Scaffold-free tissue engineering of functional corneal stromal tissue

Syed-Picard, Fatima N.; Du, Yiqin; Hertsenberg, Andrew; Palchesko, Rachelle N.; Funderburgh, Martha L.; Feinberg, Adam W.; Funderburgh, James L.

doi:10.1002/term.2363

Cited by 47 publications

(32 citation statements)

References 40 publications

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“…Moreover, the ability of curved substrates to induce cell alignment allowed the subsequent deposition of ECM to be similarly organized and create compact, robust tissues that retained their original cornea‐like shape despite their thinness and subjection to extensive handling. The diameter and d ‐spacing of collagen fibrils comprising the curved tissues closely resembled that observed in the natural cornea, as well as in aligned planar stromal tissue equivalents . Furthermore, collagen organization was probably the main factor contributing to the different mechanical properties between curved and planar tissues.…”

Section: Discussionmentioning

confidence: 53%

Template Curvature Influences Cell Alignment to Create Improved Human Corneal Tissue Equivalents

et al. 2017

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To accurately create corneal stromal equivalents with native‐like structure and composition, a new biofunctionalized, curved template is developed that allows the precise orientation of cells and of their extracellular matrix. This template is the first demonstration that curvature alone is sufficient to induce the alignment of human corneal stromal cells, which in turn are able to biofabricate stromal tissue equivalents with cornea‐like shape and composition. Specifically, tissues self‐released from curved templates show a highly organized nanostructure, comprised of aligned collagen fibrils, significantly higher expression of corneal stroma‐characteristic markers keratocan, lumican, decorin, ALDH3, and CHST6 (p = 0.012, 0.033, 0.029, 0.003, and 0.02, respectively), as well as significantly higher elastic modulus (p = 0.0001) compared with their planar counterparts. Moreover, curved tissues are shown to support the growth, stratification, and differentiation of human corneal epithelial cells in vitro, while maintaining their structural integrity and shape without any supporting carriers, scaffolds, or crosslinking agents. Together, these results demonstrate that corneal stromal cells can align and create highly organized, purposeful tissues by the influence of substrate curvature alone, and without the need of additional topographical cues. These findings can be important to further understand the mechanisms of corneal biosynthesis both in vitro and in vivo.

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Section: Discussionmentioning

confidence: 53%

Template Curvature Influences Cell Alignment to Create Improved Human Corneal Tissue Equivalents

et al. 2017

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“…MSCs can be isolated from multiple tissues, including adipose tissue and bone marrow. Our previous work has shown that human corneal stroma contains a population of cells (human corneal stem cells, CSSC), which exhibit properties of MSCs and differentiate to corneal keratocytes . These cells also block deposition of opaque scar tissue in a corneal wound healing model and elicit regeneration of normal, transparent stromal extracellular matrix .…”

Section: Introductionmentioning

confidence: 99%

Mesenchymal Stem Cells Reduce Corneal Fibrosis and Inflammation via Extracellular Vesicle-Mediated Delivery of miRNA

Shojaati

Khandaker

Funderburgh

et al. 2019

Stem Cells Translational Medicine

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Mesenchymal stem cells from corneal stromal stem cells (CSSC) prevent fibrotic scarring and stimulate regeneration of transparent stromal tissue after corneal wounding in mice. These effects rely on the ability of CSSC to block neutrophil infiltration into the damaged cornea. The current study investigated the hypothesis that tissue regeneration by CSSC is mediated by secreted extracellular vesicles (EVs). CSSC produced EVs 130–150 nm in diameter with surface proteins that include CD63, CD81, and CD9. EVs from CSSC reduced visual scarring in murine corneal wounds as effectively as did live cells, but EVs from human embryonic kidney (HEK)293T cells had no regenerative properties. CSSC EV treatment of wounds decreased expression of fibrotic genes Col3a1 and Acta2, blocked neutrophil infiltration, and restored normal tissue morphology. CSSC EVs labeled with carboxyfluorescein succinimidyl ester dye, rapidly fused with corneal epithelial and stromal cells in culture, transferring microRNA (miRNA) to the target cells. Knockdown of mRNA for Alix, a component of the endosomal sorting complex required for transport, using siRNA, resulted in an 85% reduction of miRNA in the secreted EVs. The EVs with reduced miRNA were ineffective at blocking corneal scarring. Furthermore, CSSC with reduced Alix expression also lost their regenerative function, suggesting EVs as an obligate component in the delivery of miRNA. The results of these studies support an essential role for extracellular vesicles in the process by which CSSC cells block scarring and initiate regeneration of transparent corneal tissue after wounding. EVs appear to serve as a delivery vehicle for miRNA, which affects the regenerative action. Stem Cells Translational Medicine 2019;8:1192–1201

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“…Surgical scalpel 15-ml polypropylene centrifuge tube Eppendorf 5810R centrifuge and A-4-81 rotor T25, T75, and T175 flasks Light microscope Additional reagents and equipment for limbal-rim isolation (Syed-Picard et al, 2018; NOTE: Institutional review board and federal guidelines must be followed regarding obtaining human corneal tissue.…”

Section: Methodsmentioning

confidence: 99%

“…Due to the high proliferative capabilities of hCSSCs, they are commonly used as a cell source of keratocytes by following specific differentiation protocols (Syed-Picard et al, 2018;. As keratocytes tend to proliferate very slowly in culture, the process allows one to obtain sufficient cell numbers using the stem cell progenitor prior to differentiation.…”

Section: Subculturing Of Human Corneal Stromal Stem Cellsmentioning

confidence: 99%

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Assembly and Application of a Three‐Dimensional Human Corneal Tissue Model

et al. 2019

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The cornea provides a functional barrier separating the outside environment from the intraocular environment, thereby protecting posterior segments of the eye from infection and damage. Pathological changes that compromise the structure or integrity of the cornea may occur as a result of injury or disease and can lead to debilitating effects on visual acuity. Over 10 million people worldwide are visually impaired or blind due to corneal opacity. Thus, physiologically relevant in vitro approaches to predict corneal toxicity of chemicals or effective treatments for disease prior to ocular exposure, as well as to study the corneal effects of systemic, chronic conditions, such as diabetes, are needed to reduce use of animal testing and accelerate therapeutic development. We have previously bioengineered an innervated corneal tissue model using silk protein scaffolds to recapitulate the structural and mechanical elements of the anterior cornea and to model the functional aspects of corneal sensation with the inclusion of epithelial, stromal, and neural components. The purpose of this unit is to provide a step‐by‐step guide for preparation, assembly, and application of this three‐dimensional corneal tissue system to enable the study of corneal tissue biology. © 2019 by John Wiley & Sons, Inc.

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Scaffold-free tissue engineering of functional corneal stromal tissue

Cited by 47 publications

References 40 publications

Template Curvature Influences Cell Alignment to Create Improved Human Corneal Tissue Equivalents

Template Curvature Influences Cell Alignment to Create Improved Human Corneal Tissue Equivalents

Mesenchymal Stem Cells Reduce Corneal Fibrosis and Inflammation via Extracellular Vesicle-Mediated Delivery of miRNA

Assembly and Application of a Three‐Dimensional Human Corneal Tissue Model

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