摘要
目的观察腺病毒穿梭载体介导RNA干扰对激素诱导骨髓间充质干细胞（BMSCs）成脂分化的影响。方法选取新西兰兔制备BMSCs并随机分为6组，RNA干扰1组、2组和3组（s1、s2和s3）；无关序列组（Con）；模型组（M）；正常对照组（N）。加入激素终质量浓度为1×10-7mol／L地塞米松。将重组腺病毒穿梭载体转染细胞，采用TaqMan逆转录一聚合酶链反应（RT．PCR）和Westernblot方法分别检测各组BMSCs内过氧化物酶体增殖子活化受体γ（PPARγ）mRNA及其蛋白表达，测定细胞甘油三酯含量，用苏丹Ⅲ染色细胞并计数脂肪细胞。结果处理细胞7d时，s1、s2、s3、Con、M、N组中，PPARs,mRNA相对表达值分别为0．406±0．012、0．401±0．009、0．410±0．042、0．621±0．045、0．628±0．008、0．399±0．014，PPAR＇yWesternblot灰度扫描值分别为0．246±0．027、0．235±0．015、0．257±0．025、0．800±0．017、0．793±0．019、0．244±0．010。S1、S2、S3、N组中PPARγ mRNA及其蛋白表达均明显低于M、Con组，差异均有统计学意义（P〈0．05），S1、s2、s3组和N组之间差异无统计学意义（P〉0．05）。s1、s2、s3、N组中脂肪细胞计数、甘油三酯含量均明显低于M、Con组，差异均有统计学意义（P〈0．05），s1、s2、S3组和N组之间差异无统计学意义（P〉0．05）。结论靶向PPARγ基因的腺病毒穿梭载体介导RNA干扰能够有效地抑制激素诱导的BMSCs成脂分化。
Objective To investigate the effect of adenovirns shuttle vector-mediated RNA inter- ference on steroid-induced adipogenie differentiation in the bone marrow mesenchymal stem cells （BMSCs） of rabbits. Methods BMSCs were obtained from bone marrow of New-Zealand rabbit, and divided into 6 groups at random ： RNA interference groups 1, 2 and 3 （ S1,S2 and S3 ）, irrelative sequence group （Con）, model group （M） and normal control group （N）. The BMSCs were treated with 1 x 10-7 mol/L dexamethasone and infected with small interfering RNA （siRNA） adenovirus shuttle vector. The expression of peroxisome proliferator activated receptor ~/ （PPAR-y） mRNA was determined by TaqMan RT-PCR （TaqMan probe fluorescence detection）, and expression of the PPAR~/protein was determined by Western blotting. The content of intracellular triglyceride （TG） was determined： The cells were stained with Sudan m, and adipocytes were counted under a light microscope. Results At 7th day after treatment, the ex- pression levels of PPARγmRNA in the BMSCs in S1, S2, S3, Con, M and N groups were 0. 406 ± 0. 012, 0.401±0.009, 0.410 ±0.042, 0.621 ±0.045, 0.628 ±0.008 and 0.399 ±0.014 respectively, and those of PPARγprotein in blot grayscale scanning value were 0. 246 ± 0. 027, 0. 235 ± 0. 015, 0. 257 ± 0. 025, 0. 800 ± 0. 017, 0. 793 ± 0. 019 and 0. 244 ± 0. 010 respectively. The expression levels of PPAR＇y mRNA and protein in S1, $2, $3 and N groups were significantly lower （ P 〈 0. 05 ） than those in M and Con groups. There was no significant difference （ P 〉 0. 05 ） among S1, S2, S3 and N groups. The number of adipocytes and content of TG in S1, S2, S3 and N groups were significantly lower （ P 〈 0. 05 ） than those in M and Con groups, but there was no significant difference （ P 〉 0. 05 ） among S1, S2, S3 and N groups. Conclusion The adenovirus shuttle vector targeting PPARγ gene-mediated RNA interference can effec
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2013年第7期1343-1346,共4页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金资助项目（81071520）
