Trapidil protects ischemic hearts from reperfusion injury by stimulating PKAII activity

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• 作者：Sichelschmidt ; Oliver J. ; Hahnefeld ; Claudia ; Hohlfeld ; Thomas ; Herberg ; Friedrich W. ; Schrö ; r ; Karsten
• 关键词：Ischemia ; Protein kinases ; Reperfusion ; SR (function)
• 刊名：Cardiovascular Research
• 出版年：2003
• 期刊代码：48_00086363
• 类别：med
• 出版时间：June 1, 2003
• 卷：58
• 期：3
• 页码：602-610
• 文件大小：222 K

摘要

Objective: The cardioprotective effects of trapidil on ischemic reperfused (I/R) rabbit hearts were studied. Recently, we had shown that trapidil might activate protein kinase A (PKA). In this study, we examined the exact mode of PKA stimulating activity of trapidil. Finally, we investigated the effect of trapidil on the phosphorylation state of phospholamban (PLB), a major PKA target in the heart and key regulator of Ca²⁺ sequestration via the sarcoplasmic reticulum Ca²⁺-ATPase. Methods: Langendorff-hearts of New Zealand White rabbits were perfused at constant volume and subjected to global low-flow ischemia for 2 h, followed by 1 h of reperfusion. Subsequently, hearts were used for Western blot analysis of PLB phosphorylation. Furthermore, three different regulatory subunits and one catalytic subunit of PKA were overexpressed in E. coli. These PKA subunits were purified and used in an in vitro assay system to test the impact of trapidil on PKA activities in the absence and presence of cAMP. Results: I/R resulted in a significant increase in left ventricular end-diastolic pressure and creatine kinase efflux in the hearts. Trapidil (10 μM) prevented these alterations. Using recombinant cAMP-free PKA isoforms, it was found that trapidil specifically stimulated PKAII but only did so in the presence of small amounts of added cAMP. Furthermore, the PKA-dependent ¹⁶Ser phosphorylation of PLB was markedly reduced in I/R. Trapidil largely normalized the ¹⁶Ser phosphorylation of PLB. Conclusions: The data demonstrate cardioprotective actions of trapidil in I/R and show a PKAII-dependent cAMP sensitizing effect of the compound. They also indicate PKA-dependent PLB phosphorylation as a target, suggesting an improved Ca²⁺ uptake by the sarcoplasmic reticulum. This action might be involved in the cardioprotective effects of trapidil.