Lipocalin 2 Suppresses Ocular Inflammation by Inhibiting the Activation of NF-κβ Pathway in Endotoxin-Induced Uveitis

Abstract

Background/Aims: Lipocalin 2 (LCN2), an important mediator of a variety of cellular processes, is involved in regulating the inflammatory response, but its roles in different inflammatory diseases are controversial. Because the role of LCN2 in ocular inflammation has been unclear until now, we explored the function of LCN2 in lipopolysaccharide (LPS)-induced ocular inflammation in vivo and in vitro. Methods: Endotoxin-induced uveitis (EIU) was induced in male Sprague Dawley rats by the intravitreal injection of LPS. The expression and location of LCN2 in the retina were detected with western blotting and immunohistochemistry, respectively. We determined the clinical scores for anterior inflammation, quantified the infiltrated inflammatory cells, and measured the pro-inflammatory factors to determine the anti-inflammatory effects of LCN2 in EIU eyes. Cultured primary rat Müller cells were stimulated with LPS and the expression and secretion of LCN2 were measured with real-time PCR, western blotting, and an ELISA. After Müller cells were cotreated with LPS and LCN2 or PBS, the expression and secretion of TNF-α, IL-6, and MCP-1 were examined with realtime PCR, western blotting, and ELISAs. Western blotting and immunofluorescence were used to detect the phosphorylation and cellular distribution of nuclear factor kappaB (NF-κB) subunit p65. Results: In EIU, the expression of LCN2 was significantly upregulated in the retina, especially in the outer nuclear layer (mainly composed of Müller cells). LPS stimulation of cultured Müller cells also markedly elevated LCN2 expression. Intravitreal injection of LCN2 significantly reduced the clinical scores, inflammatory infiltration, and protein leakage in EIU, which correlated with the reduced levels of proinflammatory factors in the aqueous humor and retina. LCN2 treatment also reduced the expression and secretion of TNF-α, IL-6, and MCP-1 in LPS-stimulated Müller cells. LCN2 inhibited the inflammatory response by inhibiting the phosphorylation and translocation of NF-κB p65. Conclusions: LCN2 protects against ocular inflammation, at least in part, by negatively regulating the activation of the NF-κB signaling pathway. LCN2 may be a promising anti-inflammatory therapy for ocular diseases, such as uveitis.

Plasma levels of lipocalin 2 is increased in neovascular age-related macular degeneration patients, particularly those with macular fibrosis

“…It has been reported that LCN2 participates in light-induced photoreceptor death [46] and the development of early AMD [47]. However, other studies have shown that LCN2 is also involved in retinal pro-survival and anti-oxidative responses [42] and can suppress inflammation and reduce retinal degeneration [41,42]. In this study, we observed a significantly higher plasma level of LCN2 in nAMD patients with macular fibrosis, suggesting its potential pro-fibrotic roles.…”

“…Plasma and urine levels of LCN2 have been used as a biomarker for kidney disease [38,39] and cancer progression [40]. Recent studies have shown that retinal cells including Müller glia [41][42][43], microglia and RPE [43,44] can all express LCN2 and the expression is upregulated under stress conditions [41,43,45]. Furthermore, increased LCN2 have been observed in patients with Stargardt disease, retinitis pigmentosa, and AMD [42].…”

Vitamin D promotes the cisplatin sensitivity of oral squamous cell carcinoma by inhibiting LCN2-modulated NF-κB pathway activation through RPS3

“…These observations led to the question of whether there an intermediate regulatory factor between LCN2 and NF-κB or whether LCN2 acts directly on transcriptional activation 42 . When the IP assay was performed using an endogenous LCN2 antibody and SFBfusion-LCN2, weak binding of LCN2 to NF-κB was observed, suggesting a regulatory factor between LCN2 and NF-κB.…”

The role of lipocalin-2 in age-related macular degeneration (AMD)

“…In agreement with Parmar's in vitro data, LCN-2-mediated regulation of NF-κB activation has also been documented in inflammation during progression of endotoxin-induced uveitis, a well-established model of acute ocular inflammation. LCN-2 was significantly upregulated in the retinae of the rats after induction of uveitis, and played a protective antiinflammatory role by negative regulation of phosphorylation and nuclear translocation of NF-κB p65 in Muller cells and retinal tissues [90]. An earlier study conducted using eyes from horses suffering from autoimmune equine recurrent uveitis also reported upregulation of LCN-2 expression compared to healthy eyes.…”

“…6) and in a mouse model ( Fig. 7) with AMD-like pathology [89][90][91][92]. One of the easiest ways to induce retinal degeneration is to expose dark-adapted animals to high-intensity fluorescent light right after chemical pupil dilation [89,93].…”