Project 1 - Endogenous and Exogenous Mechanisms that Promote Myocardial Remuscularization

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SUMMARY / ABSTRACT (Project 1) Endogenous and Exogenous Mechanisms that Promote Myocardial Remuscularization in Postinfarction LV Remodeling The molecular and cellular basis for progressive heart failure is the result of the inability of damaged and apoptotic myocytes to be replaced. While a number of cell- and tissue-based therapies can limit this dysfunction, the proportion of cells that survive at the site of administration for more than a few weeks after transplantation is extremely low. As such, substantial remuscularization of the infarcted region has rarely been reported; and when limited remuscularization has been reported, it is frequently accompanied by potentially lethal ventricular arrhythmias of unknown mechanism. This proposal aims at remuscularization of the injured ventricle by the definition of key endogenous factors that regulate and promote the cell cycle of the native cardiomyocyte (CM), and from exogenous transplanted bioengineered cardiac muscle patch (hCMP) that overexpresses key regulators of CM cell cycle, and will incorporate a functional vascular network and recapitulate some of the key micro environmental cues of native heart tissue. We recently established a novel hiPSC cell line with MHC-driven overexpression of a key regulator of CMs: CCND2 (hiPSC-MHC-CCND2OE), which can remuscularize injured ventricle in rodent model. The central objective of this proposal is to “turn back the clock” of the myocyte cell cycle, which will facilitate myocardial repair. The Specific Aims (SA) that will examine this objective include: SA1: To identify the key regulators that promote cell-cycle activity in the hearts of early neonatal pigs after myocardial injury. We will: 1) using state-of-the-art fate-mapping molecular biology and imaging technologies, and the single cell/nucleus RNA sequencing (scRNAseq or snRNAseq) technology demonstrate the key regulators/signaling pathways that govern the myocyte cell cycle; and 2) test the remuscularization of the injured ventricle by manipulating the key regulators using either targeted modRNA or AAV9 delivery strategies to selectively modify these regulators in adult pig hearts following ischemic injury. SA2a. To engineer hCMPs containing CMs that are capable of proliferating after transplantation, and characterized by previously unattainable size and thickness that are functionally mature and primed for in-vivo vascularization. SA2b. To evaluate the effectiveness of our hCMP constructs for myocardial recovery and remuscularization in a large-animal (pig) model of myocardial injury. We will use state-of-the-art techniques that includes optical mapping in combination with the 3- dimensional intramural cardiac mapping to delineate the potential arrhythmia mechanisms over the entire transmural and left ventricular surface. These studies will synergize with the other projects and serve as a prelude for therapeutic initiatives focused on remuscularization of the injured human heart.