title: Stromal-epithelial interaction study: The effect of corneal epithelial cells on growth factor expression in stromal cells using organotypic culture model.
abstract: Interactions between stromal and epithelial cells play important roles in the development, homeostasis, and pathological conditions of the cornea. Soluble cytokines are critical factors in stromal-epithelial interactions, and growth factors secreted from corneal stromal cells contribute to the regulation of proliferation and differentiation of corneal epithelial cells (CECs). However, the manner in which the expression of growth factors is regulated in stromal cells has not been completely determined. To study stromal-epithelial cell interactions, we used an organotypic culture model. Human or rabbit CECs (HCECs or RCECs) were cultured on amniotic membranes placed on human corneal fibroblasts (HCFs) embedded in a collagen gel. The properties of the organotypic culture were examined by hematoxylin-eosin staining and immunofluorescence. In the organotypic culture, HCECs or RCECs were stratified into two-three layers after five days and five-seven layers after nine days. However, stratification was not observed when the HCECs were seeded on a collagen gel without fibroblasts. K3/K12 were expressed on day 9. The HCF-embedded collagen gels were collected on days 3, 5, or 9 after seeding the RCECs, and mRNA expression of growth factors FGF7, HGF, NGF, EGF, TGF-α, SCF, TGF-β1, TGF-β2, and TGF-β3 were quantified by real-time PCR. mRNA expression of the growth factors in HCFs cultured with RCECs were compared with those cultured without RCECs, as well as in monolayer cultures. mRNA expression of TGF-α was markedly increased in HCFs cultured with RCECs. However, mRNA expression of the TGF-β family was suppressed in HCFs cultured with RCECs. Principal component analysis revealed that mRNA expression of the growth factors in HCFs were generally similar when they were cultured with RCECs. In organotypic cultures, the morphological changes in the CECs and the expression patterns of the growth factors in the stromal cells clearly demonstrated stromal-epithelial cell interactions, and the results suggest that stromal cells and epithelial cells may act in concert in the cornea.
authors: Department of Ophthalmology and Regenerative Medicine, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan; Department of Stem Cell Biology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
authors: Takeshi
authors: T
authors: Kobayashi
authors: Department of Ophthalmology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan; Department of Stem Cell Biology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan. Electronic address: shiraia@m.ehime-u.ac.jp.
authors: Atsushi
authors: A
authors: Shiraishi
authors: Department of Ophthalmology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
authors: Yuko
authors: Y
authors: Hara
authors: Department of Ophthalmology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
authors: Yuko
authors: Y
authors: Kadota
authors: Department of Dermatology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
authors: Lujun
authors: L
authors: Yang
authors: Department of Ophthalmology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
authors: Tomoyuki
authors: T
authors: Inoue
authors: Department of Dermatology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
authors: Yuji
authors: Y
authors: Shirakata
authors: Department of Ophthalmology, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan; Department of Infectious Diseases, Ehime University Graduate School of Medicine, Shitsukawa, Toon, Ehime 791-0295, Japan.
authors: Yuichi
authors: Y
authors: Ohashi
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D002450:Cell Communication / Q000502:physiology*; D002478:Cells, Cultured; D003315:Cornea / Q000166:cytology*; D004815:Epidermal Growth Factor / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism*; D005346:Fibroblast Growth Factors / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D023421:Models, Animal; D009414:Nerve Growth Factors / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D017154:Stromal Cells / Q000378:metabolism*; D015291:Transforming Growth Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; D009414:Nerve Growth Factors; D012333:RNA, Messenger; D005346:Fibroblast Growth Factors; D004815:Epidermal Growth Factor; D015291:Transforming Growth Factors
keywords: Cornea; Growth factor; Organotypic culture; Stromal cell; Stromal–epithelial interaction