| Co-Investigator(Kenkyū-buntansha) |
MATSUBARA Hiroaki Kyoto Prefectural University of Medicine, Dept. of Medicine, Professor, 医学研究科, 教授 (10239072)
TATSUMI Tetsuya Kyoto Prefectural University of Medicine, Dept. of Medicine, Instructor, 医学研究科, 講師 (20254328)
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| Budget Amount *help |
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 2005: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2004: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Research Abstract |
The present stud was designed to examinewhether human umbilical cord blood-derived cells have a potential of differentiation into cardiac myocytes in vitro and in vivo.
In vitro study
Human umbilical cord blood cells were obtained after childbirth in maternity hospitals. Mononuclear cells were divided from cord blood by Ficoll Plaque and lineage-negative (Lin-) cells were sorted using magnet sorting system. Culture of Lin- cells : Lin- cells were incubated for one week in initiation medium containing α medium supplemented with 10% fetal bovine serum (FBS), then cells were treated with differentiation medium for another 2 weeks containing Darbecco's modified eagle medium (DMEM) with 10%FBS and 1)1%Dimethl sulfoxide, 2)insulin (200 nM), or 3)5azacytidine (1mM). Some cells changed their shapes, but none of cells expressed cardiac markers. In another experiments, Lin- cells were cocultured with neonatal cardiac myocytes for 2 weeks with DMEM containing 10%FBS. Small number of Lin- cells, whi
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ch were detected by immunocytochemical staining using anti-human nuclear antibody, expressed cardiac markers : actinin, myosin heavy chain, and troponin I. Therefore, these results show that human umbilical cord blood derived cells might retain myogenic potential.
In vivo study
NOD/SCID mice were subjected to myocardial infarction by ligating the left anterior descending coronary artery. Shortly after coronary ligation, Lin- cells (3-6 x 10^6) were injected in the contracting wall bordering the infarct. Echocardiographic studies were performed 2 weeks after ligation, and immunohistochemical examinations were performed 2, 4, and 6 weeks after coronary ligation. Human derived cells were determined by anti-HLA-DR antibody and our using cardiac markers were actinin, myosin heavy chain, and desmin Injected cells were present at 2 - 6 weeks after coronary ligation, and the number of injected cells was gradually decreased. At 2 weeks after coronary ligation, lots of injected cells were confirmed, but none of them expressed cardiac marker. At 4 weeks after coronary ligation, rare Lin- cells expressed cardiac markers : actinin, myosin heavy chain, and desmin. However, 6 weeks after ligation a small number of residual Lin- cells did not express cardiac marker any more.
In addition, 2 weeks after coronary ligation, left ventricular dilatation and %fractional shortening were comparable between groups. Less
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