Ishii K, Takekoshi K, Shibuya S, Kawakami Y, Isobe K, Nakai T
Department of Clinical Pathology, Institute of Clinical Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan.
J Hypertens. 2001 Nov;19(11):1991-9. doi: 10.1097/00004872-200111000-00009.
Two distinct types of angiotensin II (AngII) receptors, AT1 and AT2, have been cloned. We have shown previously that stimulation of AT2 reduces intracellular cyclic guanosine monophosphate (cGMP) levels in cultured porcine chromaffin cells in which AT2 is the predominantly expressed receptor. However, it has not been determined whether AT1 or AT2 affects signal transduction pathways involving mitogen-activated protein kinases (MAPKs) and signal transducers and activators of transcription (STATs) in chromaffin cells. Also, it is unclear whether cGMP/protein kinase G (PKG) is involved in the regulation of MAPKs and STATs in these cells.
Chromaffin cells were derived from porcine adrenal medulla. The effects of AngII alone (representing physiological conditions), AngII plus CV-11974 (an AT1 antagonist, which simulates specific AT2 stimulation), AngII plus PD 123319 (an AT2 antagonist, which simulates specific AT1 stimulation), and 8-Br-cGMP (a membrane-permeable cGMP analogue) alone on MAPKs (ERKs, JNK, p-38 MAPK) and STATs (STATs 1, 3 and 5) activity were measured.
Phosphorylated MAPKs (extracellular signal-related kinases (ERKs), c-jun N-terminal kinase (JNK) and p38 MAPK) and STATs (STATs 1, 3 and 5) were measured by immunoprecipitation-Western blot analysis (IP-Western blot).
AT1 stimulation markedly increased expression of ERKs, JNK, p38 MAPK via Ca2+-dependent protein kinase C (PKC) isoforms (cPKC), as well as STATs 1, 3 and 5 in cultured porcine chromaffin cells. In contrast, AT2 stimulation markedly decreased the expression of these signaling molecules. Also, 8-Br-cGMP alone induced increases in ERKs, JNK, p38 MAPK, and STATs 1, 3 and 5. Because AT2 inhibits cGMP production, we speculate that AT2 may act to suppress cGMP production, which in turn reduces the activity of both MAPKs and STATs in chromaffin cells.
AT2 negatively regulates AT1 in signal transduction pathways in chromaffin cells.
已克隆出两种不同类型的血管紧张素II（AngII）受体，即AT1和AT2。我们之前已经表明，在主要表达AT2受体的培养猪嗜铬细胞中，刺激AT2会降低细胞内环状鸟苷单磷酸（cGMP）水平。然而，尚未确定AT1或AT2是否会影响嗜铬细胞中涉及丝裂原活化蛋白激酶（MAPK）和信号转导及转录激活因子（STAT）的信号转导途径。此外，尚不清楚cGMP/蛋白激酶G（PKG）是否参与这些细胞中MAPK和STAT的调节。
嗜铬细胞取自猪肾上腺髓质。测量单独的AngII（代表生理条件）、AngII加CV-11974（一种AT1拮抗剂，模拟特异性AT2刺激）、AngII加PD 123319（一种AT2拮抗剂，模拟特异性AT1刺激）以及单独的8-溴-cGMP（一种可透过膜的cGMP类似物）对MAPK（细胞外信号调节激酶（ERK）、c-jun氨基末端激酶（JNK）、p-38 MAPK）和STAT（STAT1、3和5）活性的影响。
通过免疫沉淀-蛋白质印迹分析（IP-蛋白质印迹）测量磷酸化的MAPK（细胞外信号相关激酶（ERK）、c-jun N末端激酶（JNK）和p38 MAPK）以及STAT（STAT1、3和5）。
在培养的猪嗜铬细胞中，AT1刺激通过钙依赖性蛋白激酶C（PKC）同工型（cPKC）显著增加ERK、JNK、p38 MAPK以及STAT1、3和5的表达。相反，AT2刺激显著降低这些信号分子的表达。此外，单独的8-溴-cGMP可诱导ERK、JNK、p38 MAPK以及STAT1、3和5增加。由于AT2抑制cGMP产生，我们推测AT2可能通过抑制cGMP产生来发挥作用，进而降低嗜铬细胞中MAPK和STAT的活性。
在嗜铬细胞的信号转导途径中，AT2对AT1起负调节作用。
