Defined three-dimensional culture conditions mediate efficient induction of definitive endoderm lineage from human umbilical cord Wharton’s jelly mesenchymal stem cells

Abstract

BackgroundWharton’s jelly-derived mesenchymal stem cells (WJ-MSCs) are gaining increasing interest as an alternative source of stem cells for regenerative medicine applications. Definitive endoderm (DE) specification is a prerequisite for the development of vital organs such as liver and pancreas. Hence, efficient induction of the DE lineage from stem cells is crucial for subsequent generation of clinically relevant cell types. Here we present a defined 3D differentiation protocol of WJ-MSCs into DE cells.MethodsWJ-MSCs were cultured in suspension to generate spheroids, about 1500 cells each, for 7 days. The serum-free differentiation media contained specific growth factors, cytokines, and small molecules that specifically regulate signaling pathways including sonic hedgehog, bone morphogenetic protein, Activin/Wnt, and Notch.ResultsWe obtained more than 85 % DE cells as shown with FACS analysis using antibodies directed against the DE marker CXCR4. In addition, biochemical and molecular analysis of bona-fide DE markers revealed a time-course induction of Sox17, CXCR4, and FoxA2. Focused PCR-based array also indicated a specific induction into the DE lineage.ConclusionsIn this study, we report an efficient serum-free protocol to differentiate WJ-MSCs into DE cells utilizing 3D spheroid formation. Our approach might aid in the development of new protocols to obtain DE-derivative lineages including liver-like and pancreatic insulin-producing cells.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-016-0426-9) contains supplementary material, which is available to authorized users.

Xeno-free pre-vascularized spheroids for therapeutic applications

“…Summing up, formation of 3D cellular spheroids under FBS-free conditions was reported before, but mostly in applications other than therapeutic vascularization 39 , 47 , 55 . Here, we show that XF spheroids present no differences in terms of morphology, size, ECM deposition and OEC organization, as compared to FBS-grown spheroids, while exhibiting enhanced angiogenic potential.…”

Soft Matrix Combined With BMPR Inhibition Regulates Neurogenic Differentiation of Human Umbilical Cord Mesenchymal Stem Cells

“…Al Madhoun et al, 2016). The best concentration of BMPR inhibitor was detected using western blotting.…”

Fetal bovine serum-free culture of endothelial progenitor cells-progress and challenges

“…With a growing understanding of the advantages of 3D culture in the field of regenerative medicine, several protocols for the generation of 3D constructs under defined conditions have been developed, mostly for pluripotent stem cells (PSC) and MSC (Al Madhoun et al, ; Bazhanov, Mohammadipoor, & Bartosh, ; Duque‐Fernandez et al, ; Miranda, Fernandes, Diogo, & Cabral, ).…”

“…lated from UCB and grown in 3D collagen matrices. In the presence of commercial platelet lysate, ECFC formed more vascular structures both in vitro and in vivo, as revealed by the quantification of perfused human microvessels, in comparison to ECFC grown in control scaffolds without platelet supplementation.With a growing understanding of the advantages of 3D culture in the field of regenerative medicine, several protocols for the generation of 3D constructs under defined conditions have been developed, mostly for pluripotent stem cells (PSC) and MSC (AlMadhoun et al, 2016;Bazhanov, Mohammadipoor, & Bartosh, 2017;Duque- Fernandez et al, 2016;Miranda, Fernandes, Diogo, & Cabral, 2016).Another humanized large-scale strategy for the propagation of ECFC was proposed byReinisch et al (2009). The cells were successfully isolated from both UCB and PB under XF conditions and further expanded to achieve clinically relevant doses of up to 150 million cells.The proposed XF formulation was based on the standard endothelial growth medium (EGM; basal medium containing GF cocktail), further supplemented with 10% human platelet lysate (HPL) pooled from a minimum of 40 donors, in place of the standard 5% FBS.…”