Bevacizumab Diminishes Inflammation in an Acute Endotoxin-Induced Uveitis Model

Abstract

Introduction: Uveitis is an eye disease characterized by inflammation of the uvea and an early and exhaustive diagnosis is essential for its treatment. The aim of our study is to assess the potential toxicity and anti-inflammatory efficacy of Bevacizumab in an experimental uveitis model by subcutaneously injecting lipopolysaccharide into Lewis rats and to clarify its mechanism. Material and Methods: Blood-aqueous barrier integrity was assessed 24 h after endotoxin-induced uveitis (EIU) by analyzing two parameters: cell count and protein concentration in aqueous humors. Histopathology of all eye structures was also studied. Enzyme-linked immunosorbent analyses of the aqueous humor samples were performed in order to calculate the diverse chemokine and cytokine protein levels and oxidative stress-related markers were also evaluated. Results: The aqueous humor's cellular content significantly increased in the group treated with only Bevacizumab, but it had no effect on retina histopathological grading. Nevertheless, the inflammation noted in ocular structures when administering Bevacizumab with endotoxin was mostly prevented since aqueous humor cell content considerably lowered, and concomitantly with a sharp drop in uveal, vitreous, and retina histopathological grading. The values of the multi-faceted cytokine IL-2 also significantly decreased (p < 0.05 vs. endotoxin group), and the protective IL-6 and IL-10 cytokines values rose with related anti-oxidant system recovery (p < 0.05 vs. endotoxin group). Concurrently, some related M1 macrophage chemokines substantially increased, e.g., GRO/KC, a chemokine that also displays any kind of protective role. Conclusion: All these results revealed that 24 h after being administered, Bevacizumab treatment in EIU significantly prevented inflammation in various eye structures and correct results in efficacy vs. toxicity balance were obtained.

Keywords: bevacizumab; chemokines; cytokines; endotoxin-induced uveitis; inflammation; oxidative stress.

FIGURE 1

Effect of endotoxin and/or Bevacizumab on cellular content (A) and protein concentration (B) in the aqueous humors collected 24 h following treatment, and on the histopathological score of anterior chamber (C), posterior chamber (D), ciliary body (E), vitreous chamber (F), and retina (G) cellular infiltration. Each value is shown as mean ± SE. ^∗p ≤ 0.05 vs. the Control group, ⁺p ≤ 0.05 vs. the Saline group and ^#p ≤ 0.05 vs. the Endotoxin group.

FIGURE 2

Histopathological study of ocular structures 24 h after Bevacizumab and/or endotoxin treatment, stained with HE. Picture number 1: Ciliary body, anterior and posterior chambers, 20x. Picture number 2: Inset of picture number 1, 63x. Picture number 3: Vitreous chamber, 63x. Picture number 4: Retina, 63x. Inflammatory cells were not observed in either the Control (C) or the Saline (S) group. Inflammatory cells were seen to infiltrate extravascular uveal tissue in Bevacizumab-treated eyes (B), and reached all the studied structures, apart from the retina (ciliary body, anterior, posterior, and vitreous chambers). Many inflammatory cells neutrophils (black arrow) and monocytes/macrophages (white arrow), were found to infiltrate extravascular uveal tissue in the endotoxin-treated eyes (E) and reached all the structures under study (ciliary body, anterior, posterior and vitreous chambers, and retina). A significant reduction was noted when Bevacizumab was also injected (E+B, ciliary body, anterior, posterior and vitreous chambers, and retina).

FIGURE 3

Effect of Bevacizumab on endotoxin-induced oxidative stress in rat eyes. Glutathione peroxidase activity (A), glutathione (B), and malondialdehyde (C) concentrations were measured in the eye homogenates of all the study groups. Each value is shown as mean ± SE. ^∗p ≤ 0.05 vs. the Control and Saline groups, and ^#p ≤ 0.05 vs. the Endotoxin group.

FIGURE 4

Effects of Bevacizumab on cytokine concentration (pg/mL) in the aqueous humors of all the study groups. IL-1b (A), IL-2 (B), IL-6 (C), IL-10 (D), TNFα (E), and INFγ (F) concentrations were measured. Each value is shown as mean ± SE. ^∗p ≤ 0.05 vs. the Control group, ˆp ≤ 0.05 vs. the Saline group, ⁺p ≤ 0.05 vs. the Bevacizumab group and ^#p ≤ 0.05 vs. the Endotoxin group.

FIGURE 5

Effects of Bevacizumab on chemokine concentration (pg/mL) in the aqueous humors of all the study groups. MIP-2 (A), MIP-3a (B), and GRO/KC (C) concentrations were measured. Each value is shown as mean ± SE. ^∗p ≤ 0.05 vs. the Control, Saline solution and Bevacizumab groups, and ^#p ≤ 0.05 vs. the Endotoxin group.