New insights to vascular smooth muscle cell and pericyte differentiation of mouse embryonic stem cells in vitro.

• OBJECTIVE: The molecular mechanisms that regulate pericyte differentiation are not well understood, partly because of the lack of well-characterized in vitro systems that model this process. In this article, we develop a mouse embryonic stem (ES) cell-based angiogenesis/vasculogenesis assay and characterize the system for vascular smooth muscle cell (VSMC) and pericyte differentiation. METHODS AND RESULTS: ES cells that were cultured for 5 days on OP9 stroma cells upregulated their transcription of VSMC and pericyte selective genes. Other SMC marker genes were induced at a later time point, which suggests that vascular SMC/pericyte genes are regulated by a separate mechanism. Moreover, sequence analysis failed to identify any conserved CArG elements in the vascular SMC and pericyte gene promoters, which indicates that serum response factor is not involved in their regulation. Gleevec, a tyrosine kinase inhibitor that blocks platelet-derived growth factor (PDGF) spell-receptor signaling, and a neutralizing antibody against transforming growth factor (TGF) beta1, beta2, and beta3 failed to inhibit the induction of vascular SMC/pericyte genes. Finally, ES-derived vascular sprouts recruited cocultured MEF cells to pericyte-typical locations. The recruited cells activated expression of a VSMC- and pericyte-specific reporter gene. CONCLUSIONS: We conclude that OP9 stroma cells induce pericyte differentiation of cocultured mouse ES cells. The induction of pericyte marker genes is temporally separated from the induction of SMC genes and does not require platelet-derived growth factor B or TGFbeta1 signaling.

Nyckelord

Animals

Cell Differentiation

Cells

Cultured

Coculture Techniques

Embryo

cytology

Genetic Markers

Mice

Muscle

Smooth

Vascular

cytology

Myocytes

Smooth Muscle

cytology

Pericytes

cytology

Proto-Oncogene Proteins c-sis

physiology

Reproducibility of Results

Stem Cells

cytology

metabolism

Stromal Cells

physiology

Transcription

Genetic

physiology

Transforming Growth Factor beta

physiology

Transforming Growth Factor beta1

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