# Signaling pathway-based culture condition improves differentiation potential of canine induced pluripotent stem cells

## TL;DR

Researchers improved the culture conditions for canine stem cells by optimizing signaling pathways, enabling better cell maintenance and differentiation into heart cells.

## Contribution

A signaling pathway-based medium that stabilizes canine iPSCs and enhances their differentiation potential is developed.

## Key findings

- FGF, activin/TGF-β, and WNT signals are critical for maintaining canine iPSCs in culture.
- The optimized medium enables differentiation into cardiomyocytes with functional traits like homogenous contraction.
- The medium reduces cell heterogeneity and enhances developmental potency across different cell lines.

## Abstract

Naturally occurring diseases in companion dogs are increasingly being recognized as valuable translational disease models. While induced pluripotent stem cell (iPSC) technology had revolutionized the field of human bio-medical research, canine iPSC (ciPSC) technology is still in its infancy, and robust canine-specific iPSC medium formulations and differentiation protocols are lacking. Here, we have established NANOG-reporter ciPSC lines and found that fibroblast growth factor (FGF), activin/transforming growth factor (TGF)-β, and WNT signals were critical for the robust maintenance of ciPSCs. Manipulating these signaling pathways stabilized the culture of ciPSC regardless of the cell line or basal medium. ciPSCs cultured in the optimized medium showed a homogenized global gene expression pattern. Furthermore, the ciPSCs cultured in this medium successfully differentiated into cardiomyocytes displaying homogenous contraction as well as sarcomere alignment. This robust culture condition provides a valuable resource to facilitate the utilization of ciPSCs for various studies, including human disease modeling.

•FGF, activin/TGF-β, and WNT signals are critical for ciPSC maintenance in culture•Activin + WNT inhibitor (AR) medium stabilizes ciPSCs and reduces heterogeneity•AR medium enhances ciPSC developmental potency regardless of the cell line•AR medium enables ciPSC differentiation into cardiomyocytes with functional traits

FGF, activin/TGF-β, and WNT signals are critical for ciPSC maintenance in culture

Activin + WNT inhibitor (AR) medium stabilizes ciPSCs and reduces heterogeneity

AR medium enhances ciPSC developmental potency regardless of the cell line

AR medium enables ciPSC differentiation into cardiomyocytes with functional traits

Nishimura and colleagues demonstrate that the modulation of activin/TGF beta and WNT signaling pathways enables the robust maintenance and stable culture of ciPSCs across multiple cell lines and basal media. Using this optimized medium, ciPSCs successfully differentiate into cardiomyocytes displaying homogenous contractions and sarcomere organization. These findings represent key steps toward overcoming long-standing barriers in ciPSC cultures and applications.