Abstract
Abstract: :
Purpose: The tight regulation of lens cell proliferation in the eye is controlled by the presence of growth factors in the ocular media, namely the aqueous humour. A number of mitogens such as FGF, IGF, PDGF and EGF have been shown to induce cell proliferation in vitro. The mitogen(s) responsible for the maintenance of lens cell proliferation in vivo and the signalling molecules activated in response to these have yet to be elucidated. Previous studies in our laboratory have demonstrated a key role for extracellular–signal regulated kinase (ERK1/2) in FGF–induced lens cell proliferation. To further understand the signalling pathways involved in regulating lens cell proliferation, the present study aimed to determine the role of ERK signalling downstream of different lens mitogens. This information may potentially provide us with the means of identifying mitogen(s) responsible for lens cell proliferation in vivo. Methods: Rat lens epithelial explants were treated with IGF–1, FGF–2 or bovine aqueous humour for up to 48hrs. Cell proliferation was assayed using immunolabelling for the incorporation of BrdU. Activation of ERK was assayed using immunofluorescence and Western blotting. Blocking experiments were performed using UO126 (a specific inhibitor of ERK activation). Results: Within a 24hr culture period, aqueous, IGF and FGF were all shown to stimulate the proliferation of lens epithelial cells. Blocking experiments showed this proliferation to be dependent on the activation of ERK1/2. Interestingly, the ERK profiles for each treatment differed. Examination of the pattern of phosphorylated ERK elicited over 24hrs revealed that IGF induced transient activation of ERK, peaking at 20mins of culture. In contrast, FGF, like aqueous, stimulated maximal ERK activation within 10mins which was sustained for up to 6hrs. In addition, these ERK profiles correlated with the different proliferative responses of lens epithelial cells to short term growth factor exposures. Exposure of lens cells to IGF for up to 10mins was sufficient to induce comparable levels of cell proliferation to that of cells exposed to IGF for up to 24hrs. In contrast, similar to aqueous, a short–term pulse of FGF was unable to induce cell proliferation after 24hrs. Conclusions: Characterising growth factor–induced signalling pathways in vitro may lead to the identification and better understanding of the roles these specific growth factors play in regulating lens cell behaviour in vivo.
Keywords: signal transduction • growth factors/growth factor receptors • proliferation