Characterization of Human Cardiac Progenitor Cell Secretome

Abstract

Heart Disease (HD) remains the greatest cause of death worldwide. Due to the heart’s poor capacity to regenerate after damage, there is a critical need for treatments that can restore cardiac cell integrity and function in HD patients. Preclinical and clinical evidence has highlighted the potential role of Cardiac Progenitor Cells (CPCs) in regulation of cardiac repair after injury, with an emerging role of CPC secreted proteins in this process. We have been studying CPCs isolated from human left ventricular myocardium characterized by their high proliferative capacity in vitro. These cells (human highly proliferative clones, hHiPC) are characterized by high expression of CD105/Endoglin and show varying potential for differentiating into endothelial cells in vitro. When injected into the mouse myocardium after surgically induced myocardial infarction (MI), individual hHiPC clones show variable effects on heart remodeling. For instance, when clone 11 was injected into the infarct zone post‐MI we saw a trend towards improvement in the fractional shortening (%) compared to clone 22 and saline injected controls. We hypothesized that hHiPCs associated with improvement in cardiac remodeling after MI will have a secretome characterized by higher expression of pro‐angiogenic and pro‐proliferative proteins, and these are regulated by the BMP/CD105 pathway. An unbiased and global proteomics approach (LC‐MS/MS) was used in our lab to identify proteins secreted from hHiPCs pre‐treated with the CD105 ligand BMP9 (5 ng/mL) compared to non‐treated controls. Our secretome data identified ~700 proteins in which 57% were characterized from UniProt as having a signal peptide sequence. Several pro‐angiogenic secreted proteins were identified in clone 11 including CCN2, CXCL6, and TGFB2. We found a >2.5‐fold significant increase in the secretion of Sclerostin (SOST) in BMP9 treated hHiPC compared to non‐treated controls. SOST is a known BMP‐target protein and WNT inhibitor. Previous evidence has highlighted the role of WNT‐signaling in heart remodeling after myocardial induced injury, but the role of Sclerostin in this process has yet to be elucidated. To further characterize the relationship between BMP9 and the secretion of SOST we performed a Real‐Time q‐PCR and ELISA assay of cell lysates and conditioned media between BMP9 treated and non‐treated controls and found a dramatic increase in the expression of SOST in BMP9 treated clone 11. We also found an increase (4.5‐fold) in CD105 mRNA expression after BMP9 treatment. Similarly, using LC‐MS/MS we found a significant increase in CD105 expression (2.5‐fold, p‐value = 7.39x10‐6) in the membrane fraction of clone 11 compared to clone 22 indicating its potential role in positive heart remodeling effects. Future studies will be conducted in vivo to determine the role SOST/WNT signaling has in heart remodeling after ischemic injury. Further, the increase in pro‐angiogenic factors after BMP9 treatment may suggest the potential relevance of BMP9 treatment in cardiac progenitor cell secretome mediated repair and we will test this hypothesis in the future.