The Potential Reversible Transition between Stem Cells and Transient-Amplifying Cells: The Limbal Epithelial Stem Cell Perspective.

Stem cells (SCs) undergo asymmetric division, producing transit-amplifying cells (TACs) with increased proliferative potential that move into tissues and ultimately differentiate into a specialized cell type. Thus, TACs represent an intermediary state between stem cells and differentiated cells. In the cornea, a population of stem cells resides in the limbal region, named the limbal epithelial stem cells (LESCs). As LESCs proliferate, they generate TACs that move centripetally into the cornea and differentiate into corneal epithelial cells. Upon limbal injury, research suggests a population of progenitor-like cells that exists within the cornea can move centrifugally into the limbus, where they dedifferentiate into LESCs. Herein, we summarize recent advances made in understanding the mechanism that governs the differentiation of LESCs into TACs, and thereafter, into corneal epithelial cells. We also outline the evidence in support of the existence of progenitor-like cells in the cornea and whether TACs could represent a population of cells with progenitor-like capabilities within the cornea. Furthermore, to gain further insights into the dynamics of TACs in the cornea, we outline the most recent findings in other organ systems that support the hypothesis that TACs can dedifferentiate into SCs.

In vitro Assay to Examine the Function of Tears on Corneal Epithelial Cells.

Tears contain numerous secreted factors, enzymes, and proteins that help in maintaining the homeostatic condition of the eye and also protect it from the external environment. However, alterations to these enzymes and/or proteins during pathologies such as mechanical injury and viral or fungal infections can disrupt the normal ocular homeostasis, further contributing to disease development. Several tear film components have a significant role in curbing disease progression and promoting corneal regeneration. Additionally, several factors related to disease progression are secreted into the tear film, thereby serving as a valuable reservoir of biomarkers. Tears are readily available and can be collected via non-invasive techniques or simply from contact lenses. Tears can thus serve as a valuable and easy source for studying disease-specific biomarkers. Significant advancements have been made in recent years in the field of tear film proteomics, lipidomics, and transcriptomics to allow a better understanding of how tears can be utilized to gain insight into the etiology of diseases. These advancements have enabled us to study the pathophysiology of various disease states using tear samples. However, the mechanisms by which tears help to maintain corneal homeostasis and how they are able to form the first line of defense against pathogens remain poorly understood and warrant detailed in vitro studies. Herein, we have developed an in vitro assay to characterize the functional importance of patient isolated tears and their components on corneal epithelial cells. This novel approach closely mimics real physiological conditions and could help the researchers gain insight into the underlying mechanisms of ocular pathologies and develop new treatments. Key features • This method provides a new technique for analyzing the effect of tear components on human corneal epithelial cells. • The components of the tears that are altered in response to diseases can be used as a biomarker for detecting ocular complications. • This procedure can be further employed as an in vitro model for assessing the efficacy of drugs and discover potential therapeutic interventions.

Endogenous TSG-6 modulates corneal inflammation following chemical injury.

PURPOSE: Tumor necrosis factor (TNF)-stimulated gene-6 (TSG-6) is upregulated in various pathophysiological contexts, where it has a diverse repertoire of immunoregulatory functions. Herein, we investigated the expression and function of TSG-6 during corneal homeostasis and after injury. METHODS: Human corneas, eyeballs from BALB/c (TSG-6+/+), TSG-6+/- and TSG-6-/- mice, human immortalized corneal epithelial cells and murine corneal epithelial progenitor cells were prepared for immunostaining and real time PCR analysis of endogenous expression of TSG-6. Mice were subjected to unilateral corneal debridement or alkali burn (AB) injuries and wound healing assessed over time using fluorescein stain, in vivo confocal microscopy and histology. RESULTS: TSG-6 is endogenously expressed in the human and mouse cornea and established corneal epithelial cell lines and is upregulated after injury. A loss of TSG-6 has no structural and functional effect in the cornea during homeostasis. No differences were noted in the rate of corneal epithelial wound closure between BALB/c, TSG-6+/- and TSG-6-/- mice. TSG-6-/- mice presented decreased inflammatory response within the first 24 h of injury and accelerated corneal wound healing following AB when compared to control mice. CONCLUSION: TSG-6 is endogenously expressed in the cornea and upregulated after injury where it propagates the inflammatory response following chemical injury.

