约显影外,其余时间均未见显影.RT-PCR证实HSV1-tk在注射部位表达.不同的Ad-tk滴度,SPECT靶/非靶比值与γ计数值旱正相关(r2=0.933,P＜0.01),与HSV1-tk的表达亦呈正相关(r2=0.877,P＜0.01).SPECT可检测到滴度低至1×107pfu的Ad-tk在心肌内的表达.结论应用HSV1-tk为报告基因,FIAU为报告探针的SPECT心肌报告基因显像是可行的.SPECT心肌报告基因显像可半定量地监测报告基因在体内的表达.
목적 이단순포진병독1-흉감격매(HSV1-tk)위보고기인,이131I표기적불-전아당기뇨밀정(FIAU)위보고탐침,대심기내전염HSV1-tk적가토행심장보고기인단광자발사계산궤단층(SPECT)현상,이탐토해보고계통응용우SPECT심장보고기인현상적가행성,위SPECT보고기인심장현상최가전염적도、현상시간선택제공의거.방법 제작선병독(Ad)위재체휴대HSV1-tk기인적중조체Ad-tk,가토심기내주사Ad-tk.실험분2부분:(1)전염보고기인후불동시간시현상적영향:가토심기주사불동적도적Ad-tk(1×109、5×108、1×108、5×107、1×107 pfu),2 d후이연정맥주사131IFIAU 600 μCi행6、24、48화72 h심기SPECT현상,주사1×109 pfu Ad-tk적가토계속행96화120 h현상.(2)불동Ad-tk적도대현상적영향:가토심기주사불동적도적Ad-tk(1×109、5×108、1×108、5×107、1×107pfu),2 d후이연정맥주사131I-FIAU 600 μCi행6、24화48 h심기SPECT현상,현상결속후처사가토,취출심장,측량γ계수치,행역전록-취합매련반응(RT-PCR)연구,병대SPECT도상화RT-PCR도상행반정량분석.결과 SPECT도상가견Ad-tk재주사국부표체,이비주사부위미견표체,대조조심기제6 h가은약현영외,기여시간균미견현영.RT-PCR증실HSV1-tk재주사부위표체.불동적Ad-tk적도,SPECT파/비파비치여γ계수치한정상관(r2=0.933,P＜0.01),여HSV1-tk적표체역정정상관(r2=0.877,P＜0.01).SPECT가검측도적도저지1×107pfu적Ad-tk재심기내적표체.결론 응용HSV1-tk위보고기인,FIAU위보고탐침적SPECT심기보고기인현상시가행적.SPECT심기보고기인현상가반정량지감측보고기인재체내적표체.
Objective To explore the feasibility of single photon emission computed tomography (SPECT)detection of heart reporter gene expression and observed the optimal thansfecting titer and imaging time by using herpes simplex virus 1-thymidine kinase(HSV1-tk)as reporter gene and 131I-2'-fluoro-2'-deoxy-1-β-D-arabinofuranosyl-5-iodouracil(131I-FIAU)as reporter probe in rabbit myocardium.Methods The recombinant Ad.tk carrying HSV1-tk gene and adenovirus(Ad)as vector was constructed and intramyocardially injected to rabbits at various concentrations(1×109 pfu,5×108 pfu,1×108 pfu,5×107 pfu,1×107 pfu).Two days later,rabbits were injected with 600 μCi131I-FIAU in ear-nlargin vein and then underwent SPECT myocardium imaging for detection of HSV1-tk expression at 6 h,24 h.48 h and 72 h after injection.rabbits with 1×109 pfu Ad-tk injeetion were imaged at 96 h and 120 h.Rabbits were sacrificed after imaging and the total myocardial 131I-FIAU accumulation was quantified in percent of injeeted dose per gram myocardium(%ID/g).The myocardial Ad-tk expression was determined with RT-PCR.Results Reporter gene was detected by SPECT imaging in the iniection site while not detected in the control myecardium and site remote from injection.RT-PCR results also evidenced HSVl.tk express in the injection site.The SPECT target/nontarget ratio was correlated with ex vivo γ-counting (r2=0.933,P＜0.01) and expression of HSV1-tk (r2 =0.877,P＜0.01).Myocardial accumulation could be identified at viral titers as low as 1×107 pfu by SPECT imaging.Conclusion The cardiac SPECT reporter gene imaging with HSV1-tk as reporter gene and 131 I-FIAU as reporter probe is feasible.