Ethanol enhances thymocyte apoptosis and autophagy in macrophages of rat thymi
Ethanol enhances thymocyte apoptosis and autophagy in macrophages of rat thymi
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DOI：
10.14670/hh-11-861
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发表时间：
2017-09-01
影响因子：
2
通讯作者：
Kondo, Yoichi
中科院分区：
文献类型：
--
作者：
Betsuyaku, Tsubasa;Eid, Nabil;Kondo, Yoichi
黑色体巨噬细胞在吞噬凋亡淋巴细胞的过程中发挥着重要作用，尤其是在各种应激源的作用下。虽然过量的酒精摄入可能会促进胸腺细胞的凋亡，但研究胸腺对酒精毒性的自噬反应的报道仍然很少。我们研究了狂饮酒精暴露的动物模型胸腺中的细胞凋亡和自噬。成年雄性Wistar大鼠腹腔注射5g/kg乙醇或磷酸盐缓冲盐水(对照组)，分别于注射后0、3、6、24小时处死动物。光镜和透射电子显微镜观察发现，乙醇处理大鼠胸腺皮质内有大量胸腺细胞吞噬TbM的形成，这种形成在24 h时最为明显，经免疫荧光和巨噬细胞标志物RM4和iNOS双重标记后，发现巨噬细胞有活化迹象。此外，与对照组相比，ETR胸腺TbM的自噬作用增强，超微结构表现为自噬空泡的积累，Lc3斑点的免疫组织化学增加，后者蛋白的Western印迹分析，以及免疫荧光双标记中Lc3和RM4的共存。免疫电子显微镜还显示ETR-TbM内有Lc3标记的自噬空泡和凋亡的细胞吞噬小体，提示可能存在Lc3相关的吞噬作用。这一点通过LC 3与溶酶体组织蛋白在双重标记中的增强共定位得到了证实。这些结果表明，ETR胸腺TBM的自噬增强不仅是一种细胞保护机制，而且可能参与清除胸腺细胞的凋亡，从而防止自身免疫反应和抑制炎症反应。
Tingible body macrophages (TBMs) play essential roles in the phagocytosis of apoptotic lymphocytes, specifically under exposure to various stressors. Although excessive ethanol consumption may enhance thymocyte apoptosis, reports investigating the autophagic response of the thymus to ethanol toxicity are still lacking. We investigated apoptosis and autophagy in thymi of an animal model of binge ethanol exposure. Adult male Wistar rats were injected intraperitoneally either with 5 g/kg ethanol or phosphate buffer saline (for the control group) and sacrificed 0, 3, 6 and 24 hours after injection. Light and transmission electron microscopy (TEM) studies revealed enhanced formation of TBMs phagocytosing many apoptotic thymocytes in the thymic cortex of the ethanol-treated rats (ETRs), and this formation was particularly marked at 24 h. The macrophages showed signs of activation under TEM and immunofluorescence double labeling with RM4 (a macrophage marker) and iNOS. Additionally, in comparison to the control group, autophagy was enhanced in ETR thymic TBMs as evidenced ultrastructurally by accumulation of autophagic vacuoles, immunohistochemical increases in LC3 puncta, Western blot analysis of the latter protein, and colocalization of LC3 and RM4 in immunofluorescence double labeling. Immunoelectron microscopy also revealed LC3-labeled autophagic vacuoles and apoptotic cell phagosomes in ETR TBMs, suggesting the possibility of LC3-related phagocytosis. This was confirmed by enhanced colocalization of LC3 with lysosomal cathepsins in double labeling. These results indicate that enhanced autophagy in ETR thymic TBMs is not only a cytoprotective mechanism but could also be involved in the clearance of apoptotic thymocytes, thus preventing autoimmune reactions and suppressing inflammatory response.