The effect of dietary salt intake on proximal tubular endocytic function

Albuminuria is strongly and independently associated with the risk of end-stage
renal disease. Higher sodium intake is associated with increases in urine
albumin excretion both in healthy subjects and in chronic kidney disease, and
dietary sodium restriction reduces albuminuria, partially independently of
blood pressure. A healthy person's urine is virtually devoid of the relatively
large serum protein albumin, which was classically attributed to optimal
glomerular filtration barrier function. In contrast, recent reports indicate
significant filtration of this protein and a greater quantitative role for
proximal tubule cells in reabsorbing filtered albumin to minimise albuminuria.
The endocytic receptors megalin and cubilin mediate proximal tubular
reabsorption of numerous filtered proteins. These include albumin and smaller
proteins that are more readily filtered by virtue of their low molecular
weight, such as retinol-binding protein (RBP), vitamin d-binding protein (DBP)
and *2-microglobulin (*2M). The very few factors that are known to regulate
megalin and cubilin expression include tumour growth factor-* (TGF-*) and
angiotensin II. Preliminary results from animal experiments by our group and
others indicate that the renal expression of megalin and cubilin is inversely
correlated with dietary sodium intake.This remarkable finding could offer an
additional explanation for the antiproteinuric effect of dietary sodium
restriction beyond differences in protein filtration. To our knowledge, the
effects of interventions in dietary sodium intake on proximal tubular endocytic
function have not been studied in humans.
Urinary extracellular vesicles (uEVs) are constantly excreted by tubular
epithelial cells, and they can be readily isolated from spot urine samples. It
is assumed that their composition mimics the membrane expression of proteins in
the cells they derive from. Hence, isolation and analysis of uEVs could allow
for a *liquid kidney biopsy* under various experimental conditions at no risk
or significant burden to the study participant. Cubilin and megalin can be
detected in uEVs.

To investigate whether dietary salt intake influences proximal tubular
endocytic function.

Intervention study

All participants will be sequentially placed on a low salt and a high salt
diet.

All participants are placed on a dietician-prescribed liquid low salt diet
during 8 days. In this period, participants are asked not to consume other
drinks than water, tea or a specific lemonade provided by the investigators. In
the last 4 days, dietary salt is supplemented via 1000-mg NaCl capsules, 14 per
day, to reach a high salt intake. On the day before commencement of the diet
(day 0), days 4 and 8, participants collect 24-hour urine, and blood pressure
is measured during 24 hours using a wearable device (ABPM). To avoid potential
interference, participants are asked to refrain from strenuous physical
exercise (sports) and sexual intercourse on these specific days. After
finishing 24-hour urine collection and ABPM, participants drink 300 mL of
water. Upon arrival in the Erasmus MC, a fasted blood sample is drawn, spot
urine is collected, participants are weighed and a body composition measurement
is performed. For the latter, we use the Body Composition Monitor device, which
quickly measures fluid distribution via electrodes placed on hands and feet in
a non-painful, noninvasive manner. After this, participants can commence their
next dietary phase (day 5) or leave the study (day 9).