scholarly journals Fabrication and Biocompatibility of Electroconductive Silk Fibroin/PEDOT: PSS Composites for Corneal Epithelial Regeneration

Polymers ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 3028
Author(s):  
Promita Bhattacharjee ◽  
Mark Ahearne
Keyword(s):  
Silk Fibroin ◽  
Crosslinking Agent ◽  
Light Transmittance ◽  
Corneal Epithelial Cell ◽  
Corneal Epithelial ◽  
Epithelial Regeneration ◽  
Human Corneal Epithelial Cells ◽  
Epithelium Regeneration ◽  
Poly Styrene Sulfonate ◽  
The Impact

The aim of this study was to develop matrices that can support human corneal epithelial cells and innervation by incorporating a conducting polymer, poly(3,4-ethylenedioxythiophene) poly(styrene sulfonate) (PEDOT:PSS), into silk fibroin (SF). Polyvinyl alcohol (PVA) was used as a crosslinking agent to enhance the mechanical properties of the matrices. The impact of PEDOT:PSS on the materials’ physical properties and cellular responses was examined. The electrical impedance of matrices decreased with increasing concentration of PEDOT:PSS suggesting improved electroconductivity. However, light transmittance also decreased with increasing PEDOT:PSS. Young’s modulus was unaffected by PEDOT:PSS but was increased by PVA. The viability of corneal epithelial cell on the matrices was unaffected by the incorporation of PEDOT:PSS except at the highest concentration tested 0.3% (w/v), which led to a cytotoxic response. These findings suggest that SF/PEDOT:PSS with a PEDOT:PSS concentration of 0.1–0.2% would be a suitable biomaterial for epithelium regeneration.

scholarly journals Optimization of Corneal Epithelial Progenitor Cell Growth onBombyx moriSilk Fibroin Membranes

2016 ◽  
Vol 2016 ◽  
pp. 1-11 ◽  
Author(s):  
Thomas A. Hogerheyde ◽  
Shuko Suzuki ◽  
Jennifer Walshe ◽  
Laura J. Bray ◽  
Sally A. Stephenson ◽  
...  
Keyword(s):  
Silk Fibroin ◽  
Cell Morphology ◽  
Kinase Inhibitor ◽  
Keratinocyte Growth Factor ◽  
Rho Kinase ◽  
Corneal Epithelial Cell ◽  
Differentiation Markers ◽  
Corneal Epithelial ◽  
Collagen Iii

Scaffolds prepared from silk fibroin derived from cocoons of the domesticated silkworm mothBombyx morihave demonstrated potential to support the attachment and growth of human limbal epithelial (HLE) cellsin vitro. In this study, we attempted to further optimize protocols to promote the expansion of HLE cells onB. morisilk fibroin- (BMSF-) based scaffolds. BMSF films were initially coated with different extracellular matrix proteins and then analysed for their impact on corneal epithelial cell adhesion, cell morphology, and culture confluency. Results showed that collagen I, collagen III, and collagen IV consistently improved HCE-T cell adherence, promoted an elongated cell morphology, and increased culture confluency. By contrast, ECM coating had no significant effect on the performance of primary HLE cells cultured on BMSF films. In the second part of this study, primary HLE cells were grown on BMSF films in the presence of medium (SHEM) supplemented with keratinocyte growth factor (KGF) and the Rho kinase inhibitor, Y-27632. The results demonstrated that SHEM medium supplemented with KGF and Y-27632 dramatically increased expression of corneal differentiation markers, keratin 3 and keratin 12, whereas expression of the progenitor marker, p63, did not appear to be significantly influenced by the choice of culture medium.

scholarly journals The O-GlcNAc modification promotes terminal differentiation of human corneal epithelial cells

Glycobiology ◽  
2020 ◽  
Vol 30 (11) ◽  
pp. 872-880 ◽  
Author(s):  
Nicole M McColgan ◽  
Marissa N Feeley ◽  
Ashley M Woodward ◽  
Damien Guindolet ◽  
Pablo Argüeso
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Ocular Surface ◽  
Barrier Function ◽  
Terminal Differentiation ◽  
Corneal Epithelial Cell ◽  
Corneal Epithelial Cells ◽  
Corneal Epithelial ◽  
Human Corneal Epithelial Cells ◽  
Glcnac Transferase

