RTN4/Nogo-A-S1PR2 negatively regulates angiogenesis and secondary neural repair through enhancing vascular autophagy in the thalamus after cerebral cortical infarction
RTN4/Nogo-A-S1PR2 negatively regulates angiogenesis and secondary neural repair through enhancing vascular autophagy in the thalamus after cerebral cortical infarction
复制标题
RTN 4/Nogo-A-S1 PR 2通过增强脑梗死后丘脑血管自噬负性调节血管生成和继发性神经修复
DOI：
10.1080/15548627.2022.2047344
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发表时间：
2022-03
期刊：
影响因子：
13.3
通讯作者：
Peiying Xiao;Jinmin Gu;Wei Xu;Xingyang Niu;Jian Zhang;Jingjing Li;Yicong Chen;Z. Pei;
中科院分区：
文献类型：
--
作者：
Peiying Xiao;Jinmin Gu;Wei Xu;Xingyang Niu;Jian Zhang;Jingjing Li;Yicong Chen;Z. Pei;
脑梗死可诱导丘脑血管生成，影响功能恢复。血管生成的机制尚不清楚。本研究旨在探讨RTN 4/Nogo-A在大脑中动脉闭塞（MCAO）后丘脑中介导大自噬/自噬和血管生成中的作用。我们评估了丘脑中的继发性神经元损伤、血管生成、血管自噬、RTN 4和S1 PR 2信号传导。RTN 4-S1 PR 2对血管自噬和血管生成的作用使用慢病毒和药理学方法进行评估。结果表明，RTN 4和S1 PR 2信号分子的上调与MCAO后同侧丘脑中的血管生成平行。通过siRNA敲低Rtn 4显著降低了血管中MAP 1 LC 3B-II的转化和BECN 1和SQSTM 1的水平，与同侧丘脑中增强的血管生成相一致。这种效果与挽救丘脑神经元损失和改善认知功能相一致。相反，激活S1 PR 2增强血管自噬，沿着抑制血管生成和加重丘脑神经元损伤。用3-甲基腺嘌呤或spautin-1进一步抑制自噬起始增强血管生成，而用巴弗洛霉素A1阻断溶酶体降解抑制同侧丘脑中的血管生成。RTN 4-S1 PR 2对自噬通量的控制在体外得到验证。此外，在MCAO后的丘脑血管中鉴定了ROCK 1-BECN 1相互作用沿着BECN 1磷酸化（Thr 119）。在血管中，Rtn 4的敲低显著降低BECN 1磷酸化，而激活S1 PR 2增加其磷酸化。这些结果表明，RTN 4-S1 PR 2相互作用的阻断通过抑制自噬激活和减轻脑梗死后血管中溶酶体降解的功能障碍来促进丘脑中的血管生成和次级神经修复。缩略语：3-甲基丙烯酸：3-甲基腺嘌呤; ACTA2/ACT-SMA：肌动蛋白α 2，平滑肌，主动脉; AIF 1/Iba 1：同种异体移植炎性因子1; BafA 1：巴弗洛霉素A1; BMVEC：脑微血管内皮细胞; BrdU：5-溴-2 '-脱氧尿苷; CLDN 11/OSP：封闭蛋白11; GFAP：胶质细胞酸性蛋白; HUVEC：人脐静脉内皮细胞; LAMA 1：层粘连蛋白α 1; MAP 2：微管相关蛋白2; MBP 2：髓鞘碱性蛋白2; MCAO：大脑中动脉闭塞; PDGFRB/PDGFRβ：血小板衍生生长因子受体，β多肽; RECA-1：大鼠内皮细胞抗原-1; RHOA：ras同系物家族成员A; RHRSP：易卒中肾血管性高血压大鼠; ROCK 1：Rho相关卷曲螺旋蛋白激酶1; RTN 4/Nogo-A：reticulon 4; RTN 4 R/NgR 1：网状蛋白4受体; S1 PR 2：鞘氨醇-1-磷酸受体2; SQSTM 1：螯合体1。
ABSTRACT Cerebral infarction induces angiogenesis in the thalamus and influences functional recovery. The mechanisms underlying angiogenesis remain unclear. This study aimed to investigate the role of RTN4/Nogo-A in mediating macroautophagy/autophagy and angiogenesis in the thalamus following middle cerebral artery occlusion (MCAO). We assessed secondary neuronal damage, angiogenesis, vascular autophagy, RTN4 and S1PR2 signaling in the thalamus. The effects of RTN4-S1PR2 on vascular autophagy and angiogenesis were evaluated using lentiviral and pharmacological approaches. The results showed that RTN4 and S1PR2 signaling molecules were upregulated in parallel with angiogenesis in the ipsilateral thalamus after MCAO. Knockdown of Rtn4 by siRNA markedly reduced MAP1LC3B-II conversion and levels of BECN1 and SQSTM1 in vessels, coinciding with enhanced angiogenesis in the ipsilateral thalamus. This effect coincided with rescued neuronal loss of the thalamus and improved cognitive function. Conversely, activating S1PR2 augmented vascular autophagy, along with suppressed angiogenesis and aggravated neuronal damage of the thalamus. Further inhibition of autophagic initiation with 3-methyladenine or spautin-1 enhanced angiogenesis while blockade of lysosomal degradation by bafilomycin A1 suppressed angiogenesis in the ipsilateral thalamus. The control of autophagic flux by RTN4-S1PR2 was verified in vitro. Additionally, ROCK1-BECN1 interaction along with phosphorylation of BECN1 (Thr119) was identified in the thalamic vessels after MCAO. Knockdown of Rtn4 markedly reduced BECN1 phosphorylation whereas activating S1PR2 increased its phosphorylation in vessels. These results suggest that blockade of RTN4-S1PR2 interaction promotes angiogenesis and secondary neural repair in the thalamus by suppressing autophagic activation and alleviating dysfunction of lysosomal degradation in vessels after cerebral infarction. Abbreviations: 3-MA: 3-methyladenine; ACTA2/ɑ-SMA: actin alpha 2, smooth muscle, aorta; AIF1/Iba1: allograft inflammatory factor 1; BafA1: bafilomycin A1; BMVECs: brain microvascular endothelial cells; BrdU: 5-bromo-2’-deoxyuridine; CLDN11/OSP: claudin 11; GFAP: glial fibrillary acidic protein; HUVECs: human umbilical vein endothelial cells; LAMA1: laminin, alpha 1; MAP2: microtubule-associated protein 2; MBP2: myelin basic protein 2; MCAO: middle cerebral artery occlusion; PDGFRB/PDGFRβ: platelet derived growth factor receptor, beta polypeptide; RECA-1: rat endothelial cell antigen-1; RHOA: ras homolog family member A; RHRSP: stroke-prone renovascular hypertensive rats; ROCK1: Rho-associated coiled-coil containing protein kinase 1; RTN4/Nogo-A: reticulon 4; RTN4R/NgR1: reticulon 4 receptor; S1PR2: sphingosine-1-phosphate receptor 2; SQSTM1: sequestosome 1.