Expression pattern of Ccr2 and Cx3cr1 in inherited retinal degeneration.
Expression pattern of Ccr2 and Cx3cr1 in inherited retinal degeneration.
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DOI：
10.1186/s12974-015-0408-3
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发表时间：
2015-10-12
影响因子：
9.3
通讯作者：
Sakai T
中科院分区：
文献类型：
--
作者：
Kohno H;Koso H;Okano K;Sundermeier TR;Saito S;Watanabe S;Tsuneoka H;Sakai T
尽管越来越多的证据表明小胶质细胞、视网膜中的常驻巨噬细胞和骨髓源性巨噬细胞可引起视网膜炎症，从而加速感光细胞死亡，但这些细胞在视网膜变性 (RD) 过程中如何被激活的细节仍不确定。因此，澄清哪些细胞在加剧视网膜炎症中起主导作用非常重要。然而，使用细胞标记物（例如 Iba-1）等传统技术很难区分小胶质细胞和巨噬细胞。最近，建立了两种用于可视化趋化因子受体的小鼠模型：Cx3cr1GFP/GFP 和 Ccr2RFP/RFP 小鼠。由于 Cx3cr1 在小胶质细胞中表达，而 Ccr2 据报道在激活的巨噬细胞中表达，这些小鼠有可能区分小胶质细胞和巨噬细胞，从而产生有关这些炎症细胞的激活及其在视网膜炎症中各自作用的新信息。在这项研究中，c-mer原癌基因酪氨酸激酶（Mertk）−/−小鼠被用作RD动物模型，该小鼠由于视网膜色素上皮吞噬作用缺陷而表现出感光细胞死亡。 Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ 小鼠是通过繁殖 Mertk−/−、Cx3cr1GFP/GFP 和 Ccr2RFP/RFP 小鼠建立的。使用视网膜和视网膜色素上皮 (RPE) 平片、视网膜切片和流式细胞术评估 Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ 小鼠的视网膜形态和炎症细胞激活和侵袭模式。 4 周大的 Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ 小鼠在视网膜内层显示出 Cx3cr1-GFP 阳性小胶质细胞。在 6 周和 8 周龄动物的外视网膜和视网膜下间隙中观察到 Cx3cr1-GFP 和 Ccr2-RFP 双阳性激活的小胶质细胞。观察到 Ccr2-RFP 单阳性骨髓源性巨噬细胞迁移到 Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ 小鼠的视网膜中。在 18 周龄动物的视网膜下腔中仍然观察到这些入侵细胞。 Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ 小鼠的视网膜中可区分 Cx3cr1-GFP 阳性小胶质细胞和 Ccr2-RFP 阳性巨噬细胞。此外，Cx3cr1 阳性小胶质细胞中的 Ccr2 表达是 RD 中小胶质细胞激活的一个特征。 Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ 小鼠能够在 RD 期间观察小胶质细胞随时间的激活，并且可能有助于将来开发针对 RD 的炎症靶向治疗策略。
Though accumulating evidence suggests that microglia, resident macrophages in the retina, and bone marrow-derived macrophages can cause retinal inflammation which accelerates photoreceptor cell death, the details of how these cells are activated during retinal degeneration (RD) remain uncertain. Therefore, it is important to clarify which cells play a dominant role in fueling retinal inflammation. However, distinguishing between microglia and macrophages is difficult using conventional techniques such as cell markers (e.g., Iba-1). Recently, two mouse models for visualizing chemokine receptors were established, Cx3cr1GFP/GFP and Ccr2RFP/RFP mice. As Cx3cr1 is expressed in microglia and Ccr2 is reportedly expressed in activated macrophages, these mice have the potential to distinguish microglia and macrophages, yielding novel information about the activation of these inflammatory cells and their individual roles in retinal inflammation. In this study, c-mer proto-oncogene tyrosine kinase (Mertk)−/− mice, which show photoreceptor cell death due to defective retinal pigment epithelium phagocytosis, were employed as an animal model of RD. Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ mice were established by breeding Mertk−/−, Cx3cr1GFP/GFP, and Ccr2RFP/RFP mice. The retinal morphology and pattern of inflammatory cell activation and invasion of Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ mice were evaluated using retina and retinal pigment epithelium (RPE) flat mounts, retinal sections, and flow cytometry. Four-week-old Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ mice showed Cx3cr1-GFP-positive microglia in the inner retina. Cx3cr1-GFP and Ccr2-RFP dual positive activated microglia were observed in the outer retina and subretinal space of 6- and 8-week-old animals. Ccr2-RFP single positive bone marrow-derived macrophages were observed to migrate into the retina of Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ mice. These invading cells were still observed in the subretinal space in 18-week-old animals. Cx3cr1-GFP-positive microglia and Ccr2-RFP-positive macrophages were distinguishable in the retinas of Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ mice. In addition, Ccr2 expression in Cx3cr1 positive microglia is a feature of microglial activation in RD. Mertk−/−Cx3cr1GFP/+Ccr2RFP/+ mice enabled observation of microglial activation over time during RD and may be useful for developing inflammation-targeted treatment strategies for RD in the future.