Abstract Dynamic modification of nuclear and cytoplasmic proteins with O-linked β-N-acetylglucosamine (O-GlcNAc) plays an important role in orchestrating the transcriptional activity of eukaryotic cells. Here, we report that the O-GlcNAc modification contributes to maintaining ocular surface epithelial homeostasis by promoting mucin biosynthesis and barrier function. We found that induction of human corneal epithelial cell differentiation stimulated the global transfer of O-GlcNAc to both nuclear and cytosolic proteins. Inflammatory conditions, on the other hand, were associated with a reduction in the expression of O-GlcNAc transferase at the ocular surface epithelia. Loss- and gain-of-function studies using small interfering RNA targeting O-GlcNAc transferase, or Thiamet G, a selective inhibitor of O-GlcNAc hydrolase, respectively, revealed that the presence of O-GlcNAc was necessary to promote glycocalyx barrier function. Moreover, we found that Thiamet G triggered a correlative increase in both surface expression of MUC16 and apical epithelial cell area while reducing paracellular permeability. Collectively, these results identify intracellular protein O-glycosylation as a novel pathway responsible for promoting the terminal differentiation of human corneal epithelial cells.

scholarly journals Biocompatibility of Nanocellulose-Reinforced PVA Hydrogel with Human Corneal Epithelial Cells for Ophthalmic Applications

2019 ◽  
Vol 10 (3) ◽  
pp. 35 ◽  
Author(s):  
Tummala ◽  
Lopes ◽  
Mihranyan ◽  
Ferraz
Keyword(s):  
Epithelial Cells ◽  
Cell Viability ◽  
Protein Adsorption ◽  
Direct Contact ◽  
Corneal Epithelial Cell ◽  
Corneal Epithelial Cells ◽  
Corneal Epithelial ◽  
Human Corneal Epithelial Cells ◽  
Hydrogel Surface

Transparent composite hydrogel in the form of a contact lens made from poly(vinyl alcohol) (PVA) and cellulose nanocrystals (CNCs) was subjected to in vitro biocompatibility evaluation with human corneal epithelial cells (HCE-2 cells). The cell response to direct contact with the hydrogels was investigated by placing the samples on top of confluent cell layers and evaluating cell viability, morphology, and cell layer integrity subsequent to 24 h culture and removal of the hydrogels. To further characterize the lens–cell interactions, HCE-2 cells were seeded on the hydrogels, with and without simulated tear fluid (STF) pre-conditioning, and cell viability and morphology were evaluated. Furthermore, protein adsorption on the hydrogel surface was investigated by incubating the materials with STF, followed by protein elution and quantification. The hydrogel material was found to have affinity towards protein adsorption, most probably due to the interactions between the positively charged lysozyme and the negatively charged CNCs embedded in the PVA matrix. The direct contact experiment demonstrated that the physical presence of the lenses did not affect corneal epithelial cell monolayers in terms of integrity nor cell metabolic activity. Moreover, it was found that viable corneal cells adhered to the hydrogel, showing the typical morphology of epithelial cells and that such response was not influenced by the STF pre-conditioning of the hydrogel surface. The results of the study confirm that PVA-CNC hydrogel is a promising ophthalmic biomaterial, motivating future in vitro and in vivo biocompatibility studies.

scholarly journals Expression of immune response genes in human corneal epithelial cells interacting with Aspergillus flavus conidia

BMC Genomics ◽  
2022 ◽  
Vol 23 (1) ◽  
Author(s):  
Divya Arunachalam ◽  
Shruthi Mahalakshmi Ramanathan ◽  
Athul Menon ◽  
Lekshmi Madhav ◽  
Gopalakrishna Ramaswamy ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cell Line ◽  
Differentially Expressed Genes ◽  
Primary Cultures ◽  
Differentially Expressed ◽  
Corneal Epithelial Cell ◽  
Epithelial Cell Line ◽  
Corneal Epithelial Cells ◽  
Corneal Epithelial ◽  
Human Corneal Epithelial Cells

