tTG硫代反义寡核苷酸抑制牛眼小梁细胞tTG的表达△  

Inhibiting effect of tissue transglutaminase (tTG) antisense oligodeoxynucleotides on tTG expression in cultured bovine trabecular meshwork cells

作　　者：胡义珍[1] 张海江[1] 熊新春[1] 曹阳[1] 席祖莲[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院眼科,湖北省武汉市430022

出　　处：《眼科新进展》2006年第3期180-183,共4页Recent Advances in Ophthalmology

基　　金：湖北省自然科学基金资助(编号:2003ABA147)~~

摘　　要：目的观察组织型谷氨酰胺转移酶(tissue transglutaminase,tTG)硫代磷酸化修饰反义寡核苷酸(antisense oligodeoxynucleotide,ASODN)对体外培养牛眼小梁细胞(bovine trabecular meshwork cell,BTMC)tTG在mRNA水平和蛋白水平表达的影响,初步探讨tTG-ASODN在治疗原发性开角型青光眼中的可能性。方法根据牛tTG mRNA的二级结构设计并合成ASODN1、ASODN2,用阳离子脂质体Lipofectamin包埋后转染BTMC,以半定量逆转录-聚合酶链反应法检测转染后tTG mRNA表达的变化,以免疫组织化学SP法检测转染后tTG蛋白表达的变化。结果ASODN1组和ASOND2组的tTG/GAPDH灰度比值分别为1.0695±0.0009、1.0662±0.0007,空白对照组和错义寡核苷酸组分别为1.0185±0.0024、1.0167±0.0012;tTG-ASODN1、tTG-ASODN2对BTMC的tTG mRNA和蛋白表达均有明显的抑制作用,与空白对照组和错义寡核苷酸组相比有显著性差异(P<0.05);tTG-ASODN1与tTG-ASODN22组间无显著性差异(P>0.05)。结论tTG-ASODN可明显下调BTMC的tTG在mRNA和蛋白水平的表达,提示tTG-ASODN有望在原发性开角型青光眼的治疗中发挥重要作用。Objective To study the effect of tTG fully phosphorothioated antisense oligodeoxynucleotides （tTG-ASODN） on tTG expression in cultured bovine trabecular meshwork cells in vitro and explore a new possibility for anti-POAG therapy. Methods According to the secondary structure of tTG, the ASODNI and ASODN2 complementary to the protein codogram region of tTG were designed, synthesized and phesphorothioated. The ASODNI and ASODN2 were embedded in Lipofectamine and transfected into bovine trabecular meshwork cells. The untreated group and the nonsense oligodeoxynucleotides group were negative controls. The expression of tTG in the mRNA and protein level was measured by semi-quantitative RT-PCR and immunohistochemical technique-Supervision method respectively. Results The gray scale rate in ASODNI group and ASODN2 group respectively was 1.069 5 ± 0.000 9 and 1.066 2±0.000 7,which was 1.018 5 ±0.002 4 and 1.0167 ±0.001 2 in untreated group and nonsense oligodeoxynucleotides group. Both the mRNA and the protein of tTG with tTG-ASODNI and tTG-ASODN2 were significantly decreased compared with that of the controls（P 〈 0.05）. However, there was no significantly difference between the group of ASODNI and the group of A- SODN2（P 〉 0.05）. Conclusion The expression of tTG mRNA and protein in cultured BTMC are down-regulated by tTG- ASODN. As a result, tTG-ASODN may be expected to treat POAG on its pathogenesis through inhibition of expression of tTG. [ Rec Adv Ophthaimol 2006; 26 （3） ： 180-183 ]

关 键 词：组织型谷氨酰胺转移酶 反义寡核苷酸 小梁细胞 开角型青光眼 

分 类 号：R775.2[医药卫生—眼科]