Abstract
Purpose: :
Ciliary neurotrophic factor (CNTF) has been shown to protect photoreceptors in several models of retinal degeneration. In this work, we examined the effects of CNTF, cardiotrophin 1 (CT1), oncostatin M (OsM), and leukemia inhibitory factor (LIF) on RPE signaling and physiology.
Methods: :
Gene expression of CNTF, CT1, OsM, LIF and their receptor subunit expression and localization were examined by quantitative RT-PCR, immunoblot, and immunofluorescence analysis using confluent monolayers of human fetal retinal pigment epithelium (hfRPE) primary cultures. Signal transduction was studied by measuring the phosphorylation of STAT3 and ERK in hfRPE as well as in adult RPE (ARPE-19). Cell proliferation was assessed by BrdU incorporation. Fluid absorption (JV) across RPE (retina to choroidal side) was determined by a capacitance probe.
Results: :
The expression of CNTF, CT1, OsM and LIF, as well as receptor subunits, including CNTFRα, LIFRβ, gp130 and OsMRβ were all detected in hfRPE by RT-PCR and immunoblotting. The amounts of LIF and CT1 mRNA are more than CNTF and OsM, and CNTFRα protein is below the detection level of immunoblotting. All of these receptor subunits, including CNTFRα, LIFRβ, gp130, and OSMRβ are mainly localized at the apical membrane. Treatments of CNTF, CT1 and OsM induced phosporylation of STAT3 in hfRPE and ARPE-19. In addition, OsM significantly activated P44/P42 (ERK) MAP kinase pathway, but not CNTF and CT1. CT1 shows a stimulatory effect (25%) on RPE proliferation at 50 - 100 ng/ml whereas OsM significantly inhibited hfRPE proliferation between 10 and 100 ng/ml. CNTF had no significant effect on hfRPE proliferation. Furthermore, CNTF significantly increased fluid transport (JV) from 8.7 ± 0.7 to 20.7 ± 3.3 µl·cm-2· hr-1 (n= 3; P < 0.05).
Conclusions: :
These studies demonstrate that CNTF, CT1, and OsM can activate the JAK/STAT3 signaling pathway in human RPE (Li et al., AJP cell physiology, 2009), raising the possibility that the neuroprotective effects of these factors on photoreceptors are partially mediated by RPE activity.
Keywords: retinal pigment epithelium • cytokines/chemokines • signal transduction