Biphasic Role of Chondroitin Sulfate in Cardiac Differentiation of Embryonic Stem Cells through Inhibition of Wnt/b-Catenin Signaling

The glycosaminoglycan chondroitin sulfate is a critical component of proteoglycans on the cell surface and in the extracellular matrix. As such, chondroitin sulfate side chains and the sulfation balance of chondroitin play important roles in the control of signaling pathways, and have a functional importance in human disease. In contrast, very little is known about the roles of chondroitin sulfate molecules and sulfation patterns during mammalian development and cell lineage specification. Here, we report a novel biphasic role of chondroitin sulfate in the specification of the cardiac cell lineage during embryonic stem cell differentiation through modulation of Wnt/beta-catenin signaling. Lineage marker analysis demonstrates that enzymatic elimination of endogenous chondroitin sulfates leads to defects specifically in cardiac differentiation. This is accompanied by a reduction in the number of beating cardiac foci. Mechanistically, we show that endogenous chondroitin sulfate controls cardiac differentiation in a temporal biphasic manner through inhibition of the Wnt/beta-catenin pathway, a known regulatory pathway for the cardiac lineage. Treatment with a specific exogenous chondroitin sulfate, CS-E, could mimic these biphasic effects on cardiac differentiation and Wnt/beta-catenin signaling. These results establish chondroitin sulfate and its sulfation balance as important regulators of cardiac cell lineage decisions through control of the Wnt/beta-catenin pathway. Our work suggests that targeting the chondroitin biosynthesis and sulfation machinery is a novel promising avenue in regenerative strategies after heart injury. Citation: Prinz RD, Willis CM, van Kuppevelt TH, Klüppel M (2014) Biphasic Role of Chondroitin Sulfate in Cardiac Differentiation of Embryonic Stem Cells through Inhibition of Wnt/b-Catenin Signaling. PLoS ONE 9(3): e92381. doi:10.1371/journal.pone.0092381 Editor: Adam J. Engler, University of California, San Diego, United States of America Received May 29, 2013; Accepted February 21, 2014; Published March 25, 2014 Copyright: 2014 Prinz et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The authors wish to acknowledge financial support from the Ann and Robert H. Lurie Children’s Hospital of Chicago Research Center. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: [email protected] ¤a Current address: I. Medizinischen Klinik und Poliklinik, Universitätsmedizin Mainz, Mainz, Germany ¤b Current address: Department of Developmental Neurobiology, St. Jude’s Children’s Research Hospital, Memphis, Tennessee, United States of America