Abstract

Background and Aims: Alterations in the resorptive
capacity of renal epithelia to sodium can have severe
implications for the normal functioning of the nephron and
are most likely to be prominent in the development and
pathogenesis of secondary hypertension, a condition associated
with a number of renal diseases including diabetic
nephropathy and glomerulonephritis. One of the major
regulators of sodium reabsorption in the nephron is the
serum and glucocorticoid induced kinase (SGK), an
aldosterone regulated gene, which mediates sodium reabsorption
via its actions on the epithelial sodium channel
(ENaC). Recent reports have identified SGK as a key
signaling element whose level of expression appears to be
upregulated in kidneys of diabetic humans and diabetic
nephropathy. Hyperglycamia is believed to be the key
pathophysiological component in promoting intercellular
changes ultimately leading to the deranged transcriptional
regulation of SGK.
Materials and Methods: RT-PCR, immunocytochemistry
and western blot analysis confirmed that human cortical
collecting duct (HCD) cells express SGK and alpha ENaC.
For glucose, experiments, HCD cells were treated with
(25 mM) glucose for 24 and 48 hours. For TGF-beta (2 nm)
and Ionomycin (1 2m) experiments, HCD cells were treated
for 4,6,8,12 and 24 hrs. A functional correlate for these
increased levels of expression was provided by sodium
microfluorimetry.
Results: Incubation of HCD cells with high glucose
(25 mM) for 24 and 48 hours increased expression of
SGK and Alpha ENaC protein (SGK, 261.7T14.9% of
control (5 mM) at 48 hours; n=3, p<0.01%; alpha ENaC,
316.5%T9.9% of control (5 mM) at 48 hours n=6, p<0.01).
Hyperglycaemia has been linked to both raised calcium and
TGF-beta levels. Application of TGF-beta or Ionomycin
alone induced a significant increase in SGK protein
expression (TGF-beta, 125.5%T7.5% of control at 8 hours;
n=3 p<0.001%; Ionomycin 263.7%T8.6% of control at
8 hours; n=3 p<0.001) suggesting a possible role for both
TGF-beta and calcium in mediating downstream effects of
hyperglycaemia. Intracellular sodium levels were found to
be significantly elevated following 24 (107%T0.88%) and
48 (114%T0.93%) hours exposure to 25 mM glucose, as
compared to cells cultured in low (5 mM) glucose.
Conclusion: This data taken in conjunction with that
described in previous literature suggests that increases in
intracellular sodium in response to high glucose after 48
hours may be mediated via the increased expression of both
SGK and alpha-ENaC. This data highlights a possible link
between hyperglycaemia and the deranged sodium reabsorption
observed in cases of diabetic nephropathy, and
may lead to identification of potential therapeutic targets.