Age related changes in hyaluronan expression leads to Meibomian gland dysfunction.

The prevalence of dry eye disease (DED) ranges from ∼5 to 50 % and its associated symptoms decrease productivity and reduce the quality of life. Approximately 85 % of all DED cases are caused by Meibomian gland dysfunction (MGD). As humans and mice age, their Meibomian glands (MGs) undergo age-related changes resulting in age related-MGD (ARMGD). The precise cause of ARMGD remains elusive, which makes developing therapies extremely challenging. We previously demonstrated that a hyaluronan (HA)-rich matrix exists surrounding the MG, regulating MG morphogenesis and homeostasis. Herein, we investigated whether changes to the HA matrix in the MG throughout life contributes towards ARMGD, and whether altering this HA matrix can prevent ARMGD. For such, HA synthase (Has) knockout mice were aged and compared to age matched wild type (wt) mice. MG morphology, lipid production, PPARγ expression, basal cell proliferation, stem cells, presence of atrophic glands and MG dropout were analyzed at 8 weeks, 6 months, 1 year and 2 years of age and correlated with the composition of the HA matrix. We found that as mice age, there is a loss of HA expression in and surrounding the MGs of wt mice, while, in contrast, Has1-/-Has3-/- mice present a significant increase in HA expression through Has2 upregulation. At 1 year, Has1-/-Has3-/- mice present significantly enlarged MGs, compared to age-matched wt mice and compared to all adult mice. Thus, Has1-/-Has3-/- mice continue to develop new glandular tissue as they age, instead of suffering MG atrophy. At 2 years, Has1-/-Has3-/- mice continue to present significantly larger MGs compared to age-matched wt mice. Has1-/-Has3-/- mice present increased lipid production, increased PPARγ expression and an increase in the number of proliferating cells when compared to wt mice at all-time points analyzed. Taken together, our data shows that a loss of the HA matrix surrounding the MG as mice age contributes towards ARMGD, and increasing Has2 expression, and consequently HA levels, prevents ARMGD in mice.

Recent advances in age-related meibomian gland dysfunction (ARMGD).

Meibomian glands (MGs), located within the tarsal plate of the eyelid, secrete meibum which is the lipid-rich secretion necessary for stabilizing the tear film and preventing tear evaporation. Changes in the quality and quantity of meibum produced causes MG dysfunction (MGD), the leading cause of evaporative dry eye disease (EDED). MGD is an underdiagnosed disease and it is estimated that, in the US, approximately 70 % of the population over 60 have MGD. Three forms of MGD occur based on their meibum secretion: hyposecretory, obstructive, and hypersecretory MGD. The pathophysiology of MGD remains poorly understood, however aging is the primary risk factor. With age, MGs undergo various age-related changes, including decreased acinar basal cell proliferation, hyperkeratinization, MG atrophy, and eventual MG drop-out, leading to age-related MGD (ARMGD). Additionally, studies have suggested that MGs can suffer inflammatory cell infiltration and changes innervation patterns with aging, which could also contribute towards ARMGD. This review focuses on how the aging process affects the MG, and more importantly, how age-related changes to the MG can lead to MG atrophy and MG drop-out, ultimately leading to ARMGD. This review also highlights the most recent developments in potential therapeutic interventions for ARMGD.

Characterization of the Limbal Epithelial Stem Cell Niche.

