Corneal collagen as a potential therapeutic target in dry eye disease

Open AccessPublished:April 18, 2021DOI:https://doi.org/10.1016/j.survophthal.2021.04.006

      Abstract

      Dry eye disease (DED) is a major cause of ocular discomfort, inflammation and dysfunction worldwide. Tear film instability in DED both causes and is exacerbated by disruption of the corneal epithelium. This tandem leads to a cycle of inflammation at the corneal surface involving immune cell dysregulation and increased chemokines and cytokines, which activate mitogen-activated protein kinases in the epithelium and elevates matrix metalloproteinases (MMPs). We review evidence suggesting that corneal collagen might be highly susceptible in DED to MMP-induced disruption, digestion, and thinning. We also summarize that collagen is far from inert and contains binding sites that serve as ligands for multiple inflammatory and immune regulators. Fragmented collagen not only challenges these receptor-ligand binding relationships, but also can promote recruitment and motility of pro-inflammatory immune cells. Current physician-directed therapies for DED focus on reducing inflammation, but do not directly ameliorate the underlying corneal damage that could exacerbate surface inflammation. We argue that an important gap in practice is lack of a direct therapeutic reparative for damaged corneal collagen, which is slow to heal, and likely amplifies sight-threatening inflammation. Healing fragmented collagen in the cornea may represent a more effective means to interrupt the “vicious cycle” of inflammation in DED and other conditions that damages, sometimes irreversibly, the ocular surface.

      Keywords

      1. Introduction

      Dry eye disease (DED) afflicts an estimated 5-30% of the world's population and over 15 million people in the United States alone.
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      While lubricants may have an incremental positive influence on corneal structure,
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      a better approach to DED, as well as other forms of ocular surface disease, may be a therapy that addresses the structural damage to the cornea in DED as a means of suppressing or even eliminating the cycle of inflammation that is so deleterious to positive clinical outcomes. In the ensuing sections we will focus on the inflammatory etiology of DED, its influence on corneal structures, and the role that collagen plays as a mediator of inflammation at the ocular surface and in the cornea.

      2. Collagen composition differs across major corneal structures

      A signature feature of all collagens is their triple helical structure– a set of three polypeptide chains comprising repeating sequences of glycine-x-y triplets where x and y often (but not always) represent proline and hydroxyproline.
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      The cornea is a highly collagenous structure in which several types of collagen distribute across five primary layers (Fig. 1). On the anterior side, the corneal epithelium consists of multiple layers of non-keratinized squamous (or flattened) epithelial cells that serve as a cellular barrier to environmental, microbial, and inflammatory insults.
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      They produce a basement membrane that, like all basement membranes, is a specialized extracellular matrix that provides scaffolding during migration, differentiation, and maintenance of the epithelium. In a healthy eye the basement membrane is renewed by the basal epithelial cells as they regenerate and migrate from the outside edge of the cornea towards the center on a continuous basis. This process is challenged in many diseases and conditions, including keratoconus, corneal dystrophies, and recurrent corneal erosion.
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      Fig 1
      Fig. 1Distribution of Collagen in the Cornea. Schematic representation of the basic structure of the cornea (left) shows primary layers from epithelium (anterior) to endothelium (posterior). Several major collagen types distribute differentially across layers (right). Basal epithelial cells produce a basement membrane (type IV collagen) that adheres via hemidesmosome attachments to underlying Bowman layer, with type VII collagen-rich fibrils extending to extracellular matrix within the stroma, which contains collagen-producing keratocytes and type I collagen-rich lamellae. Endothelial-derived Descemet membrane comprises not only type IV, but also type VIII collagen, which is specific to Descemet membrane in the cornea.
      The epithelial basement membrane is comprised primarily of four components: collagens, laminins, proteoglycans, and nidogens. Of these, the non-fibrillar type IV collagen is the most abundant and provides the greatest adherence for the epithelium; laminins represent the most prevalent of the non-collagenous proteins.
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      Proteoglycans are known to interact with collagen fibrils in the corneal stroma and to organize the collagen stroma. Indeed, a mutation in the gene encoding the core protein of a particular dermatan sulfate proteoglycan located in the cornea (decorin) is associated with stromal corneal dystrophy.
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      Nidogens interact with other basement components in the cornea, such as laminins and collagens, to organize and stabilize the basement membrane.
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      Sometimes called Bowman membrane, this layer consists of randomly oriented collagen fibrils spanning the basement membrane anteriorly and the collagen lamellae of the corneal stroma posteriorly. The major component is type I collagen, followed by types III, V, and XII,
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      with type XII contributing to the surface stability of the type I fibrils, as indicated by in vitro studies.
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      The integrity and shape of the stroma is maintained by keratocytes, which are specialized fibroblasts that produce collagen, glycosaminoglycans, and matrix metalloproteinases (or MMPs).
      The posterior-most layer of the cornea is the endothelium, which separates the stroma from the aqueous chamber of the eye. The endothelium contains a single layer of flat, polygonal specialized epithelial cells that maintain the relatively dehydrated state of the stroma through a series of ionic pumps.
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      In contrast to the basal epithelium, cell replacement in the endothelium following insult or apoptosis does not occur via mitosis, but rather through centripetal migration. The endothelium is separated from the overlying stroma by its own basement membrane called Descemet membrane. Like other basement membranes, Descemet membrane is primarily type IV collagen, but also contains type VIII collagen, which enables hexagonal lattice structures.
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      3. Epithelial inflammation increases matrix metalloproteinase activity

