Cardiac regeneration offers great promise for repair of injured myocardium and heart failure (HF), however it is not yet reality. This is due to cells not truly regenerating the heart as replacement myocytes. However, some benefit appears to occur in the post-ischemic heart with different stem cell injections, and data supports paracrine effects. Recent advances have suggested that the paracrine effects for any improvement in heart function after stem cell delivery is due to secreted exosomes from these cells. Exosomes are small vesicles containing specific cargo and it appears that they can carry potentially therapeutic molecules including growth factors and microRNAs (miRs). Data from our lab over the last two decades has implicated G protein-coupled receptor (GPCR) kinases (GRKs), especially GRK2, as key molecules in HF development following cardiac injury. Further, GRK2, which is up-regulated in failing human hearts, has emerged as a potential novel therapeutic target in HF. GRK2 has important effects on cardiac ?-adrenergic receptor (?AR) signaling that has crucial implications in HF. Importantly, inhibition of GRK2 with a peptide inhibitor (?ARKct) or knock-down of its expression leads to significant prevention and rescue of animal models of HF and preliminary data presented in this proposal shows that mechanisms of this repair includes increased markers of myocyte regeneration. Thus, we are interested in whether GRK2 or ?AR manipulation in myocytes or cardiac precursor cells (CPCs) may affect reparative or regenerative properties of the injured heart including examining paracrine, autocrine or exosome-based properties. We have started to investigate whether manipulation of GRK2 activity might alter exosomes and their cargo from either myocytes or CPCs, and whether exosomes from CPCs have improved regenerative properties. Indeed, we have recently found that introduction of the ?ARKct into CPCs improves survival and metabolism of these cells. ?ARKct-containing CPCs, as mediators of cardiac regeneration, will be employed to investigate whether manipulation of ?AR signaling and GRK2 activity may influence exosome-mediated cardiac repair in vivo. We will use mouse models and also move to the pig in order to carry out pre-clinical studies with the goal of future clinical translation. Specifically, the Central Hypothesis of this proposal is that inhibition or lowering of GRK2 in cardiac stem/precursor cells and/or cardiac myocytes improves myocardial repair through mechanisms that include favorable alterations of the content of secreted exosomes. This may include direct enhancement of ?-adrenergic signaling that improves survival and proliferation of these cells to positively affect cardiac regeneration.
Specific Aims are:  To characterize the content of exosomes secreted from CPCs and myocytes with altered GRK2 expression and activity that can lead to changes in ?AR signaling, and to determine if altered exosome properties/cargo are responsible for the improved survival, metabolism and regenerative capacity of these cells;  To determine whether inhibiting GRK2 in CPCs improves exosome-based in vivo myocardial repair/regeneration and whether specific miR cargo in these GRK2-altered exosomes are involved in any therapeutic process;  To determine whether CPC exosomes from cells with lower GRK2 activity and ?ARKct-specific miR cargo enhance myocardial repair in a large animal, pre-clinical model of ischemic injury.
Project 2 (Koch) Narrative (relevance): Elucidating novel roles for GRK2 and b-adrenergic regulation of secreted exosomes from myocytes or cardiac precursor cells will lead to a broader understanding of how GRK2 may be involved in cardiac regeneration. This may lead to innovative molecular treatments for cardiac repair of the post-ischemic and failing heart including exosomes loaded with GRK2-specific microRNAs and also the ?ARKct as a GRK2 inhibitor. Working with this PPG group will allow for robust study of exosome-mediated cardiac repair and the complementing targets have the potential for clinical translation. Importantly, elucidating these targets and mechanisms will lead to a broader understanding of the pathogenesis of myocardial dysfunction, which may provide targets for new therapeutic strategies to combat heart failure.
|Sluijter, Joost Petrus Gerardus; Davidson, Sean Michael; Boulanger, Chantal M et al. (2018) Extracellular vesicles in diagnostics and therapy of the ischaemic heart: Position Paper from the Working Group on Cellular Biology of the Heart of the European Society of Cardiology. Cardiovasc Res 114:19-34|
|Garikipati, Venkata Naga Srikanth; Kishore, Raj (2018) Induced Pluripotent Stem Cells Derived Extracellular Vesicles: A Potential Therapy for Cardiac Repair. Circ Res 122:197-198|
|Cheng, Zhongjian; Shen, Xinggui; Jiang, Xiaohua et al. (2018) Hyperhomocysteinemia potentiates diabetes-impaired EDHF-induced vascular relaxation: Role of insufficient hydrogen sulfide. Redox Biol 16:215-225|
|Gupta, Rajesh; Mackie, Alexander R; Misener, Sol et al. (2018) Endothelial smoothened-dependent hedgehog signaling is not required for sonic hedgehog induced angiogenesis or ischemic tissue repair. Lab Invest 98:682-691|
|Garikipati, Venkata Naga Srikanth; Kishore, Raj (2017) Cardiac progenitor cells: old is not always gold. J Physiol 595:6221-6222|
|Yue, Yujia; Garikipati, Venkata Naga Srikanth; Verma, Suresh Kumar et al. (2017) Interleukin-10 Deficiency Impairs Reparative Properties of Bone Marrow-Derived Endothelial Progenitor Cell Exosomes. Tissue Eng Part A 23:1241-1250|