Purpose: Limbal epithelial stem cells (LESCs) reside within a LSC niche (LSCN). We recently identified that hyaluronan (HA) is a major constituent of the LSCN, and that HA is necessary for maintaining LESCs in the "stem cell" state, both in vitro and in vivo. Herein, we characterized the LSCN to identify key components of the HA-specific LSCN. Methods: The cornea and limbal rim were dissected from mouse corneas, subjected to mRNA extraction, and sequenced using a NextSeq 500 (Illumina) and data processed using CLC Genomics Workbench 20 (Qiagen) and the STRING database to identify key components of the LSCN. Their expression was confirmed by real-time PCR, Western blotting, and immunohistochemistry. Furthermore, the differential expression of key compounds in different corneal cell types were determined with single-cell RNA sequencing. Results: We identified that the hyaladherins inter-alpha-inhibitor (IαI), TSG-6 and versican are highly expressed in the limbus. Specifically, HA/HC complexes are present in the LSCN, in the stroma underlying the limbal epithelium, and surrounding the limbal vasculature. For IαI, heavy chains 5 and 2 (HC5 and HC2) were found to be the most highly expressed HCs in the mouse and human limbus and were associate with HA-forming HA/HC-specific matrices. Conclusions: The LSCN contains HA/HC complexes, which have been previously correlated with stem cell niches. The identification of HA/HC complexes in the LSCN could serve as a new therapeutic avenue for treating corneal pathology. Additionally, HA/HC complexes could be used as a substrate for culturing LESCs before LESC transplantation.

Rational design and synthesis of lumican stapled peptides for promoting corneal wound healing.

PURPOSE: Lumican is a major extracellular matrix (ECM) component in the cornea that is upregulated after injury and promotes corneal wound healing. We have recently shown that peptides designed based on the 13 C-terminal amino acids of lumican (LumC13 and LumC13C-A) are able to recapitulate the effects of lumican on promoting corneal wound healing. Herein we used computational chemistry to develop peptide mimetics derived from LumC13C-A with increased stability and half-life that are biologically active and non-toxic, thereby promoting corneal wound healing with increased pharmacological potential. METHODS: Different peptides staples were rationalized using LumC13C-A sequence by computational chemistry, docked to TGFßRI and the interface binding energies compared. Lowest scoring peptides were synthesized, and the toxicity of peptides tested using CCK8-based cell viability assay. The efficacy of the stapled peptides at promoting corneal wound healing was tested using a proliferation assay, an in vitro scratch assay using human corneal epithelial cells and an in vivo murine corneal debridement wound healing model. RESULTS: Binding free energies were calculated using MMGBSA algorithm, and peptides LumC13C and LumC13S5 displayed superior binding to ALK5 compared to the non-stapled peptide LumC13C-A. The presence of the hydrocarbon staple in LumC13C enhances the stability of the α-helical conformation, thereby facilitating more optimal interactions with the ALK5 receptor. The stapled peptides do not present cytotoxic effects on human corneal epithelial cells at a 300 nM concentration. Similar to lumican and LumC13C-A, both C13C and LumC13S5 significantly promote corneal wound healing both in vitro and in vivo. CONCLUSIONS: Highly stable and non-toxic stapled peptides designed based on LumC13, significantly promote corneal wound healing. As a proof of principle, our data shows that more stable and pharmacologically relevant peptides can be designed based on endogenous peptide sequences for treating various corneal pathologies.

ROS-Mediated Fragmentation Alters the Effects of Hyaluronan on Corneal Epithelial Wound Healing.

