Differentiation of Mouse Embryonic Stem Cells into Dorsal Interneurons of the Spinal Cord Using BMP4 and Activin A

Objective: In vertebrates, bone morphogenetic proteins (BMPs) and activin signals play multiple roles in dorso-ventral patterning and development of the spinal cord. Here the inductions of BMP 4 and activin A on embryonic stem cells (ESCs) into dorsal interneurons have been studied. Materials and Methods: Four different treatments have been used for mESC derived neural precursors; they include BMP4 (1ng/ml and 10ng/ml), activin A (100 ng/ml), and activin A+BMP4 (100ng/ml, 10ng/ml). Induction’s effect on expression of specific dorsal interneuron markers in mature neurons have been evaluated by the use of immunocytochemistry and RT-PCR. Results: Treatment of ESC-derived neural precursors with BMP4, activin A, or both showed an increased generation of both dI1 and dI3 interneurons (Lhx2 and Isl-1-positive cells) compared to the control group. However, the synergistic effect in generation of dI3 was not observed when both factors were used. Moreover, RT-PCR analysis of differentiated cells showed the expression of Lhx9, Lhx1, and Isl1, the transcription factors that are markers of dI1, dI2, and dI3 interneurons respectively. Conclusion: Our results showed that specific combination of developmental signaling molecules can direct the differentiation of ESCs into dorsal interneurons. Furthermore, qualitative and quantitative differences in signaling by different members of the TGF-B superfamily may play a role in the specification of different types of dorsal interneurons.