Roles of Cx43-associated protein kinases in suppression of gap junction-mediated chemical coupling by ischemic preconditioning.
Roles of Cx43-associated protein kinases in suppression of gap junction-mediated chemical coupling by ischemic preconditioning.
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DOI：
10.1152/ajpheart.00448.2008
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发表时间：
2009-02
期刊：
影响因子：
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通讯作者：
K. Naitoh;T. Yano;T. Miura;T. Itoh;T. Miki;M. Tanno;Takahiro Sato;Hiroyuki Hotta;Y. Terashima;K. Shimamoto
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文献类型：
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作者：
K. Naitoh;T. Yano;T. Miura;T. Itoh;T. Miki;M. Tanno;Takahiro Sato;Hiroyuki Hotta;Y. Terashima;K. Shimamoto
缺血预适应(PC)抑制心肌细胞在缺血时通过缝隙连接(GJS)的化学偶联，这是一种辅助的保护机制。本研究的目的是研究蛋白激酶在PC诱导的GJ调节中的作用。在大鼠离体心中，分别在缺血前和缺血后分别采集心肌组织标本，分离插入的富含圆盘的组份，进行免疫沉淀和免疫印迹。脑缺血后蛋白激酶C(PKC)-epsilon、p38丝裂原活化蛋白激酶(MAPK)-α及与连接蛋白-43(Cx43)共沉淀的Src表达增加，而Cx43免疫共沉淀物中未检测到p38MAPKβ。PC不改变缺血后Cx43-Src复合体的水平。PC可促进Cx43-PKCepsilon复合体的形成，该复合体可被PKCepsilon转位抑制肽(TIP)所阻断。相反，PC可降低缺血后Cx43免疫沉淀物中Cx43-p38MAPKalpha复合体水平和P38MAPK活性。用p38MAPK抑制剂SB-203580模拟PC对Cx43-p38MAPKalpha相互作用的影响。PC可降低心肌缺血25min时GJS对荧光黄的通透性，这种作用可被PKCepsilon-TIP所阻断。SB-203580在缺血15min时较未处理的对照组增加，但在缺血后25min时差异不显著。综上所述，PC对缺血时GJ Cx43与PKCepsilon、p38MAPKalpha、Src的相互作用有明显影响。PC对缺血时GJ通透性的抑制主要是通过增强Cx43与PKCepsilon的相互作用来实现的，这一作用压倒了Cx43-p38MAPKalpha相互作用减弱的平衡作用。
Ischemic preconditioning (PC) suppresses chemical coupling of cardiomyocytes via gap junctions (GJs) during ischemia, which is an adjunct mechanism of protection. The aim of this study was to characterize roles of protein kinases in PC-induced GJ modulation. In isolated rat hearts, ventricular tissues were sampled before and after ischemia with or without PC, and intercalated disc-rich fractions were separated for immunoprecipitation and immunoblotting. Levels of protein kinase C (PKC)-epsilon, p38mitogen-activated protein kinase (MAPK)-alpha, and Src coimmunoprecipitated with connexin-43 (Cx43) were increased after ischemia, whereas p38MAPKbeta was not detected in the Cx43 immunoprecipitates. PC did not modify the level of Cx43-Src complex after ischemia. However, PC enhanced Cx43-PKCepsilon complex formation, which was abolished by PKCepsilon translocation inhibitory peptide (TIP). In contrast, PC reduced Cx43-p38MAPKalpha complex level and p38MAPK activity in the Cx43 immunoprecipitates after ischemia. The effect of PC on Cx43-p38MAPKalpha interaction was mimicked by SB-203580, a p38MAPK inhibitor. PC reduced permeability of GJs to Lucifer yellow in the myocardium at 25 min after ischemia, and this effect was abolished by PKCepsilon-TIP. SB-203580 increased the GJ permeability at 15 min after ischemia compared with that in untreated controls, but the difference became insignificant 25 min after ischemia. In conclusion, PC has distinct effects on interaction of GJ Cx43 with PKCepsilon, p38MAPKalpha, and Src during ischemia. Suppression of GJ permeability during ischemia by PC is primarily achieved by enhanced interaction of Cx43 with PKCepsilon, which overwhelms the counterbalancing effect of reduced Cx43-p38MAPKalpha interaction.