Keywords
indomethacin - nephrocalcinosis - neonatal Bartter syndrome - trophoblast basement
membrane
Bartter syndrome is an inherited renal tubular disorder associated with hypokalemic
alkalosis. The symptoms of neonatal Bartter syndrome (NBS) are polyuria, polyhydramnios,
and premature birth. Characteristic laboratory findings include hyperreninemia, hyperaldosteronism,
and high prostaglandin E2 (PGE2) production.[1]
[2]
[3] In NBS, activation of cyclooxygenase (COX) increases PGE2 production. Thus, in addition
to water and electrolyte supplementation, the COX inhibitor indomethacin (IND) has
been used to treat NBS.[4]
[5]
[6]
[7]
Here, we discuss nephrocalcinosis, one of the clinical characteristics and placental
findings in a patient with NBS. Written informed parental consent was obtained for
this study.
Case Report
The 27-year-old mother had two healthy children from a nonconsanguineous marriage.
Polyhydramnios was noted at 26 weeks of gestation. No distinct anomaly was detected
by ultrasound. Amniotic fluid was removed three times (1500, 1300, and 2800 mL) and
was not bloody. Amniotic fluid analysis revealed a 46,XY karyotype. Labor began just
after the third removal of amniotic fluid. Cesarean section was performed as the baby
was in the transverse position. The boy was born at 31 weeks and 1 day, weighed 1344 g,
and had an Apgar score of 8/8. No anomalies were detected, and we were able to insert
a tube into the stomach.
At 4 hours of age, polyuria (9.8 mL/kg/h) was detected. We also observed extensive
loss of electrolytes into the urine: Na 112 mEq/L; K 5.0 mEq/L; Cl 106 mEq/L; Ca 7.9
mg/dL; and creatinine (Cr) 1.67 mg/dL ([Table 1]). Similar proportions of electrolytes were detected in the amniotic fluid. He was
given sufficient water and electrolytes. NBS was strongly suspected due to the presence
of renin at >400 pg/mL, aldosterone at 2020 pg/mL, and antidiuretic hormone at 1 pg/mL
in the baby's blood on day 0.
Table 1
Laboratory Findings at Birth
Blood
|
Urine
|
Amniotic fluid
|
WBC
|
5900/μL
|
pH
|
7.292
|
Na
|
112 mEq/L
|
Na
|
131 mEq/L
|
RBC
|
3.95 × 106/μL
|
pCO2
|
56.1 mm Hg
|
K
|
5.0 mEq/L
|
K
|
4.1 mEq/L
|
Hb
|
15.0 g/dL
|
pO2
|
34.5 mm Hg
|
Cl
|
106 mEq/L
|
Cl
|
106 mEq/L
|
Ht
|
43.7%
|
HCO3
|
26.3 mEq/L
|
Ca
|
7.9 mg/dL
|
Ca
|
6.1 mg/dL
|
PLT
|
168 × 103/μL
|
BE
|
−1.0 mEq/L
|
P
|
6.5 mg/dL
|
P
|
3.8 mg/dL
|
CRP
|
0.05 mg/dL
|
Na
|
130.8 mEq/L
|
Cr
|
1.67 mg/dL
|
Cr
|
0.59 mg/dL
|
BUN
|
4 mg/dL
|
K
|
3.94 mEq/L
|
Osm
|
229 mOsm
|
Osm
|
257 mOsm
|
Cr
|
0.43 mg/dL
|
Cl
|
96.0 mEq/L
|
|
|
|
|
Renin
|
>400 pg/mL
|
iCa
|
1.44 mg/dL
|
|
|
|
|
Aldosterone
|
2020 pg/mL
|
Lactate
|
24 mg/dL
|
|
|
|
|
ADH
|
1.0 pg/mL
|
Glu
|
55 mg/dL
|
|
|
|
|
WBC, white blood cell; RBC, red blood cell; Hb, hemoglobin; Ht, hematocrit; PLT, platelets;
CRP, C-reactive protein; BUN, blood urea nitrogen; Cr, creatinine; ADH, antidiuretic
hormone; BE, base excess; Osm, osmolarity.
