TGF-β2 Induces Gli1 in a Smad3-Dependent Manner Against Cerebral Ischemia/Reperfusion Injury After Isoflurane Post-conditioning in Rats
TGF-β2 Induces Gli1 in a Smad3-Dependent Manner Against Cerebral Ischemia/Reperfusion Injury After Isoflurane Post-conditioning in Rats
复制标题
TGF-β 2 以 Smad3 依赖性方式诱导 Gli1 对抗大鼠异氟烷后处理后的脑缺血/再灌注损伤
DOI：
10.3389/fnins.2019.00636
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发表时间：
2019-06-26
影响因子：
4.3
通讯作者：
Ma, Ketao
中科院分区：
文献类型：
--
作者：
Peng, Li;Yang, Chengwei;Ma, Ketao
异氟醚（ISO）后适应减轻脑缺血/再灌注（I/R）损伤，但其机制尚不完全清楚。转化生长因子β (tgf - β)和hedgehog （Hh）信号通路在中枢神经系统中控制着广泛的机制。我们的目的是研究tgf - β 2/Smad3和音猬(Shh)/胶质母细胞瘤（Gli）信号通路及其串音在脑I/R损伤后ISO大鼠海马中的影响。成年雄性Sprague-Dawley大鼠进行大脑中动脉阻断（MCAO），阻断1.5 h，再灌注24 h （MCAO/R）。采用神经行为实验、TTC染色、HE染色、Nissl染色、TUNEL染色、免疫荧光（IF）、定量实时聚合酶链反应（qRT-PCR）和Western blot等方法评价I/R损伤后ISO的作用。ISO后适应组（ISO组）在再灌注开始时接受1小时的ISO后适应，导致梗死体积和神经功能缺损评分降低，存活神经元更多，损伤和凋亡神经元更少。IF染色、qRT-PCR和Western blot显示，ISO组海马CA1中tgf - β 2、Shh和Gli1表达水平较高。磷酸化Smad3 （p-Smad3）、Patched （Ptch）和Smoothed （Smo）在蛋白水平上也有所增加，而Smad3的总表达在所有组中都没有变化。当给予tgf - β 2抑制剂吡非尼酮或Smad3抑制剂SIS3 HCl时，p-Smad3和Gli1的表达水平降低，存活的锥体神经元减少。相比之下，预注射Smo抑制剂环巴胺后，tgf - β 2和p-Smad3的表达水平没有明显变化，但降低了Shh、Ptch和Gli1的表达水平。吡非尼酮联合环巴胺组Gli表达量最低。这些发现表明，tgf - β和hedgehog信号通路介导脑I/R损伤后ISO后适应的神经保护，并在Gli1水平上相互作用。
Isoflurane (ISO) post-conditioning attenuates cerebral ischemia/reperfusion (I/R) injury, but the underlying mechanism is incompletely elucidated. Transforming growth factor beta (TGF-beta) and hedgehog (Hh) signaling pathways govern a wide range of mechanisms in the central nervous system. We aimed to investigate the effect of the TGF-beta 2/Smad3 and sonic hedgehog (Shh)/Glioblastoma (Gli) signaling pathway and their crosstalk in the hippocampus of rats with ISO post-conditioning after cerebral I/R injury. Adult male Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAO), 1.5 h occlusion and 24 h reperfusion (MCAO/R). To assess the effect of ISO after I/R injury, various approaches were used, including neurobehavioral tests, TTC staining, HE staining, Nissl staining, TUNEL staining, immunofluorescence (IF), qRT-PCR (quantitative real-time polymerase chain reaction) and Western blot. The ISO post-conditioning group (ISO group) received 1 h ISO post-conditioning when reperfusion was initiated, leading to lower infarct volumes and neurologic deficit scores, more surviving neurons, and less damaged and apoptotic neurons. IF staining, qRT-PCR and Western blot showed high expression levels of TGF-beta 2, Shh and Gli1 in the hippocampal CA1 of the ISO group. Phosphorylated Smad3 (p-Smad3), Patched (Ptch), and Smoothed (Smo) were also increased at protein level in the ISO group, whereas total Smad3 expression did not change in all groups. When TGF-beta 2 inhibitor, pirfenidone, or Smad3 inhibitor, SIS3 HCl, were administered, the expression levels of p-Smad3 and Gli1 were reduced, and surviving pyramidal neurons decreased. By contrast, the expression levels of TGF-beta 2 and p-Smad3 did not change significantly after pre-injection of Smo inhibitor cyclopamine, but reduced the expression levels of Shh, Ptch, and Gli1. Moreover, Gli showed the lowest expression levels with pirfenidone combined with cyclopamine. These findings indicate that the TGF-beta and hedgehog signaling pathways mediate the neuroprotection of ISO post-conditioning after cerebral I/R injury, and crosstalk between two pathways at the Gli1 level.