Iwami K, Ashizawa N, Do Y S, Graf K, Hsueh W A
Department of Medicine, University of Southern California School of Medicine, Los Angeles 90033, USA.
Am J Physiol. 1996 Jun;270(6 Pt 2):H2100-7. doi: 10.1152/ajpheart.1996.270.6.H2100.
The purpose of the present investigation was to compare the effects of angiotensin II (ANG II) other growth factors implicated to play a role in ventricular hypertrophy on cardiac fibroblast changes associated with cardiac remodeling. These changes included induction of early growth response (Egr-1) gene and increases in message levels of extracellular matrix proteins. ANG II treatment (10(-10)-10(-6) M) of rat cardiac fibroblasts induced 1) Egr-1 and 2) a fourfold (P < 0.02) increase in fibronectin and a twofold (P = 0.05) increase in laminin mRNA levels but no increases in that of collagens I, III, or IV at 24-48 h, and 3) a decrease in AT1-receptor mRNA levels to 26% (P < 0.001) of basal at 4-6 h. These effects were all inhibited by the AT1-receptor blocker, losartan, but not AT2-receptor blockers. Immunostaining of cultured cells with antibody against rat fibronectin demonstrated positive staining of cells in serum-free medium; staining was more intense in cells treated with ANG II (10(-6) M, 48 h). Fluorescent-activated cell sorting using an antibody against rat AT1 receptor demonstrated a receptor signal in cells maintained in serum-free medium; however, the receptor signal was not detectable in ANG II-treated cells. Serum and epidermal growth factor (EGF) also induced Egr-1, but norepinephrine (NE) and endothelin (ET) had no effect. Serum increased fibronectin mRNA levels by twofold (P < 0.05). EGF, NE, and ET had no effect on matrix gene expression. Serum, EGF, and NE also transiently downregulated AT1-receptor mRNA levels at 4-6 h of treatment. These results demonstrate that 1) ANG II both induces protooncogene expression and enhances fibronectin mRNA levels in cultured cardiac fibroblasts, whereas EGF only induces Egr-1, and NE and ET have no effects on either function; 2) ANG II effects are primarily mediated by the AT1 receptor; and 3) growth factors can regulate AT1-receptor mRNA levels. Thus ANG II, relative to NE, ET, and EGF, appears to play a prominent and direct role in fibroblast changes associated with cardiac hypertrophy.
本研究的目的是比较血管紧张素II（ANG II）与其他被认为在心室肥大中起作用的生长因子对与心脏重塑相关的心脏成纤维细胞变化的影响。这些变化包括早期生长反应（Egr-1）基因的诱导以及细胞外基质蛋白信使水平的增加。用ANG II（10^(-10)-10^(-6) M）处理大鼠心脏成纤维细胞，在24至48小时内可诱导：1）Egr-1；2）纤连蛋白增加四倍（P < 0.02），层粘连蛋白信使水平增加两倍（P = 0.05），但I、III或IV型胶原蛋白的信使水平无增加；3）在4至6小时时，AT1受体信使水平降至基础水平的26%（P < 0.001）。这些作用均被AT1受体阻滞剂氯沙坦抑制，但不被AT2受体阻滞剂抑制。用抗大鼠纤连蛋白抗体对培养细胞进行免疫染色，结果显示在无血清培养基中的细胞呈阳性染色；在经ANG II（10^(-6) M，48小时）处理的细胞中染色更强烈。使用抗大鼠AT1受体抗体进行荧光激活细胞分选，结果显示在无血清培养基中培养的细胞存在受体信号；然而，在经ANG II处理的细胞中未检测到受体信号。血清和表皮生长因子（EGF）也诱导Egr-1，但去甲肾上腺素（NE）和内皮素（ET）无作用。血清使纤连蛋白信使水平增加两倍（P < 0.05）。EGF、NE和ET对基质基因表达无作用。血清、EGF和NE在处理4至6小时时也会短暂下调AT1受体信使水平。这些结果表明：1）ANG II既能诱导原癌基因表达，又能增强培养的心脏成纤维细胞中纤连蛋白信使水平，而EGF仅诱导Egr-1，NE和ET对这两种功能均无作用；2）ANG II的作用主要由AT1受体介导；3）生长因子可调节AT1受体信使水平。因此，相对于NE、ET和EGF，ANG II似乎在与心脏肥大相关的成纤维细胞变化中起突出且直接的作用。
