Robust and Highly Efficient Protocol for Differentiation of Human Pluripotent Stem Cells into Mesenchymal Stem Cells
Mesenchymal stem cells (MSCs) can be isolated from different sources, such as bone marrow, cord blood, and adipose tissue; however, there are variations in MSC capabilities based on their origin, donor age, and culturing method. Recently, human pluripoten
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Robust and Highly Efficient Protocol for Differentiation of Human Pluripotent Stem Cells into Mesenchymal Stem Cells Manale Karam and Essam M. Abdelalim Abstract Mesenchymal stem cells (MSCs) can be isolated from different sources, such as bone marrow, cord blood, and adipose tissue; however, there are variations in MSC capabilities based on their origin, donor age, and culturing method. Recently, human pluripotent stem cells (hPSCs) have been proposed as an alternative renewal source for generating MSCs with large number. Herein, we describe our recently established All-trans retinoic acid (RA)-based approach for generating a scalable number of MSCs from hPSCs. Our protocol generates highly proliferating MSCs that have all MSC characteristics, including fibroblast-like morphology, expression of the key MSC markers, lack of the hematopoietic markers, and ability to differentiate into the three mesodermal lineages. This RA-based method provides a protocol for generating an unlimited number of hPSC-derived MSCs that could be useful for cell therapy, drug screening, and disease modeling applications. Keywords Stem cells, hPSCs, MSCs, Protocol, Retinoic acid, Cell therapy, Disease modeling
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Introduction Mesenchymal stem cells (MSCs) have been demonstrated as a promising source for cell-based therapies due to their unique characteristics, such as immunomodulation, homing, and poor immunogenicity [1]. Although MSCs can be isolated from several tissues in the body, such as bone marrow, fat tissue, and cord blood, the obtained number is always small and requires in vitro propagation [2]. Also, the process of MSC isolation from human body is invasive and sometimes painful [2]. Moreover, the MSCs isolated from different sources display heterogeneity in their expansion and differentiation abilities, which may be due to the differences in the tissue micro-environment, age of the donor, and in vitro culturing method [3]. The use of human pluripotent stem cells (hPSCs), such as human embryonic stem cells (hESCs) and human-induced PSCs (iPSCs) to generate large number of MSCs, has been previously reported [4–8]. hPSC-derived MSCs show typical MSC characteristics, including fibroblast-like morphology, adherence to plastic plates, expression of the MSC markers, and absence of the
Manale Karam and Essam M. Abdelalim
Fig. 1 Generation of mesenchymal stem cells (MSCs) from hESCs. (a) A schematic diagram for the protocol of hPSC differentiation into MSCs. (b) Cell morphology of different stages during MSC differentiation
hematopoietic stem cell (HSC) markers as well as their ability to generate the three mesodermal lineages: adipocytes, osteocytes, and chondrocytes [9, 10]. Recently, we have established a robust and highly efficient All-trans retinoic acid (RA)-based protocol to generate an offthe-shelf and large number of MSCs from hPSCs [8]. Our method demonstrated that short exposure of embryoid bodies (EBs) derived from hPSCs to 10 μM RA significantly increases EB numbers, reduces cell death, and increases cell proliferation
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