Cholera Toxin‐Enhanced Apical Localization of β‐ENaC Contributes to Elevated Mammary Na + Absorption

Abstract

Cellular mechanisms to account for the low Na+ concentration in human milk are poorly defined. MCF10A cells, a cell line derived from human mammary epithelium and grown on permeable supports, exhibited amiloride- and benzamil-sensitive ion transport (short circuit current, Isc), suggesting activity of the epithelial Na+ channel, ENaC. When cultured in the presence of cholera toxin (Ctx), MCF10A cells exhibited greater amiloride sensitive Isc at all time points tested (2h to 7d), an effect that was not reversed with Ctx washout for 12 hours. Previous data showed that the effects of Ctx on Isc were independent of cAMP and PKA. Additionally, the Ctx B subunit, alone, did not replicate such effects. RT-PCR and western blot analysis showed no significant increase in either the mRNA coding for ENaC or total protein expression for either α, β, or, γ-ENaC subunits. Likewise, the abundance of Nedd4-2 was not changed. Biotinylation analysis showed that Ctx increased β-ENaC expression on the apical cell surface. These results demonstrate that human mammary epithelia express ENaC, which can account for low milk Na+ concentration, and that Ctx enhances ENaC localization at the apical membrane. Elevated Na+ absorption, which has been associated with an enhanced barrier of bovine mammary epithelial cells, may augment epithelial repair following exposure to enterotoxins during lactation. [NIH P20-RR017686 & KS Ag Exp Stn support]