Increased Hydrostatic Pressure Promotes Primary M1 Reaction and Secondary M2 Polarization in Macrophages

Abstract

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 in vitro 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: p = 0.0034) with less extra cellular matrix (ECM) production and secondary to an M2 phenotype (medium, 60 vs. 0 mmHg, d7: p = 0.0089) (medium, 60 vs. 20 mmHg, d7: p = 0.0433) with enhanced ECM production. Dex induces an M2 phenotype (Dex, medium vs. Dex, d2: p < 0.0001; d7: p < 0.0001) with more ECM production. Higher HP further increased M2 polarization of Dex-treated macrophages (Dex, 60 vs. 0 mmHg, d2: p = 0.0417; d7: p = 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.

Immune responses to injury and their links to eye disease

“…However, the likelihood of a significant immune component in glaucoma, including as a potential cause of the stiffness of the trabecular meshwork, can be drawn from studies showing the presence of proinflammatory molecules in the aqueous humor proteome of patients with POAG [91,122]. Studies of uveitic glaucoma, a rarer form of the disease, provide more direct evidence of a causative link between immune cells and the induction of glaucoma in the anterior chamber [123]. Uveitis is inflammation of the middle layer of the eye, the uvea, which functions to provide the vascular support required for phototransduction.…”

Conditional Ablation of Glucocorticoid and Mineralocorticoid Receptors from Cochlear Supporting Cells Reveals Their Differential Roles for Hearing Sensitivity and Dynamics of Recovery from Noise-Induced Hearing Loss

“…A prominent role of HPA-like activity is immunomodulation via its modulated release of GCs and activation of GC-sensitive receptors. Innate immune cell phenotypic differentiation and proinflammatory macrophage clearance of debris is sensitive to GC signaling [ 120 , 121 , 122 ]. Our studies showed that in the absence of GR expression in cochlear-supporting cells, Iba1 + cells of the cochlea at baseline, no noise conditions presented with a hypertrophied ramification morphology that indicated a chronic stress condition was underway.…”

Glycoproteins From Rabdosia japonica var. glaucocalyx Regulate Macrophage Polarization and Alleviate Lipopolysaccharide-Induced Acute Lung Injury in Mice via TLR4/NF-κB Pathway

“…ANA-1 cells were cultured as previously described and expression of CD11b + , F4/80 + , CD86 + , and CD206 + was detected ( Wang B. et al, 2020 ). ANA-1 cells were all marked with fluorescently labeled antibodies targeting F4/80 and CD11b.…”

“…The supernatant of water extract was combined and concentrated under reduced pressure. After centrifugation, four times 95% ethanol was added and the precipitates were collected and freezedried to obtain XPS (315 g, yield rate 6.3%) (Wang et al, 2020 b;Wang et al, 2020c). XPS (5 g) was dissolved in 40 ml water and centrifuged.…”