Louie Nathaniel Pinpin

The cardiac conduction system (CCS) consists of specialized cells that coordinate the heart's rhythmic contraction. The CCS is composed of multiple components, including the sinoatrial node (SAN), or cardiac pacemaker, which initiates the electrical depolarization of each heartbeat. The molecular regulators underlying the CCS remain relatively unknown due to: (1) the relatively low number of CCS cells; (2) its complex three-dimensional shape; and (3) the heterogeneity of cell types within CCS substructures, including the SAN.

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As a result, in vitro model systems have long been sought after to better interrogate the development, function, and disease processes of the SAN. However, currently, there are limited protocols for differentiating human induced pluripotent stem cell-derived SAN-like cardiomyocytes (hiPSC-SANCMs), with existing protocols yielding at most 10-15% of pure nodal-like cells. To address this unmet need, our objective is to develop an improved hiPSC-SANCM differentiation protocol as well as a novel method for purification of SANCMs. Specifically, to improve nodal differentiations, we will optimize existing hiPSC-SANCM protocols by testing a series of small molecules that alter known molecular pathways of SAN development. To purify SANCMs, we will use CRISPR-Cas9 to generate an hiPSC line that has antibiotic resistance driven by the promoter of an established SAN-specific gene. Improved differentiation and purification will be validated by single-cell RNA sequencing and electrophysiology studies, using miceroelectrode array plates. Through these efforts, we aim to generate a pure population of SAN-like cells that can subsequently be used for elucidating SAN function, disease modeling, and even developming novel therapeutics.