A buildup of reactive oxygen species (ROS) occurs in virtually all pathological conditions. Hyaluronan (HA) is a major extracellular matrix component and is susceptible to oxidation by reactive oxygen species (ROS), yet the precise chemical structures of oxidized HA products (oxHA) and their physiological properties remain largely unknown. This study characterized the molecular weight (MW), structures, and physiological properties of oxHA. For this, high-molecular-weight HA (HMWHA) was oxidized using increasing molar ratios of hydrogen peroxide (H2O2) or hypochlorous acid (HOCl). ROS lead to the fragmentation of HA, with the oxHA products produced by HOCl exhibiting an altered chemical structure while those produced by H2O2 do not. HMWHA promotes the viability of human corneal epithelial cells (hTCEpi), while low MWHA (LMWHA), ultra-LMWHA (ULMWHA), and most forms of oxHA do not. HMWHA and LMWHA promote hTCEpi proliferation, while ULMWHA and all forms of oxHA do not. LMWHA and some forms of oxHA promote hTCEpi migration, while HMWHA does not. Finally, all native forms of HA and oxHA produced by HOCl promote in vivo corneal wound healing, while oxHA produced by H2O2 does not. Taken together, our results show that HA fragmentation by ROS can alter the physiological activity of HA by altering its MW and structure.

Novel Peptides with Dual Properties for Treating Pseudomonas aeruginosa Keratitis: Antibacterial and Corneal Wound Healing.

The corneal epithelium is a layer in the anterior part of eye that contributes to light refraction onto the retina and to the ocular immune defense. Although an intact corneal epithelium is an excellent barrier against microbial pathogens and injuries, corneal abrasions can lead to devastating eye infections. Among them, Pseudomonas aeruginosa-associated keratitis often results in severe deterioration of the corneal tissue and even blindness. Hence, the discovery of new drugs able not only to eradicate ocular infections, which are often resistant to antibiotics, but also to elicit corneal wound repair is highly demanded. Recently, we demonstrated the potent antipseudomonal activity of two peptides, Esc(1-21) and its diastereomer Esc(1-21)-1c. In this study, by means of a mouse model of P. aeruginosa keratitis and an in vivo corneal debridement wound, we discovered the efficacy of these peptides, particularly Esc(1-21)-1c, to cure keratitis and to promote corneal wound healing. This latter property was also supported by in vitro cell scratch and ELISA assays. Overall, the current study highlights Esc peptides as novel ophthalmic agents for treating corneal infection and injury, being able to display a dual function, antimicrobial and wound healing, rarely identified in a single peptide at the same micromolar concentration range.

Conditional deletion of CD25 in the corneal epithelium reveals sex differences in barrier disruption.

PURPOSE: IL-2 promotes activation, clonal expansion, and deletion of T cells. IL-2 signals through its heterotrimeric receptor (IL-2R) consisting of the CD25, CD122 and CD132 chains. CD25 knockout (KO) mice develop Sjögren Syndrome-like disease. This study investigates whether corneal CD25/IL-2 signaling is critical for ocular health. METHODS: Eyes from C57BL/6 mice were collected and prepared for immunostaining or in-situ hybridization. Bulk RNA sequencing was performed on the corneal epithelium from wild-type and CD25KO mice. We generated a conditional corneal-specific deletion of CD25 in the corneal epithelium (CD25Δ/ΔCEpi). Corneal barrier function was evaluated based on the uptake of a fluorescent dye. Mice were subjected to unilateral corneal debridement, followed by epithelial closure over time. RESULTS: In C57BL/6 mice, CD25 mRNA was expressed in ocular tissues. Protein expression of CD25, CD122, and CD132 was confirmed in the corneal epithelium. Delayed corneal re-epithelization was seen in female but not male CD25KO mice. There were 771 differentially expressed genes in the corneal epithelium of CD25KO compared to wild-type mice. While barrier function is disrupted in CD25Δ/ΔCEpi mice, re-epithelialization rates are not delayed. CONCLUSIONS: All three chains of the IL-2R are expressed in the corneal epithelium. Our results indicate for the first time, deleting CD25 systemically in all tissues in the mouse and deleting CD25 locally in just the corneal epithelium compromises corneal epithelial barrier function, leading to dry eye disease in female mice. Future studies are needed to delineate the pathways used by IL-2 signaling to influence cornea homeostasis.

Small leucine rich proteoglycans: Biology, function and their therapeutic potential in the ocular surface.

