The applicants' long-term aim is to gain insights into mechanisms and therapies of heart failure (HF). In this study, we will examine the way that cellular architecture and calcium (Ca) handling proteins involved in excitation-contraction coupling are modified in human HF. Further studies will be performed on patients that undergo implantation of left ventricular assist devices (LVADs) to establish (1) a microstructural basis for functional remodeling in HF cells, and (2) if microstructure of HF cells can predict long-term sustainability of cardiac recovery by LVAD unloading. Our studies are motivated by the finding that nearly 20% of our patients respond to unloading with cardiac recovery and that recovery in patients required integrity of the transverse tubular system (t-system) before unloading.
In Specific Aim 1, three-dimensional reconstructions of the t- system and associated ryanodine receptor (RyR) clusters will be obtained with scanning confocal microscopy. We will investigate ventricular tissues from normal donor hearts and hearts from patients in end stage HF. The studies are based on the hypothesis that structures in control myocytes are altered in HF cells. We found that human cells exhibit a very different phenotype in HF than previously described in animal models. In particular, we found that the t-system remodels to axial, sheet-like invaginations of the sarcolemma in HF. We will then establish the extent to which RyR clusters are not associated with sarcolemma (primarily t-system) for control and HF cells. Findings in intact tissue will be compared with isolated cells.
In Specific Aim 2, we will study the changes in Ca transients that produce contractions as a consequence of alterations in the t-system and proteins associated with EC coupling. This includes measuring Ca movements in normal and HF cells with rapid two-dimensional confocal microscopy. We will test the hypothesis that Ca transients in HF are disorganized compared to control. We will measure L-type Ca currents and calculate the gain of EC coupling. We expect that remodeling of the t-system leads to decreased gain in HF cells. Also, we will measure Ca extrusion from the cell and Ca uptake into the sarcoplasmic reticulum, and relate those to t-system remodeling.
In Specific Aim 3, we will test the hypothesis that alterations that we observe at the time of LVAD implantation predict cardiac recovery. Integrity of t-system and improvement of functional properties induced by LVAD unloading are expected to underlie cardiac recovery. We will investigate whether LVAD unloading can reverse effects of HF on protein densities, t-system, Ca release, gain of EC coupling and decline of Ca transients. Finally, we will test the hypothesis that microstructural alterations observed at time of LVAD implantation predict long-term sustainability of recovery in patients undergoing LVAD explantation. Together, the proposed studies constitute a crucial step towards understanding effects of human HF and LVAD unloading at the cellular level. Clinical applications of our results and conclusions include the identification of patients that are likely to be sustained responders based on cardiac biopsies that are acquired at time of LVAD implantation.
Left ventricular assist devices commonly serve as a bridge to transplantation for end stage heart failure patients. In this project we investigate if remodeling of microstructure and function of ventricular muscle cells in patients undergoing left ventricular assist device therapy can predict sustained cardiac recovery.