      The tear film itself is highly complex, comprising multiple layers that originate as secretions from adjacent tissues.
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      In DED, apoptotic loss of conjunctiva goblet cells reduces mucin biosynthesis and disrupts the protective barrier over the apical epithelium and the superficial epithelial microvilli.
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      This reduction also depletes the tear film of critical immune regulators like transforming growth factor-β2 (TGF-β2), which inhibits activation of antigen-presenting dendritic cells on the ocular surface.
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      Tears also contain interleukin- 1-receptor antagonist (IL-1RA), which in vitro studies show blocks inflammatory signals initiated by IL-1 expressed constitutively by apical corneal epithelial cells.
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      The tear film contains several other anti-inflammatory factors, including tissue inhibitor of matrix metalloproteinase (TIMP-1), a glycoprotein that counters the action of MMPs in degradation of extracellular matrix and has anti-nociceptive effects.
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      Thus, loss of tear film in DED disrupts homeostasis of the ocular surface in part by reducing regulators of inflammation and immunity.
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      A secondary but equally pathogenic ramification of DED is thinning of the corneal epithelium itself, the degree of which reflects disease severity, as shown by human tissue and animal studies.
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      Evaporative stress also causes gaps to form between adjacent epithelial cells, evident with the finding of superficial punctate keratitis (SPK) in human patients.
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      The degree of SPK is reflected directly in severity of symptoms that include ocular burning, scratchiness and irritation, and blurred vision. Unfortunately, many topical drugs used to treat DED contain preservatives that are cytotoxic to the epithelium.
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      Damage to the basement membrane of the epithelial layer allows penetration of inflammatory cytokines, chemokines and other signaling molecules (including TGF-β) to the corneal stroma that can induce the differentiation of myofibroblast precursor cells, promoting haze, opacity and surface irregularity.
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      Stressed epithelial cells in DED release IL-1 (IL-1α and IL-1β) and IL-2, which can induce increased levels of other interleukins involved in inflammatory signaling, including IL-6 and IL-8,
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      Focus on molecules: interleukin-1: a master regulator of the corneal response to injury.
      as well as tumor necrosis factor α (TNFα), which perpetuates epithelial apoptosis and inhibits renewal and migration.
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      In DED epithelial lesions from chronic desiccation result in multiple local inflammatory and immune-activating reactions that lead to increased secretion of MMPs.
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      For example, IL-8 in DED patients promotes the recruitment of T lymphocytes that can damage the ocular surface by producing cytokines such as IL-17 and interferon gamma (IFN-γ), both of which are linked to increased MMPs.
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      Also, evidence primarily from animal models of DED indicates that apoptotic thinning of the epithelial layer in DED involves activation of mitogen-activated protein kinases (MAPKs), nuclear factor κB (NFκB), and IFNγ-dependent pathways.
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      Experimental dry eye stimulates production of inflammatory cytokines and MMP-9 and activates MAPK signaling pathways on the ocular surface.
      ,
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      What we have learned from animal models of dry eye.
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      Dry eye as a mucosal autoimmune disease.
      In turn, activation of MAPKs and increased release of additional cytokines like IL-1α and IL-6 from dying epithelial cells stimulate production of a number of MMPs.
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      Of these, biochemical studies from mouse tissue indicate that MMP-1, -3, -9, -10, and -13 in particular contribute to the inflammatory milieu.
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      TIMP-1 attenuates the development of inflammatory pain through MMP-dependent and receptor-mediated cell signaling mechanisms.
      In the next section we will examine the ramifications of increased MMP levels and activation on collagen throughout the cornea.

