MicroRNA Profiling of Highly Enriched Human Corneal Epithelial Stem Cells by Small RNA Sequencing

Kalaimani, Lavanya; Devarajan, Bharanidharan; Umadevi, S.; Vanniarajan, Ayyasamy; Namperumalsamy, Venkatesh Prajna; Muthukkaruppan, Veerappan; Chidambaranathan, Gowri Priya

doi:10.1038/s41598-020-64273-0

Cited by 10 publications

(10 citation statements)

References 64 publications

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“…Our previous study on miRNA profiling of enriched CESCs 9 suggested the possible role of hsa-miR-143-3p in the maintenance of stemness in CESCs. In continuation, the ectopic expression of hsa-miR-143-3p in this study, increased the colony forming potential of cultured limbal epithelial cells along with increased number of holoclone-like colonies based on the size and morphology.…”

Section: Discussionmentioning

confidence: 98%

“…Six miRNAs- hsa-miR-3168, hsa-miR-21-5p, hsa-miR-143-3p, hsa-miR-150-5p, hsa-miR-10a-5p and hsa-miR-1910-5p were identified to be highly expressed in the enriched CESCs and validated through qPCR. Interestingly, based on the locked nucleic acid in-situ hybridization, hsa-miR-143-3p was found be expressed in the basal layer of the limbal epithelium, exclusively in the clusters of small cells indicating its potential association with the CESCs 9 . In continuation, the functional role of hsa-miR-143-3p in CESCs was evaluated in this study.…”

Section: Introductionmentioning

confidence: 99%

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Hsa-miR-143-3p inhibits Wnt-β-catenin and MAPK signaling in human corneal epithelial stem cells

Kalaimani

Devarajan²,

Namperumalsamy

et al. 2022

Sci Rep

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Our previous study demonstrated hsa-miR-143-3p as one of the highly expressed miRNAs in enriched corneal epithelial stem cells (CESCs). Hence this study aims to elucidate the regulatory role of hsa-miR-143-3p in the maintenance of stemness in CESCs. The target genes of hsa-miR-143-3p were predicted and subjected to pathway analysis to select the targets for functional studies. Primary cultured limbal epithelial cells were transfected with hsa-miR-143-3p mimic, inhibitor or scrambled sequence using Lipofectamine 3000. The transfected cells were analysed for (i) colony forming potential, (ii) expression of stem cell (SC) markers/ transcription factors (ABCG2, NANOG, OCT4, KLF4, ΔNp63), (iii) differentiation marker (Cx43), (iv) predicted five targets of hsa-miR-143-3p (DVL3, MAPK1, MAPK14, KRAS and KAT6A), (v) MAPK signaling regulators and (vi) Wnt-β-catenin signaling regulators by qPCR, immunofluorescence staining and/or Western blotting. High expression of hsa-miR-143-3p increased the colony forming potential (10.04 ± 1.35%, p < 0.001) with the ability to form holoclone-like colonies in comparison to control (3.33 ± 0.71%). The mimic treated cells had increased expression of SC markers but reduced expression of Cx43 and hsa-miR-143-3p targets involved in Wnt-β-catenin and MAPK signaling pathways. The expression of β-catenin, active β-catenin and ERK2 in hsa-miR-143-3p inhibitor transfected cells were higher than the control cells and the localized nuclear expression indicated the activation of Wnt and MAPK signaling. Thus, the probable association of hsa-miR-143-3p in the maintenance of CESCs through inhibition of Wnt and MAPK signaling pathways was thus indicated.

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Section: Discussionmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

Hsa-miR-143-3p inhibits Wnt-β-catenin and MAPK signaling in human corneal epithelial stem cells

Kalaimani

Devarajan²,

Namperumalsamy

et al. 2022

Sci Rep

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“…miR-10b and miR-184 have been identified in the limbus and central cornea, respectively [ 37 ], and several miRNAs are dysregulated in diabetic cornea [ 21 , 38 ]. More recently, small RNA sequencing identified miRNA profiles in enriched corneal epithelial stem cells and mature central corneal epithelial cells [ 39 ]. Altogether, these findings suggest that miRNA signatures are associated with specific cell types or diseases.…”

Section: Discussionmentioning

confidence: 99%

Extracellular miR-6723-5p could serve as a biomarker of limbal epithelial stem/progenitor cell population

et al. 2022

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Background Dysfunction or loss of limbal stem cells can result in limbal stem cell deficiency (LSCD), a disease that cause corneal opacity, pain, and loss of vision. Cultivated limbal epithelial transplantation (CLET) can be used to restore stem cell niche homeostasis and replenish the progenitor pool. Transplantation has been reported with high success rate, but there is an unmet need of prognostic markers that correlate with clinical outcomes. To date, the progenitor content in the graft is the only parameter that has been retrospectively linked to success. Methods In this study, we investigate extracellular micro RNAs (miRNAs) associated with stem/progenitor cells in cultivated limbal epithelial cells (cLECs). Using micro RNA sequencing and linear regression modelling, we identify a miRNA signature in cultures containing high proportion of stem/progenitor cells. We then develop a robust RNA extraction workflow from culture media to confirm a positive miRNA correlation with stem/progenitor cell proportion. Results miR-6723-5p is associated with cultures containing high proportion of stem/progenitor cells, and is detected in the basal layer of corneal epithelium. Conclusions These results indicate that miR-6723-5p could potentially serve as a stem/progenitor cell marker in cLECs.

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“…TCF4, a transcription factor that interacts with β-catenin in the nucleus, is also expressed on the basal layer of the limbal epithelium, colocalizing with the stemness markers p63 and ABCG2 [ 56 , 57 ]. MicroRNAs (miRs) such as miR-10b, miR-150-5p, miR-21-5p, miR-1910-5p, miR-10a-5p, and miR-103/107 family are differentially expressed in the basal limbal epithelium and target components of Wnt signaling [ 58 , 59 , 60 ].…”

Section: Signaling Cascades In the Lsc Niche And In The Regulation Of Lscsmentioning

confidence: 99%

Regulation of Limbal Epithelial Stem Cells: Importance of the Niche

Robertson

Roberts

Deng

2021

IJMS

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Limbal epithelial stem/progenitor cells (LSCs) reside in a niche that contains finely tuned balances of various signaling pathways including Wnt, Notch, BMP, Shh, YAP, and TGFβ. The activation or inhibition of these pathways is frequently dependent on the interactions of LSCs with various niche cell types and extracellular substrates. In addition to receiving molecular signals from growth factors, cytokines, and other soluble molecules, LSCs also respond to their surrounding physical structure via mechanotransduction, interaction with the ECM, and interactions with other cell types. Damage to LSCs or their niche leads to limbal stem cell deficiency (LSCD). The field of LSCD treatment would greatly benefit from an understanding of the molecular regulation of LSCs in vitro and in vivo. This review synthesizes current literature around the niche factors and signaling pathways that influence LSC function. Future development of LSCD therapies should consider all these niche factors to achieve improved long-term restoration of the LSC population.

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MicroRNA Profiling of Highly Enriched Human Corneal Epithelial Stem Cells by Small RNA Sequencing

Cited by 10 publications

References 64 publications

Hsa-miR-143-3p inhibits Wnt-β-catenin and MAPK signaling in human corneal epithelial stem cells

Hsa-miR-143-3p inhibits Wnt-β-catenin and MAPK signaling in human corneal epithelial stem cells

Extracellular miR-6723-5p could serve as a biomarker of limbal epithelial stem/progenitor cell population

Regulation of Limbal Epithelial Stem Cells: Importance of the Niche

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