Association of Hemopexin in Tear Film and Conjunctival Macrophages With Vernal Keratoconjunctivitis

Abstract

Vernal keratoconjunctivitis (VKC) is a chronic allergic inflammatory disease with unclear etiology and pathogenesis. We investigated the tear film proteome of patients with VKC to understand the pathologic characteristics of VKC. Methods: Tear samples were collected from healthy volunteers and patients with VKC. Electrophoresis was performed to display the tear proteomic profiles according to VKC severity. The identities of differentially expressed proteins were analyzed by mass spectrometry and quantified by enzyme-linked immunosorbent assay. Impression cytology was performed on VKC conjunctival samples to demonstrate the cellular protein expression. Allergic sensitization was performed in mice to study the pathologic role of these proteins in VKC. Results: Hemopexin, an inflammatory protein, was elevated in the tear film of patients with VKC. The increased hemopexin concentration in VKC tears was significantly associated with disease severity. Impression cytology showed specific high hemopexin expression in dekeratinized conjunctival epithelium and necrotic macrophages in patients with VKC. Immunohistochemical examination of normal lacrimal tissues from mice showed that hemopexin was not expressed in any lacrimal apparatus. Under systemic and topical sensitization and challenge using hemopexin in mice, the affected eye had mild to moderate bead discharge, chemosis, and edema with excessive macrophage infiltration and conjunctival necrosis. Conclusion: An association exists between tear hemopexin and the development and pathologic effects of VKC. Clinical Relevance: Increased hemopexin may have a role in the development of VKC.

Effects of Benzalkonium Chloride on THP-1 Differentiated Macrophages In Vitro

“…In the conjunctiva, macrophages are localized in the stroma and can interact closely with conjunctival epithelial cells, playing an important role in ocular surface diseases. For example, allergic conjunctivitis has been shown to be associated with excessive macrophage infiltration in the conjunctiva [3], [4]. In trachoma, they are known to increase local production of connective tissue growth factor (CTGF), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) [5].…”

Tears and ocular surface disorders: Usefulness of biomarkers

“…Recently reported, the hemopexin (a plasma protein having highest binding affinity to heme) has been quantified in VKC tears, particularly in association with disease severity, suggesting hemopexin as monitoring biomarker in addition to a potential diagnostic one (Pong et al, 2011). The main increased inflammatory factors detected in VKC tears are schematized in Table 2.…”

Biomarkers of Ocular Surface Disease Using Impression Cytology

“…Increased expression of this eotaxin subgroup in VKC indicated their potential as pathophysiological biomarkers of inflammatory cell infiltrates in this specific demographic of ocular surface allergy. While later work on VKC by Pong et al [52] analyzed IC-derived human conjunctival cells (alongside tear proteomic assays) and demonstrated high levels of hemopexin expression (a class I acute-phase protein which is secreted after inflammatory events) via immunocytochemistry. Specifically in VKC patients, they reported increased cytoplasmic hemopexin in dekeratinized conjunctival epithelium and necrotic macrophages, but not in other inflammatory cells (eosinophils and neutrophils), thus indicating a role for this protein in the pathology of VKC [52].…”

“…While later work on VKC by Pong et al [52] analyzed IC-derived human conjunctival cells (alongside tear proteomic assays) and demonstrated high levels of hemopexin expression (a class I acute-phase protein which is secreted after inflammatory events) via immunocytochemistry. Specifically in VKC patients, they reported increased cytoplasmic hemopexin in dekeratinized conjunctival epithelium and necrotic macrophages, but not in other inflammatory cells (eosinophils and neutrophils), thus indicating a role for this protein in the pathology of VKC [52]. Additionally, work by Inada et al [53] utilized IC (nitrocellulose membranes) to analyze conjunctival epithelial cell mRNA of patients with VKC (n = 11), versus normal healthy controls (n = 11), alongside in vitro assays.…”

“…By contrast, significantly increased expression of CCL-20 was observed in the severe VKC group, versus the mild VKC and control groups (p < 0.05). The authors surmised that CCL-20, which is associated with inflammatory cell migration and the presence of inflammation, may mediate ocular surface homeostasis, hence worsening of VKC presentation [52]. Similarly, a recent IC study by Shiraki et al [54] enrolled 18 patients with VKC or AKC (and 12 healthy controls) to assess eotaxin-2 mRNA levels by qPCR.…”