腺病毒介导hTGF-β1基因体内转染兔腰椎间盘髓核细胞对蛋白多糖含量的影响

Adenovirus-mediated transfer of hTGF-β1 encoding gene to the nucleus pulposus cells of lumbar disc of the rabbit: the effect on proteoglycan level

目的观察构建腺病毒载体介导外源性人转化生长因子(humantransforminggrowthfac-tor,hTGF-β1)基因(Ad/CMV-hTGF-β1)转染到兔椎间盘髓核细胞后,髓核组织中蛋白多糖含量的变化。方法(1)纯种成年新西兰大白兔35只,其中25只对腰椎间盘髓核组织注射Ad/CMV-hTGF-β1,每只注射2个椎间盘作为实验组,注射量为每个腰椎间盘20μl(6×106pfu);同时每只取2个未做注射的腰椎间盘作为自身空白对照组。其余10只兔每只注射2个腰椎间盘各20μl磷酸盐缓冲液(PBS)作为实验对照组。全组共120个椎间盘。(2)手术后1～12周的不同时间段分别取出各组椎间盘组织,采用间苯三酚分光光度法测定髓核组织中蛋白多糖的含量,所得数据经SPSS10.0统计学软件进行统计学处理。结果(1)术后1周,实验组与自身空白对照组蛋白多糖测定值经配对t检验,t=3.968,P<0.05。(2)术后2周,三组结果经方差分析显示,F=17.871,P<0.01;各组间再行两两q检验,实验组与自身空白对照组间q=7.686,P<0.01;实验组与实验对照组间q=6.894,P<0.01;实验对照组与自身空白对照组间q=0.792, P>0.05。实验组与自身空白对照组间配对t检验,t=5.276,P<0.01。(3)术后4周,实验组与自身空白对照组经配对t检验,t=8.352,P<0.01。(4)术后8周,实验组与自身空白对照组经配对t检验,t=7.086,

Objective To investigate the effect on the proteoglycan level after delivering an exogenous therapeutic gene-hTGF-β1 mediated by adenovirus vector to rabbit lumbar disc in vivo. Methods 0.01 mol/L PBS 20 μl with or without TGF-β1 gene Ad/CMV-hTGF-β1 carrying adenovirus 6×106 pfu, was injected directly into nucleus pulposus of lumbar discs of 35 mature New Zealand white rabbits. The TGF-β1 gene Ad/CMV-hTGF-β1 was constructed in authors' lab. All of the examined discs (120 discs) were divided into three groups, including experimental group, self-control group and control group. The proteoglycan of lumbar discs was quantitated in different periods (1st week, 2nd week, 4th week, 8th week and 12th week) by the phloroglucin spectrophotometry. The experimental data were analyzed by SPSS 10.0 soft ware. Results 1) There was significant difference of the proteoglycan quantity between the experimental groups and self-control groups from 1st week to 12th week (1st week: t=3.968, P< 0.05; 2nd week: t=5.276, P< 0.01; 4th week: t=8.352, P< 0.01; 8th week: t=7.086, P< 0.01 and 12th week: t=4.875, P< 0.01). 2) The difference of the proteoglycan quantity analysed by a one-way ANOVA among the experimental groups, self-control groups and control groups was significant at 2nd week and 8th week (2nd week: F=17.871, P< 0.01; 12th week: F=10.67, P< 0.05). The q-test results in different periods (2nd week and 12th week) were displayed as the following: the difference between experimental groups and self-control groups was significant (2nd week: q=7.686, P< 0.01; 12th week: q=5.710, P< 0.01); the difference between experimental groups and control groups was also significant (2nd week: q=6.894, P< 0.01; 12th week: q=6.304, P< 0.01); there was no significant difference between self-control groups and control groups (2nd week: q=0.792, P > 0.05; 12th week: q=0.594, P > 0.05). Conclusion The proteoglycan level in rabbit nucleus pulposus can be increased by trensformatiom of Ad/CMV- hTGF-β1 of disc in a certain period.