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NKCC-1 and ENaC are down-regulated in nitrofen-induced hypoplastic lungs with congenital diaphragmatic hernia

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Abstract

Congenital diaphragmatic hernia (CDH) is accompanied by pulmonary hypoplasia and pulmonary hypertension. Fetal lung growth is dependent on the secretion of lung liquid, which normally is absorbed at partus. The ion channel NKCC-1 is involved in this secretory process, but has recently also been reported to be implicated in absorption. CDH patients show a disturbed transition from secretion to absorption. α- and β-ENaC are essential for lung liquid absorption. Common for all transcellular ion transport is the need for Na/K-ATPase as a primary driving force. The aim of the study was first to map the normal pulmonary expression of the above proteins during late gestation and secondly to see if the expression was affected in a CDH rat model. Pregnant Sprague–Dawley rat dams were given nitrofen on gestational day 9.5 to induce CDH. The fetuses were removed on gestational days E18 and E21. In addition, newborn rats were harvested postpartum on day P2. The fetuses were put into one of two groups: hypoplastic lungs without CDH (N−CDH) and hypoplastic lungs with CDH (N+CDH). The pulmonary expression of NKCC-1, α-/β-ENaC and Na/K-ATPase was then analyzed using Western blot. We found that the protein levels of NKCC-1 on gestational days E18 and E21 were significantly lower among fetuses with N+CDH as well as N−CDH compared to controls. The expression of β-ENaC was also significantly down-regulated in both the groups on E18 and E21. The protein levels of α-ENaC and Na/K-ATPase were not found to be significantly decreased, but both showed a tendency towards down-regulation. The marked down-regulation of NKCC-1 in fetal hypoplastic lungs with CDH indicates a possibly decreased lung liquid production. This may be one of the mechanisms behind the disturbed pulmonary development in CDH. We also show that β-ENaC is down-regulated. Down-regulation of β-ENaC may result in abnormal lung liquid absorption, which could be one of the mechanisms behind the respiratory distress seen in CDH patients postpartum.

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Abbreviations

CDH:

Congenital diaphragmatic hernia

ECMO:

Extracorporeal membrane oxygenation

ENaC:

Epithelial sodium channel

FGF:

Fibroblast growth factor

NKCC-1:

Sodium-potassium-2-chloride co-transporter 1

RDS:

Respiratory distress syndrome

TO:

Tracheal occlusion

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Acknowledgments

This work was supported by grants from the HRH Crown Princess Lovisa and Theilmans Foundation, the Swedish Freemasons Childhood Foundation, the Swedish Heart-Lung Foundation and Karolinska Institutet. We thank Prof. S. Nielsen, Faculty of Health Sciences, University of Aarhus, Denmark for kindly providing the β-ENaC antibody and Y. Li for valuable technical assistance. The experiments comply with the current laws of Sweden.

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Authors and Affiliations

  1. Department of Woman and Child Health, Astrid Lindgren Children’s Hospital Research Laboratory Q2:09, Nordic Centre of Excellence for Research in Water Imbalance Related Disorders (WIRED), Karolinska Institutet, 171 76, Stockholm, Sweden

    Andreas Ringman, Marina Zelenina, Ann-Christine Eklöf & Anita Aperia

  2. Division of Pediatric Surgery, Department of Woman and Child Health, Karolinska Institutet, Stockholm, Sweden

    Andreas Ringman & Björn Frenckner

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  1. Andreas Ringman
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Ringman, A., Zelenina, M., Eklöf, AC. et al. NKCC-1 and ENaC are down-regulated in nitrofen-induced hypoplastic lungs with congenital diaphragmatic hernia. Pediatr Surg Int 24, 993–1000 (2008). https://doi.org/10.1007/s00383-008-2209-6

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