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Endoplasmic reticulum stress response is activated in pulmonary hypoplasia secondary to congenital diaphragmatic hernia, but is decreased by administration of amniotic fluid stem cells

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Abstract

Purpose

Pulmonary hypoplasia secondary to congenital diaphragmatic hernia (CDH) is characterized by impaired epithelial homeostasis. Recently, amniotic fluid stem cells (AFSCs) have been shown to promote growth in hypoplastic lungs of rat fetuses with CDH. Herein, we investigated whether CDH hypoplastic lungs mount an endoplasmic reticulum (ER) stress response and whether AFSCs could re-establish pulmonary epithelial homeostasis.

Methods

Primary epithelial cells were isolated from fetal rat lungs at E14.5 from control and nitrofen-exposed dams at E9.5. Nitrofen-exposed epithelial cells were grown in medium alone or co-cultured with AFSCs. Epithelial cell cultures were compared for apoptosis (TUNEL), cytotoxicity (LIVE/DEAD assay), proliferation (5′EdU), and ER stress (CHOP, Bcl-2) using one-way ANOVA (Dunn’s post-test).

Results

Compared to control, nitrofen-exposed epithelial cells had increased cytotoxicity and apoptosis, reduced proliferation, and activated ER stress. AFSCs restored apoptosis, proliferation, and ER stress back to control levels, and significantly reduced cytotoxicity.

Conclusions

This study shows for the first time that ER stress-induced apoptosis is activated in the pulmonary epithelium of hypoplastic lungs from fetuses with CDH. AFSC treatment restores epithelial cellular homeostasis by attenuating the ER stress response and apoptosis, by increasing proliferation and migration ability, and by reducing cytotoxicity.

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Acknowledgements

The authors are indebted to Prof. Paolo De Coppi for providing the amniotic fluid stem cells used in this study.

Funding

This work was supported by SickKids start-up funds and by the Canadian Institutes of Health Research (CIHR)—SickKids Foundation New Investigator Research Grant (NI18—1270R).

Author information

Authors and Affiliations

  1. Developmental and Stem Cell Biology Program, PGCRL, The Hospital for Sick Children, Toronto, ON, Canada

    Areti Tzanetakis, Lina Antounians, Alyssa Belfiore, Qi Ma, Mark Stasiewicz, Ornella Pellerito & Augusto Zani

  2. Division of General and Thoracic Surgery, Department of Surgery, The Hospital for Sick Children, 555 University Avenue, Toronto, ON, M5G 1X8, Canada

    Areti Tzanetakis, Lina Antounians, Alyssa Belfiore, Qi Ma, Mark Stasiewicz, Ornella Pellerito & Augusto Zani

Authors
  1. Areti Tzanetakis
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  2. Lina Antounians
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Correspondence to Augusto Zani.

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The authors declare that they have no conflict of interest.

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Ethical approval for experiments conducted AUP #39168.

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Tzanetakis, A., Antounians, L., Belfiore, A. et al. Endoplasmic reticulum stress response is activated in pulmonary hypoplasia secondary to congenital diaphragmatic hernia, but is decreased by administration of amniotic fluid stem cells. Pediatr Surg Int 35, 63–69 (2019). https://doi.org/10.1007/s00383-018-4376-4

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