Touyz R M, Deng L Y, He G, Wu X H, Schiffrin E L
MRC Multidisciplinary Research Group on Hypertension, Clinical Research Institute of Montreal, University of Montreal, Quebec, Canada.
J Hypertens. 1999 Jul;17(7):907-16. doi: 10.1097/00004872-199917070-00006.
This study investigates the growth effects and associated signaling pathways of angiotensin II (Ang II) in human vascular smooth muscle cells.
Cultured vascular smooth muscle cells derived from resistance arteries (< 300 microm diameter) from subcutaneous gluteal biopsies of healthy subjects (n = 6) and human aortic vascular smooth muscle cells were used. Cells were studied between passages 3 and 6. Both 3H-thymidine and 3H-leucine incorporation were measured as indices of vascular smooth muscle cell hyperplasia (DNA synthesis) and cell hypertrophy (protein synthesis), respectively. Growth effects of Ang II (10(-12) - 10(-6) mol/l), in the absence and presence of 10(-5) mol/l losartan (AT1 antagonist) and PD123319 (AT2 antagonist), were determined. Ang II-induced effects were compared to those of endothelin-1. To determine whether extracellular signal-regulated kinase (ERK)-dependent pathways play a role in Ang II-mediated growth, cells were pretreated with the selective ERK kinase (MEK) inhibitor, PD98059 (10(-5) mol/l). ERK activation was determined by Western blot in the absence and presence of PD98059.
Ang II dose-dependently increased 3H-thymidine incorporation in cells from aorta (Emax = 276 +/- 10.4% of control) and resistance arteries (Emax = 284 +/- 5.1% of control). Ang II also stimulated 3H-leucine incorporation in cells from aorta (Emax = 162 +/- 11.6 of control) and resistance arteries (Emax 175 +/- 10% of control). Unlike Ang II, endothelin-1 failed to significantly alter cellular growth, except at high concentrations (> 10(-7) mol/l), where it had a weak stimulatory effect Losartan, but not PD123319, blocked Ang II-stimulated growth responses. Ang II significantly increased phosphorylation of ERK-1 and ERK-2, with maximum responses obtained at 5 min. PD98059 inhibited Ang II-stimulated ERK activity and abrogated agonist-induced DNA and protein synthesis. Losartan, but not PD123319 inhibited Ang II-induced phosphorylation of ERK-1 and ERK-2.
Ang II stimulates both hyperplasia and hypertrophy in vascular smooth muscle cells from human arteries. These growth effects are mediated via Ang II receptors of the AT1 subtype that are linked to ERK-dependent signaling pathways.
本研究调查血管紧张素II（Ang II）对人血管平滑肌细胞的生长影响及相关信号通路。
使用从健康受试者（n = 6）臀下皮下活检获取的直径小于300微米的阻力动脉来源的培养血管平滑肌细胞以及人主动脉血管平滑肌细胞。细胞在第3至6代之间进行研究。分别测量3H-胸腺嘧啶核苷和3H-亮氨酸掺入量，作为血管平滑肌细胞增生（DNA合成）和细胞肥大（蛋白质合成）的指标。测定了在不存在和存在10(-5)摩尔/升氯沙坦（AT1拮抗剂）和PD123319（AT2拮抗剂）的情况下，Ang II（10(-12) - 10(-6)摩尔/升）的生长影响。将Ang II诱导的效应与内皮素-1的效应进行比较。为了确定细胞外信号调节激酶（ERK）依赖性途径是否在Ang II介导的生长中起作用，细胞用选择性ERK激酶（MEK）抑制剂PD98059（[10(-5)摩尔/升]）预处理。在不存在和存在PD98059的情况下，通过蛋白质印迹法测定ERK激活情况。
Ang II剂量依赖性地增加主动脉细胞（Emax =对照的276 +/- 10.4%）和阻力动脉细胞（Emax =对照的284 +/- 5.1%）中的3H-胸腺嘧啶核苷掺入量。Ang II还刺激主动脉细胞（Emax =对照的162 +/- 11.6）和阻力动脉细胞（Emax =对照的175 +/- 10%）中的3H-亮氨酸掺入量。与Ang II不同，内皮素-1除了在高浓度（> 10(-7)摩尔/升）时有微弱的刺激作用外，未能显著改变细胞生长。氯沙坦而非PD123319阻断了Ang II刺激的生长反应。Ang II显著增加ERK-1和ERK-2的磷酸化，在5分钟时获得最大反应。PD98059抑制Ang II刺激的ERK活性并消除激动剂诱导的DNA和蛋白质合成。氯沙坦而非PD123319抑制Ang II诱导的ERK-1和ERK-2磷酸化。
Ang II刺激人动脉血管平滑肌细胞的增生和肥大。这些生长效应是通过与ERK依赖性信号通路相关的AT1亚型的Ang II受体介导的。
