Keratoconus patients exhibit a distinct ocular surface immune cell and inflammatory profile

Sharon D’Souza,Swaminathan Sethu,Rudy M M A Nuijts,Ganesh Ram Sahu,Pooja Khamar,Tanuja Vaidya,Archana Padmanabhan Nair,Sneha Gupta,Ritika Mullick,Rohit Shetty,Mor M Dickman,Arkasubhra Ghosh,Rajiv R Mohan

doi:10.1038/s41598-021-99805-9

Abstract

Inflammatory factors have been considered to contribute to keratoconus (KC) pathogenesis. This study aims to determine the immune cells subsets and soluble inflammatory factor profile on the ocular surface of KC patients. 32 KC subjects (51 eyes) across different grades of severity and 15 healthy controls (23 eyes) were included in the study. Keratometry and pachymetry measurements were recorded. Ocular surface immune cells (collected by ocular surface wash) immunophenotyped using flow cytometry include leukocytes, neutrophils, macrophages, natural killer (NK) cells, pan-T cells, gamma delta T (γδT) cells and NKT cells. Tear fluid collected using Schirmer’s strip was used to measure 50 soluble factors by multiplex ELISA. Proportions of activated neutrophils, NK cells and γδT cells were significantly increased in KC patients. Significantly higher levels of tear fluid IL-1β, IL-6, LIF, IL-17A, TNFα, IFNα/β/γ, EPO, TGFβ1, PDGF-BB, sVCAM, sL-selectin, granzyme-B, perforin, MMP2, sFasL and IgE, along with significantly lower levels of IL-1α and IL-9 were observed in KC patients. Alterations observed in few of the immuno-inflammatory parameters correlated with grades of disease, allergy, eye rubbing and keratometry or pachymetry measurements. The observation implies a distinct immuno-inflammatory component in KC pathogenesis and its potential as an additional therapeutic target in KC management.

Highlights

Inflammatory factors have been considered to contribute to keratoconus (KC) pathogenesis
Higher levels of tear fluid IL-1β, IL-6, LIF, IL-17A, TNFα, IFNα/β/γ, EPO, TGFβ1, platelet-derived growth factor-BB (PDGF-BB), soluble vascular cell adhesion molecule (sVCAM), sL-selectin, granzyme-B, perforin, matrix metalloproteinase 2 (MMP2), soluble Fas ligand (sFasL) and Immunoglobulin E (IgE), along with significantly lower levels of IL-1α and IL-9 were observed in KC patients
We determined the levels of a variety of secreted factors in the tear fluid, concurrently with proportions of leukocytes and various types of immune cell subsets relevant in mucosal biology on the ocular surface of KC patients

Summary

Introduction

Inflammatory factors have been considered to contribute to keratoconus (KC) pathogenesis. A variety of inflammatory factors, including IL-1β, TNFα, IL-6, IL-17A, IFNγ and MMP9 with an ability to influence ECM remodelling, are elevated in the tear fluid and/or corneal tissue of KC patients[5,6] These could be contributed either by corneal structural cells or immune cells. We used non-invasive methods for sample collection, such as open eye ocular surface wash for immune cell proportions d etermination[10] and Schirmer’s strip-based tear fluid collection for secreted factor levels quantification[11]. These methods could be applied in the clinic to determine the immune-inflammatory status in KC patients, assist in disease stratification and guide targeted therapeutics

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