The in聽vitro preconditioning of myoblasts to enhance subsequent survival in an in聽vivo tissue engineering chamber model

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• 作者：Daniel J. Tilkorn^b ; E. Michele Davies^c ; Effie Keramidaris^a ; Aaron M. Dingle^a ; Yi-Wen Gerrand^a ; Caroline J. Taylor^a ; Xiao Lian Han^a ; Jason A. Palmer^a ; Anthony J. Penington^a ; Christina A. Mitchell^c ; Wayne A. Morrison^a ; Gregory J. Dusting^d ; Geraldine M. Mitchell^a ; ^{geraldine.mitchell@svhm.org.au}
• 关键词：Cell preconditioning ; In vitro test of cell survival ; Implantation in an in vivo tissue engineering chamber ; Morphometric assessment of cell survival
• 刊名：Biomaterials
• 出版年：2012
• 期刊代码：7_01429612
• 类别：ms
• 出版时间：May, 2012
• 卷：33
• 期：15
• 页码：3868-3879
• 文件大小：2384 K

摘要

The effects of in聽vitro preconditioning protocols on the ultimate survival of myoblasts implanted in an in聽vivo tissue engineering chamber were examined. In聽vitro testing: L6 myoblasts were preconditioned by heat (42聽掳C; 1.5聽h); hypoxia (<8%O₂; 1.5聽h); or nitric oxide donors: S-nitroso-N-acetylpenicillamine (SNAP, 200聽渭M, 1.5聽h) or 1-[N-(2-aminoethyl)-N-(2-aminoethyl)amino]-diazen-1-ium-1,2-diolate (DETA-NONOate, 500聽渭M, 7聽h). Following a rest phase preconditioned cells were exposed to 24聽h hypoxia, and demonstrated minimal overall cell loss, whilst controls (not preconditioned, but exposed to 24聽h hypoxia) demonstrated a 44%cell loss. Phosphoimmunoblot analysis of pro-survival signaling pathways revealed significant activation of serine threonine kinase Akt with DETA-NONOate (p聽<聽0.01) and heat preconditioning (p聽<聽0.05). DETA-NONOate also activated ERK 1/2 signaling (p聽<聽0.05). In聽vivo implantation: 100,000 preconditioned (heat, hypoxia, or DETA-NONOate) myoblasts were implanted in SCID mouse tissue engineering chambers. 100,000 (not preconditioned) myoblasts were implanted in control chambers. At 3 weeks, morphometric assessment of surviving myoblasts indicated myoblast percent volume (p聽=聽0.012) and myoblasts/mm² (p聽=聽0.0005) overall significantly increased in preconditioned myoblast chambers compared to control, with DETA-NONOate-preconditioned myoblasts demonstrating the greatest increase in survival (p聽=聽0.007 and p聽=聽0.001 respectively). DETA-NONOate therefore has potential therapeutic benefits to significantly improve survival of transplanted cells.