Differentiation of Pluripotent Stem Cells for Disease Modeling: Learning from Heart Development.

Heart disease is a pressing public health problem and the leading cause of death worldwide. The heart is the first organ to gain function during embryogenesis in mammals. Heart development involves cell determination, expansion, migration, and crosstalk, which are orchestrated by numerous signaling pathways, such as the Wnt, TGF-ß, IGF, and Retinoic acid signaling pathways. Human-induced pluripotent stem cell-based platforms are emerging as promising approaches for modeling heart disease in vitro. Understanding the signaling pathways that are essential for cardiac development has shed light on the molecular mechanisms of congenital heart defects and postnatal heart diseases, significantly advancing stem cell-based platforms to model heart diseases. This review summarizes signaling pathways that are crucial for heart development and discusses how these findings improve the strategies for modeling human heart disease in vitro.

Advances in the Generation of Constructed Cardiac Tissue Derived from Induced Pluripotent Stem Cells for Disease Modeling and Therapeutic Discovery.

The human heart lacks significant regenerative capacity; thus, the solution to heart failure (HF) remains organ donation, requiring surgery and immunosuppression. The demand for constructed cardiac tissues (CCTs) to model and treat disease continues to grow. Recent advances in induced pluripotent stem cell (iPSC) manipulation, CRISPR gene editing, and 3D tissue culture have enabled a boom in iPSC-derived CCTs (iPSC-CCTs) with diverse cell types and architecture. Compared with 2D-cultured cells, iPSC-CCTs better recapitulate heart biology, demonstrating the potential to advance organ modeling, drug discovery, and regenerative medicine, though iPSC-CCTs could benefit from better methods to faithfully mimic heart physiology and electrophysiology. Here, we summarize advances in iPSC-CCTs and future developments in the vascularization, immunization, and maturation of iPSC-CCTs for study and therapy.

Isoelectric point (pI)-based phase separation (pI-BPS) purification of elastin-like polypeptides (ELPs) containing charged, biologically active fusion proteins (ELP-FPs).

Elastin-like polypeptides (ELPs) are peptide-based biomaterials with residue sequence (VPGXG)n where X is any residue except proline. ELPs are a useful modality for delivering biologically active proteins (growth factors, protease inhibitors, anti-inflammatory peptides, etc.) as fusion proteins (ELP-FP). ELP-FPs are particularly cost-effective because they can be rapidly purified using Inverse Temperature Cycling (ITC) via the reversible formation and precipitation of entropically driven aggregates above a transition temperature (Tt ). When ELP fusion proteins (ELP-FPs) contain significant charge density at physiological pH, electrostatic repulsion between them severely inhibits aggregate formation. The literature does not currently describe methods for purifying ELP-FPs containing charged proteins on either side of the ELP sequence as fusion partners without organic solvents. Here, the isoelectric point (pI) of ELP-FPs is discussed as a means of neutralizing surface charges on ELP-FPs and increasing ITC yield to dramatically high levels. We use pI-based phase separation (pI-BPS) to purify ELP-FPs containing cationic and anionic fusion proteins. We report a dramatic increase in protein yield when using pI-BPS for purification of ELP-FPs. Proteins purified by this method also retain the functional activity of the protein present in the ELP-FP. Techniques developed here enable significant diversification of possible fusion proteins delivered by ELPs as ELP-FPs by allowing them to be produced and purified at higher quantities and yields.