Neuroprotection and neuroregeneration of retinal ganglion cells after intravitreal carbon monoxide release.
Neuroprotection and neuroregeneration of retinal ganglion cells after intravitreal carbon monoxide release.
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DOI：
10.1371/journal.pone.0188444
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Biermann J
中科院分区：
文献类型：
--
作者：
Stifter J;Ulbrich F;Goebel U;Böhringer D;Lagrèze WA;Biermann J
视网膜缺血诱导细胞凋亡，导致神经变性和视力损害。气态一氧化碳（CO）在视网膜缺血再灌注损伤（IRI）后具有细胞保护和抗炎作用。静脉内施用的CO释放分子（CORM）ALF-186也证明了这些作用。本文总结了玻璃体内释放CO的结果，以评估其作为神经保护和神经再生剂的适用性。将水溶性CORM ALF-186（25 μg）、PBS或灭活ALF（iALF）（均为5 μl）在视网膜IRI后直接玻璃体内应用于大鼠左眼1 h。他们的右眼不受影响，并用于比较。干预后24 h收集视网膜组织，分析Caspase-3、pERK 1/2、p38、HSP 70/90、NF-κ B、AIF-1（同种异体移植炎性因子）、TNF-α和GAP-43的mRNA或蛋白表达。荧光金预标记的视网膜神经节细胞（RGC）的密度在IRI后7天在平板安装的视网膜中进行了检查，并表示为平均值/mm 2。在IRI后2天和7天，使用层粘连蛋白-1包被的培养物中的视网膜外植体评估RGC再生其轴突的能力。免疫组织化学方法分析视网膜组织块中生长的不同类型细胞。与对侧右眼中的RGC密度（2804±214 RGC/mm 2;数据为平均值±SD）相比，IRI+PBS注射导致RGC的显著损失（1554±159 RGC/mm 2），p<0.001。IRI后立即玻璃体内注射ALF-186提供了RGC保护并降低了RGC损伤的程度（IRI+PBS 1554±159对IRI+ALF 2179±286，p<0.001）。ALF-186增加了IRI介导的MAP激酶p38磷酸化。与IRI+PBS或IRI+iALF相比，IRI+ALF后Caspase-3、NF-κ B、TNF-α和AIF-1表达显著降低，可检测到抗凋亡和抗炎作用。IRI+ALF后Gap-43表达显著增加。iALF显示与PBS相似的效果。在生长许可条件下，IRI和ALF或iALF处理后，RGC轴突的内在再生潜力被诱导到几乎相同的水平，尽管两组的RGC活力显著不同。玻璃体内一氧化碳进一步增加了视网膜移植物中GFAP阳性细胞的迁移和转分化，这是通过β-III微管蛋白和巢蛋白的再表达来检测的。玻璃体内注射CORM ALF-186可保护IRI后的RGC，并刺激其轴突再生。ALF在IRI后传递抗凋亡、抗炎和生长相关信号。CO在神经再生中的作用及其对视网膜神经胶质细胞的影响需要进一步研究。
Retinal ischemia induces apoptosis leading to neurodegeneration and vision impairment. Carbon monoxide (CO) in gaseous form showed cell-protective and anti-inflammatory effects after retinal ischemia-reperfusion-injury (IRI). These effects were also demonstrated for the intravenously administered CO-releasing molecule (CORM) ALF-186. This article summarizes the results of intravitreally released CO to assess its suitability as a neuroprotective and neuroregenerative agent. Water-soluble CORM ALF-186 (25 μg), PBS, or inactivated ALF (iALF) (all 5 μl) were intravitreally applied into the left eyes of rats directly after retinal IRI for 1 h. Their right eyes remained unaffected and were used for comparison. Retinal tissue was harvested 24 h after intervention to analyze mRNA or protein expression of Caspase-3, pERK1/2, p38, HSP70/90, NF-kappaB, AIF-1 (allograft inflammatory factor), TNF-α, and GAP-43. Densities of fluorogold-prelabeled retinal ganglion cells (RGC) were examined in flat-mounted retinae seven days after IRI and were expressed as mean/mm2. The ability of RGC to regenerate their axon was evaluated two and seven days after IRI using retinal explants in laminin-1-coated cultures. Immunohistochemistry was used to analyze the different cell types growing out of the retinal explants. Compared to the RGC-density in the contralateral right eyes (2804±214 RGC/mm2; data are mean±SD), IRI+PBS injection resulted in a remarkable loss of RGC (1554±159 RGC/mm2), p<0.001. Intravitreally injected ALF-186 immediately after IRI provided RGC protection and reduced the extent of RGC-damage (IRI+PBS 1554±159 vs. IRI+ALF 2179±286, p<0.001). ALF-186 increased the IRI-mediated phosphorylation of MAP-kinase p38. Anti-apoptotic and anti-inflammatory effects were detectable as Caspase-3, NF-kappaB, TNF-α, and AIF-1 expression were significantly reduced after IRI+ALF in comparison to IRI+PBS or IRI+iALF. Gap-43 expression was significantly increased after IRI+ALF. iALF showed effects similar to PBS. The intrinsic regenerative potential of RGC-axons was induced to nearly identical levels after IRI and ALF or iALF-treatment under growth-permissive conditions, although RGC viability differed significantly in both groups. Intravitreal CO further increased the IRI-induced migration of GFAP-positive cells out of retinal explants and their transdifferentiation, which was detected by re-expression of beta-III tubulin and nestin. Intravitreal CORM ALF-186 protected RGC after IRI and stimulated their axons to regenerate in vitro. ALF conveyed anti-apoptotic, anti-inflammatory, and growth-associated signaling after IRI. CO’s role in neuroregeneration and its effect on retinal glial cells needs further investigation.
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