Prospective Surface Marker-Based Isolation and Expansion of Fetal Endothelial Colony-Forming Cells From Human Term Placenta

Abstract

The term placenta is a highly vascularized tissue and is usually discarded upon birth. Our objective was to isolate clinically relevant quantities of fetal endothelial colony-forming cells (ECFCs) from human term placenta and to compare them to the well-established donor-matched umbilical cord blood (UCB)-derived ECFCs. A sorting strategy was devised to enrich for CD45-CD34+CD31Lo cells prior to primary plating to obtain pure placental ECFCs (PL-ECFCs) upon culture. UCB-ECFCs were derived using a well-described assay. PL-ECFCs were fetal in origin and expressed the same cell surface markers as UCB-ECFCs. Most importantly, a single term placenta could yield as many ECFCs as 27 UCB donors. PL-ECFCs and UCB-ECFCs had similar in vitro and in vivo vessel forming capacities and restored mouse hind limb ischemia in similar proportions. Gene expression profiles were only minimally divergent between PL-ECFCs and UCB-ECFCs, probably reflecting a vascular source versus a circulating source. Finally, PL-ECFCs and UCB-ECFCs displayed similar hierarchies between high and low proliferative colonies. We report a robust strategy to isolate ECFCs from human term placentas based on their cell surface expression. This yielded much larger quantities of ECFCs than UCB, but the cells were comparable in immunophenotype, gene expression, and in vivo functional ability. We conclude that PL-ECFCs have significant bio-banking and clinical translatability potential.

Avoidance of Maternal Cell Contamination and Overgrowth in Isolating Fetal Chorionic Villi Mesenchymal Stem Cells from Human Term Placenta

“…The CD34+CD31+CD45− population comprised the placental endothelial colony forming cells (PL‐EPC). Thus, the isolation of placental EPC in EGM2 + 10 led serendipitously to the method previously reported by our group for isolating fetal and maternal MSC from human term placental villi . This CD34+ method was being carried out concurrently but independently to the experiments described in the current manuscript.…”

Concise Review: Functional Definition of Endothelial Progenitor Cells: A Molecular Perspective

“…Importantly, ECFCs have demonstrated a single-cell self-renewal capacity and form characteristic high-proliferativepotential (HPP) colonies, with each having a significant capacity for long-term culture (.15 passages) and expansion of cells with clear characteristics of endothelial cells [14]. ECFCs can be readily isolated from adult and fetal (umbilical cord blood) circulation but also in greater quantities from vascular structures, particularly those of the placenta, as our group has previously demonstrated [14][15][16][17][18][19] (Fig. 1).…”

“…Assessing candidate EPC populations by quantification has also been attempted in pathologies such as diabetes, preeclampsia, and bronchopulmonary dysplasia [42][43][44][45][46]. Since their discovery in 2004, there has been a growing body of evidence to suggest that ECFCs play a role in regenerating vascular structures and can form de novo vessels when injected into an ischemic site [15,[47][48][49]. To this end, although methods have been put forward in attempting to functionally define EPCs, none of these methods provides a consensus across the field.…”

Vasculogenesis and Its Cellular Therapeutic Applications

“…Rat Ratajska et al Cells Tissues Organs 2017;203:141-152 DOI: 10.1159/000448551 146 (and human) pulmonary microvascular ECs contain a subset of CD31 + cells of high proliferative and vasculogenic activity which form perfused vessels when implanted on Matrigel plugs in vivo [Schniedermann et al, 2010;Basile and Yoder, 2014;Alphonse et al, 2015]. These cells are more numerous than the EPCs found in the rat pulmonary artery [Lin et al, 2013;Patel et al, 2013;Alphonse et al, 2014Alphonse et al, , 2015Basile and Yoder, 2014]. It is assumed that EPCs are easily mobilized from the BM or the CB to these niche locations in solid tissues [Hirschi et al, 2008].…”

“…Another source of tissue EPCs is the microvessels of the lung, placenta and white adipose tissue [Lin et al, 2013;Patel et al, 2013;Alphonse et al, 2014Alphonse et al, , 2015. Rat Ratajska et al Cells Tissues Organs 2017;203:141-152 DOI: 10.1159/000448551 146 (and human) pulmonary microvascular ECs contain a subset of CD31 + cells of high proliferative and vasculogenic activity which form perfused vessels when implanted on Matrigel plugs in vivo [Schniedermann et al, 2010;Basile and Yoder, 2014;Alphonse et al, 2015].…”