Fukuda N, Kubo A, Watanabe Y, Nakayama T, Soma M, Izumi Y, Kanmatsuse K
Second Department of Internal Medicine, Nihon University School of Medicine, Tokyo, Japan.
J Hypertens. 1997 Oct;15(10):1123-36. doi: 10.1097/00004872-199715100-00010.
To evaluate the effects of the antisense oligodeoxynucleotide (ODN) to platelet-derived growth factor (PDGF) A-chain messenger RNA (mRNA) on the growth of cardiovascular organs in hypertension.
15-Mer antisense ODN complementary to the initiation codon region of rat PDGF-A chain mRNA and non-sense ODN of identical proportion but with a random order of bases relative to that of antisense ODN were synthesized with a DNA synthesizer.
We examined the effects of the antisense ODN on the growth of vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats, and on the expression of PDGF A-chain mRNA by reverse transcription and polymerase chain reaction and PDGF A-chain protein by Western blot analysis in vitro. We evaluated the distribution of 32P-labeled antisense ODN and examined the effects of the antisense ODN on the growth of cardiovascular organs in vivo.
The antisense ODN reduced the basal DNA synthesis of VSMC from SHR significantly, but did not do so in cells from Wistar-Kyoto rats. Mutations in the antisense ODN sequence reduced the ODN-induced inhibition of DNA synthesis. Addition of serum or transforming growth factor-beta 1 increased the DNA synthesis in the SHR-derived VSMC that was inhibited by the antisense ODN. The antisense ODN inhibited the production of PDGF A-chain protein, but not of the PDGF A-chain mRNA. The injection of 32P-antisense ODN in vivo led to a greater accumulation of radioactivity in the aorta than in other organs. Infusion of antisense ODN for 28 days did not alter the systolic blood pressure appreciably in rats of either strain. However, in SHR, it reduced markedly the elevated DNA content, [3H]-thymidine uptake, and incorporation of [3H]-thymidine into aortic DNA, and suppressed the production of aortic PDGF A-chain protein. These results indicated that the PDGF A-chain is involved in the exaggerated growth of VSMC from SHR by which inhibition of the translation of PDGF A-chain mRNA to the protein with antisense ODN occurs in vitro, and that antisense ODN to PDGF A-chain suppresses the exaggerated arterial proliferation in SHR without altering the high blood pressure in vivo.
These results imply that inhibition of the final responsible growth factor PDGF A-chain by antisense ODN can suppress the arterial proliferation in hypertension without altering the blood pressure, suggesting that the arterial proliferation in hypertension is independent of the high blood pressure in part, and that antisense therapy could be feasible for treating hypertension.
评估血小板源性生长因子（PDGF）A链信使核糖核酸（mRNA）的反义寡脱氧核苷酸（ODN）对高血压状态下心血管器官生长的影响。
用DNA合成仪合成与大鼠PDGF - A链mRNA起始密码子区域互补的15聚体反义ODN，以及碱基比例相同但相对于反义ODN碱基顺序随机的无义ODN。
我们在体外检测了反义ODN对自发性高血压大鼠（SHR）和Wistar - Kyoto大鼠血管平滑肌细胞（VSMC）生长的影响，以及通过逆转录聚合酶链反应检测其对PDGF A链mRNA表达的影响，通过蛋白质免疫印迹分析检测对PDGF A链蛋白表达的影响。我们评估了32P标记的反义ODN的分布，并检测了反义ODN对体内心血管器官生长的影响。
反义ODN显著降低了SHR来源的VSMC的基础DNA合成，但对Wistar - Kyoto大鼠来源的细胞没有此作用。反义ODN序列中的突变降低了ODN诱导的DNA合成抑制作用。添加血清或转化生长因子 - β1可增加被反义ODN抑制的SHR来源的VSMC中的DNA合成。反义ODN抑制了PDGF A链蛋白的产生，但不抑制PDGF A链mRNA的产生。体内注射32P - 反义ODN导致主动脉中放射性的积累比其他器官更多。输注反义ODN 28天对两种品系大鼠的收缩压均无明显改变。然而，在SHR中，它显著降低了升高的DNA含量、[3H] - 胸腺嘧啶摄取以及[3H] - 胸腺嘧啶掺入主动脉DNA，并抑制了主动脉PDGF A链蛋白的产生。这些结果表明，PDGF A链参与了SHR来源的VSMC过度生长，体外反义ODN可抑制PDGF A链mRNA向蛋白质的翻译，并且PDGF A链的反义ODN可抑制SHR中过度的动脉增殖，而不改变体内的高血压状态。
这些结果表明，反义ODN抑制最终起作用的生长因子PDGF A链可抑制高血压中的动脉增殖而不改变血压，提示高血压中的动脉增殖部分独立于高血压，反义疗法可能对治疗高血压可行。
