Neurohumoral inflammatory mediators promote calcium entry through store operated calcium entry channels, and the resulting rise in cytosolic calcium disrupts the endothelial cell barrier and causes pulmonary edema. Canonical transient receptor potential proteins 1 and 4 contribute subunits that form the endothelial cell store operated calcium entry channel (TRPC1/4). Orai1 interacts with the this channel, and controls TRPC1/4 ion selectivity, as determined by whole cell electrophysiology. Our work has recently shown that in the absence of Orai1, more sodium and less calcium permeates through TRPC1/4. Because calcium is a principal second messenger that activates intracellular enzymes and regulatory proteins, considerable work has focused on the determinants of calcium influx. However, these new results suggest a potentially important role for sodium influx through TRPC1/4. We therefore measured the cytosolic calcium and sodium responses following activation of store operated calcium entry channels. Similar to our patch clamp results, we found that thapsigargin increases cytosolic sodium. Orai1 silencing substantially increases this cytosolic sodium response, but has a relatively small impact on the global cytosolic calcium response. Orai1 silencing prevented thapsigargin-induced gap formation. We interpreted these data to mean that elevated cytosolic sodium is a critical determinant of cellular movement. This idea was tested further in preliminary studies using migration and angiogenesis assays, where we observed that increased cytosolic sodium impairs cellular motility and angiogenesis. Hence, we now test the HYPOTHESIS that Orai1 interacts with TRPC1/4 channels and modulates the channel's calcium and sodium selectivity important for control of endothelial cell barrier function, migration and angiogenesis.
Specific aims test the related hypotheses that: (1) Orai1 interaction with the TRPC1/4 channel modulates the magnitude and spatial spread ofthe cytosolic sodium and calcium signals; and (2) sodium influx through the Orai1-TRPC1/4 channel strengthens the endothelial cell barrier and reduces cell migration and angiogenesis.

Public Health Relevance

Pulmonary endothelial cell permeability increases during inflammation, and is an important cause of patient morbidity and mortality. Here, we identify an endothelial cell calcium channel that contributes to permeability, and may therefore represent a novel anti-inflammatory target.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37HL060024-21
Application #
9538765
Study Section
Special Emphasis Panel (NSS)
Program Officer
Xiao, Lei
Project Start
1998-04-10
Project End
2020-06-30
Budget Start
2018-07-01
Budget End
2019-06-30
Support Year
21
Fiscal Year
2018
Total Cost
Indirect Cost
Name
University of South Alabama
Department
Type
DUNS #
172750234
City
Mobile
State
AL
Country
United States
Zip Code
36688
Lin, Mike T; Balczon, Ron; Pittet, Jean-Francois et al. (2018) Nosocomial Pneumonia Elicits an Endothelial Proteinopathy: Evidence for a Source of Neurotoxic Amyloids in Critically Ill Patients. Am J Respir Crit Care Med :
Lee, Ji Young; McMurtry, Sarah A; Stevens, Troy (2017) Single cell cloning generates lung endothelial colonies with conserved growth, angiogenic, and bioenergetic characteristics. Pulm Circ 7:777-792
Balczon, Ron; Morrow, K Adam; Zhou, Chun et al. (2017) Pseudomonas aeruginosa infection liberates transmissible, cytotoxic prion amyloids. FASEB J 31:2785-2796
Morrow, K Adam; Frank, Dara W; Balczon, Ron et al. (2017) The Pseudomonas aeruginosa Exoenzyme Y: A Promiscuous Nucleotidyl Cyclase Edema Factor and Virulence Determinant. Handb Exp Pharmacol 238:67-85
Francis, Michael; Xu, Ningyong; Zhou, Chun et al. (2016) Transient Receptor Potential Channel 4 Encodes a Vascular Permeability Defect and High-Frequency Ca(2+) Transients in Severe Pulmonary Arterial Hypertension. Am J Pathol 186:1701-9
Morrow, K Adam; Ochoa, Cristhiaan D; Balczon, Ron et al. (2016) Pseudomonas aeruginosa exoenzymes U and Y induce a transmissible endothelial proteinopathy. Am J Physiol Lung Cell Mol Physiol 310:L337-53
Zhou, Chun; Townsley, Mary I; Alexeyev, Mikhail et al. (2016) Endothelial hyperpermeability in severe pulmonary arterial hypertension: role of store-operated calcium entry. Am J Physiol Lung Cell Mol Physiol 311:L560-9
Xu, Ningyong; Cioffi, Donna L; Alexeyev, Mikhail et al. (2015) Sodium entry through endothelial store-operated calcium entry channels: regulation by Orai1. Am J Physiol Cell Physiol 308:C277-88
Gopal, Sandeep; Søgaard, Pernille; Multhaupt, Hinke A B et al. (2015) Transmembrane proteoglycans control stretch-activated channels to set cytosolic calcium levels. J Cell Biol 210:1199-211
Morrow, K A; Seifert, R; Kaever, V et al. (2015) Heterogeneity of pulmonary endothelial cyclic nucleotide response to Pseudomonas aeruginosa ExoY infection. Am J Physiol Lung Cell Mol Physiol 309:L1199-207

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