Acute death of astrocytes in blast-exposed rat organotypic hippocampal slice cultures.
Acute death of astrocytes in blast-exposed rat organotypic hippocampal slice cultures.
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DOI：
10.1371/journal.pone.0173167
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Glavaski-Joksimovic A
中科院分区：
文献类型：
--
作者：
Miller AP;Shah AS;Aperi BV;Kurpad SN;Stemper BD;Glavaski-Joksimovic A
爆炸性创伤性脑损伤（bTBI）影响着全世界的平民、士兵和退伍军人，并带来了重大的健康问题。bTBI后神经退行性变的机制仍然难以捉摸，目前的治疗方法在很大程度上是无效的。为了开发更有效的治疗方法，更好地表征爆炸诱发的细胞变化和潜在机制是很重要的。在本研究中，我们的小组利用大鼠器官型海马脑片培养物（OHC）作为体外系统的模型bTBI。OHC暴露于138 ± 22 kPa（低）或273 ± 23 kPa（高）的超压，使用开放式氦驱动激波管，或被分配到假对照组。在受伤后2小时（h），我们描述了星形胶质细胞对爆炸超压的反应。对星形胶质细胞标记物胶质细胞酸性蛋白（GFAP）的免疫染色显示急性剪切和星形胶质细胞的形态学变化，包括clasmatodendrosis。此外，GFAP免疫染色和碘化丙啶（PI）的重叠表明星形胶质细胞死亡。海马角Ammonis 1区（CA 1）每个计数区域死亡星形胶质细胞数量的定量表明，与假手术对照OHC相比，低原始细胞和高原始细胞中死亡星形胶质细胞的数量显著增加。然而，只有少量的GFAP表达星形胶质细胞与凋亡标记物膜联蛋白V共标记，表明坏死是冲击波暴露后急性期细胞死亡的主要类型。此外，蛋白质印迹分析显示钙蛋白酶介导的GFAP的分解。葡聚糖排除法还表明膜破坏是急性星形胶质细胞死亡的潜在机制。此外，虽然冲击波暴露没有引起谷氨酸转运蛋白1（GLT-1）表达的显着变化，GLT-1表达的星形胶质细胞的损失表明损伤后谷氨酸摄取失调。我们的数据说明冲击波超压对OHCs中星形胶质细胞在损伤后2小时的深刻影响，并表明钙蛋白酶活性增加和膜破坏是潜在的潜在机制。
Blast traumatic brain injury (bTBI) affects civilians, soldiers, and veterans worldwide and presents significant health concerns. The mechanisms of neurodegeneration following bTBI remain elusive and current therapies are largely ineffective. It is important to better characterize blast-evoked cellular changes and underlying mechanisms in order to develop more effective therapies. In the present study, our group utilized rat organotypic hippocampal slice cultures (OHCs) as an in vitro system to model bTBI. OHCs were exposed to either 138 ± 22 kPa (low) or 273 ± 23 kPa (high) overpressures using an open-ended helium-driven shock tube, or were assigned to sham control group. At 2 hours (h) following injury, we have characterized the astrocytic response to a blast overpressure. Immunostaining against the astrocytic marker glial fibrillary acidic protein (GFAP) revealed acute shearing and morphological changes in astrocytes, including clasmatodendrosis. Moreover, overlap of GFAP immunostaining and propidium iodide (PI) indicated astrocytic death. Quantification of the number of dead astrocytes per counting area in the hippocampal cornu Ammonis 1 region (CA1), demonstrated a significant increase in dead astrocytes in the low- and high-blast, compared to sham control OHCs. However only a small number of GFAP-expressing astrocytes were co-labeled with the apoptotic marker Annexin V, suggesting necrosis as the primary type of cell death in the acute phase following blast exposure. Moreover, western blot analyses revealed calpain mediated breakdown of GFAP. The dextran exclusion additionally indicated membrane disruption as a potential mechanism of acute astrocytic death. Furthermore, although blast exposure did not evoke significant changes in glutamate transporter 1 (GLT-1) expression, loss of GLT-1-expressing astrocytes suggests dysregulation of glutamate uptake following injury. Our data illustrate the profound effect of blast overpressure on astrocytes in OHCs at 2 h following injury and suggest increased calpain activity and membrane disruption as potential underlying mechanisms.