In vivo blockade of KCa3.1 ion channel alleviates ER stress in type II alveolar epithelial cells and macrophages in pulmonary fibrosis

Abstract Background Idiopathic pulmonary fibrosis (IPF) is a fatal interstitial lung disease, characterized by progressive damage to the lung tissues. Apoptosis and endoplasmic reticulum stress (ER stress) in type II alveolar epithelial cells (AECs) and lung macrophages has been linked with the development of IPF. Therefore, apoptosis- and ER stress-targeted therapies have drawn attention as potential avenues for treatment of IPF. The calcium-activated potassium ion channel KCa3.1 has been proposed as a potential therapeutic target for fibrotic diseases including IPF. While KCa3.1 is expressed in AECs and macrophages, its influence on ER stress and apoptosis during the disease process is unclear. Methods We utilized a novel sheep model of pulmonary fibrosis to demonstrate that apoptosis and ER stress occurs in type II AECs and macrophages in sheep with bleomycin-induced lung fibrosis. Apoptosis in type II AEC and macrophages was identified using the TUNEL method of tagging fragmented nuclear DNA, while ER stress was characterized by increased expression of GRP-78 ER chaperone proteins. Results We demonstrated that apoptosis and ER stress in type II AECs and macrophages increased significantly 2 weeks after the final bleomycin infusion and remained high for up to 7 weeks post-bleomycin injury. Senicapoc treatment significantly reduced the rates of ER stress in type II AECs and macrophages that were resident in bleomycin-infused lung segments. There were also significant reductions in the rates of apoptosis of type II AECs and macrophages in the lung segments of senicapoc-treated sheep. Conclusion In-vivo blockade of the KCa3.1 ion channel alleviates the ER stress and apoptosis in type II AECs and macrophages, and this effect potentially contributes to the antifibrotic effects of senicapoc.

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Increased Levels of ER Stress and Apoptosis in a Sheep Model for Pulmonary Fibrosis Are Alleviated by In Vivo Blockade of the KCa3.1 Ion Channel

Canadian Respiratory Journal ◽

10.1155/2021/6683195 ◽

2021 ◽

Vol 2021 ◽

pp. 1-11

Author(s):

Udari E. Perera ◽

Louise Organ ◽

Sasika N. V. Dewage ◽

Habtamu B. Derseh ◽

Andrew Stent ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Ion Channel ◽

Er Stress ◽

Nuclear Dna ◽

Disease Process ◽

Alveolar Epithelial Cells ◽

Type Ii ◽

Sheep Model ◽

Alveolar Epithelial

Idiopathic pulmonary fibrosis (IPF) is a fatal interstitial lung disease, characterized by progressive damage to the lung tissues. Apoptosis and endoplasmic reticulum stress (ER stress) in type II alveolar epithelial cells (AECs) and lung macrophages have been linked with the development of IPF. Therefore, apoptosis- and ER stress-targeted therapies have drawn attention as potential avenues for treatment of IPF. The calcium-activated potassium ion channel KCa3.1 has been proposed as a potential therapeutic target for fibrotic diseases including IPF. While KCa3.1 is expressed in AECs and macrophages, its influence on ER stress and apoptosis during the disease process is unclear. We utilized a novel sheep model of pulmonary fibrosis to demonstrate that apoptosis and ER stress occur in type II AECs and macrophages in sheep with bleomycin-induced lung fibrosis. Apoptosis in type II AEC and macrophages was identified using the TUNEL method of tagging fragmented nuclear DNA, while ER stress was characterized by increased expression of GRP-78 ER chaperone proteins. We demonstrated that apoptosis and ER stress in type II AECs and macrophages increased significantly 2 weeks after the final bleomycin infusion and remained high for up to 7 weeks post-bleomycin injury. Senicapoc treatment significantly reduced the rates of ER stress in type II AECs and macrophages that were resident in bleomycin-infused lung segments. There were also significant reductions in the rates of apoptosis of type II AECs and macrophages in the lung segments of senicapoc-treated sheep. In vivo blockade of the KCa3.1 ion channel alleviates the ER stress and apoptosis in type II AECs and macrophages, and this effect potentially contributes to the anti-fibrotic effects of senicapoc.