      4. Matrix metalloproteinase activity in DED degrades corneal architecture

      The sequelae of events that disrupt the corneal surface in DED clearly are inflammatory in nature, involving increased levels of MMPs, cytokines, and chemokines, all of which have secondary pro-inflammatory effects that often cascade and amplify each other through positive feedback. These relationships are summarized in a recent review of clinical and experimental studies.
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      While it is widely acknowledged that inflammation in DED influences the integrity and function of cellular structures (conjunctival goblet cells, corneal epithelium, stromal keratocytes, and so on), inflammation also could directly challenge integrity of the collagen substrate that forms the backbone of the cornea and the basis of the integrity of the epithelial cell layer.
      The degradation of collagens depends on an initial enzymatic break of at least one of the strands forming the collagen trimer. Such a break causes uncoiling of the triple helical structure that in turn allows for subsequent cleavage and degradation. Like elastin, collagens are resistant to most proteolytic enzymes, thus adding to their general stability, but are susceptible to cleavage by MMPs, which comprise many types.
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      Of the MMPs notably elevated at the corneal surface in humans and animal models of DED (MMP-1, -3, -9, -10, and -1334,70), the collagenases MMP-1 and MMP-13 cleave triple-helical fibrillar collagens, while the gelatinase MMP-9 digests unwound collagen.
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      The effects of MMPs released by injured corneal epithelium in DED could act immediately at the surface. The type IV collagen so critical to the integrity of the epithelial basement membrane is susceptible to digestion by MMP-9 and the stromelysins MMP-3 and -10.
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      MMP-9 and -13 also degrade laminin, which is abundant in the basement membrane.
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      While MMP-9 is important to the cleavage of basement membrane components that enables migration of new cells under normal conditions, in recurrent injury, increased MMP-9 impedes reattachment of the collagen fibrils of migrating epithelial cells to underlying Bowman layer.
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      Also, human tissue studies indicate that an aberrant basement membrane can prohibit migration of epithelial cells to the apical surface for shedding, leading to cystic accumulation of cellular debris that compromises adherence of the epithelium to underlying Bowman layer.
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      In other corneal dystrophies, degradation of the epithelial basement membrane directly leads to destruction of Bowman layer, as shown by ultrastructural analysis of human cornea.
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      Type VII collagen that forms the fibrils of the epithelial basement membrane and Descemet membrane is susceptible to the collagenase MMP-1, which also cleaves type VIII collagen found in Descemet' membrane.
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      MMP-1 also disturbs the helical region of both type I collagen and corneal stroma and type III collagen found in the Bowman layer. This initial injury increases susceptibility of types I and V collagen to further digestion via MMP-9,
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      consistent with evidence from human patients that DED includes breaks in the Bowman layer associated with increased density of dendritic cells on the ocular surface.
      • Shetty R
      • Deshpande K
      • Deshmukh R
      • et al.
      Bowman break and subbasal nerve plexus changes in a patient with dry eye presenting with chronic ocular pain and vitamin D deficiency.
      Increased levels of collagenases in DED (MMP-1 and MMP-13) can also induce apoptosis of collagen-producing keratocytes (specialized corneal fibroblasts) in the corneal stroma,
      • Meyer LM
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      • Wegener AR
      [Schleimpflug photography detects alterations in corneal density and thickness in patients with dry eye disease].
      ,
      • Zhou HY
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      Role of corneal collagen fibrils in corneal disorders and related pathological conditions.
      further exacerbating loss of collagen.
      Thus, recurrent stress to the corneal epithelium layer is compounded by MMP- induced damage to the basement membrane, which in turn perpetuates injury to underlying collagen-rich tissue. These interactions likely contribute to (or even explain) thinning of the stroma, of Descemet membrane, and of the endothelial layer in DED, all of which track closely with severity of symptoms.
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      Effect of viscous agents on corneal density in dry eye disease.

