Submitted on May 7, 2008
Revised on January 21, 2009
Accepted on February 5, 2009
Tissue inhibitor of metalloproteinase-3 inhibits neonatal mouse cardiomyocyte proliferation via EGFR/JNK/SP-1 signaling
Lamis Hammoud1, Dylan Burger1, Xiangru Lu2, and Qingping Feng1*
1 University of Western Ontario
2 Lawson Health Research Institute
* To whom correspondence should be addressed. E-mail: qfeng{at}uwo.ca.
We have recently demonstrated that tissue inhibitor of metalloproteinase-3 (TIMP-3) decreases neonatal cardiomyocyte proliferation. The aim of this study was to delineate a pathway through which TIMP-3 exerts its anti-proliferative effect. Experiments were conducted on neonatal cardiomyocyte cultures and heart tissues isolated from wild-type (WT) and TIMP-3-/- mice. Deficiency in TIMP-3 decreased p27 expression and increased cardiomyocyte proliferation in cardiomyocytes and neonatal hearts. A TIMP-3/epidermal growth factor receptor (EGFR)/Jun N-terminal kinase (JNK)/SP-1/p27 pathway was investigated. JNK phosphorylation and EGFR protein levels were increased in TIMP-3-/- cardiomyocytes and heart tissues. Treatment with recombinant TIMP-3 decreased JNK phosphorylation and EGFR expression/phosphorylation. Inhibition of JNK activity using SP600125 decreased SP-1 phosphorylation, increased p27 expression, and decreased cardiomyocyte proliferation. Furthermore, treatment with the EGFR specific inhibitor PD168393 or the EGF neutralizing antibody decreased cardiomyocyte proliferation as well as phosphorylation of JNK and SP-1 in both WT and TIMP-3-/- cardiomyocytes. We conclude that TIMP-3 inhibits neonatal mouse cardiomyocyte proliferation by upregulating p27 expression. The effects of TIMP-3 are mediated via inhibition of EGFR expression/phosphorylation, and decreases in JNK and SP-1 signaling.
