The K+ Channel Opener 1-EBIO Potentiates Residual Function of Mutant CFTR in Rectal Biopsies from Cystic Fibrosis Patients

BACKGROUND: The identification of strategies to improve mutant CFTR function remains a key priority in the development of new treatments for cystic fibrosis (CF). Previous studies demonstrated that the K⁺ channel opener 1-ethyl-2-benzimidazolone (1-EBIO) potentiates CFTR-mediated Cl⁻ secretion in cultured cells and mouse colon. However, the effects of 1-EBIO on wild-type and mutant CFTR function in native human colonic tissues remain unknown. METHODS: We studied the effects of 1-EBIO on CFTR-mediated Cl⁻ secretion in rectal biopsies from 47 CF patients carrying a wide spectrum of CFTR mutations and 57 age-matched controls. Rectal tissues were mounted in perfused micro-Ussing chambers and the effects of 1-EBIO were compared in control tissues, CF tissues expressing residual CFTR function and CF tissues with no detectable Cl⁻ secretion. RESULTS: Studies in control tissues demonstrate that 1-EBIO activated CFTR-mediated Cl⁻ secretion in the absence of cAMP-mediated stimulation and potentiated cAMP-induced Cl⁻ secretion by 39.2±6.7% (P<0.001) via activation of basolateral Ca²⁺-activated and clotrimazole-sensitive KCNN4 K⁺ channels. In CF specimens, 1-EBIO potentiated cAMP-induced Cl⁻ secretion in tissues with residual CFTR function by 44.4±11.5% (P<0.001), but had no effect on tissues lacking CFTR-mediated Cl⁻ conductance. CONCLUSIONS: We conclude that 1-EBIO potentiates Cl⁻secretion in native CF tissues expressing CFTR mutants with residual Cl⁻ channel function by activation of basolateral KCNN4 K⁺ channels that increase the driving force for luminal Cl⁻ exit. This mechanism may augment effects of CFTR correctors and potentiators that increase the number and/or activity of mutant CFTR channels at the cell surface and suggests KCNN4 as a therapeutic target for CF

CNTF Mediates Neurotrophic Factor Secretion and Fluid Absorption in Human Retinal Pigment Epithelium

Ciliary neurotrophic factor (CNTF) protects photoreceptors and regulates their phototransduction machinery, but little is known about CNTF's effects on retinal pigment epithelial (RPE) physiology. Therefore, we determined the expression and localization of CNTF receptors and the physiological consequence of their activation in primary cultures of human fetal RPE (hfRPE). Cultured hfRPE express CNTF, CT1, and OsM and their receptors, including CNTFRα, LIFRβ, gp130, and OsMRβ, all localized mainly at the apical membrane. Exogenous CNTF, CT1, or OsM induces STAT3 phosphorylation, and OsM also induces the phosphorylation of ERK1/2 (p44/42 MAP kinase). CNTF increases RPE survivability, but not rates of phagocytosis. CNTF increases secretion of NT3 to the apical bath and decreases that of VEGF, IL8, and TGFβ2. It also significantly increases fluid absorption (JV) across intact monolayers of hfRPE by activating CFTR chloride channels at the basolateral membrane. CNTF induces profound changes in RPE cell biology, biochemistry, and physiology, including the increase in cell survival, polarized secretion of cytokines/neurotrophic factors, and the increase in steady-state fluid absorption mediated by JAK/STAT3 signaling. In vivo, these changes, taken together, could serve to regulate the microenvironment around the distal retinal/RPE/Bruch's membrane complex and provide protection against neurodegenerative disease

Expression of CNTF receptor-alpha in chick violet-sensitive cones with unique morphologic properties.

Journal ArticlePURPOSE: Application of ciliary neurotrophic factor (CNTF) can rescue mature photoreceptors from lesion-induced and hereditary degeneration. In the chick retina, expression of the CNTF receptor is present in a subpopulation of photoreceptor cells. The present study was undertaken to identify the CNTF receptor-expressing photoreceptors and to describe the subcellular localization of the receptor protein. METHODS: The localization of the CNTF receptor was analyzed by light and electron microscopic immunocytochemistry in chick retinal wholemount preparations, with an antibody for CNTF receptor alpha (CNTFRalpha). Immunoreactive cells were identified by double labeling with immunocytochemical markers for photoreceptor subpopulations. RESULTS: The CNTFRalpha antibody labeled evenly distributed outer segments (OS) of a photoreceptor subpopulation. CNTFRalpha-positive OS were associated with oil droplets of uniform size. Receptor immunoreactivity did not colocalize with markers for rods and red-green cones. Complete overlap was found after double labeling with the antibody CERN 933, which recognizes violet-sensitive cones in the chick retina. Ultrastructurally, the CNTFRalpha-immunoreactive OS showed rodlike properties: an elongated shape and stacks of membrane discs separated from the plasma membrane. Immunoreactivity was completely restricted to the plasma membrane of the OS and the inner membrane sheet of the photoreceptor calices present in avian retinas. CONCLUSIONS: CNTFRalpha expression identifies a unique type of photoreceptors in the avian retina which does not fit into the classic morphologic definition of rods and cones. The specific expression in violet-sensitive photoreceptors suggests that CNTF may have a neuroprotective role related to the specific function of these cells

Expression of CNTF receptor-alpha in chick violet-sensitive cones with unique morphologic properties.

Contains fulltext : 58499.pdf (publisher's version ) (Closed access)PURPOSE: Application of ciliary neurotrophic factor (CNTF) can rescue mature photoreceptors from lesion-induced and hereditary degeneration. In the chick retina, expression of the CNTF receptor is present in a subpopulation of photoreceptor cells. The present study was undertaken to identify the CNTF receptor-expressing photoreceptors and to describe the subcellular localization of the receptor protein. METHODS: The localization of the CNTF receptor was analyzed by light and electron microscopic immunocytochemistry in chick retinal wholemount preparations, with an antibody for CNTF receptor alpha (CNTFRalpha). Immunoreactive cells were identified by double labeling with immunocytochemical markers for photoreceptor subpopulations. RESULTS: The CNTFRalpha antibody labeled evenly distributed outer segments (OS) of a photoreceptor subpopulation. CNTFRalpha-positive OS were associated with oil droplets of uniform size. Receptor immunoreactivity did not colocalize with markers for rods and red-green cones. Complete overlap was found after double labeling with the antibody CERN 933, which recognizes violet-sensitive cones in the chick retina. Ultrastructurally, the CNTFRalpha-immunoreactive OS showed rodlike properties: an elongated shape and stacks of membrane discs separated from the plasma membrane. Immunoreactivity was completely restricted to the plasma membrane of the OS and the inner membrane sheet of the photoreceptor calices present in avian retinas. CONCLUSIONS: CNTFRalpha expression identifies a unique type of photoreceptors in the avian retina which does not fit into the classic morphologic definition of rods and cones. The specific expression in violet-sensitive photoreceptors suggests that CNTF may have a neuroprotective role related to the specific function of these cells