      5. Damaged collagen in DED could amplify inflammation

      Inflammation in DED likely degrades corneal structure by increasing collagen exposure to the activity of MMPs, but in turn could also be exacerbated by that very damage, leading to a destructive cycle of inflammation and structural damage to the cornea. Far from inert, collagen within the layers of the cornea – like collagen in cellular networks in other tissues – is involved in a multitude of physiological processes, including mediation of cell proliferation, migration and adhesion, and inflammation. Collagen remodeling is a crucial process in tissue homeostasis and many other constitutive physiological events, and collagen is the major extracellular matrix component that interacts with almost all cell types. These functionalities arise from collagen's natural structure.
      During normal turnover, intact collagen in its stable triple-helix structure is incorporated into the extracellular matrix, presenting multiple binding sites that serve as ligands for several cell surface receptors, including integrins, discoidin domain receptors, glycoprotein VI, and proteoglycan receptors.
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      Collagens are functional, high affinity ligands for the inhibitory immune receptor LAIR-1.
      Upon binding, such receptors modulate other molecules and cellular pathways related to remodeling, inflammation and immune regulation. Thus, aside from their structural role as scaffolding proteins, nearly all collagens initiate cellular signaling cascades by activating specific receptors on the cell surface. Signaling events mediated by these binding interactions are necessary for proliferation, motility, adhesion and survival of overlying epithelial cells as they regenerate and form new apical tissue, as in embryonic development.
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      Integrins, for example, are cell adhesion structures that play critical roles in cellular signaling, migration and survival, through their interactions with extracellular matrix components and other intra- and extracellular signaling moieties (e.g.,
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      ). They function as heterodimers, some of which bind to types I and IV collagen.
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      • Wang J
      • Mort JS
      • Komarova SV
      Collagen type I as a ligand for receptor-mediated signaling.
      Intact collagen, however, also provides ligand sites for signaling pathways involved in inflammation, including the leukocyte receptor complex (LRC), which comprises a diverse group of cell surface receptors primarily expressed by immune cells. Interestingly, the most highly expressed gene in cells of human Meibomian glands encodes a member of this group, the leukocyte-associated immunoglobulin-like receptor or LAIR-1,
      • Bron AJ
      • de Paiva CS
      • Chauhan SK
      • et al.
      TFOS DEWS II pathophysiology report.
      ,
      • Liu S
      • Richards SM
      • Lo K
      • et al.
      Changes in gene expression in human meibomian gland dysfunction.
      LAIR-1 (also called CD305) is an inhibitory receptor that attenuates activation of most immune cells, including T cells, B cells, mast cells, eosinophils, basophils and monocytes/macrophages, while suppressing proinflammatory cytokine production upon binding ligand sites on intact triple helical collagen.
      • Lebbink RJ
      • van den Berg MC
      • de Ruiter T
      • et al.
      The soluble leukocyte-associated Ig-like receptor (LAIR)-2 antagonizes the collagen/LAIR-1 inhibitory immune interaction.
      ,
      The epidemiology of dry eye disease: report of the epidemiology subcommittee of the international dry eye workShop.
      Both type I and type III collagen contain the Gly-Pro-Hyp motif with capacity for high-affinity binding to LAIR-1, for which they serve as functional inhibitory ligands.
      • Coelho NM
      • McCulloch CA
      Contribution of collagen adhesion receptors to tissue fibrosis.
      ,
      • Meyaard L
      The inhibitory collagen receptor LAIR-1 (CD305).
      There is also evidence that type IV collagen binds LAIR-1.
