Investigation of neural crest stem cell therapy and niche factors for Hirschsprung's disease

2010 Fiscal Year Final Research Report

• Project/Area Number: 21791736
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Single-year Grants
• Research Field: Pediatric surgery
• Research Institution: Keio University
• Principal Investigator: NISHIKAWA Ryuhei Keio University, 医学部, 助教 (30534531)
• Project Period (FY): 2009 – 2010
• Keywords: neural crest-derived stem cells NCSCs / Hirschsprung's disease / Neurosphere-like Bodies NLBs / Ret

Research Abstract

Hirschsprung's disease, also called aganglionosis or megacolon, is a developmental disorder of large intestine (colon) which occurs in 1 in 5000 children. Ret KO (Ret^<[-/-]>) mouse were one of the Hirschsprung's disease model mouse which have no intestinal nerve systems. An object of this NLB co-culture study with Ret^<[-/-]>KO mouse aganglionic colon is to research regenerative medicine tactics for Hirschsprung's disease using de novo Neurosphere-like Bodies (NLBs) generation protocol with serum free culture to regenerate enteric nervous system.
Although there are several report described Generation of NLBs in gut but serum free culture procedure have not reported.
P0-Cre/floxed-EGFP mice are known to be used to target neural crest derived stem cells (NCSCs) population using EGFP fluorescent and could separate NCSCs rich cell population using FACS cell sorter.
We generated Neurosphere-like Bodies (NLBs) from the E13.5 embryo gut of transgenic mice P0-Cre/floxp-EGFP and then cells were cultured in the serum-free medium successfully. After 10 days to 3 w cultures, serum-Free-cultured EGFP-positive cells formed Neurosphere-like structures that expressed NCSC genes. Neurons, glial cells, and myofibroblasts marker protein expression were confirmed after 4days culture of NLBs using poly-D-lysine-laminin coated slide chamber and 10% FBS contained medium.

Research Products

• [Journal Article] ヒルシュスプルング病の神経堤幹細胞移植治療に必要とされるニッチ因子の研究 (2009)
  • Author(s): 西川竜平, 下島, 直樹, 森川, 康英, 森, 昌玄, 山本, 裕輝, 山田, 洋平, 堀田, 亮, 渕本, 康史, 星野, 健, 名越, 慈人, 中村, 雅也, 芝田, 晋介, 岡野, ジェイムス洋尚, 岡野, 栄之
  • Journal Title: 日本小児外科学会雑誌 第45巻5号 Pages: 884-884
• [Presentation] Isolation, proliferation, and differentiation of neural crest stem/progenitor cells from human gut in normal and motility disorder patients (2010)
  • Author(s): Naoki Shimojima, Yasuhide Morikawa, Ryuhei Nishikawa
  • Organizer: ハンガリー外科学会
  • Place of Presentation: Szeged, Hungary
  • Year and Date: 20100917-20100918
• [Presentation] ヒルシュスプルング病および類縁疾患の病態解析と腸管神経再生治療に関する基礎的研究 (2010)
  • Author(s): 下島直樹、森川康英、西川竜平
  • Organizer: 第47回日本小児外科学会学術集会
  • Place of Presentation: 慶應義塾大学、東京
  • Year and Date: 20100617-20100618
• [Presentation] Isoolation, proliferation, and differentiation of neural crest stem cells from human gut (2010)
  • Author(s): Naoki Shimojima, Yasuhide Morikawa, Ryuhei Nishikawa
  • Organizer: 43th Annual meeting of Pacific Association of Pediatric Surgery
  • Place of Presentation: Kobe, Japan
  • Year and Date: 20100523-20100527
• [Presentation] ヒト腸管を用いた腸管神経再生治療の可能性に関する研究(続報) (2010)
  • Author(s): 下島直樹、森川康英、西川竜平
  • Organizer: 第22回小腸移植研究会、招待講演
  • Place of Presentation: 慶應義塾大学、東京、三田講堂
  • Year and Date: 20100305-20100306
• [Presentation] 腸管神経再生治療の実験的検討-胎仔無神経節腸管への神経堤幹細胞移植 (2010)
  • Author(s): 下島直樹、森川康英、西川竜平
  • Organizer: 第46回日本周産期・新生児医学会学術集会
  • Place of Presentation: 神戸国際会議場、兵庫
  • Year and Date: 2010-07-12
• [Presentation] Investigation of neural crest stem cell therapy for Hirschsprung's disease (2009)
  • Author(s): Ryuhei Nishikawa,Naoki Shimojima, Yasuhide Morikawa
  • Organizer: 第7回幹細胞シンポジウム
  • Place of Presentation: 東京都港区六本木Izumi GardenGallery
  • Year and Date: 20090515-20090516
• [Remarks] ホームページ等
  • URL: http://web.sc.itc.keio.ac.jp/surgery/ps/staff/studies.html