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 tTG反义寡核苷酸对小梁细胞tTG表达的抑制作用
 张海江;胡义珍;熊新春
 　　目的通过计算机程序模拟组织型谷氨酰胺转移酶(tissue transglutaminase,tTG)基因的mRNA二级结构,优化硫代反义寡核苷酸(antisense oligodeoxynucleotide,A-SODN)药物设计,为有效抑制小梁细胞表达tTG提供新的途径。方法根据牛tTG mRNA的二级结构,选择环与茎结合区(280～300bp)和茎区(2720～2740bp)作为反义药物作用的靶点,分别合成ASODN1、ASODN2,用脂质体包埋后转染牛眼小梁细胞。通过半定量RT-PCR法检测转染后tTG mRNA表达的变化来评价tTG-ASODN的生物学效应。结果ASODN1、ASODN2对tTG mRNA的表达均有抑制作用,与空白对照组相比差异有显著性(P<0.05),且tTG-A-SODN1抑制作用明显强于tTG-ASODN2(P<0.05)。结论选择环与茎连接的部位作为反义作用的靶点,其反义作用明显强于茎区。从机制上讲,可能是因为环与茎连接的部位的形成需要吸收能量,是整个tTG mRNA序列中最不稳定的区域,ASODN容易接近而发挥作用,提示通过tTG mRNA二级结构的模拟而选择ASODN是设计反义药物的新途径。
【作者单位】：三峡大学第一临床医学院湖北省宜昌市中心人民医院眼科 湖北宜昌443000(张海江);华中科技大学附属协和医院眼科 湖北武汉430022(胡义珍;熊新春)
【关键词】：组织型谷氨酰胺转移酶;反义药物;小梁细胞
【基金】：湖北省自然科学基金资助项目(2003ABA147)
【分类号】：R775.2
【DOI】：CNKI:SUN:ZXYK.0.2007-04-004
【正文快照】：
　　组织型谷氨酰胺转移酶(tissue transglutami-nase,tTG)作为在体内分布最为广泛的谷氨酰胺转移酶家族成员之一,具有催化赖氨酸残基与谷氨酰胺残基交联的活性,在细胞凋亡、细胞分化、基质的稳定、信号传导、伤口愈合等许多生理和病理中起着重要作用[1]。已有研究表明,在眼部,tTG
 
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 Inhibiting effect of tTG antisense oligodeoxynucleotides on its expression of trabecular meshwork cells
 ZHANG Haijiang;HU Yizhen;XIONG Xinchun Department of Ophthalmology;the First Clinical Medical College of Three Gorges University;Yichang Central Hospital;Yichang China;44300
 　Objective According to the secondary structure of tissue transglutaminase (tTG),design and select the best antisense oligodeoxynucleotide(ASODN) and to reduce the expression of tTG in cultured bovine trabecular meshwork cells. Methods We selected two bands which were complementary to the combined position of ring and caudex (ASODN1,280~300bp) and the caudex region (ASODN2,2720~2740bp). Both were designed,synthesized,phosphorothioated and embedded in Lipofectamine to transfect bovine trabecular meshwork cells. Through measuring the expression of tTG in mRNA level by semi-Quantitative RT-PCR,we estimate the inhibiting effect of tTG-ASODN. Results The tTG-ASODN were significantly decreased the expression of tTG,but the inhibiting effect of tTG-ASODN1 was stronger than tTG-ASODN2. Moreover,there had significantly difference. Conclusion The ASODN1 which was located on the combined position of ring and caudex is better than the ASODN2. The reason is that that position was very unstable because of its wasting energy for maintaining the ring structure,inducing the band to approximate mRNA of tTG easily. So this study may be exploring a new way for designing anti-tTG through the secondary structure of tTG.
【Keyword】：tissue transglutaminase;antisense medicine;trabecular meshwork cell
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