Umbilical cord blood-derived non-hematopoietic stem cells retrieved and expanded on bone marrow-derived extracellular matrix display pluripotent characteristics

Abstract

BackgroundUmbilical cord blood (UCB) not only contains hematopoietic stem cells (HSCs), but also non-hematopoietic stem cells (NHSCs) that are able to differentiate into a number of distinct cell types. Based on studies published to date, the frequency of NHSCs in UCB is believed to be very low. However, the isolation of these cells is primarily based on their adhesion to tissue culture plastic surfaces.Methods and resultsIn the current study, we demonstrate that this approach overlooks some of the extremely immature NHSCs because they lack the ability to adhere to plastic. Using a native extracellular matrix (ECM), produced by bone marrow (BM) stromal cells, the majority of the UCB-NHSCs attached within 4 h. The colony-forming unit fibroblast frequency of these cells was 1.5 × 104/108 mononuclear cells, which is at least 4000-fold greater than previously reported for UCB-NHSCs. The phenotype of these cells was fibroblast-like and different from those obtained by plastic adhesion; they formed embryonic body-like clusters that were OCT4-positive and expressed other human embryonic stem cell-related markers. Importantly, when implanted subcutaneously for 8 weeks into immunocompromised mice, these ECM-adherent and expanded NHSCs generated three germ layer-derived human tissues including muscle, fat, blood vessel, bone, gland, and nerve. Moreover, injection of these cells into muscle damaged by cryoinjury significantly accelerated muscle regeneration.ConclusionsThese results indicate that UCB may be a virtually unlimited source of NHSCs when combined with isolation and expansion on ECM. NHSCs may be a practical alternative to embryonic stem cells for a number of therapeutic applications.

Mesenchymal Stromal Cell-Produced Components of Extracellular Matrix Potentiate Multipotent Stem Cell Response to Differentiation Stimuli

“…Extracellular matrix production by MSCs is enhanced if cells are cultured as cell sheets which can be used as cell-derived matrices after decellularization. MSCproduced dECM has recently emerged as a promising substrate for the improved expansion of different cells (Chen et al, 2007;Lai et al, 2010;Ng et al, 2014;Wu et al, 2016). It was FIGURE 10 | Induction of osteogenic differentiation of hMSCs cultured on plastic, plastic covered by fibronectin, or laminin, or type I collagen and dECM produced by human fibroblasts or MSCs for 4 days assessed by histological staining with Alizarin Red.…”

“…Extracellular matrix production by MSCs is enhanced if cells are cultured as cell sheets which can be used as cell-derived matrices after decellularization. MSC-produced dECM has recently emerged as a promising substrate for the improved expansion of different cells ( Chen et al, 2007 ; Lai et al, 2010 ; Ng et al, 2014 ; Wu et al, 2016 ). It was shown that MSCs cultured on these surfaces exhibited improved proliferation capacity, maintenance of phenotype, and increased differentiation potential ( Shakouri-Motlagh et al, 2017 ; Sart et al, 2020 ).…”

Restoring the quantity and quality of elderly human mesenchymal stem cells for autologous cell-based therapies

“…As reported by others [ 25 , 26 , 29 ], and shown in the present study, these criteria are inadequate. First, plastic adhesion of very early-stage MSCs is poor [ 29 , 30 ] and accumulating evidence indicates that culture on TCP alters the MSC phenotype [ 10 , 31 , 32 ]. Second, cell surface marker expression, which is routinely used to define MSCs, does not correlate well with the differentiation state of the cells or their potency, and does not distinguish between MSCs obtained from young versus old donors (Additional file 1 : Figure S1).…”

Células madre mesenquimatosas y sus diferentes usos

“…Por último, las células adheridas deben ser separadas, lo cual se realiza con tripsina o colagenasa. 20,21 Para la visualización de su morfología, se usa un microscopio electrónico de barrido y emisión para poder evaluar la viabilidad que estas potencialmente tendrían. 22 Para asegurar esa viabilidad se implementa una coloración con calceína-ester de acetoximetilo y homodímero de etidio que permite visualizar en un microscopio de barrido de laser focal la cantidad de fluorescencia de las células, la cual se relaciona de manera directamente proporcional con la viabilidad de las mismas.…”