Ophthalmology and Ophtha-Lab at St. Franziskus Hospital, Münster, Germany.
Institution of Neurology and Institution for Translational Neurology, Universitätsklinikum Münster, Münster, Germany.
Front Immunol. 2020 Oct 14;11:573955. doi: 10.3389/fimmu.2020.573955. eCollection 2020.
Patients with chronic anterior uveitis are at particularly high risk of developing secondary glaucoma when corticosteroids [e.g., dexamethasone (Dex)] are used or when inflammatory activity has regressed. Macrophage migration into the eye increases when secondary glaucoma develops and may play an important role in the development of secondary glaucoma. Our aim was to evaluate if increased hydrostatic pressure and corticosteroids could induce changes in macrophages phenotype. By using a pressure chamber cell culture system, we assessed the effect of increased hydrostatic pressure (HP), inflammation, and immunosuppression (Dex) on the M1/M2 phenotype of macrophages. Bone marrow-derived macrophages (BMDMs) were stimulated with medium, lipopolysaccharide (LPS, 100 ng/ml), Dex (200 ng/ml), or LPS + Dex and incubated with different HP (0, 20, or 60 mmHg) for 2 or 7 days. The numbers of CD86+/CD206- (M1 phenotype), CD86-/CD206+ (M2 phenotype), CD86+/CD206+ (intermediate phenotype), F4/80+/TNF-α+, and F4/80+/IL-10+ macrophages were determined by flow cytometry. TNF-α and IL-10 levels in cell culture supernatants were quantified by ELISA. TNF-α, IL-10, fibronectin, and collagen IV expression in BMDMs were detected by immunofluorescence microscopy. Higher HP polarizes macrophages primarily to an M1 phenotype (LPS, 60 vs. 0 mmHg, d2: = 0.0034) with less extra cellular matrix (ECM) production and secondary to an M2 phenotype (medium, 60 vs. 0 mmHg, d7: = 0.0089) (medium, 60 vs. 20 mmHg, d7: = 0.0433) with enhanced ECM production. Dex induces an M2 phenotype (Dex, medium vs. Dex, d2: < 0.0001; d7: < 0.0001) with more ECM production. Higher HP further increased M2 polarization of Dex-treated macrophages (Dex, 60 vs. 0 mmHg, d2: = 0.0417; d7: = 0.0454). These changes in the M1/M2 phenotype by high HP or Dex treatment may play a role in the pathogenesis of secondary uveitic glaucoma- or glucocorticoid (GC)-induced glaucoma.
患有慢性前葡萄膜炎的患者在使用皮质类固醇（例如地塞米松（Dex））或炎症活动消退时，发生继发性青光眼的风险特别高。当继发性青光眼发展时，巨噬细胞向眼内迁移增加，并且可能在继发性青光眼的发展中发挥重要作用。我们的目的是评估增加的静水压力和皮质类固醇是否会引起巨噬细胞表型的变化。通过使用压力室细胞培养系统，我们评估了增加的静水压力（HP），炎症和免疫抑制（Dex）对巨噬细胞的 M1 / M2表型的影响。骨髓来源的巨噬细胞（BMDM）用培养基，脂多糖（LPS，100ng/ml），Dex（200ng/ml）或 LPS + Dex刺激，并在不同的 HP（0、20 或 60mmHg）下孵育 2 或 7 天。通过流式细胞术确定 CD86 + / CD206-（M1 表型），CD86-/ CD206+（M2 表型），CD86 + / CD206+（中间表型），F4 / 80 + / TNF-α+和 F4 / 80 + / IL-10+巨噬细胞的数量。通过 ELISA 定量细胞培养上清液中的 TNF-α和 IL-10 水平。通过免疫荧光显微镜检测 BMDM 中 TNF-α，IL-10，纤连蛋白和胶原蛋白 IV 的表达。较高的 HP 主要使巨噬细胞向 M1 表型极化（LPS，60 与 0mmHg，d2： = 0.0034），并减少细胞外基质（ECM）的产生，并继发于 M2 表型（培养基，60 与 0mmHg，d7： = 0.0089）（培养基，60 与 20mmHg，d7： = 0.0433），增强 ECM 的产生。 Dex 诱导 M2 表型（Dex，培养基与 Dex，d2：<0.0001;d7：<0.0001），并产生更多的 ECM。较高的 HP 进一步增加了 Dex 处理的巨噬细胞的 M2 极化（Dex，60 与 0mmHg，d2：= 0.0417;d7：= 0.0454）。 HP 升高或 Dex 治疗引起的 M1 / M2 表型的这些变化可能在继发性葡萄膜炎性青光眼或糖皮质激素（GC）诱导性青光眼的发病机制中起作用。
