Liu Jie, Wu Li-ling, Li Li, Zhang Liang, Song Zong-En
Department of Physiology and Pathophysiology, Peking University Health Science Center, Beijing 100083, China.
Regul Pept. 2005 Apr 15;127(1-3):11-8. doi: 10.1016/j.regpep.2004.10.018.
Platelet-derived growth factor (PDGF) is a dimeric molecule consisting of disulfide-bonded A- and B-polypeptide chains. Homodimeric (PDGF-AA, PDGF-BB) as well as heterodimeric (PDGF-AB) isoforms exert their effects on target cells by binding with different specificities to two structurally related protein tyrosine kinase receptors, denoted alpha- and beta-receptors. PDGF stimulates growth in various cell types, but little is known about its effect on mammalian cardiomyocytes. Therefore, growth-promoting effect of PDGF on rat cardiomyocytes was investigated. Primary culture of neonatal rat ventricular myocytes was prepared and cellular growth was estimated by [3H]-leucine incorporation assay. Tyrosine-phosphorylated PDGF-beta receptor of cardiomyocytes was determined by immunoblotting analysis after immunoprecipitation. PDGF-beta receptor, extracellular signal-regulated kinase (ERK) 1/2 and phosphorylated ERK1/2 of cardiomyocytes were measured by immunoblotting analysis. [3H]-leucine incorporation into the cultured myocytes was increased in a time- and dose-dependent manner after PDGF-BB stimulation. Phosphorylation of PDGF-beta receptor and ERK1/2 in cardiomyocytes was increased after short-term stimulation of PDGF-BB. Protein expression of PDGF-beta receptor and ERK1/2 was increased after long-term stimulation of PDGF-BB. [(3)H]-leucine incorporation into the cultured myocytes induced by PDGF-BB was partly blocked by mitogen-activated ERK-activating kinase (MEK) inhibitor PD98059, phospholipase C (PLC) inhibitor U73122, and protein kinase C (PKC) inhibitor staurosporin aglycone, respectively. Therefore, PDGF beta receptor, ERK1/2, PLC and PKC are involved in the signal transduction of PDGF-induced growth response of rat cardiac myocytes.
血小板衍生生长因子（PDGF）是一种二聚体分子，由通过二硫键连接的A和B多肽链组成。同二聚体（PDGF-AA、PDGF-BB）以及异二聚体（PDGF-AB）亚型通过与两种结构相关的蛋白酪氨酸激酶受体（分别称为α受体和β受体）以不同的特异性结合，对靶细胞发挥作用。PDGF可刺激多种细胞类型的生长，但对其对哺乳动物心肌细胞的影响了解甚少。因此，研究了PDGF对大鼠心肌细胞的促生长作用。制备新生大鼠心室肌细胞的原代培养物，并通过[3H]-亮氨酸掺入试验评估细胞生长。免疫沉淀后通过免疫印迹分析测定心肌细胞酪氨酸磷酸化的PDGF-β受体。通过免疫印迹分析测量心肌细胞的PDGF-β受体、细胞外信号调节激酶（ERK）1/2和磷酸化的ERK1/2。PDGF-BB刺激后，[3H]-亮氨酸掺入培养的心肌细胞呈时间和剂量依赖性增加。短期刺激PDGF-BB后，心肌细胞中PDGF-β受体和ERK1/2的磷酸化增加。长期刺激PDGF-BB后，PDGF-β受体和ERK1/2的蛋白表达增加。PDGF-BB诱导的[3H]-亮氨酸掺入培养的心肌细胞分别被丝裂原活化的ERK激活激酶（MEK）抑制剂PD98059、磷脂酶C（PLC）抑制剂U73122和蛋白激酶C（PKC）抑制剂星形孢菌素苷元部分阻断。因此，PDGF-β受体、ERK1/2、PLC和PKC参与了PDGF诱导的大鼠心肌细胞生长反应的信号转导。
