CAJ | 학술논문

目的探讨反义寡核昔酸多位点封闭HBx基因对人类肝癌组织在裸小鼠皮下移植瘤发生的影响.方法 50只裸小鼠随机等分为5组,1组为对照组,其余4组为实验组(AS1～AS4组),取对数期生长的内源性表达HBx蛋白的人类肝癌细胞系(Hep3B细胞)皮下注射,从第2天开始实验组分别通过腹膜腔内注射反义寡核苷酸AS1～AS4片段(剂量为10 μg/g),隔日注射,共注射5次,对照组注射无菌双蒸馏水,观察裸小鼠皮下移植瘤生长情况,记录移植瘤潜伏期(从皮下注射肿瘤细胞至首次观察到皮下瘤结节),满30 d实验终止,比较实验组和对照组移植瘤发生率,以生存分析模型分析移植瘤潜伏期在各组间的差异.结果实验组(AS1～AS3组)及对照组移植瘤发生率均为100%,AS4组10只小鼠中有1只小鼠实验结束时仍未观察到移植瘤的形成,各组小鼠移植瘤发生率差异无统计学意义.AS1～AS4组及对照组移植瘤潜伏期中位时间分别为19、12、11、21、10 d,AS1,AS4组与对照组之间,差异有统计学意义(χ2=40.94,P<0.01).结论反义寡核苷酸在恰当位点封闭HBx基因可以有效延长人类肝癌组织(Hep3B细胞系)在裸小鼠皮下移植瘤形成的潜伏期;但有限次数的干扰不能完全阻断移植瘤的发生;腹膜腔注射是反义寡核苷酸进入活体裸小鼠并发挥干扰作用的一条有效途径.
목적 탐토반의과핵석산다위점봉폐HBx기인대인류간암조직재라소서피하이식류발생적영향.방법 50지라소서수궤등분위5조,1조위대조조,기여4조위실험조(AS1～AS4조),취대수기생장적내원성표체HBx단백적인류간암세포계(Hep3B세포)피하주사,종제2천개시실험조분별통과복막강내주사반의과핵감산AS1～AS4편단(제량위10 μg/g),격일주사,공주사5차,대조조주사무균쌍증류수,관찰라소서피하이식류생장정황,기록이식류잠복기(종피하주사종류세포지수차관찰도피하류결절),만30 d실험종지,비교실험조화대조조이식류발생솔,이생존분석모형분석이식류잠복기재각조간적차이.결과 실험조(AS1～AS3조)급대조조이식류발생솔균위100%,AS4조10지소서중유1지소서실험결속시잉미관찰도이식류적형성,각조소서이식류발생솔차이무통계학의의.AS1～AS4조급대조조이식류잠복기중위시간분별위19、12、11、21、10 d,AS1,AS4조여대조조지간,차이유통계학의의(χ2=40.94,P<0.01).결론 반의과핵감산재흡당위점봉폐HBx기인가이유효연장인류간암조직(Hep3B세포계)재라소서피하이식류형성적잠복기;단유한차수적간우불능완전조단이식류적발생;복막강주사시반의과핵감산진입활체라소서병발휘간우작용적일조유효도경.
Objective To study the inhibitory effect of HBx antisense oligodeoxynuclentide on the formation of transplanted hepatocellular carcinoma in nude mice.Method 50 nude mice were randomly divided into 5 groups:1 control group and 4 experimental groups.Log-phase Hep3B cells endogenouslyexpressing HBX were injected subcutaneously in nude mice.From the second day,the PAGE purified AS1,AS2,AS3 and AS4 HBx antisense oligodeoxynuclentides were injected intraperitoneally into the 4 experi-mental groups,respectively,on alternate days for 5 times,and distilled water was injected into the control group.Growth information of subcutaneous transplantation tumor in nude mice was recorded for 30 days.Incidence rate of transplanted tumor in different groups was compared and analyzed by survival analysis.Statistics software SPSS 12.0 was used to analyze the data.Results Incidence rate of transplanted tumor was 100% in AS1,AS2,AS3 and control groups,and 90% in AS4 group(χ2= 3.995,P = 1.0).The median latency period for transplanted tumor formation was 19 days(17.48-20.52),12 days(9.93-14.07),11 days(9.45-12.55),21 days(19.48-22.52),and 10 days(8.99-11.01)in AS1,AS2,AS3,AS4 and control group,respectively.The latency period for tumor formation was prolonged by treatment of mice with AS1 and AS4 antisense oligodeoxynucleotide(P < 0.01).Conclusion Antisense oligodeoxynucleotide targeting to the appropriate sites of HBx gene can prolong the latency period of subeutaneously transplanted tumor in nude mice,however,the formation of transplanted tumor can not be completely blocked by limited treatment with these antisense oligos.In addition,our results suggest that peritoneal injection may be an effective way to deliver antisense oligodeoxynucleotide to living organisms.