Cell and tissue specific expression of human Krüppel-liketranscription factors in human ocular surface

[摘要] Purpose: The ocular surface, composed of the conjunctiva and thecornea, is essential for vision. Its integrity depends on numerousmolecular and cellular processes such as proliferation, differentiation,apoptosis, adhesion, and extracellular matrix homeostasis, whosederegulation can induce ophthalmological pathologies. The Krüppel-liketranscription factors (KLFs) family is made up of 15 C2H2zinc-finger proteins involved in vertebrate development and able tocontrol cell proliferation, growth, and differentiation. In order tobetter define their respective roles in the human ocular surface, wedecided to determine their pattern of expression in ocular tissues. Wethen focused on the expression of KLF4 and some of its target genes toestablish KLF4's biological activities in human ocular surface.Methods: Firstly, total mRNA was extracted from human total cornea,conjunctiva, corneal epithelial cells (primary culture and establishedcell line), corneal keratocytes (primary culture), corneal endothelialcells (established cell line), and conjunctival epithelial cells(established cell line) and submitted to RT-PCR experiments in order todetermine the expression patterns of the different KLFs. Secondly, KLF4protein localization was visualized by immunofluorescence assays attissue and cell levels. Finally, KLF4 target genes (endoglin, ornithinedecarboxylase) mRNA expression levels were determined bysemi-quantitative RT-PCR, after KLF4 transient transfection in humancorneal epithelium (HCE) cells.Results: We detected the presence of twelve transcripts of KLFs inthe cornea (KLF2, KLF3, KLF4, KLF5, KLF6, KLF7, KLF8, KLF10, KLF11,KLF12, KLF13, and KLF16) and eight in the conjunctiva (KLF2, KLF3, KLF4,KLF6, KLF7, KLF10, KLF11, and KLF12). Under our conditions, thetranscripts encoding KLF1 and KLF9 were never detected. Specificexpression patterns of each KLF were also determined for the majorcellular components of the human cornea and conjunctiva. KLF4immunolocalization assays indicated its presence in both the cytoplasmicand nuclear compartments of conjunctival and corneal cells. KLF4transient overexpression in HCE cells down regulated both endoglin andODC mRNA levels.Conclusions: For the first time, we established the presence of aKLF network in the human ocular surface and illustrated the conservationof KLF4's biological properties in a corneal derived epithelial cellline.

[学科分类] 生物化学/生物物理