Fibrosis is the outcome of excess deposition of extracellular matrix components and underlies diastolic and systolic heart failure. Metalloproteinases (MMPs and ADAMs) and their inhibitors (TIMPs) are critical regulators of ECM turnover and fibrosis. TIMP3 is a potent inhibitor of MMPs and ADAMs, whereby it regulates tissue proteolysis and cytokine bioavailability. TIMP3 levels are reduced in human heart disease, and we found that mice lacking Timp3 (KO) exhibit severe dilated cardiomyopathy (DCM), extensive interstitial fibrosis and early heart failure (HF) following pressure overload compared to wild type (WT) mice. We showed upregulated TNF signaling within 6 hrs of aortic banding (AB) which when blocked, significantly improved DCM and prevented HF. We then investigated the mechanism underlying the extensive fibrosis despite the increased MMP activity in the TIMP3KO mice. Comparative microarray analyses show a significant upregulation of the TGFβ1 signaling pathway within 6 hrs of AB in KO vs WT hearts.
In vivo
treatment of KO-AB mice with a TGFβ1-neutralizing antibody (1D11) completely abolishes fibrosis, markedly improves left ventricular dilation, systolic and diastolic heart function (LVEDD(mm): 4.3 ± 0.1 in WT, 5.4 ± 0.1 in KO, 4.7 ± 0.1 in KO+ 1D11; fractional shortening(%): 40.2 ± 1.7 in WT, 19.6 ± 2.2 in KO, 36.2 ± 0.7 in KO+ 1D11; E/Ac ratio: 2.5 ± 0.3 in WT, 1.8 ± 0.3 in KO, 2.4 ± 0.2 in KO + 1D11; n = 22, P < 0.05). Interestingly, blocking TGFβ1 also prevented the early rise in TNF in KO hearts. Using primary neonatal culture systems, we found that cardiomyocyte-fibroblast interaction is essential for a fibrotic response to agonists (Ang II, PE). Further, KO co-cultures generate a 5-fold greater fibrosis (P < 0.05) through activation of Smad2/3 pathway compared to WT. TGFβ also regulates TNF transcription more potently in KO cells since a 3-fold higher TNF induction occurred in response to recombinant TGFβ1 in KO vs WT co-cultures (P < 0.05). In conclusion, TIMP3 is a critical regulator of TGFβ1 and TNF, and its deficiency induces parallel dysregulations in the signaling of these cytokines very early after pressure overload, leading to severe interstitial fibrosis and DCM, hence, TIMP3-based therapies can simultaneously impact fibrosis and DCM.
Cardiac Insulin-Resistance and Decreased Mitochondrial Energy Production Precede the Development of Systolic Heart Failure After Pressure-Overload Hypertrophy
