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Depletion of Caveolin‐1 in Lung Vasculature and Increase in Caveolin‐1 Positive Circulating Extracellular Vesicles as Early Biomarkers of Endothelial Injury
Author(s) -
Oliveira Suellen Darc Santos,
Castellon Maricela,
Machado Roberto F,
Elliott Michael H,
Bonini Marcelo G,
Minshall Richard D
Publication year - 2017
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.31.1_supplement.1015.25
Subject(s) - caveolin 1 , enos , lung , lipopolysaccharide , extracellular vesicle , medicine , western blot , inflammation , immunology , endothelial dysfunction , endocrinology , andrology , chemistry , nitric oxide , nitric oxide synthase , microvesicles , biochemistry , microrna , gene
Rationale Endothelial cell activation and dysfunction are hallmark features of Acute Lung Injury and Acute Respiratory Distress Syndrome (ALI/ARDS). Caveolin‐1 (Cav‐1) is highly expressed in pulmonary microvascular endothelial cells (PMVECs) and plays a key role in maintaining vascular homeostasis. The aim of this study was to determine whether the lung inflammatory response to endotoxin promotes PMVEC Cav‐1 depletion and appearance of Cav‐1 positive circulating extracellular vesicles (Cav‐1 + EVs) and whether this contributes to ALI. Methods ALI was induced in C57BL6 WT mice (male; 2.5–3 months old) by nebulized Escherichia coli lipopolysaccharide (LPS; 10 mg over 1 hr daily for 1 or 4 days). Lung pathology, Cav‐1 and eNOS expression, Cav‐1 ubiquitination, and Cav‐1 + EVs were quantified in total lung or platelet‐free plasma by immunoshistochemistry and western blot, respectively. Broncho‐alveolar lavage fluid (BALF) obtained from control and LPS‐treated mice was used to measure inflammatory cell, cytokine, and albumin levels. Results Alveolar accumulation of plasma albumin, immune cells, and pro‐inflammatory and regulatory cytokines (TNF‐α, IL‐1β, IL‐6, TGF‐β) was signficantly elevated in LPS‐treated WT mice. Intermittent airway LPS exposure for 96 hrs also increased Cav‐1 phosphorylation and ubiquitination (Ub) and decreased total lung Cav‐1 expression by 48 ± 4.16 % (n=3–4 mice; P <0.05) when compared to untreated WT mice. Depletion of EC Cav‐1 was accompanied by a decrease in eNOS dimerization indicative of endothelial dysfunction. In addition, circulating Cav‐1 + EVs increased by 38% after 24 hrs of LPS exposure (0.61 ± 0.05 to 0.98 ± 0.16 a.u.) in control and LPS‐treated mice, respectively (n= 8–10 mice; P <0.05) and decreased below basal level after 96 hrs indicating PMVEC Cav‐1 depletion may be due to both Cav‐1 ubiquitination and proteosomal degradation as well as release Cav‐1 + EVs into the circulation. PMVEC Cav‐1 depletion in response to endotoxin exposure was associated with endothelial dysfunction and vascular injury suggesting EC Cav‐1 depletion may play a significant role in the onset of ALI. Similar lung pathology was observed in EC‐specific Cav‐1 −/− mice. Conclusion Endothelial cell Cav‐1 depletion and Cav‐1 + EVs may be an early biomarker of pulmonary vascular injury that may predict pulmonary vascular inflammatory disease due to endothelial dysfunction. Support or Funding Information Financial support: CNPq Fellowship‐CsF/Brazil (SDO) and HL125356 and DOD W911NF‐15‐R‐002 (RDM, MGB).