GENE-TRANSFER IN REGENERATING MUSCLE
GENE-TRANSFER IN REGENERATING MUSCLE
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DOI：
10.1089/hum.1994.5.1-11
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发表时间：
1994-01-01
影响因子：
4.2
通讯作者：
SCHIAFFINO, S
中科院分区：
文献类型：
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作者：
VITADELLO, M;SCHIAFFINO, MV;SCHIAFFINO, S
我们比较了直接基因转移在正常和再生大鼠骨骼肌中的效率。在成年大鼠比目鱼肌内肌肉注射布比卡因，引起肌肉坏死和再生。将含有由病毒启动子驱动的β-半乳糖苷酶（β-gal）或氯霉素乙酰转移酶（CAT）基因的质粒在布比卡因治疗后3天注射到再生和对侧未受伤的肌肉中。在转染后7天，与对照肌肉相比，再生中CAT活性的表达高出> 80倍，但在30和60天时降低。Southern印迹分析表明，CAT DNA的主要形式是在转染的肌肉游离型，然而，CAT活性测量进行相同的转染肌肉的酶活性和质粒DNA的量之间没有精确的相关性。在转染后7天，在受损比目鱼肌的许多再生纤维中检测到β-gal的表达;相反，在对照肌肉中仅发现罕见的阳性纤维。在转染后30天观察到单核细胞的局灶性浸润，其选择性地包围和侵入β-gal阳性纤维片段，这表明免疫机制与转基因随时间的进行性损失有关。再生肌纤维显示更高的转染效率的发现可能与杜氏肌营养不良症的基因治疗有关，因为再生纤维在疾病的早期阶段是大量的。
We have compared the efficiency of direct gene transfer in normal and regenerating rat skeletal muscle. Muscle necrosis and regeneration was induced by intramuscular injection of bupivacaine in the soleus muscle of adult rats. Plasmids containing beta-galactosidase (beta-gal) or chloramphenicol acetyltransferase (CAT) genes driven by viral promoters were injected 3 days after bupivacaine treatment into the regenerating and the contralateral uninjured muscles. Expression of CAT activity was > 80-fold higher in regenerating compared to control muscles at 7 days post-transfection, but decreased at 30 and 60 days. Southern blot analysis showed that the predominant form of CAT DNA was episomal in transfected muscles; however, CAT activity measurements performed on the same transfected muscles showed no precise correlation between enzymatic activity and amount of plasmid DNA. Expression of beta-gal was detected in numerous regenerating fibers of the injured soleus muscles at 7 days post-transfection; in contrast, only rare positive fibers were found in control muscles. Focal infiltrates of mononuclear cells, which surround and invade selectively beta-gal-positive fiber segments, were observed at 30 days post-transfection, suggesting that immune mechanisms are implicated in the progressive loss of transgenes with time. The finding that regenerating muscle fibers display a higher efficiency of transfection may be relevant to gene therapy of Duchenne muscular dystrophy, because regenerating fibers are numerous in the early stages of the disease.