Novel Evidence That Human Umbilical Cord Blood-Purified CD133+ cells Secrete Several Soluble Factors and Microvesicles/Exosomes That Mediate Paracrine, Pro-Angiopoietic Effects Of These Cells – Implications For and Important Role Of Paracrine Effects in stem Cell Therapies In Regenerative Medicine

• Published in: Blood 2013-11, Vol.122 (21), p.1216-1216
• Main Authors: Ratajczak, Janina, Mierzejewska, Kasia, Borkowska, Sylwia, Kucia, Magdalena, Ratajczak, Mariusz Z
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V122.21.1216.1216

Populations of CD34+, CD34+CXCR4+, and CD133+ cells are currently employed in the clinic to treat damaged organs (e.g., heart after myocardial infarction [AMI]). These cells are highly enriched, primarily for hematopoietic stem/progenitor cells (HSPCs), and for many years it has been wrongly supposed that HSPCs can trans-dedifferentiate into tissue-specific cells. However, even when improvement of organ function is observed after employing these cells in therapy, the lack of a convincing demonstration of significant donor-recipient chimerism in treated tissues in most of the studies performed indicates that mechanisms other than trans-dedifferentiation play a significant role in positive clinical outcomes. In support of this conclusion, we have already reported that CD34+ cells secrete a variety of growth factors, cytokines, chemokines, and bioactive lipids that interact with the surrounding microenvironment (Blood 2001;97:3075). Furthermore, microvesicles (MVs) or exosomes shed from the cell surface or derived from the intracellular membrane compartment (respectively) are important mediators in cell-to-cell communication and, as we demonstrated, may affect the biology of target cells by horizontal transfer of mRNA and proteins (Leukemia 2006;20:847). We hypothesized that some reported positive outcomes in adult stem cell therapies (e.g., when employing CD133+ cells) can be explained by the paracrine effects of these cells, involving both soluble factors as well as cell membrane-derived MVs. CD133+ cells were purified from UCB (>95% purity as checked by FACS) and incubated for 24 hours in RPMI at 37°C in a small volume of medium supplemented with 0.5% albumin. Subsequently, we harvested conditioned media (CM) from these cells and isolated CD133+ cell-shed microvesicles (MVs) by high-speed centrifugation. We then employed sensitive ELISA assays to measure the concentration of important pro-angiopoietic and anti-apoptotic factors in CD133+ cell-derived CM and isolated mRNA from both CD133+ cells and CD133+ cell-derived MVs for RQ-PCR analysis of gene expression. Subsequently, the chemotactic activity of CD133+ cell-derived CM and MVs was tested against human UCB endothelial cells (HUVECs), and in parallel we tested whether CD133+ cell-derived CM and MVs induce major signaling pathways in HUVECs. Finally, in in vitro functional assays, we tested the ability of CD133+ cell-derived CM and MVs to induce tube formation by HUVECs and the ability of in vivo Matrige