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Investigative Ophthalmology & Visual Science | 2010

Human Corneal Fibrosis: An In Vitro Model

Dimitris Karamichos; Xiaoqing Q Guo; Audrey E. K. Hutcheon; James D. Zieske

PURPOSE Corneal injury may ultimately lead to a scar by way of corneal fibrosis, which is characterized by the presence of myofibroblasts and improper deposition of extracellular matrix (ECM) components. TGF-beta1 is known to stimulate overproduction and deposition of ECM components. Previously, an in vitro three-dimensional (3-D) model of a corneal stroma was developed by using primary human corneal fibroblasts (HCFs) stimulated with stable vitamin C (VitC). This model mimics corneal development. The authors postulate that with the addition of TGF-beta1, a 3-D corneal scar model can be generated. METHODS HCFs were grown in four media conditions for 4 or 8 weeks: VitC only; VitC+TGF-beta1 for the entire time; VitC+TGF-beta1 for 1 week, then VitC only for 3 or 7 weeks; and VitC for 4 weeks, then VitC+TGF-beta1 for 4 weeks. Cultures were analyzed with TEM and indirect immunofluorescence. RESULTS Compared with the control, addition of TGF-beta1 increased construct thickness significantly, with maximum increase in constructs with TGF-beta1 present for the entire time-2.1- to 3.2-fold at 4 and 8 weeks, respectively. In all TGF-beta-treated cultures, cells became long and flat, numerous filamentous cells were seen, collagen levels increased, and long collagen fibrils were visible. Smooth muscle actin, cellular fibronectin, and type III collagen expression all appeared to increase. Cultures between weeks 4 and 8 showed minimal differences. CONCLUSIONS Human corneal fibroblasts stimulated by VitC and TGF-beta1 appear to generate a model that resembles processes observed in human corneal fibrosis. This model should be useful in examining matrix deposition and assembly in a wound-healing situation.


Developmental Dynamics | 2008

Human primary corneal fibroblasts synthesize and deposit proteoglycans in long-term 3-D cultures

Ruiyi Ren; Audrey E. K. Hutcheon; Xiaoqing Q Guo; Nima Saeidi; Suzanna A. Melotti; Jeffrey W. Ruberti; James D. Zieske; Vickery Trinkaus-Randall

Our goal was to develop a 3‐D multi‐cellular construct using primary human corneal fibroblasts cultured on a disorganized collagen substrate in a scaffold‐free environment and to use it to determine the regulation of proteoglycans over an extended period of time (11 weeks). Electron micrographs revealed multi‐layered constructs with cells present in between alternating parallel and perpendicular arrays of fibrils. Type I collagen increased 2–4‐fold. Stromal proteoglycans including lumican, syndecan4, decorin, biglycan, mimecan, and perlecan were expressed. The presence of glycosaminoglycan chains was demonstrated for a subset of the core proteins (lumican, biglycan, and decorin) using lyase digestion. Cuprolinic blue–stained cultures showed that sulfated proteoglycans were present throughout the construct and most prominent in its mid‐region. The size of the Cuprolinic‐positive filaments resembled those previously reported in a human corneal stroma. Under the current culture conditions, the cells mimic a development or nonfibrotic repair phenotype. Developmental Dynamics 237:2705–2715, 2008.


Investigative Ophthalmology & Visual Science | 2005

Effect of wound type on smad 2 and 4 translocation

Audrey E. K. Hutcheon; Xiaoqing Q Guo; Mary Ann Stepp; Kenneth J. Simon; Paul H. Weinreb; Shelia M. Violette; James D. Zieske


Investigative Ophthalmology & Visual Science | 2017

Potential Approach to Reverse Corneal Myofibroblast Formation

Xiaoqing Q Guo; Sriniwas Sriram; Jennifer A Tran; Audrey E. K. Hutcheon; James D. Zieske


Investigative Ophthalmology & Visual Science | 2017

FAK Signaling Regulates the Difference in T1 and T3’s Ability to Stimulate SMA

Sriniwas Sriram; Xiaoqing Q Guo; Audrey E. K. Hutcheon; James D. Zieske


Investigative Ophthalmology & Visual Science | 2016

p38/TGFβ-signaling pathway involved in induction of thrombospondin-1 in human corneal fibroblasts

Audrey E. K. Hutcheon; Xiaoqing Q Guo; Sriniwas Sriram; Jennifer A Tran; James D. Zieske


Investigative Ophthalmology & Visual Science | 2016

Role of epithelial cell-derived exosomes in corneal wound healing

Jennifer A Tran; Xiaoqing Q Guo; Sriniwas Sriram; Audrey E. K. Hutcheon; James D. Zieske


Investigative Ophthalmology & Visual Science | 2016

Potential Approach to Prevent Corneal Fibrosis

Xiaoqing Q Guo; Sriniwas Sriram; Jennifer A Tran; Audrey E. K. Hutcheon; James D. Zieske


Investigative Ophthalmology & Visual Science | 2015

PDGFRα is required for the anti-fibrotic action of TGFβ3

Sriniwas Sriram; Jennifer A Tran; Xiaoqing Q Guo; Andrius Kazlauskas; James D. Zieske


Investigative Ophthalmology & Visual Science | 2015

Transplantation of 3D construct onto wounded mouse cornea

Audrey E. K. Hutcheon; Xiaoqing Q Guo; Dimitrios Karamichos; Rose Mathew; Joan Stein-Streilein; James D. Zieske

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