Cathepsin S activation contributes to elevated CX3CL1 (fractalkine) levels in tears of a Sj\xf6gren\u2019s syndrome murine model

Runzhong Fu,Minchang Choi,Maria C Edman,Hao Guo,Wannita Klinngam,Sarah F Hamm-Alvarez,Srikanth Janga

doi:10.1038/s41598-020-58337-4

Runzhong Fu, Minchang Choi + Show 5 more

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https://doi.org/10.1038/s41598-020-58337-4

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Abstract

Autoimmune dacryoadenitis and altered lacrimal gland (LG) secretion are features of Sjögren’s syndrome (SS). Activity of cathepsin S (CTSS), a cysteine protease, is significantly and specifically increased in SS patient tears. The soluble chemokine, CX3CL1 (fractalkine), is cleaved from membrane-bound CX3CL1 by proteases including CTSS. We show that CX3CL1 is significantly elevated by 2.5-fold in tears (p = 0.0116) and 1.4-fold in LG acinar cells (LGAC)(p = 0.0026) from male NOD mice, a model of autoimmune dacryoadenitis in SS, relative to BALB/c controls. Primary mouse LGAC and human corneal epithelial cells (HCE-T cells) exposed to interferon-gamma, a cytokine elevated in SS, showed up to 9.6-fold (p ≤ 0.0001) and 25-fold (p ≤ 0.0001) increases in CX3CL1 gene expression, and 1.9-fold (p = 0.0005) and 196-fold (p ≤ 0.0001) increases in CX3CL1 protein expression, respectively. Moreover, exposure of HCE-T cells to recombinant human CTSS at activity equivalent to that in SS patient tears increased cellular CX3CL1 gene and protein expression by 2.8-fold (p = 0.0021) and 5.1-fold (p ≤ 0.0001), while increasing CX3CL1 in culture medium by 5.8-fold (p ≤ 0.0001). Flow cytometry demonstrated a 4.5-fold increase in CX3CR1-expressing immune cells (p ≤ 0.0001), including increased T-cells and macrophages, in LG from NOD mice relative to BALB/c. CTSS-mediated induction/cleavage of CX3CL1 may contribute to ocular surface and LG inflammation in SS.

Highlights

Sjögren’s syndrome (SS) is a systemic autoimmune disease associated with lymphocytic infiltration of lacrimal glands (LG) and salivary glands (SG), associated with dacryoadenitis and sialoadenitis, respectively[1]
Cathepsin S (CTSS) activity was measured in the same tear samples in Fig. 1A, revealing a marked increase in non-obese diabetic mouse (NOD) mice (Fig. 1B) paralleling increased CX3CL1
In NOD mice, autoimmune dacryoadenitis associated with elevated CTSS activity in LG and tears is correlated with increased CX3CL1 in LG, cornea and tears, and with increased CX3CR1+ immune cells in LG

Summary

Introduction

Sjögren’s syndrome (SS) is a systemic autoimmune disease associated with lymphocytic infiltration of lacrimal glands (LG) and salivary glands (SG), associated with dacryoadenitis and sialoadenitis, respectively[1]. The male NOD mice share many ocular surface system manifestations with SS patients including lymphocytic infiltration of the LG7,8, reduced tear flow[7,8,9,10], generation of a proteolytic tear film[7,9,10], altered distribution and expression of Rab3D9,11, reduced myoepithelial cells[12], loss of extracellular matrix[8] and elevated cytokines in LG and tears[13]. In vitro studies suggest that elevated tear CTSS may affect ocular surface homeostasis, since it can induce expression and secretion of pro-inflammatory cytokines and matrix metallopeptidase 9 (MMP-9) in a corneal cell line[18]. We hypothesised that the elevated CTSS in tears and LG in SS might participate in the generation of soluble CX3CL1, contributing to pathogenesis in the lacrimal functional unit

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