Proceedings of The Physiological Society
University of Oxford (2011) Proc Physiol Soc 23, PC65
Haplotype deletion of the gene encoding 11-beta-hydroxysteroid dehydronase type 2 (11-β-HSD2+/-) alters the activity of renal serum / glucocorticoid-inducible kinase 1 (SGK1) in mice fed a high Na+ diet
M. A. Bailey2, S. H. Rankin1, J. M. Hendry1, N. A. Ismail1, S. M. Wilson1
1. Centre for Cardiovascular and Lung Biology, University of Dundee, Dundee, United Kingdom. 2. University/BHF Centre for Cardiovascular Science, University of Edinburgh, Edinburgh, United Kingdom.
Na+ re-absorption within the aldosterone-sensitive distal nephron (ASDN) determines the amount of Na+ lost in urine and is therefore is critical to whole body Na+ and water balance, and crucial to the control of blood pressure (Loffing & Korbmacher, 2009). This process depends upon epithelial Na+ channels (ENaC) which provide a tightly regulated route for apical Na+ entry in absorptive epithelia. Mice with a haplotype deletion of 11-β-HSD2 display inappropriately high levels of ENaC-dependent Na+ absorption when fed a high Na+ diet and this excessive retention of Na+ leads to the development of severe hypertension. Since the physiological role of 11-β-HSD2 is to “protect” the ASDN from circulating cortisol, this abnormally high level of ENaC activity appears to be an inappropriate response to this hormone (Bailey et al., 2010). Such steroid hormones control ENaC via a mechanism that is dependent, at least in part, upon SGK1, and so we have now assayed the activity of this regulatory kinase in renal tissue from 11-β-HSD2+/- mice and from wild type litter mates that had been maintained on a high Na+ diet for 21 days. Kidneys that had been removed from the mice used in an earlier, physiological study (Bailey et al., 2010) and stored at -80°C, were therefore homogenized in the presence of protease and phosphatase inhibitors and aliquots (80 μg) of extracted protein fractionated by SDS-polyacrylamide gel electrophoresis and subject to Western analysis using antibodies against the total and Thr346/356/366-phosphorylated forms of the protein encoded by the N-myc downstreame reguylated gene 1 (NDRG1), and β-actin. These analyses clearly showed (Fig. 1) that Thr346/356/366-phosphorylated NDRG1 was less abundant in the 11-β-HSD2+/- mice (Fig. 1) and this effect cannot be attributed to differences in the amout of protein loaded onto the gels since the overall abundance of NDRG1, and the abundance of beta-actin were very similar (Fig. 1). Since NDRG1-Thr346/356/366 is a physiological substrate for SGK1 but not for other, closely related kinases (Murray et al., 2004), the excessive ENaC activity seen in the kidneys of 11-β-HSD2+/- mice (Bailey et al., 2010) now appear to be associated abnormally low levels of SGK1 activity. Whilst this finding was not anticipated, It is interesting that airway epithelial cells chronically (6 - 7 days) exposed to glucocorticoids also display persistent ENaC activity that is accompanied by a loss of SGK1 activity (Inglis et al., 2009). Glucocorticoid-induced SGK1 activation cannot, therefore account for the sustained activation of ENaC seen in 11-β-HSD2+/- mice.
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