Inhibition of CLC-2 chloride channel expression interrupts expansion of fetal lung cysts

Abstract

Normal lung morphogenesis is dependent on chloride-driven fluid transport. The molecular identity of essential fetal lung chloride channel(s) has not been elucidated. CLC-2 is a chloride channel, which is expressed on the apical surface of the developing respiratory epithelium. CLC-2-like pH-dependent chloride secretion exists in fetal airway cells. We used a 14-day fetal rat lung submersion culture model to examine the role of CLC-2 in lung development. In this model, the excised fetal lung continues to grow, secrete fluid, and become progressively cystic in morphology (26). We inhibited CLC-2 expression in these explants, using antisense oligonucleotides, and found that lung cyst morphology was disrupted. In addition, transepithelial voltage (V(t)) of lung explants transfected with antisense CLC-2 was inhibited with V(t) = -1.5 +/- 0.2 mV (means + SE) compared with -3.7 +/- 0.3 mV (means + SE) for mock-transfected controls and -3.3 +/- 0.3 mV (means + SE) for nonsense oligodeoxynucleotide-transfected controls. This suggests that CLC-2 is important for fetal lung fluid production and that it may play a role in normal lung morphogenesis.

The extracellular calcium-sensing receptor, CaSR, in fetal development

“…Cl − influx raises its intracellular concentration above equilibrium to facilitate its luminal exit through apical Cl − channels [58], followed by iso-osmotic water transport [55]. Although several Cl − channels are expressed in lung epithelial cells including the cystic fibrosis conductance regulator (CFTR) [59] and several members of the Chloride Channel (CLC) family [60], the identities of the channels are not yet clear.…”

Sp1: Regulation of gene expression by phosphorylation

“…At birth, the epithelial cells switch from fluid secretion to fluid absorption (Strang, 1991). The fluid secretion and absorption across lung epithelia are driven by Cl À secretion and Na + absorption, respectively, via multiple ion channels (Blaisdell et al, 2004;Folkesson et al, 1998;O'Brodovich, 1996;Pitkanen, 2001). Accordingly, Cl À channel ClC-2 and Na + channel ENaC are transcriptionally down-and up-regulated, respectively, in the perinatal lung (Murray et al, 1995(Murray et al, , 1996Talbot et al, 1999;Watanabe et al, 1998).…”

Additional Disruption of the ClC-2 Cl- Channel Does Not Exacerbate the Cystic Fibrosis Phenotype of Cystic Fibrosis Transmembrane Conductance Regulator Mouse Models

“…This work shows that, even in a CFTRdeficient background, lung morphology is normal. A recent paper has again raised this issue using ClC-2 antisense oligonucleotides (38). However, oligonucleotides with identical sequences were used in a study on choroid plexus chloride channels (39), and they later turned out to down-regulate a chloride channel not related to ClC-2 (40).…”