Small leucine rich proteoglycans (SLRPs) are the largest family of proteoglycans, with 18 members that are subdivided into five classes. SLRPs are small in size and can be present in tissues as glycosylated and non-glycosylated proteins, and the most studied SLRPs include decorin, biglycan, lumican, keratocan and fibromodulin. SLRPs specifically bind to collagen fibrils, regulating collagen fibrillogenesis and the biomechanical properties of tissues, and are expressed at particularly high levels in fibrous tissues, such as the cornea. However, SLRPs are also very active components of the ECM, interacting with numerous growth factors, cytokines and cell surface receptors. Therefore, SLRPs regulate major cellular processes and have a central role in major fundamental biological processes, such as maintaining corneal homeostasis and transparency and regulating corneal wound healing. Over the years, mutations and/or altered expression of SLRPs have been associated with various corneal diseases, such as congenital stromal corneal dystrophy and cornea plana. Recently, there has been great interest in harnessing the various functions of SLRPs for therapeutic purposes. In this comprehensive review, we describe the structural features and the related functions of SLRPs, and how these affect the therapeutic potential of SLRPs, with special emphasis on the use of SLRPs for treating ocular surface pathologies.

Meibomian gland development: Where, when and how?

The Meibomian gland (MG) is an indispensable adnexal structure of eye that produces meibum, an important defensive component for maintaining ocular homeostasis. Normal development and maintenance of the MGs is required for ocular health since atrophic MGs and disturbances in composition and/or secretion of meibum result in major ocular pathologies, collectively termed as Meibomian gland dysfunction (MGD). Currently available therapies for MGD merely provide symptomatic relief and do not treat the underlying deficiency of the MGs. Hence, a thorough understanding of the timeline of MG development, maturation and aging is required for regenerative purposes along with signaling molecules & pathways controlling proper differentiation of MG lineage in mammalian eye. Understanding the factors that contribute to the development of MGs, developmental abnormalities of MGs, and changes in the quality & quantity of meibum with developing phases of MGs are essential for developing potential treatments for MGD. In this review, we compiled a timeline of events and the factors involved in the structural and functional development of MGs and the associated developmental defects of MGs during development, maturation and aging.

Characterization of the Molecular Weight of Hyaluronan in Eye Products Using a Novel Method of Size Exclusion High-Pressure Liquid Chromatography.

Purpose: Hyaluronan (HA) exists in two forms, high molecular weight HA (HMWHA) and low molecular weight HA (LMWHA), which have distinct physiological functions. Therefore it is imperative to know the form of HA within pharmaceutical products, including eye products. This study developed an accurate, sensitive, and quantitative method to characterize the form of HA in eye products. Thereafter, the effects of the HA-containing eye products on corneal wound healing were investigated. Methods: The MW distributions and concentrations of HA in over the counter eye products were determined by size exclusion chromatography (SEC) high-pressure liquid chromatography (HPLC). The effects of the eye products containing HA on corneal wound healing were characterized both in vitro and in vivo using the scratch assay and the debridement wound model, respectively. Results: The concentrations and MWs of HA were successfully determined within a range of 0.014 to 0.25 mg/mL using SEC HPLC. The concentrations of HA in the ophthalmic products varied from 0.14 to 4.0 mg/mL and the MWs varied from ∼100 kDa to >2500 kDa. All but one HA-containing eye product had an inhibitory effect on corneal wound healing, whereas pure HA promoted corneal wound healing. Conclusions: A novel SEC-HPLC method was developed for quantifying and characterizing the MW of HA in eye products. Although HA promoted corneal wound healing, HA-containing eye products inhibited corneal wound healing, likely caused by preservatives. Translational Relevance: SEC-HPLC could be implemented as a routine method for determining the form of HA in commercially available ophthalmic products.

Hyaluronan Modulates the Biomechanical Properties of the Cornea.

