シングルドナー由来ヒト歯科間葉系幹細胞の膵β細胞様細胞への分化

differentiation of single donor derived human dental mesenchymal stem cells into pancreatic β cell-like cells.

• Sharath B Shivakumar
• Hyeon-Jeong Lee
• Young-Bum Son
• Dinesh Bharti
• Sun A Ock
• Sung-Lim Lee
• Young-Hoon Kang
• Bong-Wook Park
• Gyu-Jin Rho

抄録

本研究では、同一ドナーのヒト歯科組織（歯髄、乳頭、卵胞）から単離した間葉系幹細胞（MSC）の分化能を調べ、比較検討した。消化法に従って歯科組織（歯髄、乳頭、卵胞）から単離したMSCを、多能性マーカーと細胞表面マーカーの発現を解析した。3種類のMSCはすべて間葉系に分化する可能性があるかどうかを評価した。さらに、多段階プロトコールを用いて膵臓β細胞様細胞に分化させ、膵臓系統特異的マーカーの発現を調べた。また、分化した膵β細胞様細胞の機能特性は、ジチゾン染色とグルコースチャレンジ試験により評価した。3種類のMSCはすべて培養時に線維芽細胞様の形態を示し、多能性および間葉系細胞表面マーカーを発現していた。これらのMSCは間葉系への分化に成功し、膵臓β細胞様細胞への形質転換に成功した。その中でも、歯科用卵胞由来のMSCは、膵臓系に特異的なマーカーをmRNAとタンパク質レベルで発現することで評価される膵臓系への高い分化能を示し、グルコース負荷時にはより高いインスリン分泌を示した。さらに、卵胞由来のMSCは分化時に高いジチゾン染色を示しました。単一ドナー由来の3種類のMSCはいずれも同様の細胞特性を有しており、膵臓系への分化が可能である。しかし、歯胚葉由来のMSCは、歯髄・乳頭由来のMSCよりも膵臓系への分化能が高く、将来の糖尿病治療における幹細胞治療への応用の可能性を示唆しています。

The present study was carried out to investigate and compare the differentiation potential of mesenchymal stem cells (MSCs) isolated from human dental tissues (pulp, papilla, and follicle) of the same donor. MSCs were isolated from dental tissues (pulp, papilla, and follicle) following digestion method and were analyzed for the expression of pluripotent markers and cell surface markers. All three types of MSCs were evaluated for their potential to differentiate into mesenchymal lineages. Further, the MSCs were differentiated into pancreatic β cell-like cells using multistep protocol and characterized for the expression of pancreatic lineage specific markers. Functional properties of differentiated pancreatic β cell-like cells were assessed by dithizone staining and glucose challenge test. All three types of MSCs showed fibroblast-like morphology upon culture and expressed pluripotent, and mesenchymal cell surface markers. These MSCs were successfully differentiated into mesenchymal lineages and transdifferentiated into pancreatic β cell-like cells. Among them, dental follicle derived MSCs exhibits higher transdifferentiation potency toward pancreatic lineage as evaluated by the expression of pancreatic lineage specific markers both at mRNA and protein level, and secreted higher insulin upon glucose challenge. Additionally, follicle-derived MSCs showed higher dithizone staining upon differentiation. All three types of MSCs from a single donor possess similar cellular properties and can differentiate into pancreatic lineage. However, dental follicle derived MSCs showed higher potency toward pancreatic lineage than pulp and papilla derived MSCs, suggesting their potential application in future stem cell based therapy for the treatment of diabetes.

© 2019 The Author(s).