title: Three-dimensional cell culture environment promotes partial recovery of the native corneal keratocyte phenotype from a subcultured population.
abstract: Corneal disease is the fourth leading cause of blindness. According to the World Health Organization, roughly 1.6 million people globally are blind as a result of this disease. The only current treatment for corneal opacity is a corneal tissue transplant. Unfortunately, the demand for tissue exceeds supply, making a tissue-engineered in vitro cornea highly desirable. For an in vitro cornea to be useful, it must be transparent, which requires downregulation of the light-scattering intracellular protein alpha-smooth muscle actin (αSMA) and upregulation of the native corneal marker, aldehyde dehydrogenase 1A1 (ALDH1A1). This study focuses on the effects of a three-dimensional (3D) matrix on the expression levels of αSMA and ALDH1A1 by a subcultured population of rabbit corneal keratocytes and the comparison of the 3D matrix effects to other culture conditions. We show that, through western blot and quantitative real-time PCR, the presence of collagen strongly downregulates αSMA. Further, 3D cultures maintain low actin expression even in the presence of a proinflammatory cytokine, transforming growth factor-beta (TGF-β). Finally, 3D culture conditions show a partial recovery of ALDH1A1 expression, which has never been previously observed in a serum-exposed subcultured cell population. Overall, this study suggests that 3D culture is not only a relatively stronger signal than both collagen and TGF-β, it is also sufficient to induce some recovery of ALDH1A1 and the native corneal phenotype despite the presence of serum.
authors: Department of Biology, Harvey Mudd College, Claremont, CA 91711, USA.
authors: Russell E
authors: RE
authors: Thompson
authors: Liana C
authors: LC
authors: Boraas
authors: Miranda
authors: M
authors: Sowder
authors: Marta K
authors: MK
authors: Bechtel
authors: Elizabeth J
authors: EJ
authors: Orwin
mesh_terms: D000444:Aldehyde Dehydrogenase / Q000378:metabolism; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002478:Cells, Cultured; D060527:Corneal Keratocytes / Q000166:cytology* / Q000378:metabolism; D011817:Rabbits
chemical_list: D000444:Aldehyde Dehydrogenase