CAJ | 학술논문

目的探讨Toll样受体4(TLR4)/NF-κB信号转导通路在大鼠骨癌痛形成及维持中的作用.方法健康雌性SD大鼠81只,5～6周龄,体重150～ 180 g,采用随机数字表法,将其分为8组,骨癌痛组(BCP组)18只,其他7组每组9只,对照组(C组)不作任何处理；骨癌痛组大鼠胫骨注射l×105 Walker 256乳腺癌细胞；Ⅴ1组、TLR4siRNA1组和TLR4msiRNA1组分别于造模后4d(预注射)鞘内注射溶媒、TLR4小干扰RNA (TLR4siRNA)和TLR4错义siRNA(TLR4msiRNA),Ⅴ2组、TLR4siRNA2组和TL-R4msiRNA2组分别于造模后9d(后注射)鞘内注射溶媒、TLR4siRNA和TLR4msiRNA.TLR4siRNA和TLR4msiRNA 2μg用5％葡萄糖和jetPEITM稀释到5μl,1次/d,连续3d,最后1次注射后24 h处死大鼠,BCP组分别于造模后7、12d处死大鼠,取L4-6脊髓,采用RT-PCR法检测NF-κBp65 mRNA、IL-6 mRNA和TNF-α mRNA表达,采用免疫组化法和Western blot法检测NF-κBp65表达.结果预注射TLR4siRNA可完全抑制骨癌痛大鼠脊髓组织NF-κBp65 mRNA、TNF-α mRNA和NF-κBp65表达上调,而后注射TLR-4siRNA可部分抑制骨癌痛大鼠脊髓组织NF-κBp65 mRNA、IL-6 mRNA、TNF-α mRNA和NF-κB表达上调(P＜ 0.05或0.01).结论 TLR4/NF-κB信号转导通路介导了大鼠骨癌痛的形成和维持.
목적 탐토Toll양수체4(TLR4)/NF-κB신호전도통로재대서골암통형성급유지중적작용.방법 건강자성SD대서81지,5～6주령,체중150～ 180 g,채용수궤수자표법,장기분위8조,골암통조(BCP조)18지,기타7조매조9지,대조조(C조)불작임하처리；골암통조대서경골주사l×105 Walker 256유선암세포；Ⅴ1조、TLR4siRNA1조화TLR4msiRNA1조분별우조모후4d(예주사)초내주사용매、TLR4소간우RNA (TLR4siRNA)화TLR4착의siRNA(TLR4msiRNA),Ⅴ2조、TLR4siRNA2조화TL-R4msiRNA2조분별우조모후9d(후주사)초내주사용매、TLR4siRNA화TLR4msiRNA.TLR4siRNA화TLR4msiRNA 2μg용5％포도당화jetPEITM희석도5μl,1차/d,련속3d,최후1차주사후24 h처사대서,BCP조분별우조모후7、12d처사대서,취L4-6척수,채용RT-PCR법검측NF-κBp65 mRNA、IL-6 mRNA화TNF-α mRNA표체,채용면역조화법화Western blot법검측NF-κBp65표체.결과 예주사TLR4siRNA가완전억제골암통대서척수조직NF-κBp65 mRNA、TNF-α mRNA화NF-κBp65표체상조,이후주사TLR-4siRNA가부분억제골암통대서척수조직NF-κBp65 mRNA、IL-6 mRNA、TNF-α mRNA화NF-κB표체상조(P＜ 0.05혹0.01).결론 TLR4/NF-κB신호전도통로개도료대서골암통적형성화유지.
Objective To evaluate the role of Toll-like receptor 4 (TLR4)/NF-κB signal transduction pathway in development and maintenance of bone cancer pain (BCP) in rats.Methods Eighty-one female Sprague-Dawley rats,aged 5-6 weeks,weighing 150-180 g,were randomly divided into 8 groups:control group (group C),BCP group,vehicle 1 group (Ⅴ1 group),TLR4 small interfering RNA (siRNA) 1 group (TLR4siRNA1 group),TLR4 mismatch siRNA (msiRNA) 1 group (TLR4msiRNA1 group),Ⅴ2 group,TLR4siRNA2 group and TLR4msiRNA2 group.There were 18 rats in group BCP and 9 rats in each group of the left 7 groups.BCP was induced by intra-tibial inoculation of 1 × 105 Walker 256 breast cancer cells.In Ⅴ1,TLR4siRNA1 and TL-R4msiRNA1 groups,vehicle,TLR4siRNA and TLR4msiRNA were intrathecally injected,respectively,on 4th day after inoculation (pre-injection).In Ⅴ2,TLR4siRNA2 and TLR4msiRNA2 groups,vehicle,TLR4siRNA and TLR4msiRNA were intrathecally injected,respectively,on 9th day after inoculation (post-injection).TLR4siRNA and TLR4msiRNA 2 μg were diluted with 5％ glucose and jetPEITM to 5 μl,and then given once a day for 3 consecutive days,and the rats were sacrificed at 24 h after the last injection.The rats in group BCP were sacrificed ondays 7 and 12 after inoculation.The lumbar segments (L4-6) of the spinal cord were removed for determination of the expression of NF-κBp65 mRNA,IL-6 mRNA and TNF-α mRNA (by real-time PCR) and NF-κBp65 (by Western blot and immuno-histochemistry).Results Pre-injection of TLR4siRNA could completely inhibit up-regulation of the expression of NF-κBp65 mRNA,NF-κBp65 and TNF-α mRNA in the spinal cord tissues,however,post-injection of TLR4siRNA could partially inhibit up-regulation of the expression of NF-κBp65 mRNA,NF-κBp65,IL-6 mRNA and TNF-α mRNA in the spinal cord tissues (P ＜ 0.05 or 0.01).Conclusion TLR4/NF-κB signaling pathway mediates the development and maintenance of BCP in rats.