Differentiation of Induced Pluripotent Stem Cells towards Mesenchymal Stromal Cells is Hampered by Culture in 3D Hydrogels

Abstract

Directed differentiation of induced pluripotent stem cells (iPSCs) towards specific lineages remains a major challenge in regenerative medicine, while there is a growing perception that this process can be influenced by the three-dimensional environment. In this study, we investigated whether iPSCs can differentiate towards mesenchymal stromal cells (MSCs) when embedded into fibrin hydrogels to enable a one-step differentiation procedure within a scaffold. Differentiation of iPSCs on tissue culture plastic or on top of fibrin hydrogels resulted in a typical MSC-like phenotype. In contrast, iPSCs embedded into fibrin gel gave rise to much smaller cells with heterogeneous growth patterns, absence of fibronectin, faint expression of CD73 and CD105, and reduced differentiation potential towards osteogenic and adipogenic lineage. Transcriptomic analysis demonstrated that characteristic genes for MSCs and extracellular matrix were upregulated on flat substrates, whereas genes of neural development were upregulated in 3D culture. Furthermore, the 3D culture had major effects on DNA methylation profiles, particularly within genes for neuronal and cardiovascular development, while there was no evidence for epigenetic maturation towards MSCs. Taken together, iPSCs could be differentiated towards MSCs on tissue culture plastic or on a flat fibrin hydrogel. In contrast, the differentiation process was heterogeneous and not directed towards MSCs when iPSCs were embedded into the hydrogel.

Engineering Tissue-Informed Biomaterials to Advance Pulmonary Regenerative Medicine

“…In fact, iPSCs differentiated into induced mesenchymal stem cells (iMSCs) in the 2D condition and were a closer match to primary MSC than the 3D differentiated cells. The cells differentiated in 3D had a notable upregulation of genes related to the cardiovascular system and neurogenesis (61). These results indicate that even though 3D systems are more physiologically relevant and represent an increased complexity compared to 2D systems, these platforms still need further optimization to be more physiologically precise.…”

How the mechanical microenvironment of stem cell growth affects their differentiation: a review

“…Induced pluripotent stem cell (iPSC) differentiates into a typical MSC-like phenotype on tissue culture plastic or on the surface of fibrin hydrogels. In contrast, iPSCs embedded in a 3D environment do not differentiate towards MSC and have reduced differentiation potential for osteogenic and lipogenic lineages [ 27 ].…”

Polysaccharide-Based Hydrogels for Microencapsulation of Stem Cells in Regenerative Medicine

“…However, directing iPSCs to a specific cell lineage remains a challenge and each differentiation progress likely requires unique features in the culture system. While a 3D culture system is favorable to the generation of many other cell types, a recent study has shown that it may impair the differentiation of iPSCs towards mesenchymal stem cells-like phenotype as compared to a 2D culture system ( Goetzke et al, 2019 ). iPSCs are equally suitable for all the biomedical applications of ESCs, such as drug screening, toxicological studies, disease modeling and cell therapies.…”