Distal renal tubular acidosis in primary Sjögren syndrome

Prevalence of dRTA in pSS
The association between dRTA and pSS has been described in various case
reports. It is unclear what the true prevalence of dRTA in pSS is. In our
previous study, we found a prevalence of 39%. A study which used both urinary
acidification tests (the ammonium chloride loading test and the
furosemide/fludrocortisone test) to detect dRTA in a large group of pSS
patients is lacking. Furthermore, the prevalence of complications of dRTA is
poorly described and is based at retrospective data.

Role for auto-antibodies
Why patients with pSS develop dRTA is unknown. Autoantibody formation may play
a role as was suggested by the study of Takemoto et al. We postulate that
antibodies against carbonic anhydrase type 2 might be involved in the
development of dRTA. Carbonic anhydrase type 2 is a member of the α-
carbonic-anhydrase family and the enzyme is expressed in the cytosol of both
proximal and distal renal tubulus, the aqueous chamber of the eye and
erythrocytes. Activity of carbonic anhydrase type 2 catalyzes carbondioxide to
generate a proton and a bicarbonate ion, which is an important process in the
acid-base metabolism.

Role for urinary exosomes
In 2004 the presence of exosomes in urine was first reported, including a
reproducible method to isolate these exosomes. One of the investigators worked
in the laboratory where the method to isolate urinary exosomes was developed
(National Institutes of Health, USA) and also proposed a protocol to use
urinary exosomes as biomarkers. Mass spectrometry analysis revealed that
urinary exosomes contained several disease-associated proteins that originated
from the renal tubular epithelium. Among these proteins was several transport
proteins located at the apical plasma membrane. The question remained, however,
if the presence of these proteins in urinary exosomes correlated with disease
processes in the kidney. We recently addressed this question and showed that
during the disease process of aldosteronism, the abundance of an
aldosterone-sensitive sodium transporter (the sodium chloride cotransporter)
was increased in urinary exosomes of experimental animals and patients. In
analogy with these results, we propose that the abundance of acid-base
transporters in the distal tubule in urinary exosomes correlates with dRTA in
pSS.

The end-points of this study are:

1. To evaluate the prevalence of dRTA in Sjögren using a new screening test
with furosemide and fludrocortisone and compare this against the gold standard
test with ammonium chloride
2. To determine the prevalence of disorders in calcium metabolism in patients
with pSS and dRTA
3. To investigate whether auto-antibodies against carbonic anhydrase are
involved in the pathogenesis of dRTA in pSS
4. To investigate the hypothesis that renal acid-base transporters are absent
in urinary exosomes

The study is an observational study with invasive measurements. All 33 subjects
(with an abnormal urinary acidificaiton test in the past) will be seen twice in
the hospital, with minimal one week in between. On the first day, they will
undergo the ammonium chloride loading test. On the second day, the furosemide
and fludrocortisone test will be repeated. During the time they are in the
hospital, the DEXA scan and ultrasound examination will be performed.
The 105 control subjects will only undergo the radiographic examinations. Both
tests will be performed on one day.

We collected and stored blood and urine of pSS patients in our previous study.
In a later stage of the present study, we will perform indirect
immunofluorescence on patients* sera to evaluate the presence of
auto-antibodies against kidney tissue. Urine will be stored to perform urinary
exosome analysis. Also this part will be performed in a later stage of the
study.

The urinary acidification test by a single simultaneous oral furosemide and
fludrocortisone treatment is an easy, effective and well-tolerated test. In our
previous study, we performed this test on 96 subjects and no side effects were
described. The ammonium chloride test has some side effects (e.g. vomitting,
bloating). This side effects are temporarely. Although the test is safe, all
participants are insured for adverse events.
The patients with an abnormal urinay acidification test (with furosemide and
fludrocortisone) in the previous study maybe have (not) dRTA.

According to the literature, we thought that this test was specific to detect
dRTA, but the high amount of abnormal tests in the controlgroup challenges
this. Therefore, we made this follow-up study to be sure of the test results
and give patients the results they deserve. Additionally, this study provides
a load of information about this diagnostic tests and the prevalences of dRTA
and its complications.