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Lipopolysaccharide and Isoproterenol Alter Sodium Bicarbonate Cotransporter Phosphorylation in Pulmonary Microvascular Endothelial Cells
Author(s) -
Sayner Sarah L.
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.980.5
Subject(s) - phosphorylation , forskolin , rolipram , chemistry , lipopolysaccharide , medicine , bicarbonate , endothelial stem cell , endocrinology , stimulation , biochemistry , phosphodiesterase , biology , enzyme , in vitro
The hallmark of acute respiratory distress syndrome (ARDS) is damage to alveolar capillaries leading to increased endothelial permeability. Bacterial induced sepsis is a primary cause of pulmonary endothelial barrier dysfunction. Our recent studies reveal that the lipopolysaccharide (LPS)‐induced increase in pulmonary endothelial permeability is dependent upon bicarbonate stimulation of soluble adenylyl cyclase isoform 10 (AC10 a.k.a. sAC). Further, bicarbonate stimulation of AC10 is dependent upon sodium, suggesting a role for Na + ‐bicarbonate cotransporters (NBCs). Further, it has been reported that forskolin stimulation of transmembrane ACs and generation of subplasma membrane cAMP inactivates NBCn2 via PKA, potentially reducing the availability of bicarbonate for stimulation of AC10. Thus, we investigated the expression of NBCs in the lung and pulmonary endothelium and the phosphorylation status of NBCn2 in response to barrier disruptive (LPS) or barrier protective (isoproterenol) agents. To examine the phosphorylation status of NBCn2, endothelial cells were treated with either LPS or isoproterenol and rolipram, the phosphodiesterase 4 inhibitor. Immunoprecipitation was performed on whole cell lysate with the NBCn2 or control IgG antibodies and western blot analysis with PKA Phospho‐Ser/Thr antibody. Our results revealed the differential expression of NBCs in the pulmonary circulation. Further, while LPS led to a decrease in the phosphorylation of NBCn2 at a PKA specific site, isoproterenol and rolipram increased PKA‐phosphorylation of NBCn2. Thus, while transmembrane AC activity increases NBCn2 phosphorylation, LPS decreases NBCn2 phosphorylation and could act as a mechanism to regulate the bicarbonate source necessary to stimulate AC10. Support or Funding Information This work is supported by the Parker B. Francis Fellowship Foundation Award (SLS) and HL121513 (SLS)