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Faculty Opinions recommendation of Targeted injury of type II alveolar epithelial cells induces pulmonary fibrosis.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1166509.627502 ◽

2009 ◽

Author(s):

Gregory Cosgrove

Keyword(s):

Epithelial Cells ◽

Pulmonary Fibrosis ◽

Alveolar Epithelial Cells ◽

Type Ii ◽

Alveolar Epithelial

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Faculty Opinions recommendation of Targeted injury of type II alveolar epithelial cells induces pulmonary fibrosis.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1166509.2474068 ◽

2010 ◽

Author(s):

Moisés Selman ◽

Carolina Garcia de Alba

Keyword(s):

Epithelial Cells ◽

Pulmonary Fibrosis ◽

Alveolar Epithelial Cells ◽

Type Ii ◽

Alveolar Epithelial

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Dome formation in primary cultured monolayers of alveolar epithelial cells

AJP Cell Physiology ◽

10.1152/ajpcell.1982.243.1.c96 ◽

1982 ◽

Vol 243 (1) ◽

pp. C96-C100 ◽

Cited By ~ 86

Author(s):

B. E. Goodman ◽

E. D. Crandall

Keyword(s):

Epithelial Cells ◽

Life Span ◽

Fluid Balance ◽

Alveolar Epithelial Cells ◽

Type Ii ◽

Transport Function ◽

Dome Formation ◽

Alveolar Epithelial ◽

Free Air

We have observed the formation of domes by type II alveolar epithelial cells harvested from rat lungs. The cells were harvested using elastase and grew to confluence in 3-4 days after plating on plastic. Numerous domes were observed in the monolayers 4-18 days after plating, with peak dome density occurring at days 6-9. When trypsin was used instead of elastase as the harvesting enzyme, many fewer domes were formed by the monolayers, with peak dome density observed at day 5 and no domes seen after 8 days. The life span of an individual dome was about 3-4 h. The presence of domes indicates an intact active transport function of the cells in the monolayer, which may represent an important mechanism for the maintenance of fluid-free air spaces and normal alveolar fluid balance in mammalian lungs in vivo.

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miR-34 miRNAs Regulate Cellular Senescence in Type II Alveolar Epithelial Cells of Patients with Idiopathic Pulmonary Fibrosis

PLoS ONE ◽

10.1371/journal.pone.0158367 ◽

2016 ◽

Vol 11 (6) ◽

pp. e0158367 ◽

Cited By ~ 56

Author(s):

Supparerk Disayabutr ◽

Eun Kyung Kim ◽

Seung-Ick Cha ◽

Gary Green ◽

Ram P. Naikawadi ◽

...

Keyword(s):

Epithelial Cells ◽

Idiopathic Pulmonary Fibrosis ◽

Pulmonary Fibrosis ◽

Cellular Senescence ◽

Alveolar Epithelial Cells ◽

Type Ii ◽

Alveolar Epithelial

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Role of diminished epithelial GM-CSF in the pathogenesis of bleomycin-induced pulmonary fibrosis

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.2000.279.3.l487 ◽

2000 ◽

Vol 279 (3) ◽

pp. L487-L495 ◽

Cited By ~ 37

Author(s):

Paul J. Christensen ◽

Marc B. Bailie ◽

Richard E. Goodman ◽

Aidan D. O'Brien ◽

Galen B. Toews ◽

...