      • Lebbink RJ
      • van den Berg MC
      • de Ruiter T
      • et al.
      The soluble leukocyte-associated Ig-like receptor (LAIR)-2 antagonizes the collagen/LAIR-1 inhibitory immune interaction.
      For LAIR-1, the strength and efficacy of inhibitory signaling depends directly on the activating strength of the cross-linking interaction with collagen.
      • Meyaard L
      The inhibitory collagen receptor LAIR-1 (CD305).
      Crosslinking of LAIR-1 by triple helical collagens at glycine-proline-hydroxyproline repeats (GPO10) inhibits immune cells while diminished LAIR-1 or its binding sites in disrupted collagen exacerbates immune cell activation and leads to chronic inflammation in tissue.
      • Lebbink RJ
      • de Ruiter T
      • Adelmeijer J
      • et al.
      Collagens are functional, high affinity ligands for the inhibitory immune receptor LAIR-1.
      Degradation of collagen produces much shorter triple-helical proteins, along with various single strand fragments. Thus, collagens through binding LAIR-1 set a threshold for inhibition or activation of immune cells entering damaged tissue based on the level of intact binding sites.
      Meyaard L: LAIR and collagens in immune regulation.
      In DED, MMP-induced damage to Bowman layer, stroma and both epithelial basement and Descemet membranes could disrupt available binding sites for LAIR-1 on the collagen comprising these layers, types I, III, and IV, thus compromising its inhibitory actions on immune cells at the ocular surface.
      The discoidin domain receptors (DDR1 and DDR2) are receptor tyrosine kinases that bind collagen and serve as cellular sensors of environmental cues involved in cellular invasion, migration and adhesion and in remodeling of extracellular matrix.
      • Boraschi-Diaz I
      • Wang J
      • Mort JS
      • Komarova SV
      Collagen type I as a ligand for receptor-mediated signaling.
      Both DDR1 and DDR2 bind to the fibrillary collagen types I-III and V, while DDR1 also binds type IV collagen.
      • Fu HL
      • Valiathan RR
      • Arkwright R
      • et al.
      Discoidin domain receptors: unique receptor tyrosine kinases in collagen-mediated signaling.
      Upon binding, DDR1 and DDR2 activate via tyrosine autophosphorylation, which is unusually slow compared to other receptor tyrosine kinases.
      • Boraschi-Diaz I
      • Wang J
      • Mort JS
      • Komarova SV
      Collagen type I as a ligand for receptor-mediated signaling.
      Of the family of receptor tyrosine kinases, only the DDRs are activated by an extracellular matrix protein.
      • Valiathan RR
      • Marco M
      • Leitinger B
      • et al.
      Discoidin domain receptor tyrosine kinases: new players in cancer progression.
      Functionally, DDR1 and DDR2 are best characterized for their role in various metastatic cancers, promoting an invasive phenotype for proliferating cells; however, through an E-cadherin-dependent pathway, activated DDR1 contributes to epithelial cell differentiation and spreading.
      • Wang CZ
      • Yeh YC
      • Tang MJ
      DDR1/E-cadherin complex regulates the activation of DDR1 and cell spreading.
      ,
      • Yeh YC
      • Wu CC
      • Wang YK
      • Tang MJ
      DDR1 triggers epithelial cell differentiation by promoting cell adhesion through stabilization of E-cadherin.
      The receptors could play a similar role in the cornea. Both DDR1 and DDR2 are expressed in all major cell types of the human cornea and in particular within the epithelium,
      • Mohan RR
      • Mohan RR
      • Wilson SE
      Discoidin domain receptor (DDR) 1 and 2: collagen-activated tyrosine kinase receptors in the cornea.
      which through type IV collagen in the basement membrane could activate DDR1. Since collagen types I and III are represented in Bowman's layer and I and V in the stroma, there is ample substrate for activation of both receptors.
      Inhibitors of receptor tyrosine kinases used in cancer therapies, including several targeting DDR1 and DDR2, often cause DED in patients in conjunction with corneal inflammation (keratitis), conjunctivitis, and corneal thinning with erosion of the epithelium.
      • Davis ME
      Ocular Toxicity of Tyrosine Kinase Inhibitors.