Purpose: Hyaluronan (HA) is a major constituent of the extracellular matrix (ECM) that has high viscosity and is essential for maintaining tissue hydration. In the cornea, HA is enriched in the limbal region and is a key component of the limbal epithelial stem cell niche. HA is upregulated after injury participating in the formation of the provisional matrix, and has a key role in regulating the wound healing process. This study investigated whether changes in the distribution of HA before and after injury affects the biomechanical properties of the cornea in vivo. Methods: Corneas of wild-type (wt) mice and mice lacking enzymes involved in the biosynthesis of HA were analyzed before, immediately after, and 7 and 14 days after a corneal alkali burn (AB). The corneas were evaluated using both a ring light and fluorescein stain by in vivo confocal microscopy, optical coherence elastography (OCE), and immunostaining of corneal whole mounts. Results: Our results show that wt mice and mice lacking HA synthase (Has)1 and 3 present an increase in corneal stiffness 7 and 14 days after AB without a significant increase in HA expression and absence of scarring at 14 days after AB. In contrast, mice lacking Has2 present a significant decrease in corneal stiffness, with a significant increase in HA expression and scarring at 14 days after AB. Conclusions: Our findings show that the mechanical properties of the cornea are significantly modulated by changes in HA distribution following alkali burn.

Plasticity of ocular surface epithelia: Using a murine model of limbal stem cell deficiency to delineate metaplasia and transdifferentiation.

Maintaining corneal health and transparency are necessary pre-requisites for exquisite vision, a function ascribed to stem cells (SCs) nestled within the limbus. Perturbations to this site or depletion of its SCs results in limbal SC deficiency. While characterizing a murine model of this disease, we discovered unusual transformation phenomena on the corneal surface including goblet cell metaplasia (GCM), conjunctival transdifferentiation, and squamous metaplasia (SQM). GCM arose from K8+ differentiated conjunctival epithelial cells when the limbus was breached and was exacerbated by neovascularization. Regions within the cornea that harbored newly transformed K12+ epithelia were void of blood vessels and GCs, suggesting that the cornea also initiated a self-repair program. Knowledge of the intrinsic circuits that contribute to cell identity change in lineage-restricted epithelia will be invaluable for designing new therapeutics for patients with blinding corneal disease.

Hyaluronan supports the limbal stem cell phenotype during ex vivo culture.

BACKGROUND: Hyaluronan (HA) has previously been identified as an integral component of the limbal stem cell niche in vivo. In this study, we investigated whether a similar HA matrix is also expressed in vitro providing a niche supporting limbal epithelial stem cells (LESCs) during ex vivo expansion. We also investigated whether providing exogenous HA in vitro is beneficial to LESCs during ex vivo expansion. METHOD: Human LESCs (hLESCs) were isolated from donor corneas and a mouse corneal epithelial progenitor cell line (TKE2) was obtained. The HA matrix was identified surrounding LESCs in vitro using immunocytochemistry, flow cytometry and red blood exclusion assay. Thereafter, LESCs were maintained on HA coated dishes or in the presence of HA supplemented in the media, and viability, proliferation, cell size, colony formation capabilities and expression of putative stem cell markers were compared with cells maintained on commonly used coated dishes. RESULTS: hLESCs and TKE2 cells express an HA-rich matrix in vitro, and this matrix is essential for maintaining LESCs. Further supplying exogenous HA, as a substrate and supplemented to the media, increases LESC proliferation, colony formation capabilities and the expression levels of putative limbal stem cell markers. CONCLUSION: Our data show that both exogenous and endogenous HA help to maintain the LESC phenotype. Exogenous HA provides improved culture conditions for LESC during ex vivo expansion. Thus, HA forms a favorable microenvironment for LESCs during ex vivo expansion and, therefore, could be considered as an easy and cost-effective substrate and/or supplement for culturing LESCs in the clinic.

Longitudinal assessment of the effect of alkali burns on corneal biomechanical properties using optical coherence elastography.

