Cellular Mechanisms of Cardiac ECM Structure and Function

Chronic heart failure (CHF) has a designated Quality Enhancement Research Initiative (QUERI) in the VA
system to address ways to improve cardiovascular healthcare for Veteran’s suffering from CHF. Success in
treatment of CHF associated with chronic pressure overload (hypertension, aortic valve stenosis) is limited by
the presence of persistent interstitial fibrosis despite our ability to normalize hemodynamic load. The
proposed studies will define abnormalities in cellular mechanisms that cause this critical clinical unmet need.
Filling this need will depend on defining the fundamental causal determinants that control both initial ECM
degradation and persistence of interstitial myocardial fibrosis following normalization of hemodynamic load.
Primary cellular regulators of ECM homeostasis are postulated to be myocardial macrophages and fibroblasts.
Our previous studies and preliminary data have led to our central hypothesis: Chronic hemodynamic
overload causes fundamental changes in both macrophage and fibroblast phenotype, the hallmark of which is
dysregulated protease homeostasis that in turn impedes cellular response to unloading and limits complete
regression of fibrosis even after normalization of hemodynamic load. To test this hypothesis, innovations in in
vivo animal models and in vitro fibroblast culture were developed. In vivo, a clinically relevant reversal of LVPO
(unloading) was created in mice by surgical removal of the transverse aortic constriction (unTAC). UnTAC was
found to initiate but lead to an incomplete regression of cardiac fibrosis. Preliminary data indicate that a
significant increase in myocardial macrophages coincides with initiation of collagen degradation following
hemodynamic unloading but these increases in macrophages are not sustained at later times after unTAC. To
address whether load-dependent changes in fibroblast phenotype were a key factor in this remodeling, a
fibroblast culture systems that mimics clinically relevant myocardial stiffness was established. In vivo,
measurements of myocardial stiffness demonstrated that fibroblasts are exposed to a force of ~8 kPA in PO
myocardium and ~2 kPA in normal myocardium. Physiologically relevant, stiffness-dependent changes in
fibroblasts phenotype were observed in fibroblasts from normal myocardium whereas fibroblasts from TAC and
unTAC myocardium exhibited a pro-fibrotic non-responsive phenotype to changes in stiffness. Preliminary data
indicate that Tissue inhibitor of metalloproteinase (TIMP)-1 was a causal factor in this pro-fibrotic persistent
phenotype. Our strong preliminary data gave rise to the following Specific Aims to test our central hypothesis:
Aim 1: Test the hypothesis that reversal of sustained hemodynamic overload shifts myocardial
macrophage phenotype to a distinct but transient anti-fibrotic (ECM-degradation) phenotype that
initiates, but does not complete, a load-dependent regression of accumulated interstitial ECM.
Aim 2: Test the hypothesis that sustained in vivo increases in hemodynamic load change myocardial
fibroblasts to a profibrotic, TIMP-1 dependent phenotype that remains profibrotic even when
hemodynamic load is reversed.

Grant Number: 5I01BX005943-03
NIH Institute/Center: VA
Principal Investigator: Amy Bradshaw