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Presence of EGF growth factor ligands and their effects on cultured rat conjunctival goblet cell proliferation
- Source :
-
Experimental Eye Research . Feb2008, Vol. 86 Issue 2, p322-334. 13p. - Publication Year :
- 2008
-
Abstract
- Abstract: The amount of mucin on the ocular surface is regulated by the rate of mucin synthesis, mucin secretion, and the number of goblet cells. We have previously shown that cholinergic agonists are potent stimuli of mucin secretion. In contrast, there have been no studies on the control of goblet cell proliferation. In this study we investigate the presence of the EGF family of growth factors and their receptors in rat conjunctiva and cultured rat conjunctival goblet cells as well as their effects on activation of signaling intermediates and goblet cell proliferation. Rat conjunctival goblet cells were grown in organ culture and identified as goblet cells by their morphology and positive staining for the lectin UEA-1 and cytokeratin 7. In the rat conjunctiva, the presence of the EGF family members epidermal growth factor (EGF), transforming growth factor α (TGF-α), heparin binding EGF (HB-EGF), and heregulin was determined by RT-PCR. The receptors for these ligands, EGF receptor (EGFR), erbB2, erbB3, and erbB4 were detected in both rat conjunctiva and goblet cells by Western blot analysis. Immunofluorescence microscopy of conjunctival tissue determined that EGFR was present as punctate staining in the cytoplasm of conjunctival goblet cells while ErbB2 was present in the basolateral and lateral membranes of goblet cells. ErbB3 was localized to the cytosol of rat conjunctival goblet cells. In cultured goblet cells, EGFR and ErbB2 were present in the perinuclear area of the cells. ErbB3 was widely distributed throughout the cytoplasm of the cells. ErbB4 was not detected in either the conjunctiva or goblet cells by immunofluorescence microscopy. Using a multiplex assay system we measured phosphorylation (activation) of p44/p42 mitogen-activated protein kinase (MAPK), also known as ERK, Jun N-terminal kinase (JNK), p38 MAPK and AKT (also known as protein kinase B), molecules known to be activated by EGF receptor members. EGF, TGF-α and HB-EGF activated the signaling intermediate proteins whereas heregulin did not. No EGF family member significantly activated AKT. Consistent with these findings, EGF, TGF-α and HB-EGF each stimulated goblet cell proliferation as measured by WST-1 assay or immunofluorescence microscopy using an antibody against Ki-67, a protein expressed in dividing cells. Heregulin did not cause goblet cell proliferation. We conclude that multiple members of the EGF family, EGF, TGF-α and HB-EGF, and heregulin are present with three of the four erbB receptor subtypes. EGF, TGF-α and HB-EGF all stimulated the activation of signaling intermediates and caused goblet cell proliferation. [Copyright &y& Elsevier]
- Subjects :
- *EYE
*GROWTH factors
*CYTOKINES
*BIOLOGICAL transport
Subjects
Details
- Language :
- English
- ISSN :
- 00144835
- Volume :
- 86
- Issue :
- 2
- Database :
- Academic Search Index
- Journal :
- Experimental Eye Research
- Publication Type :
- Academic Journal
- Accession number :
- 30028846
- Full Text :
- https://doi.org/10.1016/j.exer.2007.11.004