The pluripotency factor Oct4 as a regulator of vascular differentiation

Abstract

Rat extraembryonic endoderm precursor (XEN-P) cells express the transcription factor Oct4. Here I demonstrate continued and increased Oct4 gene expression during differentiation. High cell density rapidly reduced XEN-P cell markers while upregulating Oct4 and endothelial markers (VE-cadherin, vWF) in a LIF-dependent manner. The upregulation of Oct4 was prevented by a MAPK inhibitor but not a STAT3 inhibitor, and was associated with activation of a novel Oct4 gene enhancer. Colonies arising on feeder cells from low density-plated XEN-P cells eventually also activated this enhancer, concomitantly with the co-expression of Oct4 and VE-cadherin and the formation of vessel-like structures. A PI3K inhibitor upregulated Oct4 mRNA levels independently of, and additively with, the effect of density/LIF. The data identify high cell density and LIF as synergistic stimulators of Oct4 gene expression during the vessel-like differentiation of XEN-P cells and suggest that the effect is mediated by MAPK and a novel upstream Oct4 gene enhancer.|쥐의 배외 내배엽 전구 (XEN-P) 세포는 전사인자 Oct4 를 발현한다. 이 논문에서는 XEN-P 세포가 분화하는 동안 지속적으로 상향 조절되는 Oct4 유전자 발현에 대해 소개할 것이다. LIF가 지속적으로 존재하는 상태에서 세포 밀도를 높였을 때, XEN-P 세포 마커들은 빠르게 감소하는 반면, Oct4와 내피 마커 (VE-cadherin, vWF) 는 빠르게 상향 조절된다. 상향 조절된 Oct4는 MAPK inhibitor에 의해 차단되지만 Stat3 inhibitor에 의해서는 차단되지 않는다. 그리고 이 조건은 새로운 Oct4 유전자의 enhancer 활성과 관련이 있다. Feeder 세포 위에서 낮은 밀도로 배양하기 시작한 XEN-P 세포들도 결국은 이 enhancer를 활성화 시키며, 동시에 혈관 유사 구조를 형성하고 Oct4와 VE-cadherin를 발현 시킨다. PI3K inhibitor는 독립적으로 Oct4 mRNA 발현을 증가시키는데, LIF와 높은 세포밀도에서와 같은 효과를 나타낸다. 이 논문으로부터 XEN-P 세포가 혈관으로 분화하는 동안 높은 세포 밀도와 LIF는 Oct4 유전자 발현의 상호 자극제로 작용하는 것을 확인하였으며 또한 이 효과는 MAPK와 새로운 Oct4 유전자 enhancer 에 의해 조절됨을 제시한다.; Rat extraembryonic endoderm precursor (XEN-P) cells express the transcription factor Oct4. Here I demonstrate continued and increased Oct4 gene expression during differentiation. High cell density rapidly reduced XEN-P cell markers while upregulating Oct4 and endothelial markers (VE-cadherin, vWF) in a LIF-dependent manner. The upregulation of Oct4 was prevented by a MAPK inhibitor but not a STAT3 inhibitor, and was associated with activation of a novel Oct4 gene enhancer. Colonies arising on feeder cells from low density-plated XEN-P cells eventually also activated this enhancer, concomitantly with the co-expression of Oct4 and VE-cadherin and the formation of vessel-like structures. A PI3K inhibitor upregulated Oct4 mRNA levels independently of, and additively with, the effect of density/LIF. The data identify high cell density and LIF as synergistic stimulators of Oct4 gene expression during the vessel-like differentiation of XEN-P cells and suggest that the effect is mediated by MAPK and a novel upstream Oct4 gene enhancer.