A Small Molecule Modulating Monounsaturated Fatty Acids and Wnt Signaling Confers Maintenance to Induced Pluripotent Stem Cells Against Endodermal Differentiation

Abstract

Background: Stearoyl-coenzyme A desaturase 1 (SCD1) is required for de novo synthesis of fatty acids. This enzyme can orchestrate posttranslational modification of proteins involved in the development and differentiation of cells through the fatty acid acylation process. In this study, we evaluated whether a small molecule modulating unsaturated fatty acids influences early endodermal differentiation of induced pluripotent stem cells, using biochemical methods and immunostaining.Methods: The hiPSCs were cultured in an endoderm-inducing medium containing activin A and low defined fetal bovine serum in the presence of an SCD1 inhibitor at different time points. The yield of three germ layers endoderm, mesoderm, and ectoderm, and the cell cycle analysis were assessed using flow cytometry. The expression of endoderm and pluripotency markers, as well as the expression of Wnt signaling pathway proteins, were assessed using western blotting and RT-PCR. Total protein acylation was evaluated using a click chemistry reaction.Results: The population of cells showing endoderm features was decreased at the end of differentiation when SCD1 was inhibited on the first day. Moreover, early SCD1 inhibition preserved hiPSCs properties without a shift toward mesoderm or ectoderm. Treatment of cells with SCD1 inhibitor only on the first day decreased the β-catenin gene expression and intensity of fluorescent emission in the click chemistry. These effects were effectively rescued by cotreatment with oleate. Late treatment at two subsequent days of endoderm induction induced no significant effect on endoderm-specific markers and fluorescent intensity. Reproducible results were also obtained with a human embryonic stem cell line. Conclusion: The small molecule SCD1 inhibitor attenuates the Wnt/β-catenin signaling pathway, conferring maintenance to hiPSCs by opposing the initiation of endoderm differentiation. The immediate requirement for SCD1 activity in endoderm commitment of pluripotent stem cells may be eminent in disorders of endoderm-derived organs and dysregulated metabolism.

Fatty acids of type 2 diabetic serum decrease the stemness properties of human adipose‐derived mesenchymal stem cells

“…Wnt activity is strictly regulated by palmitoleoylation and SCD1 function 54 . The MUFAs generated by SCD1 also amplify the Wnt signaling pathway by increasing the receptor stability, 57 and drug‐mediated SCD1 inhibition has led to decreased expression and self‐renewal of stemness markers in mouse liver tumor‐initiated stem‐like cells 57–60 …”

“…54 The MUFAs generated by SCD1 also amplify the Wnt signaling pathway by increasing the receptor stability, 57 and drug-mediated SCD1 inhibition has led to decreased expression and self-renewal of stemness markers in mouse liver tumor-initiated stemlike cells. [57][58][59][60] β-catenin modulates Wnt 61 and, through binding to the TCF, induces the expression of c-Myc and Axin-2. 61 Axin-2 is the direct target gene of Wnt/β-catenin, and in a negative feedback mechanism, β-catenin destruction controls the β-catenin cytosolic level in response to Wnt signaling.…”