He suffered from neonatal toxic-shock-syndrome-like exanthematous disease on day 5
due to methicillin-resistant Staphylococcus aureus and was treated with antibiotics. On day 8, his blood pressure (65/29 to 41/16 mm
Hg) and urine output (11.6 to 3.5 mL/kg/h) decreased. There was the absence of diastolic
renal artery flow in the ultrasound, and he was diagnosed with late-onset circulatory
collapse. We initiated hydrocortisone (HDC) on day 11, and subsequently his urine
output improved. We attempted to decrease the amount of HDC twice, but without success.
On day 31, IND was initiated at 0.2 mg/kg/d and was gradually increased to 2.7 mg/kg/d.
After IND initiation, HDC was reduced and finally terminated on day 73. During his
hospital stay, plasma aldosterone levels gradually decreased from 2020 pg/mL on day
0 to 1280 and 935 pg/mL on days 31 and 69, respectively, whereas the plasma renin
levels were over the measurement range on days 0, 31, and 69. Ca was given intravenously
from day 0 (600 mg/kg/d) and decreased gradually. Breast milk was started at day 0
and reached at 92 mL/kg/d at day 7. After the late-onset circulatory collapse at day
8, breast milk decreased. Afterward, the breast milk was increased again and became
over 100 mL/kg/d at day 13. In addition to fortifier, we added calcium lactate at
day 19. We added vitamin D (0.05 μg/kg/d) at day 20. The serum level of Ca was almost
kept from 9 to 11 mg/dL, with the nadir of 8.4 mg/dL at day 21. Urinary Ca (mg/mg
Cr) remained high (2 to 3) even after IND administration ([Fig. 1]). Nephrocalcinosis was not detected by ultrasound during his hospital stay. Urinary
Fe was <0.18 mg/d on day 14.
Fig. 1 Urinary Ca (mg/mg Cr, circle marker) levels before and after indomethacin administration
(mg/kg/d, square marker).
With respect to the auditory brain stem response (ABR), V waves were not detected
at 35 dB in either ear at the corrected age of 41 weeks. Brain magnetic resonance
imaging showed no abnormal findings at the corrected age of 40 weeks.
He harbored a compound heterozygous mutation in exon 8 c.del1100t and exon 10 p.R471X
of the SLC12A1 gene encoding NKCC2. Both mutations were new to the Human Gene Mutation Database.
We did not determine the functional consequences of our mutants. However, two mutations
might cause the disease. One mutation, R471X, which was located in exon 10, is subjected
to nonsense-mediated decay. The other deletion mutation changes the open reading frame
after the deletion site. Thus NKCC2 function might be impaired by amino acids substitutions.
We provided genetic counseling for his parents; however, they did not want to get
their DNA analyzed.
The placenta weighed 700 g (+2.7 standard deviations).[8] Although the proportion of terminal villi was consistent with the gestational age,
many of them showed poorly dilated capillaries ([Fig. 2]). Hemosiderin pigment was seen throughout the amniochorionic connective tissue and
along about 50% of the trophoblast basement membrane (TBM; [Fig. 3]).Von Kossa stain also revealed the corresponding area of mineralization along the
TBM ([Fig. 3]). No vascular damage or interstitial hemorrhage was detected in the villi and the
amniochorionic connective tissue.
Fig. 2 Hematoxylin-eosin staining of the placenta (A). Although the proportion of terminal villi was consistent with the gestational age,
many of them showed poorly dilated capillaries. CD34-positive endothelial cells (B).
Fig. 3 Placental histology. Hematoxylin-eosin staining (A, D). Hemosiderin pigment deposition seen throughout the amniochorionic connective tissue
and along about 50% of trophoblast basement membrane (B, E). Von Kossa staining along the trophoblast basement membrane (C, F).
On day 77 (42 weeks old), he was discharged with IND, NaCl, and potassium glucuronate.
At 4 months of age, nephrocalcinosis was detected by ultrasound. At 7 months of age
(corrected age: 5 months), the ABR was normal. Catch-up growth and rolling over were
detected.
Discussion
Mutations in ion transport channels lead to decreased Ca reabsorption in patients
with NBS, which is similar to that observed in patients treated with loop diuretics.
The end result is hypercalciuria, leading to nephrocalcinosis.
