Novel expression cassettes for increasing apolipoprotein AI transgene expression in vascular endothelial cells.
Novel expression cassettes for increasing apolipoprotein AI transgene expression in vascular endothelial cells.
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DOI：
10.1038/s41598-022-25333-9
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发表时间：
2022-12-06
影响因子：
4.6
通讯作者：
Dichek, David A.
中科院分区：
文献类型：
--
作者：
Sethuraman, Meena;Dronadula, Nagadhara;Bi, Lianxiang;Wacker, Bradley K.;Knight, Ethan;De Bleser, Pieter;Dichek, David A.
用表达载脂蛋白A-I（APOAI）的载体转导内皮细胞（EC）可减少脂肪喂养兔动脉中的动脉粥样硬化。然而，对动脉粥样硬化的影响是部分的，如果APOAI表达增加，可能会增强。为了开发在EC中产生更高水平的APOAI mRNA的表达盒，我们测试了4种策略，主要是在体外：添加2种类型的增强子，添加计算鉴定的EC特异性顺式调节模块（CRM），以及在从培养EC中高度表达的基因克隆的序列的转录起始位点（TSS）插入兔APOAI基因。添加剪切应力响应增强子并不增加APOAI表达。添加2个拷贝的Mef 2c增强子增加了中度活性启动子/增强子的APOAI表达，但降低了高度活性启动子/增强子的APOAI表达。在11种CRM中，3种增加了中度活性启动子的APOAI表达（2-7倍; P < 0.05）;没有增加高活性启动子/增强子的表达。将APOAI基因插入到高表达EC基因的TSS中不会使表达增加到高于用中等活性启动子/增强子获得的水平。需要新的策略来进一步增加EC中的APOAI转基因表达。
Transduction of endothelial cells (EC) with a vector that expresses apolipoprotein A-I (APOAI) reduces atherosclerosis in arteries of fat-fed rabbits. However, the effects on atherosclerosis are partial and might be enhanced if APOAI expression could be increased. With a goal of developing an expression cassette that generates higher levels of APOAI mRNA in EC, we tested 4 strategies, largely in vitro: addition of 2 types of enhancers, addition of computationally identified EC-specific cis-regulatory modules (CRM), and insertion of the rabbit APOAI gene at the transcription start site (TSS) of sequences cloned from genes that are highly expressed in cultured EC. Addition of a shear stress-responsive enhancer did not increase APOAI expression. Addition of 2 copies of a Mef2c enhancer increased APOAI expression from a moderately active promoter/enhancer but decreased APOAI expression from a highly active promoter/enhancer. Of the 11 CRMs, 3 increased APOAI expression from a moderately active promoter (2–7-fold; P < 0.05); none increased expression from a highly active promoter/enhancer. Insertion of the APOAI gene into the TSS of highly expressed EC genes did not increase expression above levels obtained with a moderately active promoter/enhancer. New strategies are needed to further increase APOAI transgene expression in EC.
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影响因子：
64.8
作者：
Barzel, A.;Paulk, N. K.;Shi, Y.;Huang, Y.;Chu, K.;Zhang, F.;Valdmanis, P. N.;Spector, L. P.;Porteus, M. H.;Gaensler, K. M.;Kay, M. A.
通讯作者：
Kay, M. A.
影响因子：
3
作者：
Heskes T;Eisinga R;Breitling R
通讯作者：
Breitling R
影响因子：
14.9
作者：
Griffon A;Barbier Q;Dalino J;van Helden J;Spicuglia S;Ballester B
通讯作者：
Ballester B
影响因子：
14.9
作者：
GuhaThakurta D
通讯作者：
GuhaThakurta D
影响因子：
12.4
作者：
Flynn, Rowan;Qian, Kun;Dichek, David A.
通讯作者：
Dichek, David A.