Citrullinated protein and Anti-Citrullinated Protein Antibodies (ACPAs) are present in Tear fluid of Dry Eye Disease (DED) patients – Pathophysiological Implications

Abstract

Purpose : While T-cell contribution to Dry Eye Disease (DED) pathophysiology is relatively well known, evidence linking autoantibodies to DED pathophysiology is relatively sparse. We investigated the role of citrullinated proteins and Anti-Citrullinated Protein autoantibodies (ACPAs) in the pathology of dry eye disease (DED).

Methods : Patients with DED were examined clinically. Ocular surface washings (OSW) and impression cytology (IC) was performed, and analyzed for the presence of ACPA using commercially available ELISA kits. The pathological consequences of OSW with high ACPA using in vitro experiments and in vivo murine models were identified. Immunostaining was performed for presence of citrullinated proteins, Fc receptors, plasma cells and IGG. Dot blot analysis was performed to detect specific ACPAs.

Results : Impression cytology revealed the presence of CD4 +ive T cells, CD19 +ive B cells, and immunoglobulin containing CD138 +ive Plasma cells on the ocular surface in Sjögren’s syndrome as well as in Ocular Graft-versus-Host disease (oGVHD). Further, we found citrullinated proteins on the ocular surface and that their source was primarily neutrophils. The Protein Arginine Deaminase (PAD4) enzymes that catalyzes this post-translational modification were present significantly more in DED eyes than in healthy. We detected significantly higher immunoglobulin amount and presence of several species of ACPAs in OSW from DED patients. We also found that OSW with high ACPA contributes to production of Neutrophil Extracellular Traps (NETs), and that ACPAs cause ocular surface disease in murine eyes. Antibodies to multiple citrullinated proteins (vimentin, alpha-enolase, histone 3, histone 4 and fibrinogen) were detected. Patients with DED who had ACPA positive OSW had more severe ocular surface disease. ACPA are present in OSW despite their absence in the serum in majority of patients in all DED, however, ACPA levels were greater in OSW of patients who have ACPA in serum suggesting contribution from blood vessel filtration as well.

Conclusions : Our findings point to citrullinated proteins on the ocular surface as the self-antigens driving autoimmune-based inflammation in DED. ACPAs may be produced over the ocular surface and contribute to DED signs and symptoms.

This is a 2020 ARVO Annual Meeting abstract.