Enhanced prostacyclin synthesis in endothelial cells by retrovirus-mediated transfer of prostaglandin H synthase cDNA.
Enhanced prostacyclin synthesis in endothelial cells by retrovirus-mediated transfer of prostaglandin H synthase cDNA.
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通过逆转录病毒介导的前列腺素 H 合酶 cDNA 转移增强内皮细胞中的前列环素合成。
DOI：
10.1172/jci116398
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Wu,KK
中科院分区：
文献类型：
--
作者：
Xu,XM;Ohashi,K;Sanduja,SK;Ruan,KH;Wang,LH;Wu,KK
用逆转录病毒载体（BAG）将人前列腺素H合成酶（PGHS-1）基因导入人内皮细胞系，以提高PGI 2的合成。用含有相对于逆转录病毒启动子（PGHS（S））有义方向的PGHS-1 cDNA的BAG感染的细胞表达的mRNA增加了30倍，但由于阅读移码，没有显示PGHS蛋白或PGI 2合成的增加，而用相对于病毒启动子（PGHS（R））反向的PGHS-1感染的细胞，与对照组相比，PGHS mRNA增加> 10倍（169 +/-22vs14.8 +/阿莫尔/mg RNA），PGHS蛋白（5.82 +/-1.07vs0.23 +/-0.04ng/mg蛋白）和酶活性一致增加。PGHS（R）的引物延伸分析显示，两个转录起始位点位于与PGHS-1 cDNA相邻的SV 40晚期启动子区域。PGHS（R）细胞产生高的基础PGI 2水平，其响应于离子载体、花生四烯酸和凝血酶的刺激而增加数倍。动力学分析显示，PGI 2合成速率为14 ng/min-1/百万细胞，PGI 2合成的t1/2为13.3 min。这些发现表明，将PGHS-1基因转移到血管细胞中可增强PGI 2合成，可能是恢复受损血管血栓保护特性的有用策略。
A retroviral vector (BAG) was used to transfer human prostaglandin H synthase (PGHS-1) gene into a human endothelial cell line for enhancement of PGI2 synthesis. Cells infected with BAG containing PGHS-1 cDNA in the sense orientation relative to the retroviral promoter (PGHS(S)) expressed a 30-fold increase in mRNA but, due to a reading frame shift, did not show an increase in PGHS protein or in PGI2 synthesis, while those with PGHS-1 in reverse orientation relative to the viral promoter (PGHS(R)), produced a > 10-fold increase in PGHS mRNA over the control (169 +/- 22 vs 14.8 +/- 1.2 amol/micrograms RNA) with a concordant increase in PGHS protein (5.82 +/- 1.07 vs 0.23 +/- 0.04 ng/mg protein) and enzyme activity. Primer extension analysis of PGHS(R) revealed two transcription start sites located in the SV40 late promoter region adjacent to PGHS-1 cDNA. PGHS(R) cells produced a high basal PGI2 level which was increased by several-fold in response to stimulation by ionophore, arachidonic acid, and thrombin. Kinetic analysis revealed the PGI2 synthetic rate to be 14 ng/min-1 per million cells and t1/2 of PGI2 synthesis, 13.3 min. These findings indicate that transfer of PGHS-1 gene into vascular cells enhances PGI2 synthesis and may be a useful strategy for restoring thromboprotective property of damaged blood vessels.Images