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Differential gene expression of extracellular matrix components in dilated cardiomyopathy
S C Tyagi1, S Kumar, D J Voelker
1Department of Medicine, Dalton Cardiovascular Research Center, University of Missouri-Health Sciences Center, Columbia 65212, USA.
Abstract:
Extracellular matrix metalloproteinases (MMPs) are activated in dilated cardiomyopathic (DCM) hearts [Tyagi et al. (1996): Mol Cell Biochem 155:13-21]. To examine whether the MMP activation is occurring at the gene expression level, we performed differential display mRNA analysis on tissue from six dilated cardiomyopathy (DCM) explanted and five normal human hearts. Specifically, we identified three genes to be induced and several other genes to be repressed following DCM. Southern blot analysis of isolated cDNA using a collagenase cDNA probe indicated that one of the genes induced during DCM was interstitial collagenase (MMP-1). Northern blot analysis using MMP-1 cDNA probe indicated that MMP-1 was induced three- to fourfold in the DCM heart as compared to normal tissue. To analyze posttranslational expression of MMP and tissue inhibitor of matrix metalloproteinase (TIMP) we performed immunoblot, immunoassay, and substrate zymographic assays. TIMP-1 and MMP-1 levels were 37 +/- 8 ng/mg and 9 +/- 2 ng/mg in normal tissue specimens (P < 0.01) and 2 +/- 1 ng/mg and 45 +/- 11 ng/mg in DCM tissue (P < 0.01), respectively. Zymographic analysis demonstrated lytic bands at 66 kDa and 54 kDa in DCM tissue as compared to one band at 66 kDa in normal tissue. Incubation of zymographic gel with metal chelator (phenanthroline) abolished both bands suggesting activation of neutral MMP in DCM heart tissue. TIMP-1 was repressed approximately twentyfold in DCM hearts when compared with normal heart tissue. In situ immunolabeling of MMP-1 indicated phenotypic differences in the fibroblast cells isolated from the DCM heart as compared to normal heart. These results suggest disruption in the balance of myopathic-fibroblast cell ECM-proteinase and antiproteinase in ECM remodeling which is followed by dilated cardiomyopathy.
Insights
Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) are key in dilated cardiomyopathy (DCM). MMP-1 is upregulated, while TIMP-1 is downregulated in DCM hearts, indicating ECM remodeling disruption.
Area of Science:
- Cardiovascular Biology
- Molecular Cardiology
- Extracellular Matrix Biology
Background:
- Extracellular matrix metalloproteinases (MMPs) are implicated in dilated cardiomyopathy (DCM).
- Activation of MMPs in DCM hearts suggests a role in disease pathogenesis.
- Understanding gene expression changes is crucial for DCM research.
Purpose of the Study:
- To investigate MMP activation at the gene expression level in DCM.
- To identify specific MMPs and their inhibitors involved in DCM.
- To elucidate the role of fibroblast cells in ECM remodeling in DCM.
Main Methods:
- Differential display mRNA analysis of human heart tissues.
- Southern and Northern blot analyses for gene expression.
- Immunoblot, immunoassay, and zymography for protein analysis.
- In situ immunolabeling of MMP-1 in fibroblast cells.
Main Results:
- MMP-1 gene expression and protein levels were significantly increased in DCM hearts.
- TIMP-1 levels were markedly decreased in DCM hearts.
- Zymography revealed activated neutral MMPs in DCM tissue, with altered fibroblast phenotypes.
Conclusions:
- DCM involves a significant imbalance between MMPs and TIMPs, leading to ECM remodeling.
- Upregulation of MMP-1 and downregulation of TIMP-1 contribute to the pathogenesis of DCM.
- Altered fibroblast behavior in DCM impacts extracellular matrix homeostasis.