Laboratory of Radiobiology and Molecular Genetics, Institute Vinca, University of Belgrade, P.O. Box 522, 11001 Belgrade, Serbia.
J Physiol Biochem. 2011 Jun;67(2):195-204. doi: 10.1007/s13105-010-0063-1. Epub 2010 Nov 24.
The purpose of this study was to examine the effects of ghrelin on protein kinase B (Akt) and mitogen-activated protein kinase p42/44 (ERK1/2) activation as well as ghrelin effects on inducible nitric oxide (NO) synthase (iNOS; for gene Nos2) activity/expression in rat hearts. Male Wistar rats were treated with ghrelin (0.3 nmol/5 μl) or an equal volume of phosphate-buffered saline, injected every 24 h into the lateral cerebral ventricle for 5 days and 2 h after the last treatment the animals were sacrificed. Serum NO, L-arginine (L-Arg), and arginase activity were measured spectrophotometrically. For phosphorylation of Akt, ERK1/2, and iNOS protein expression, Western blot method was used. The expression of Nos2 mRNA was measured by the quantitative real-time polymerase chain reaction (qRT-PCR). Treatment with ghrelin significantly increased NO production in serum by 1.4-fold compared with control. The concentration of L-Arg was significantly higher in ghrelin-treated rats than in control while arginase activity was significantly lower in ghrelin-treated than in control hearts. Ghrelin treatment increased phosphorylation of Akt by 1.9-fold and ERK1/2 by 1.6-fold and increased iNOS expression by 2.5-fold compared with control. In addition, ghrelin treatment increased Nos2 gene expression by 2.2-fold as determined by qRT-PCR. These results indicate that ghrelin regulation of iNOS expression/activity is mediated via Akt/ERK1/2 signaling pathway. These results may be relevant to understanding molecular mechanisms underlying direct cardiovascular actions of ghrelin.
本研究旨在探讨 ghrelin 对蛋白激酶 B（Akt）和丝裂原活化蛋白激酶 p42/44（ERK1/2）激活的影响，以及 ghrelin 对大鼠心脏诱导型一氧化氮合酶（iNOS；基因 Nos2）活性/表达的影响。雄性 Wistar 大鼠接受 ghrelin（0.3 nmol/5 μl）或等量磷酸盐缓冲盐水处理，每隔 24 小时侧脑室注射一次，共 5 天，最后一次注射后 2 小时处死动物。采用分光光度法测定血清 NO、L-精氨酸（L-Arg）和精氨酸酶活性。采用 Western blot 法检测 Akt、ERK1/2 和 iNOS 蛋白表达的磷酸化情况。通过实时定量聚合酶链反应（qRT-PCR）测定 Nos2 mRNA 的表达。与对照组相比，ghrelin 处理使血清中 NO 生成增加 1.4 倍。ghrelin 处理组大鼠 L-Arg 浓度明显高于对照组，而 ghrelin 处理组大鼠心脏精氨酸酶活性明显低于对照组。与对照组相比，ghrelin 处理使 Akt 磷酸化增加 1.9 倍，ERK1/2 增加 1.6 倍，iNOS 表达增加 2.5 倍。此外，qRT-PCR 结果表明，ghrelin 处理使 Nos2 基因表达增加 2.2 倍。这些结果表明，ghrelin 对 iNOS 表达/活性的调节是通过 Akt/ERK1/2 信号通路介导的。这些结果可能有助于理解 ghrelin 对心血管直接作用的分子机制。