Abstract Background Aspergillus flavus, one of the causative agents of human fungal keratitis, can be phagocytosed by human corneal epithelial (HCE) cells and the conidia containing phagosomes mature into phagolysosomes. But the immunological responses of human corneal epithelial cells interacting with A. flavus are not clear. In this study, we report the expression of immune response related genes of HCE cells exposed to A. flavus spores using targeted transcriptomics. Methods Human corneal epithelial cell line and primary cultures were grown in a six-well plate and used for coculture experiments. Internalization of the conidia was confirmed by immunofluorescence microscopy of the colocalized endosomal markers CD71 and LAMP1. Total RNA was isolated, and the quantity and quality of the isolated RNA were assessed using Qubit and Bioanalyzer. NanoString nCounter platform was used for the analysis of mRNA abundance using the Human Immunology panel. R-package and nSolver software were used for data analysis. KEGG and FunRich 3.1.3 tools were used to analyze the differentially expressed genes. Results Different morphotypes of conidia were observed after 6 h of coculture with human corneal epithelial cells and found to be internalized by epithelial cells. NanoString profiling showed more than 20 differentially expressed genes in immortalized human corneal epithelial cell line and more than ten differentially expressed genes in primary corneal epithelial cells. Distinct set of genes were altered in their expression in cell line and primary corneal epithelial cells. KEGG pathway analysis revealed that genes associated with TNF signaling, NF-KB signaling, and Th17 signaling were up-regulated, and genes associated with chemokine signaling and B cell receptor signaling were down regulated. FunRich pathway analysis showed that pathways such as CDC42 signaling, PI3K signaling, and Arf6 trafficking events were activated by the clinical isolates CI1123 and CI1698 in both type of cells. Conclusions Combining the transcript analysis data from cell lines and primary cultures, we showed the up regulation of immune defense genes in A. flavus infected cells. At the same time, chemokine signaling and B cell signaling pathways are downregulated. The variability in the expression levels in the immortalized cell line and the primary cultures is likely due to the variable epigenetic reprogramming in the immortalized cells and primary cultures in the absence of any changes in the genome. It highlights the importance of using both cell types in host-pathogen interaction studies.

In vitro potential of human mesenchymal stem cells for corneal epithelial regeneration

2020 ◽  
Vol 15 (3) ◽  
pp. 1409-1426 ◽  
Author(s):  
Núria Nieto-Nicolau ◽  
Beatriz Martín-Antonio ◽  
Claudia Müller-Sánchez ◽  
Ricardo P Casaroli-Marano
Keyword(s):  
Stem Cells ◽  
Wound Healing ◽  
Mesenchymal Stem Cells ◽  
Epithelial Cells ◽  
Electric Resistance ◽  
Corneal Epithelial Cells ◽  
Corneal Epithelial ◽  
Epithelial Regeneration ◽  
Human Corneal Epithelial Cells

Aim: To determine the potential of mesenchymal stem cells (MSC) for corneal epithelial regeneration in vitro. Materials & methods: Bone marrow MSC (BM-MSC) and adipose tissue MSC were analyzed for corneal epithelial and mesenchymal markers, using limbal stem cells and corneal cells as controls. MSC with better potential were cultured with specific mediums for epithelial induction. Transepithelial electric resistance and wound healing assay with human corneal epithelial cells were performed. Results: BM-MSC showed better potential, increased corneal markers, and higher transepithelial electric resistance values when induced with limbal epithelial culture medium. Induced BM-MSC promoted better wound healing of human corneal epithelial cells by paracrine secretion. Conclusion: BM-MSC has potential for corneal epithelial induction in a protocol compatible with human application.

scholarly journals The common YAP activation mediates corneal epithelial regeneration and repair with different-sized wounds

2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Yijian Li ◽  
Lingling Ge ◽  
Xia Chen ◽  
Yumei Mao ◽  
Xianliang Gu ◽  
...  
Keyword(s):  
Stem Cells ◽  
Epithelial Cells ◽  
Adult Stem Cells ◽  
Cell Junction ◽  
Epithelial Stem Cells ◽  
Corneal Epithelial ◽  
Epithelial Regeneration ◽  
The Common ◽  
Epithelium Regeneration ◽  
Large Wound

AbstractRegeneration/repair after injury can be endowed by adult stem cells (ASCs) or lineage restricted and even terminally differentiated cells. In corneal epithelium, regeneration after a large wound depends on ASCs (limbal epithelial stem cells, LESCs), whereas repair after a small wound is LESCs-independent. Here, using rat corneal epithelial wounds with different sizes, we show that YAP activation promotes the activation and expansion of LESCs after a large wound, as well as the reprogramming of local epithelial cells (repairing epithelial cells) after a small wound, which contributes to LESCs-dependent and -independent wound healing, respectively. Mechanically, we highlight that the reciprocal regulation of YAP activity and the assembly of cell junction and cortical F-actin cytoskeleton accelerates corneal epithelial healing with different-sized wounds. Together, the common YAP activation and the underlying regulatory mechanism are harnessed by LESCs and lineage-restricted epithelial cells to cope with corneal epithelial wounds with different sizes.

