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Rong Li, Tina Banzon, Rong Wen, Arvydas Maminishkis, Sheldon S. Miller; CNTF Increases Fluid Absorption Across Human RPE. Invest. Ophthalmol. Vis. Sci. 2011;52(14):861.
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Ciliary neurotrophic factor (CNTF) protects photoreceptors and regulated phototransduction machinery in photoreceptors. Little is known about its effects on retinal pigment epithelial (RPE) cells. The present work examines the effects of CNTF and two other members of the IL-6 family of cytokines, cardiotrophin-1 (CT-1), and oncostatin M (OSM), on primary cultures of human fetal RPE (hfRPE).
Quantitative RT-PCR and immunoblotting were used to examine expression of CNTF, CT-1, OSM, and their receptors, CNTF receptor α (CNTFRα), leukemia inhibitory factor receptor β (LIFRβ), gp130, and OSM receptor β (OSMRβ) in confluent monolayers of cultured hfRPE. Receptors were localized by immunocytochemistry, and phosphorylation of STAT3 and ERK was assessed by immunoblotting. Cell proliferation and survivability were examined by BrdU incorporation and MTT assay, respectively. Polarized secretion of growth factors/cytokines was measured by ELISA. Fluid absorption (JV) was measured by capacitance probes in a modified Ussing chamber.
Cultured hfRPE expresses CNTF, CT-1 and OsM, as well as CNTFRα, LIFRβ, gp130, and OsMRβ as measured by RT-PCR. The protein of LIFRβ, gp130, and OsMRβ were detected by immunoblotting. Immunocytochemistry showed that these receptors are mainly located at the apical membrane. Treatment with recombinant CNTF, CT-1, or OsM proteins significantly increased STAT3 phosphorylation. OsM also induced a significant increase in ERK phosphorylation. Cell survival was significantly increased when treated with CNTF, along with an increase in NT3 secretion, and decreases in VEGF, IL-8, TGFβ2 secretion. CNTF also significantly increased fluid absorption (JV) by activating basolateral membrane chloride channels (CFTR). This was not observed with CT-1 or OSM.
We have demonstrated that CNTF activates the JAK/STAT3 signaling in cultured hfRPE and promotes their survival. CNTF also alters growth factor, cytokine, and neurotrophic factor secretion, as well as the steady-state fluid absorption across the RPE. These results indicate that CNTF plays a role in regulating the microenvironment around the distal retinal/RPE/Bruch’s membrane complex and could contribute to the neuroprotective effects of CNTF on photoreceptors.
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