Monocarboxylate Transporters Mediate Fluorescein Uptake in Corneal Epithelial Cells

Sun, Yichen; Liou, Hau-Min; Yeh, Po-Ting; Chen, Wei-Li; Hu, Fung-Rong

doi:10.1167/iovs.16-20998

Cited by 6 publications

(4 citation statements)

References 18 publications

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“… 15 , 16 Recent studies favor the idea that fluorescein stains individual cells, particularly those with altered physiological conditions. 14 , 17 , 26 Adhering to this idea, our study found that fluorescein ingress was reduced in HCECs that had been promoted to express MUC1, whereas a knockdown of MUC1 in HCECs enhanced fluorescein ingress. This effect was likely attributed to the MUC1 extracellular domain because overexpressing MUC1-C (the MUC1 cytoplasmic domain) failed to decrease fluorescein ingress in HCECs.…”

Section: Discussionmentioning

confidence: 57%

“… 14 – 16 Nonetheless, our previous study has shown that fluorescein ingress into the corneal epithelial cells can also occur through the monocarboxylate transporter. 17 Herein, we used both in vivo and in vitro approaches to examine whether the expression of transmembrane MUC1 is causally linked to blocking of fluorescein ingress through corneal epithelial cells as another working mechanism to explain how fluorescein punctate staining might develop in dry eye diseases.…”

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confidence: 99%

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Transmembrane Mucin 1 Blocks Fluorescein Ingress to Corneal Epithelium

Sun

Hung

et al. 2022

Invest. Ophthalmol. Vis. Sci.

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Purpose To determine the role of transmembrane mucins in blocking fluorescein ingress to the corneal epithelium and its deficiency in contributing to corneal fluorescein punctate staining. Methods A dry eye model was established by extirpating lacrimal and Harderian glands in rabbits to correlate the expression of mucins with fluorescein-stained areas on the corneal button using immunofluorescence. Expression of transmembrane mucins was promoted in human corneal epithelial cells (HCECs) by culturing with the mucin-promoting medium (MPM) or diquafosol treatment. Conversely, the expression of mucins was downregulated by knockdown with short hairpin RNA. The role of mucin1 extracellular domain in fluorescein ingress was further verified by overexpression of N-terminally truncated mucin1 in HCECs. Results In the rabbit dry eye model, the expression level of mucin1 was significantly decreased in superficial corneal epithelial cells where fluorescein punctate staining was observed. Upregulation of mucin1 and mucin16 in HCECs promoted by MPM or by diquafosol treatment impeded intracellular fluorescein ingress. Downregulation of mucin1 and mucin16 enhanced fluorescence ingress in HCECs after fluorescein staining. Overexpression of truncated mucin1 did not alter the fluorescein intensity of fluorescein-stained HCECs, supporting the notion that the ability of mucin1 to block fluorescein ingress was primarily mediated by its extracellular domain. Minimal inherent expression of mucin16 in the rabbit cornea limited the validation of its role in blocking fluorescein ingress in vivo. Conclusion Transmembrane mucin1 blocks fluorescein ingress in the corneal epithelium, explaining how fluorescein staining is positive when the level of transmembrane mucins is disturbed in dry eyes.

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Section: Discussionmentioning

confidence: 57%

mentioning

confidence: 99%

Transmembrane Mucin 1 Blocks Fluorescein Ingress to Corneal Epithelium

Sun

Hung

et al. 2022

Invest. Ophthalmol. Vis. Sci.

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“…In our transcriptomic analysis, we observed a significant upregulation in the expression of SLC16A7 gene, which encode monocarboxylate transporter 2 and has an affinity for acetoacetate and their derivatives metabolites (methyl-2-oxovaleric acid and 3-methyl-2-oxobutanoic) 28 . Considering the domination of the monocarboxylate transporters on the apical direction of the corneal epithelium 29 , DCF might have an active impact of these transporters, leading to the excessive transportation of these metabolites in the apical direction. However, the precise mechanism favoring their apical release under DCF remains to be elucidated.…”

