Toll-like receptor 4 ablation improves stem cell survival after hypoxic injury
Toll-like receptor 4 ablation improves stem cell survival after hypoxic injury
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DOI：
10.1016/j.jss.2012.04.042
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发表时间：
2012-10-01
影响因子：
2.2
通讯作者：
Meldrum, Daniel R.
中科院分区：
文献类型：
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作者：
Brewster, Benjamin D.;Rouch, Joshua D.;Meldrum, Daniel R.
简介：间充质干细胞（MSC）治疗可改善缺血/再灌注损伤后的心脏功能，但其有效性受到缺氧环境中 MSC 存活的限制。 Toll 样受体 4 (TLR4) 在缺氧条件下有助于促凋亡信号传导。细胞内 AKT 和 ERK 通路的激活会对抗该信号并提高细胞存活率。目前尚不清楚 TLR4 的消融是否会影响 MSC 缺氧损伤后的这些通路。我们假设：1）MSC 中 TLR4 敲除（TLR4KO）可提高缺氧损伤后的存活率； 2) 这种存活率差异是由于 AKT 和 ERK 通路中的信号传导改善所致。 材料和方法：从骨髓中收获小鼠野生型 (WT) 和 TLR4KO MSC，并在体外生长。与常氧对照相比，在低氧条件下孵育总共 0.1 x 10(6) 个细胞/孔。 24 小时后，通过计数检查这些组的细胞存活率，并使用 t 检验进行比较，P < 0.05 = 统计显着性。使用蛋白质印迹分析测量裂解物中的 AKT 和 ERK 浓度。结果：WT 和 TLR4KO MSC 的形态相似。与我们之前的研究结果一致，缺氧确实显着增加了 WT 细胞的细胞死亡（1.79 x 10(5) 活细胞/mL 对照与 0.88 x 10(5) 缺氧，P < 0.05）。缺氧损伤不会增加TLR4KO组的细胞死亡（1.68 x 10(5)对照与1.82 x 10(5)缺氧，P < 0.05）。在所有 TLR4KO 组中均观察到 AKT 激活增加。 TLR4 不影响磷酸化 ERK 水平。结论：由于 AKT 通路信号增加，TLR4 敲除的 MSC 在缺氧损伤后表现出更高的存活率。在缺血/再灌注研究中使用 TLR4 敲除的 MSC 可增强心脏保护作用；干细胞存活率的提高可能是一个促成因素。 (C) 2012 年，爱思唯尔公司出版。
Introduction: Mesenchymal stem cell (MSC) therapy improves cardiac function after ischemia/reperfusion injury, but its effectiveness is limited by MSC survival in hypoxic environments. Toll-like receptor 4 (TLR4) contributes to pro-apoptotic signaling under hypoxic conditions. Activation of intracellular AKT and ERK pathways opposes this signal and improves cell survival. It is unknown whether ablation of TLR4 affects these pathways after hypoxic injury in MSCs. We hypothesized that: 1) TLR4 knockout (TLR4KO) in MSCs improves survival after hypoxic injury; and 2) this survival difference is due to improved signaling in the AKT and ERK pathways.Materials and methods: Murine wild-type (WT) and TLR4KO MSCs were harvested from bone marrow and grown in vitro. A total of 0.1 x 10(6) cells/well were incubated in hypoxic conditions versus normoxic controls. After 24 h, these groups were examined for cell survival via counting and compared using a t-test with P < 0.05 = statistical significance. AKT and ERK concentrations were measured in lysate using Western blot analysis.Results: The morphology of WT and TLR4KO MSCs was similar. In line with our previous findings, hypoxia did significantly increase cell death in WT cells (1.79 x 10(5) living cells/mL control versus 0.88 x 10(5) hypoxia, P < 0.05). Hypoxic injury did not increase cell death in the TLR4KO group (1.68 x 10(5) control versus 1.82 x 10(5) hypoxia, P < 0.05). Increased AKT activation was observed in all TLR4KO groups. TLR4 did not affect phosphorylated ERK levels.Conclusion: TLR4-knockout MSCs show improved survival after hypoxic injury because of increased AKT pathway signal. Use of TLR4-knockout MSCs in ischemia/reperfusion studies results in enhanced cardioprotection; improved stem cell survival was likely a contributing factor. (C) 2012 Published by Elsevier Inc.