      That these major side effects occur indicates that collagen activation of DDR1 and DDR2 is necessary to maintain homeostasis of the immune environment of the ocular surface. In fact, neither DDR1 nor DDR2 are able to bind to degraded collagen or to individual collagen α-chains but rather only to native triple helical collagens.
      • Boraschi-Diaz I
      • Wang J
      • Mort JS
      • Komarova SV
      Collagen type I as a ligand for receptor-mediated signaling.
      ,
      • Fu HL
      • Valiathan RR
      • Arkwright R
      • et al.
      Discoidin domain receptors: unique receptor tyrosine kinases in collagen-mediated signaling.
      Thus, MMP-induced degradation of collagenous structures in the cornea during DED could very likely reduce binding of key modulators of inflammation, which may exacerbate the cycle of damage and impede repair of the corneal surface (Fig. 2). Interestingly, type IV collagen comprising the epithelial basement and Descemet' membrane is among the highly glycosylated collagens, the fragmentation of which promotes binding of the endocytic cell-surface receptor uPARAP/Endo180 (urokinase plasminogen activator receptor-associated protein
      • Jurgensen HJ
      • Madsen DH
      • Ingvarsen S
      • et al.
      A novel functional role of collagen glycosylation: interaction with the endocytic collagen receptor uparap/ENDO180.
      ). This receptor is a member of the macrophage mannose receptor family of endocytic transmembrane glycoproteins and is specifically involved in the uptake and lysosomal degradation of collagen fragments generated by initial MMP-mediated cleavage, as shown by studies of tumor cells.
      • Noel A
      • Gutierrez-Fernandez A
      • Sounni NE
      • et al.
      New and paradoxical roles of matrix metalloproteinases in the tumor microenvironment.
      uPARAP/Endo180 also facilitates adherence and migration of fibroblasts along fibrillary collagen.
      • Boraschi-Diaz I
      • Wang J
      • Mort JS
      • Komarova SV
      Collagen type I as a ligand for receptor-mediated signaling.
      ,
      • Jurgensen HJ
      • Madsen DH
      • Ingvarsen S
      • et al.
      A novel functional role of collagen glycosylation: interaction with the endocytic collagen receptor uparap/ENDO180.
      This process is involved in corneal scar formation, a common complication of dry eye.
      • Zhang X
      • VJ M
      • Qu Y
      • et al.
      Dry eye management: targeting the ocular surface microenvironment.
      ,
      • Zhou HY
      • Cao Y
      • Wu J
      • Zhang WS
      Role of corneal collagen fibrils in corneal disorders and related pathological conditions.
      Finally, in vitro work using fibroblasts demonstrates that fragments of type I collagen (as found in the Bowman layer and stroma) promote the release and activity of IL-1β,
      • Castillo-Briceno P
      • Bihan D
      • Nilges M
      • et al.
      A role for specific collagen motifs during wound healing and inflammatory response of fibroblasts in the teleost fish gilthead seabream.
      which contributes to recruitment and motility of pro-inflammatory immune cells and is implicated in corneal damage during human DED.
      • Hessen M
      • Akpek EK
      Dry eye: an inflammatory ocular disease.
      Fig 2
      Fig. 2Corneal Collagen as an Inflammatory Mediator and Therapeutic Target. In DED, instability of the tear film and thinning of the epithelium exacerbate one another, leading to inflammation and immune dysregulation of the corneal surface. Inflammation in turn leads to activation of mitogen-activated protein kinase (MAPK) and subsequent elevation of matrix metalloproteinases (MMPs), which cleave and digest collagen throughout the cornea. Collagen degradation not only impedes healing of the epithelium, but also contributes to inflammatory signaling by reducing ligand binding sites for important mediators. Repairing damaged collagen directly could interrupt (dashed lines) epithelial thinning and reduce inflammation by restoring inflammatory and immune modulator binding sites and promote migration and adherence of the corneal epithelium (dashed arrow) by restoring cell migration and proliferation.

