Protein kinase A dependent signalling mediates anti-apoptotic effects of the atrial natriuretic peptide in ischemic livers
Protein kinase A dependent signalling mediates anti-apoptotic effects of the atrial natriuretic peptide in ischemic livers
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DOI：
10.1016/j.jhep.2004.05.017
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发表时间：
2004-09-01
影响因子：
25.7
通讯作者：
Kiemer, AK
中科院分区：
文献类型：
--
作者：
Kulhanek-Heinze, S;Gerbes, AL;Kiemer, AK
背景/目的：心钠素(ANP)对肝脏的预适应可通过颗粒鸟苷环化酶减轻缺血再灌注损伤(IRI)。最近，我们发现ANP影响肝脏中的p38 MAPK信号级联反应。因此，本研究的目的是阐明cGMP和p38 MAPK依赖的信号通路在ANP介导的抗细胞凋亡效应中的作用。方法：用含或不含ANP、8-Br-cGMP(+/-激酶抑制剂)的KH缓冲液灌流大鼠肝脏，并将其保存在UW液中(4℃，24 h)。荧光分光光度法检测caspase-3样活性。RT-PCR法检测肝组织cGMP依赖性蛋白激酶(PKG)的表达，Western印迹法检测BAD磷酸化，体外磷酸化法检测cAMP依赖性蛋白激酶(PKA)活性。结果：与对照组相比，ANP预适应使缺血后caspase-3样活性降低。无论是用p38MAPK抑制剂灌流还是用PKG抑制剂灌流，都不能阻断ANP介导的抗细胞凋亡作用。这两种PKG亚型在肝脏中不表达。相反，选择性PKA抑制剂的肝脏灌流取消了ANP的抗细胞凋亡作用。ANP激活的PKA对促凋亡的BAD的磷酸化可能抑制肝细胞的凋亡。结论：ANP通过与PKA通路的串扰，介导其在缺血损伤中的抗凋亡作用。(C)2004年欧洲肝脏研究协会。爱思唯尔出版，版权所有。
Background/Aims: Preconditioning of livers with atrial natriuretic peptide (ANP) attenuates ischemia-reperfusion injury (IRI) via the particulate guanylate cyclase. Recently, we have shown that ANP affects the p38 MAPK signalling cascade in the liver. Thus, aim of the present study was to elucidate the role of cGMP- and p38 MAPK-dependent signalling pathways in ANP-mediated anti-apoptotic effects.Methods: Rat livers were perfused with KH-buffer with or without ANP, 8-Br-cGMP (+/-kinase inhibitors) and kept in UW solution (4 degreesC, 24 h). Caspase-3-like activity was measured by a fluorometric assay. Expression of cGMP-dependent protein kinases (PKG) in liver tissue was determined by RT-PCR, BAD phosphorylation by Western blot, and cAMP-dependent protein kinase (protein kinase A, PKA) activity by in vitro phosphorylation.Results: Compared to control organs, ANP-preconditioning reduced post-ischemic caspase-3-like activity. Neither perfusion with a p38 MAPK inhibitor nor with a PKG inhibitor abolished the ANP-mediated anti-apoptotic action. The two PKG isoforms were demonstrated not to be expressed in the liver. In contrast, liver perfusion with a selective PKA inhibitor abrogated the anti-apoptotic effect of ANP. Phosphorylation of pro-apoptotic BAD by ANP-activated PKA might inhibit liver cell apoptosis.Conclusions: ANP mediates its anti-apoptotic action during ischemic injury via a crosstalk with the PKA pathway. (C) 2004 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.