Keyword(s):

Epithelial Cells ◽

Pulmonary Fibrosis ◽

Animal Studies ◽

Protective Role ◽

Alveolar Epithelial Cells ◽

Type Ii ◽

Alveolar Epithelial ◽

Gm Csf ◽

High Level

Evidence derived from human and animal studies strongly supports the notion that dysfunctional alveolar epithelial cells (AECs) play a central role in determining the progression of inflammatory injury to pulmonary fibrosis. We formed the hypothesis that impaired production of the regulatory cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) by injured AECs plays a role in the development of pulmonary fibrosis. To test this hypothesis, we used the well-characterized model of bleomycin-induced pulmonary fibrosis in rats. GM-CSF mRNA is expressed at a constant high level in the lungs of untreated or saline-challenged animals. In contrast, there is a consistent reduction in expression of GM-CSF mRNA in the lung during the first week after bleomycin injury. Bleomycin-treated rats given neutralizing rabbit anti-rat GM-CSF IgG develop increased fibrosis. Type II AECs isolated from rats after bleomycin injury demonstrate diminished expression of GM-CSF mRNA immediately after isolation and in response to stimulation in vitro with endotoxin compared with that in normal type II cells. These data demonstrate a defect in the ability of type II epithelial cells from bleomycin-treated rats to express GM-CSF mRNA and a protective role for GM-CSF in the pathogenesis of bleomycin-induced pulmonary fibrosis.

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Deletion of Pou2af1 in Type II Alveolar Epithelial Cells Augments Bleomycin-Induced Pulmonary Fibrosis

10.1164/ajrccm-conference.2019.199.1_meetingabstracts.a5407 ◽

2019 ◽

Author(s):

Q. Li ◽

N. Omote ◽

Z. Chen ◽

G. DeIuliis ◽

Z. Zhu ◽

...

Keyword(s):

Epithelial Cells ◽

Pulmonary Fibrosis ◽

Alveolar Epithelial Cells ◽

Type Ii ◽

Alveolar Epithelial

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ER Stress is Involved in Epithelial-To-Mesenchymal Transition of Alveolar Epithelial Cells Exposed to a Hypoxic Microenvironment

International Journal of Molecular Sciences ◽

10.3390/ijms20061299 ◽

2019 ◽

Vol 20 (6) ◽

pp. 1299 ◽

Cited By ~ 4

Author(s):

Eva Delbrel ◽

Yurdagül Uzunhan ◽

Abdoulaye Soumare ◽

Thomas Gille ◽

Dominique Marchant ◽

...

Keyword(s):

Epithelial Cells ◽

Er Stress ◽

Epithelial To Mesenchymal Transition ◽

Alveolar Epithelial Cells ◽

Mesenchymal Transition ◽

Alveolar Epithelial ◽

Rat Lungs ◽

Fibrotic Process

Background: Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive and fatal interstitial lung disease of unknown origin. Alveolar epithelial cells (AECs) play an important role in the fibrotic process as they undergo sustained endoplasmic reticulum (ER) stress, and may acquire a mesenchymal phenotype through epithelial-to-mesenchymal transition (EMT), two phenomena that could be induced by localized alveolar hypoxia. Here we investigated the potential links between hypoxia, ER stress and EMT in AECs. Methods: ER stress and EMT markers were assessed by immunohistochemistry, western blot and qPCR analysis, both in vivo in rat lungs exposed to normoxia or hypoxia (equivalent to 8% O2) for 48 h, and in vitro in primary rat AECs exposed to normoxia or hypoxia (1.5% O2) for 2–6 days. Results: Hypoxia induced expression of mesenchymal markers, pro-EMT transcription factors, and the activation of ER stress markers both in vivo in rat lungs, and in vitro in AECs. In vitro, pharmacological inhibition of ER stress by 4-PBA limited hypoxia-induced EMT. Calcium chelation or hypoxia-inducible factor (HIF) inhibition also prevented EMT induction under hypoxic condition. Conclusions: Hypoxia and intracellular calcium are both involved in EMT induction of AECs, mainly through the activation of ER stress and HIF signaling pathways.

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