scholarly journals Treatment of Silk Fibroin with Poly(ethylene glycol) for the Enhancement of Corneal Epithelial Cell Growth

2015 ◽  
Vol 6 (2) ◽  
pp. 345-366 ◽  
Author(s):  
Shuko Suzuki ◽  
Rebecca Dawson ◽  
Traian Chirila ◽  
Audra Shadforth ◽  
Thomas Hogerheyde ◽  
...  
Keyword(s):  
Cell Growth ◽  
Epithelial Cell ◽  
Ethylene Glycol ◽  
Silk Fibroin ◽  
Corneal Epithelial Cell ◽  
Poly Ethylene Glycol ◽  
Corneal Epithelial ◽  
Poly Ethylene

Characterisation of Human Corneal Epithelial Cell Cultures Maintained under Serum-free Conditions

2008 ◽  
Vol 36 (5) ◽  
pp. 569-583 ◽  
Author(s):  
Judith W. Seeber ◽  
Michaela Zorn-Kruppa ◽  
Simone Lombardi-Borgia ◽  
Heike Scholz ◽  
Anna K. Manzer ◽  
...  
Keyword(s):  
Cell Line ◽  
Barrier Function ◽  
Culture Media ◽  
Cytokine Expression ◽  
Corneal Epithelial Cell ◽  
Antibody Array ◽  
Full Thickness ◽  
Corneal Epithelial ◽  
Serum Free ◽  
The Impact

Three-dimensional tissue constructs have been proposed as in vitro screening models for ocular irritancy. Based on our previous studies, in which a full-thickness corneal model based exclusively on SV40-immortalised cell lines was generated, we have currently evaluated the effects of a range of commercially-available cell culture media on several cellular parameters in cultures of a human corneal epithelial (HCE) cell line. This cell line was used in an attempt to establish a rational basis for the development of serum-free culture media for the assembly and long-term tissue culture of full-thickness corneal models. Briefly, we investigated the impact of serum-free culture on the proliferation, morphology, barrier function and cytokine expression of HCE cells. The number of cell layers and the epithelial differentiation were evaluated by histology. Barrier properties were characterised via the determination of transepithelial electrical resistance (TEER), fluorescein permeation, and the expression of the tight junction-related protein, zona occludin 1 (ZO-1). The cytokine expression pattern in response to serum-free culture was measured by using an antibody array system. Our results revealed that both the morphology and the barrier function of the epithelial constructs were comparable to those of human donor corneas, when serum-free media were supplemented with ascorbic acid, calcium, hydrocortisone and retinoic acid. Under these conditions, the artificial epithelium based on serum-free HCE cultures represented a valid model for the natural ocular surface.

scholarly journals Differentiation Induction of Human Stem Cells for Corneal Epithelial Regeneration

2020 ◽  
Vol 21 (21) ◽  
pp. 7834
Author(s):  
Kasem Theerakittayakorn ◽  
Hong Thi Nguyen ◽  
Jidapa Musika ◽  
Hataiwan Kunkanjanawan ◽  
Sumeth Imsoonthornruksa ◽  
...  
Keyword(s):  
Stem Cells ◽  
Epithelial Cells ◽  
Pluripotent Stem Cells ◽  
Corneal Epithelium ◽  
Embryonic Stem ◽  
Human Stem Cells ◽  
Corneal Epithelial Cells ◽  
Corneal Epithelial ◽  
Epithelial Regeneration ◽  
Epithelium Regeneration

Deficiency of corneal epithelium causes vision impairment or blindness in severe cases. Transplantation of corneal epithelial cells is an effective treatment but the availability of the tissue source for those cells is inadequate. Stem cells can be induced to differentiate to corneal epithelial cells and used in the treatment. Multipotent stem cells (mesenchymal stem cells) and pluripotent stem cells (embryonic stem cells and induced pluripotent stem cells) are promising cells to address the problem. Various protocols have been developed to induce differentiation of the stem cells into corneal epithelial cells. The feasibility and efficacy of both human stem cells and animal stem cells have been investigated for corneal epithelium regeneration. However, some physiological aspects of animal stem cells are different from those of human stem cells, the protocols suited for animal stem cells might not be suitable for human stem cells. Therefore, in this review, only the investigations of corneal epithelial differentiation of human stem cells are taken into account. The available protocols for inducing the differentiation of human stem cells into corneal epithelial cells are gathered and compared. Also, the pathways involving in the differentiation are provided to elucidate the relevant mechanisms.

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