Ca2+/calmodulin-dependent kinase II triggers cell membrane injury by inducing complement factor B gene expression in the mouse heart
Madhu V. Singh, … , Peter J. Mohler, Mark E. Anderson
Madhu V. Singh, … , Peter J. Mohler, Mark E. Anderson
Published March 9, 2009
Citation Information: J Clin Invest. 2009;119(4):986-996. https://doi.org/10.1172/JCI35814.
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Research Article Cardiology

Ca2+/calmodulin-dependent kinase II triggers cell membrane injury by inducing complement factor B gene expression in the mouse heart

Abstract

Myocardial Ca2+/calmodulin-dependent protein kinase II (CaMKII) inhibition improves cardiac function following myocardial infarction (MI), but the CaMKII-dependent pathways that participate in myocardial stress responses are incompletely understood. To address this issue, we sought to determine the transcriptional consequences of myocardial CaMKII inhibition after MI. We performed gene expression profiling in mouse hearts with cardiomyocyte-delimited transgenic expression of either a CaMKII inhibitory peptide (AC3-I) or a scrambled control peptide (AC3-C) following MI. Of the 8,600 mRNAs examined, 156 were substantially modulated by MI, and nearly half of these showed markedly altered responses to MI with CaMKII inhibition. CaMKII inhibition substantially reduced the MI-triggered upregulation of a constellation of proinflammatory genes. We studied 1 of these proinflammatory genes, complement factor B (Cfb), in detail, because complement proteins secreted by cells other than cardiomyocytes can induce sarcolemmal injury during MI. CFB protein expression in cardiomyocytes was triggered by CaMKII activation of the NF-κB pathway during both MI and exposure to bacterial endotoxin. CaMKII inhibition suppressed NF-κB activity in vitro and in vivo and reduced Cfb expression and sarcolemmal injury. The Cfb–/– mice were partially protected from the adverse consequences of MI. Our findings demonstrate what we believe is a novel target for CaMKII in myocardial injury and suggest that CaMKII is broadly important for the genetic effects of MI in cardiomyocytes.

Authors

Madhu V. Singh, Ann Kapoun, Linda Higgins, William Kutschke, Joshua M. Thurman, Rong Zhang, Minati Singh, Jinying Yang, Xiaoqun Guan, John S. Lowe, Robert M. Weiss, Kathy Zimmermann, Fiona E. Yull, Timothy S. Blackwell, Peter J. Mohler, Mark E. Anderson

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Figure 2

Cfb expression in heart.

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Cfb expression in heart. (A) RT-PCR analyses using RNA show PCR pro...
(A) RT-PCR analyses using RNA show PCR products for Cfb (137 bp). PCR amplification of hypoxanthine-guanine phosphoryltransferase (Hprt) was used as a positive control (165 bp). M, lanes containing the DNA size markers (bp). The + and – indicate RT reactions with or without the RT, respectively. (B) Immunoblotting for CFB protein expression in heart and liver. Tissue homogenates were immunoblotted for CFB. After immunoblotting the blot was stained with Coomassie blue. (C) Reduced expression of Cfb mRNA in post-MI AC3-I hearts compared with WT controls (n = 3). The qRT-PCR results were normalized to Gapdh mRNA expression. (D) Reduced CFB protein in post-MI AC3-I hearts compared with WT hearts. Homogenates from WT and AC3-I–infarcted hearts were immunoblotted for CFB (n = 3). Following immunoblotting, total protein on the blots was visualized by Coomassie staining. The ratio of the CFB band to the total protein in each lane was determined and is presented as mean ± SEM. (E) Reduced Cfb mRNA expression in isolated cardiomyocytes from post-MI AC3-I hearts. RNA was prepared from cardiomyocytes isolated from WT and AC3-I hearts 7 days after MI (n = 3) and qRT-PCR was performed. Results were normalized to Gapdh mRNA expression.