Cell therapy for diabetes applied by pancreatic duct cells.

• Project/Area Number: 13671200
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Metabolomics
• Research Institution: Jikei University School of Medicine
• Principal Investigator: NEMOTO Masami Jikei University School of Medicine, Lecturer, 医学部, 講師 (10281396)
• Co-Investigator(Kenkyū-buntansha): SASAKI Takashi Jikei University School of Medicine, Assistant Professor, 医学部, 助教授 (90205849)
• Project Period (FY): 2001 – 2004
• Project Status: Completed (Fiscal Year 2004)
• Budget Amount: ¥3,500,000 (Direct Cost: ¥3,500,000); Fiscal Year 2004: ¥600,000 (Direct Cost: ¥600,000); Fiscal Year 2003: ¥800,000 (Direct Cost: ¥800,000); Fiscal Year 2002: ¥1,300,000 (Direct Cost: ¥1,300,000); Fiscal Year 2001: ¥800,000 (Direct Cost: ¥800,000)
• Keywords: Pancreatic duct cells / Bone marrow cells / Insulin / Cytokeratin 19 / AAV / ネスチン / 再生医学 / レーザーマククロダイセクション / レーザーマイクロダイセクション / PDX-1遺伝子 / 細胞治療 / ngn-3遺伝子

Research Abstract

[Introduction]We performed bone marrow transplantation to examine whether bone marrow cells had ability to trans-differentiate insulin producing cells. 1x10^6 bone marrow cells were derived form Lac Z transgenic rats. STZ diabetic rats were irradiated for eight Gy and received these bone marrow cells by tail vein injection. After eight weeks from transplantation, the section of pancreas was fixed and stained by anti-Lac Z, aniti-nestine and anti-cytokeratin 19(CK19) antibodies. Nestin is a marker of immature neural cells and CK-19 is a marker of immature pancreatic duct cells expressing during the development of pancreas. There were observed many cells that stained by both anti-Lac Z and anti-CK19 antibody in pancreas, but there were few cells that stained by anti-Lac Z and anti-nestine antibody.
[Aim]The aim of this study is to elucidate whether anti-CK-19 antibody positive bone marrow cells would migrate and grow in the pancreas, and whether would differentiate and regenerate to insul in producing cells.
[Method]CK-promoter/enhancer and enhanced green fluorescent protein(EGFP) gene were prepared in cloning site of the adeno-associated virus vector(AAV). Bone marrow cells were transfected this vector and cultured. 1)We extracted total RNA from bone marrow cells and examined the mRNA expression of ins1 and ins2 gene. 2)Bone marrow cells were transplanted to recipient rats via tail vein. We removed the pancreas after eight weeks and stained by anti-insulin antibody.
[Results]1)The mRNA expression of ins2 gene was observed in bone marrow cells, but there was no mRNA expression of ins1,pdx-1 gene. 2)GFP positive cells were recognized in acinar glands of pancreas. These cells were also stained by anti-insulin antibody and these results indicated that bone marrow cells differentiated to insulin producing cells. However, there were no GFP positive cells in pancreatic ducts. Bone marrow cells differentiated insulin positive cells, but it was no evidence that bone marrow cells would regenerate insulin producing cells thorough the process of the development in pancreatic duct cells.