Section: Discussionmentioning

confidence: 99%

Modeling dry eye with an air–liquid interface in corneal epithelium-on-a-chip

Kado Abdalkader,

Chaleckis,

Fujita

et al. 2024

Sci Rep

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Dry eye syndrome (DES) is a complex ocular condition characterized by an unstable tear film and inadequate tear production, leading to tissue damage. Despite its common occurrence, there is currently no comprehensive in vitro model that accurately reproduce the cellular characteristics of DES. Here we modified a corneal epithelium-on-a-chip (CEpOC) model to recapitulate DES by subjecting HCE-T human corneal epithelial cells to an air–liquid (AL) interface stimulus. We then assessed the effects of AL stimulation both in the presence and absence of diclofenac (DCF), non-steroidal anti-inflammatory drug. Transcriptomic analysis revealed distinct gene expression changes in response to AL and AL_DCF, affecting pathways related to development, epithelial structure, inflammation, and extracellular matrix remodeling. Both treatments upregulated PIEZO2, linked to corneal damage signaling, while downregulating OCLN, involved in cell–cell junctions. They increased the expression of inflammatory genes (e.g., IL-6) and reduced mucin production genes (e.g., MUC16), reflecting dry eye characteristics. Metabolomic analysis showed increased secretion of metabolites associated with cell damage and inflammation (e.g., methyl-2-oxovaleric acid, 3-methyl-2-oxobutanoic acid, lauroyl-carnitine) in response to AL and even more with AL_DCF, indicating a shift in cellular metabolism. This study showcases the potential use of AL stimulus within the CEpOC to induce cellular characteristics relevant to DES.

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“…In the human cornea, to date, MCTs are critically understudied with only eleven studies published [ 4 – 7 , 12 , 17 – 22 ]. The majority of the existing literature focus on the expression of MCTs 1-4 [ 12 , 18 , 21 ] and MCT5 [ 19 ], as they relate to the corneal epithelium. Corneal endothelial cells have also been reported to express MCT1, MCT4, and MCT8 [ 17 , 22 ].…”

Section: Introductionmentioning

confidence: 99%

Monocarboxylate Transporters: Role and Regulation in Corneal Diabetes

Shrestha

Whelchel

Nicholas

et al. 2022

Analytical Cellular Pathology

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Diabetes mellitus (DM) is a group of metabolic diseases that is known to cause structural and functional ocular complications. In the human cornea, DM-related complications affect the epithelium, stroma, and nerves. Monocarboxylate transporters (MCTs) are a family of proton-linked plasma membrane transporters that carry monocarboxylates across plasma membranes. In the context of corneal health and disease, their role, presence, and function are largely undetermined and solely focused on the most common MCT isoforms, 1 through 4. In this study, we investigated the regulation of MCT1, 2, 4, 5, 8, and 10, in corneal DM, using established 3D self-assembled extracellular matrix (ECM) in vitro models. Primary stromal corneal fibroblasts were isolated from healthy (HCFs), type I (T1DMs), and type II (T2DMs) DM donors. Monoculture 3D constructs were created by stimulating stromal cells on transwells with stable vitamin C for two or four weeks. Coculture 3D constructs were created by adding SH-SY5Y neurons at two different densities, 12 k and 500 k, on top of the monocultures. Our data showed significant upregulation of MCT1 at 4 weeks for HCF, T1DM, and T2DM monocultures, as well as the 500 k nerve cocultures. MCT8 was significantly upregulated in HCF and T1DM monocultures and all of the 500 k nerve cocultures. Further, MCT10 was only expressed at 4 weeks for all cocultures and was limited to HCFs and T1DMs in monocultures. Immunofluorescence analysis showed cytoplasmic MCT expression for all cell types and significant downregulation of both MCT2 and MCT4 in HCFs, when compared to T1DMs and T2DMs. Herein, we reveal the existence and modulation of MCTs in the human diabetic cornea in vitro. Changes appeared dependent on neuronal density, suggesting that MCTs are very likely critical to the neuronal defects observed in diabetic keratopathy/neuropathy. Further studies are warranted in order to fully delineate the role of MCTs in corneal diabetes.

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Monocarboxylate Transporters Mediate Fluorescein Uptake in Corneal Epithelial Cells

Cited by 6 publications

References 18 publications

Transmembrane Mucin 1 Blocks Fluorescein Ingress to Corneal Epithelium

Transmembrane Mucin 1 Blocks Fluorescein Ingress to Corneal Epithelium

Modeling dry eye with an air–liquid interface in corneal epithelium-on-a-chip

Monocarboxylate Transporters: Role and Regulation in Corneal Diabetes

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