CAJ | 학술논문

目的：利用圈套dsODNs／载体基因技术，通过体外实验研究NF-кB decoy dsODNs／载体复合物在急性肺损伤肺泡巨噬细胞中的体外转染效率，为进一步的体内试验提供依据，从而在细胞分子水平为临床防治全血性炎症反应综合征／急性肺损伤（ SIRS／ALI）筛选精确药物靶点和寻求高效合理的治疗方案提供前瞻性的实验依据。方法制备失血性休克致ALI的实验动物模型，以Lipofectamine2000为载体，对巨噬细胞分组进行dsODNs干预：对照组、脂多糖（ lipopolysaccharide ，LPS）组、Lipofectamine2000组（ Lipo2000组）、NF-κB dsODNs／Lipofectamine2000组（ dsODNs组）。乳酸脱氢酶（ LDH）试剂盒检测巨噬细胞（ AM）毒性；观察并测定巨噬细胞的转染活性、最佳转染浓度和转染效率。结果 dsODNs／Lipofectamine2000浓度比率为1∶5，体外转染6h时，转染效率、荧光强度和细胞状态最佳，细胞毒性适中。结论 NF-кB圈套dsODNs与Lipofectamine2000在一定浓度范围内能有效提高圈套dsODNs的转染效率，并且最大限度降低各种刺激对细胞造成的损伤，为SIRS／ALI的临床防治提供了新的思路和途径，为临床治疗ALI筛选特异性的炎症治疗药物和开发临床药物确定了精确靶点。
목적：이용권투dsODNs／재체기인기술，통과체외실험연구NF-кB decoy dsODNs／재체복합물재급성폐손상폐포거서세포중적체외전염효솔，위진일보적체내시험제공의거，종이재세포분자수평위림상방치전혈성염증반응종합정／급성폐손상（ SIRS／ALI）사선정학약물파점화심구고효합리적치료방안제공전첨성적실험의거。방법제비실혈성휴극치ALI적실험동물모형，이Lipofectamine2000위재체，대거서세포분조진행dsODNs간예：대조조、지다당（ lipopolysaccharide ，LPS）조、Lipofectamine2000조（ Lipo2000조）、NF-κB dsODNs／Lipofectamine2000조（ dsODNs조）。유산탈경매（ LDH）시제합검측거서세포（ AM）독성；관찰병측정거서세포적전염활성、최가전염농도화전염효솔。결과 dsODNs／Lipofectamine2000농도비솔위1∶5，체외전염6h시，전염효솔、형광강도화세포상태최가，세포독성괄중。결론 NF-кB권투dsODNs여Lipofectamine2000재일정농도범위내능유효제고권투dsODNs적전염효솔，병차최대한도강저각충자격대세포조성적손상，위SIRS／ALI적림상방치제공료신적사로화도경，위림상치료ALI사선특이성적염증치료약물화개발림상약물학정료정학파점。
Objective To study the transfection efficiency by NF -кB decoy dsODNs/carrier in the alveo-lar macrophages in vito during acute lung injury by NF -кB decoy dsODNs/liposome technology .Intended results will provide a preliminary experimental basis in vitro .It could provide the feasibe experiment basis for screening drug targets and seek dfficient and reasonable treatment to prevent and cure SIRS /ALI.Methods Animal model of hemorrhagic shock caused acute lung injury was established .Different stimulation and complex processing were giv-en to alveolar macrophages in vitro using Lipofectamine 2000 as carrier.The transfection efficiency of each phase and the best time to determine dfficient transfection was measured .The LDH level trasformation by different pre-treatment were detected in different time points .Results When NF-кB decoy dsODNs/Lipofectamine2000 rate was 1∶5 and transfected time was six hours ,the transfection efficiency was highest ,and the cytotoxicity was moder-ate.Conclusion NF-кB decoy dsODNs could significantly improve the transfection efficiency within a proper dsODNs/Lipofectamine2000 rate and reduce injury of alveolar macrophages by different stimulations as much as pos -sible.Decoy dsODNs strategy provides new ideas and approaches for preventing and treating for SIRS /ALI, deter-mines the precise target and screening specific inflammatory therapy for the clinical treatment , and determines the precise target of the clinical drug development .