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研究発表を行った学会；
・新学術領域研究「哺乳類初期発生の細胞コミュニティー」第６回班会議
　２０１３年９月２５日〜２７日（南阿蘇、熊本）
タイトル；Redefining the in vivo Origin of Nephron Progenitors Enables Generation of Three-dimensional Kidney Structures from Pluripotent Stem Cells in vitro.
発表者；太口　敦博　氏
　　　（熊本大学　発生医学研究所　腎臓発生分野）
Abstract；
Generation of the kidney in vitro is a challenge for developmental biology and regenerative medicine, because reconstitution of the three-dimensional nephron structures including glomeruli and nephric tubules is a prerequisite for the kidney functions. Adult kidney derives from caudally located embryonic metanephros which develops by the reciprocal interaction of the metanephric mesenchyme (MM) and the ureteric bud (UB), both of which are reportedly derived from common intermediate mesoderm (IM). Most nephron components are derived from metanephric nephron progenitors in the MM. However, how the IM differentiates from nascent mesoderm and how the anteroposterior axis is formed along the IM, so that finally gives rise to the posteriorly located MM are largely unknown.
We first evaluated the functional and molecular differences between the early stage renal precursors using Osr1-GFP knock-in mice. We also tried to identify the origin of metanephric mesenchyme by lineage trace experiments utilizing T-GFPCreER mice. We identified that the MM is originated from posteriorly located T+ precursors at embryonic day (E) 8.5, which may correspond to the so-called “axial progenitor”, and that developmentally distinct from Osr1+ anteriorly located IM, namely the UB progenitors.
Next, we examined the combinations of factors which are required for metanephric nephron progenitor specification from embryonic nascent mesoderm. T+ cells sorted from mouse embryos differentiate into the metanephric mesenchyme in vitro by posteriorization with a high concentration of Wnt agonist, followed by its graded attenuation and stage-specific growth factor addition.
Finally, by applying this protocol to the mouse and human pluripotent stem cells, metanephric nephron progenitors were obtained. When the induced metanephric nephron progenitors were cocultured with Wnt4 expressing feeder cells or embryonic spinal cords, they formed the three-dimensional structures of the kidney, including glomeruli with podocytes and renal tubules with proximal and distal regions. Furthermore, the glomeruli were efficiently vascularized upon transplantation.
In conclusion, we have succeeded in inducing the metanephric nephron progenitors from both mouse embryonic stem (ES) cells and human induced pluripotent stem (iPS) cells, in vitro. The resultant progenitors readily “self-organized” the three-dimensional structures of the kidney, comprising renal tubules and glomeruli with podocytes, which has not been achieved in previous reports. Thus, by re-evaluating the developmental origins of metanephric progenitors, we have provided key insights into kidney specification in vivo and taken important steps toward kidney organogenesis in vitro.