      6. Conclusion: targeting collagen in dry eye could be therapeutic

      The evidence reviewed thus far indicates that collagen damage in the cornea could exacerbate the inflammatory “vicious cycle” of DED (Fig. 2). To date, there has not been a direct therapeutic reparative for damaged collagen in DED or other corneal conditions and injury. This is an important gap in pharmaceutical interventions, DED aside.
      • Lee T
      The Ins and Outs of Corneal Wound Healing.
      Anterior ocular surface injuries primarily involve the cornea since ocular fixation tends to direct the cornea towards sources of potential danger. These include abrasions, lacerations, ulcers, thermal injuries, chemical injuries and blunt, concussive injuries. In each of these cases, like DED, a secondary effect of the primary insult is a complex inflammatory cascade that disrupts the immune homeostasis of the ocular surface. In what we have shown here, inflammation not only could damage collagen throughout the cornea, but in turn could amplify the effects of MMP-induced digestion of collagen, which reduces critical receptor-ligand binding sites. Thus, damage to the collagen scaffold would result in slowing of the healing process (in particular, healing of the corneal epithelium) with amplification of sight-threatening inflammation that underlies aberrant healing and even scarring.
      In this sense, current physician-directed therapies for DED that rely heavily on reducing inflammation at the ocular surface have a relatively narrow range of action, since they do nothing directly to reduce the underlying structural damage arising from inflammation and possible corneal neuropathy that occurs in severe DED.
      • Shetty R
      • Deshpande K
      • Deshmukh R
      • et al.
      Bowman break and subbasal nerve plexus changes in a patient with dry eye presenting with chronic ocular pain and vitamin D deficiency.
      As well, long-term use of these traditional therapies is not without risk. For example, ocular application of corticosteroids over time is associated with both cataract and glaucoma and tissue-specific resistance, necessitating the use of alternative and more potent antiinflammatories.
      • Stephenson M
      Dry-Eye therapies: what's next?.
      Thus, there may be considerable advantage to repairing corneal collagen directly.
      As ligand binding sites on collagen are identified, so too are small biomolecules that target such sites as possible therapeutics – including sites involved in tumor progression (like DDR1 and DDR2) and in inflammatory signaling.
      • An B
      • Lin YS
      • Brodsky B
      Collagen interactions: Drug design and delivery.
      The use of collagen peptides to heal fragmented or partially digested collagen has shown promise in a variety of domains.
      • Chattopadhyay S
      • Guthrie KM
      • Teixeira L
      • et al.
      Anchoring a cytoactive factor in a wound bed promotes healing.
      For example, peptides derived from fish scales promote endothelial adherence and reduce TNF-α-induced inflammatory signaling.
      • Subhan F
      • Kang HY
      • Lim Y
      • et al.
      Fish scale collagen peptides protect against CoCl2/TNF-alpha-Induced cytotoxicity and inflammation via inhibition of ROS, MAPK, and NF-kappaB pathways in HaCaT Cells.
      Single strand forms of collagen-like peptides selectively target degraded regions of a collagen triple-helix and strongly hybridize to type I collagen fibrils under physiological conditions.
      • Wang AY
      • Mo X
      • Chen CS
      • Yu SM
      Facile modification of collagen directed by collagen mimetic peptides.
      Such peptide strands ostensibly could be applied as topical agents to heal corneal type I collagen. Collagen peptides can also be crafted to deliver custom bioactive molecules to sites of damaged and unfolded collagen triple-helices to promote healing and reduce inflammation, as in cutaneous wound beds.
      • Chattopadhyay S
      • Guthrie KM
      • Teixeira L
      • et al.
      Anchoring a cytoactive factor in a wound bed promotes healing.
      Already, synthesized collagen peptides intended to promote tear adherence to the ocular surface show promise in animal models of dry eye in reducing surface inflammation and facilitating epithelium stabilization.
      • Lee H
      • Kim CE
      • Ahn BN.
      • et al.
      Anti-inflammatory effect of hydroxyproline-GQDGLAGPK in desiccation stress-induced experimental dry eye mouse.
      These results in total suggest that healing fragmented collagen in the cornea may in the end represent a more effective means to interrupt the non-ending cycle of inflammation in DED and other conditions that damage the ocular surface.

      7. Method of literature search

      We conducted a search of the PubMed database for “dry eye disease” and each of the following keywords: cornea, ocular surface, Bowman's layer, stroma, Descemet's membrane, collagen, collagen fibrils, collagen ligand, matrix metalloproteinase, collagenase, collagen degradation, collagen receptor, inflammation, basement membrane, keratitis, cell migration, cell adhesion, and collagen fragment. We limited the search to articles published in English from 1990 to June 2020, when the search was conducted last. We screen all abstracts and included relevant articles in this review, along with older seminal papers published prior to 1990.

      Conflict of Interest

      B.O.B, E.S, B.J.D.B., S.S.D, and D.J.C. are stakeholders in Stuart Therapeutics, Inc.

      Acknowledgement

      The authors thank Jay S. Pepose, MD, PhD, for helpful comments on the manuscript.

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