From these data, we conclude that CFTR-mRNA transfection could comprise a novel alternative for gene therapy to restore impaired CFTR function.

No abstract

The epithelial sodium channel (ENaC) mediates the first step in Na+ reabsorption in epithelial cells such as kidney, colon, and airways and may consist of four homologous subunits (alpha, beta, gamma, delta). Predominantly, the alpha-subunit is expressed in these epithelia, and it usually forms functional channels with the beta- and gamma-subunits. The delta-subunit was first found in human brain and kidney, but the expression was also detected in human cell lines of lung, pancreatic, and colonic origin. When co-expressed with beta and gamma accessory subunits in heterologous systems, the two known isoforms of the delta-ENaC subunit (delta1 and delta2) can build amiloride-sensitive Na+ channels. In the present study we demonstrate the expression and function of the delta-subunit in human nasal epithelium (HNE). We cloned and sequenced the full-length cDNA of the delta-ENaC subunit and were able to show that in nasal tissue at least isoform 1 is expressed. Furthermore, we performed Western blot analyses and compared the cell surface expression of the delta-subunit with the classically expressed alpha-subunit by using immunofluorescence experiments. Thereby, we could show that the quantity of both subunits is almost similar. In addition, we show the functional expression of the delta-ENaC subunit with measurements in modified Ussing chambers, and demonstrate that in HNE a large portion of the Na+ transport is mediated by the delta-ENaC subunit. Therefore, we suppose that the delta-subunit may possess an important regulatory function and might interact with other ENaC subunits or members of the DEG/ENaC family in the human respiratory epithelium.

Background Cystic fibrosis (CF) respiratory epithelia are characterized by a defect Cl − secretion and an increased Na + absorption through epithelial Na + channels (ENaC). The present study aimed to find an effective inhibitor of human ENaC with respect to replacing amiloride therapy for CF patients. Therefore, we developed specific antisense oligonucleotides (AON) that efficiently suppress Na + hyperabsorption by inhibiting the expression of the α-ENaC subunit.

The phosphodiesterase-5 inhibitor sildenafil is an established and approved drug to treat symptoms of a variety of human diseases. In the context of cystic fibrosis (CF), a genetic disease caused by a defective CFTR gene (e.g. ΔF508-CFTR), it was assumed that sildenafil could be a promising substance to correct impaired protein expression. This study focuses on the molecular mechanisms of sildenafil on CFTR recovery. We used ΔF508-CFTR/wt-CFTR expressing Xenopus laevis oocytes and human bronchial epithelial cell lines (CFBE41o^-/16HBE14o^-) to investigate the pathways of sildenafil action. Cells were treated with sildenafil and cAMP-mediated current (I_m), conductance (G_m), and capacitance (C_m) were determined. Sildenafil increased I_m, G_m, and C_m of wt-CFTR and functionally restored ΔF508-CFTR in oocytes. These effects were also seen in CFBE41oand 16HBE14o^-cells. Transepithelial measurements revealed that sildenafil mediated increase (wt-CFTR) and restoration (ΔF508-CFTR) of channel activity. cGMP pathway blocker inhibited the activity increase but not CFTR/ΔF508-CFTR exocytosis. From these data we conclude that sildenafil mediates potentiation of CFTR activity by a cGMP-dependent and initiates cGMP-independent functional insertion of CFTR/ ΔF508-CFTR molecules into the apical membranes. Thus, sildenafil is a corrector and potentiator of CFTR/ΔF508-CFTR. Yet, the necessary high doses of the drug for CFTR recovery demonstrate that sildenafil might not be suited as a therapeutic drug for CF lung disease.