Dane et al reported two cases (33 weeks, 1640 g and 35 weeks, 2420 g) where treatment
with IND was initiated in the antenatal period. At 18 months of age, the first case
had minimal calcification and the second had no calcification,[5] suggesting that antenatal IND treatment was effective in preventing nephrocalcinosis
([Table 2]). In the present case, however, nephrocalcinosis was detected at 4 months of age,
and urinary Ca levels did not decrease despite the treatment. Although antenatal IND
could prevent nephrocalcinosis, there would be some side effects.[9] On the contrary, our findings suggest that nephrocalcinosis may be inevitable because
IND was not effective in our case. We checked the usage dose of IND in the previous
reports.[5]
[6]
[10] It was around 2 to 3 mg/kg/d, which was too much for us, because we usually use
it in dosages of 0.2 mg/kg for patent ductus arteriosus therapy. So we started at
a small dose and increased the dose, taking care of side effects.
Table 2
Indomethacin Dosages and Nephrocalcinosis in Neonatal Bartter Syndrome
Author
|
Indomethacin Dosage
|
Nephrocalcinosis
|
Nakagawa[10]
|
1st case (42 wk): 1 y old, ~3 mg/kg/d (18 mg/d); 2 y old, ~22 mg/d; 5 y old, ~25 mg/d;
16 y old, ~0.5 mg/kg/d
|
Not mentioned
|
Konrad[6]
|
1st case (28 wk, 750 g): 11 wk old, ~1.6 mg/kg/d, then 3 mg/kg/d
2nd case (35 wk, 2670 g): 7 d old, ~1 mg/kg/d, then 0.2 mg/kg/d due to oliguria, then
0.6 mg/kg/d
|
1st case: (+) at 2 mo of age
2nd case: mild (+) at 3 mo of age
|
Ernst[11]
|
1st case (30 wk) and 2nd case (32 wk); Indomethacin was used (dosage not mentioned)
|
Both cases: (+)
|
Adachi[4]
|
1st case (35 wk, 2340 g): 0.33 mg/kg/d
2nd case (33 wk; 1918 g); 0.1 mg/kg/d
|
1st case: (+) at 1.5 y of age
2nd case: (+) at 87 d of age
|
Dane[5]
|
1st case (33 wk, 1640 g): 2 mg/kg/d to mother before birth; at 2 wk of age, initiation
of indomethacin; spontaneous intestinal perforation at 3 wk of age
2nd case (35 wk, 2420 g): 1 mg/kg/d to mother before birth; at 16 d of age, initiation
of indomethacin
|
1st case: minimal calcification at 18 mo of age
2nd case: (−) at 18 mo of age
|
In the previous reports, the placental weight was high.[5]
[11] Small vessels in the villi showed reduced blood circulation. We believe that villi
may grow reactively, thereby increasing placental weight. We could not find the cause
in the previous reports. The receptors to angiotensin and PGE2 exist in the placenta.[12]
[13] We doubt that there would be the influence of them.
According to Dane et al, fetal renal excretion of Ca could lead to dystrophic calcification
with placental involvement, particularly in a subtrophoblastic pattern.[5] Ernst and Parkash hypothesized that a relatively hypocalcemic fetus would require
increased transport of Ca ions across the placenta and that excesses of Ca due to
overloading of the transport system could be deposited beneath the trophoblastic layer.[11]
These theories may explain elevated Ca levels in the amniotic fluid and fetal hypocalcemia-induced
increase in mother-to-fetus Ca transport. However, the passage of Ca through amniochorionic
connective tissue to the TBM as proposed by Dane et al[5] is difficult to understand. Only a few studies have specifically examined these
linear subtrophoblastic deposits. Krohn et al reported increased membrane calcifications
in polyhydramnios.[14] In polyhydramnios, fetal hypovolemia may be caused by increased urine output and
intestinal atresia, among others. Accordingly, maternal plasma, including Ca, may
pass through the TBM, resulting in Ca deposits.
The hemosiderin deposition pattern was similar to that of Ca, suggesting that the
same mechanism may underlie hemosiderin deposition. In other words, increased Fe in
the blood and urine caused by enhanced maternal plasma inflow may explain the hemosiderin
deposition in the amniochorionic connective tissue. However, increased renal Fe has
not been reported, and in our case, the infant did not have much urinary Fe. Furthermore,
the amniotic fluid was not bloody. Hemosiderin pigment was seen along about 50% of
the trophoblast basement membrane, which was less than 75%.[11] But this finding is nonspecific. Thus, the low percentage would not be the problem.
In conclusion, nephrocalcinosis may be inevitable in NBS. Enhanced maternal plasma
inflow through TBM could cause Ca deposition, given that the same finding was obtained
in the case with polyhydramnios. The same mechanism could account for the hemosiderin
pigment distribution.