Eye injury due to alkali burn is a severe ocular trauma that can profoundly affect corneal structure and function, including its biomechanical properties. Here, we assess the changes in the mechanical behavior of mouse corneas in response to alkali-induced injury by conducting longitudinal measurements using optical coherence elastography (OCE). A non-contact air-coupled ultrasound transducer was used to induce elastic waves in control and alkali-injured mouse corneas in vivo, which were imaged with phase-sensitive optical coherence tomography. Corneal mechanical properties were estimated using a modified Rayleigh-Lamb wave model, and results show that Young's modulus of alkali-burned corneas were significantly greater than that of their healthy counterparts on days 7 (p = 0.029) and 14 (p = 0.026) after injury. These findings, together with the changes in the shear viscosity coefficient postburn, indicate that the mechanical properties of the alkali-burned cornea are significantly modulated during the wound healing process.

Anti-inflammatory protein TNFα-stimulated gene-6 (TSG-6) reduces inflammatory response after brain injury in mice.

BACKGROUND: Current research suggests that the glial scar surrounding penetrating brain injuries is instrumental in preserving the surrounding uninjured tissue by limiting the inflammatory response to the injury site. We recently showed that tumor necrosis factor (TNF)-stimulated gene-6 (TSG-6), a well-established anti-inflammatory molecule, is present within the glial scar. In the present study we investigated the role of TSG-6 within the glial scar using TSG-6 null and littermate control mice subjected to penetrating brain injuries. RESULTS: Our findings show that mice lacking TSG-6 present a more severe inflammatory response after injury, which was correlated with an enlarged area of astrogliosis beyond the injury site. CONCLUSION: Our data provides evidence that TSG-6 has an anti-inflammatory role within the glial scar.

Extracellular Matrix Deposition and Remodeling after Corneal Alkali Burn in Mice.

Corneal transparency relies on the precise arrangement and orientation of collagen fibrils, made of mostly Type I and V collagen fibrils and proteoglycans (PGs). PGs are essential for correct collagen fibrillogenesis and maintaining corneal homeostasis. We investigated the spatial and temporal distribution of glycosaminoglycans (GAGs) and PGs after a chemical injury. The chemical composition of chondroitin sulfate (CS)/dermatan sulfate (DS) and heparan sulfate (HS) were characterized in mouse corneas 5 and 14 days after alkali burn (AB), and compared to uninjured corneas. The expression profile and corneal distribution of CS/DSPGs and keratan sulfate (KS) PGs were also analyzed. We found a significant overall increase in CS after AB, with an increase in sulfated forms of CS and a decrease in lesser sulfated forms of CS. Expression of the CSPGs biglycan and versican was increased after AB, while decorin expression was decreased. We also found an increase in KS expression 14 days after AB, with an increase in lumican and mimecan expression, and a decrease in keratocan expression. No significant changes in HS composition were noted after AB. Taken together, our study reveals significant changes in the composition of the extracellular matrix following a corneal chemical injury.

Meibomian Gland Dysfunction: What Have Animal Models Taught Us?

Studies have estimated that currently 344 million people worldwide and 16.4 million adults in the US have some form of dry eye disease (DED). It is believed that approximately 70% of DED cases are due to some form of evaporative dry eye, for which Meibomian gland dysfunction (MGD) is the major cause. Unfortunately, currently there is no effective treatment for MGD, and solely palliative care is available. Given the importance of MGD in DED, there has been a growing interest in studying Meibomian gland development, homeostasis and pathology, and, also, in developing therapies for treating and/or preventing MGD. For such, animal models have shown to be a vital tool. Much of what is known today about the Meibomian gland and MGD was learnt from these important animal models. In particular, canine and rabbit models have been essential for studying the physiopathology and progression of DED, and the mouse model, which includes different knockout strains, has enabled the identification of specific pathways potentially involved in MGD. Herein, we provide a bibliographic review on the various animal models that have been used to study Meibomian gland development, Meibomian gland homeostasis and MGD, primarily focusing on publications between 2000 and 2020.