Date of Award

5-2022

Document Type

Thesis

Degree Name

Master of Science in Biology

Department

Biology

First Reader/Committee Chair

Bournias-Vardiabasis, Nicole

Abstract

The utilization of cardiac progenitor cells (CPC) has been shown to induce favorable regenerative effects. While there are various populations of endogenous CPCs in the heart, there is no consensus regarding which population is the most ideal for cell-based regenerative therapy. Identifying an Islet-1+ (Isl-1+) early-stage progenitor population with enhanced stemness, multipotency and differentiation potential would be beneficial for regenerative therapy. Spalt-like transcription factor 4 (SALL4) plays a role in embryonic development as well as proliferation and expansion of hematopoietic progenitor cells. We hypothesize that SALL4 will be co-expressed in Isl-1+ cardiac progenitor cell clones isolated from human cardiac tissue and represent a pre-mesendodermal progenitor population. Ingenuity Pathway Analysis revealed Isl-1+ human neonates exhibit enhanced stemness properties compared to Isl-1+ adult CPCs. We compared RNA-seq based transcriptomic data from human neonatal Isl-1+ CPC clones with that of published time-course specific RNA-Seq based transcriptomic data collected from various stages of cardiac differentiation from human pluripotent stem cells (hPSCs). This approach elucidated genes that are highly expressed at different stages of cardiac development giving insight into the developmental state of SALL4 expressing Isl-1+ neonatal CPCs. In addition to SALL4, evidence suggests that SOX2, EpCAM and TBX5 are expressed early along the cardiovascular pathway. CPCs were previously derived from human cardiac tissue discarded at surgery, clonally expanded, and screened for expression of Isl-1. RNA was isolated from individual human neonatal (n=10) Isl-1+ CPCs, cDNA was synthesized, and real time PCR was done. We utilized RT-PCR to identify the expression of SALL4, SOX2, EpCAM and TBX5 in individual neonatal Isl-1+ CPC clones. Gel electrophoresis of the PCR products was used to confirm that transcripts of the correct size were amplified. Results demonstrate that 9 out of 10 Isl-1+ neonatal CPC clones tested expressed SALL4. To further substage neonatal Isl-1+ CPC clones that expressed SALL4, SOX2, EpCAM and TBX5, clones were tested for expression of TFAP2C. These features will allow for the identification and isolation of an optimal early-stage CPC clone that may be of value in regenerative therapeutic applications.

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