Modulation of neutrophil apoptosis by murine pulmonary microvascular endothelial cell inducible nitric oxide synthase

Wang, Lefeng; Mehta, Sanjay; Gillis, Chris; Law, Cedrin; Taneja, Ravi

doi:10.1016/j.bbrc.2010.09.029

Cited by 7 publications

(5 citation statements)

References 45 publications

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“…We have previously reported that iNOS plays an important role in multiple features of septic ALI in the murine CLP-sepsis model, including pulmonary microvascular endothelial barrier dysfunction and the resulting albumin hyper-permeability, pulmonary oxidant stress, pulmonary microvascular PMN sequestration and trans-PMVEC PMN migration [ 17 , 18 , 35 , 47 ]. Thus, we assessed the contribution of iNOS to septic PMVEC apoptosis in CLP-sepsis vs. sham mice.…”

Section: Resultsmentioning

confidence: 99%

Role of pulmonary microvascular endothelial cell apoptosis in murine sepsis-induced lung injury in vivo

2015

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BackgroundSepsis remains a common and serious condition with significant morbidity and mortality due to multiple organ dysfunction, especially acute lung injury (ALI) and acute respiratory distress syndrome (ARDS). Sepsis-induced ALI is characterized by injury and dysfunction of the pulmonary microvasculature and pulmonary microvascular endothelial cells (PMVEC), resulting in enhanced pulmonary microvascular sequestration and pulmonary infiltration of polymorphonuclear leukocytes (PMN) as well as disruption of the normal alveolo-capillary permeability barrier with leak of albumin-rich edema fluid into pulmonary interstitium and alveoli. The role of PMVEC death and specifically apoptosis in septic pulmonary microvascular dysfunction in vivo has not been established.MethodsIn a murine cecal ligation/perforation (CLP) model of sepsis, we quantified and correlated time-dependent changes in pulmonary microvascular Evans blue (EB)-labeled albumin permeability with (1) PMVEC death (propidium iodide [PI]-staining) by both fluorescent intravital videomicroscopy (IVVM) and histology, and (2) PMVEC apoptosis using histologic fluorescent microscopic assessment of a panel of 3 markers: cell surface phosphatidylserine (detected by Annexin V binding), caspase activation (detected by FLIVO labeling), and DNA fragmentation (TUNEL labeling).ResultsCompared to sham mice, CLP-sepsis resulted in pulmonary microvascular barrier dysfunction, quantified by increased EB-albumin leak, and PMVEC death (PI+ staining) as early as 2 h and more marked by 4 h after CLP. Septic PMVEC also exhibited increased presence of all 3 markers of apoptosis (Annexin V+, FLIVO+, TUNEL+) as early as 30 mins – 1 h after CLP-sepsis, which all similarly increased markedly until 4 h. The time-dependent changes in septic pulmonary microvascular albumin-permeability barrier dysfunction were highly correlated with PMVEC death (PI+; r = 0.976, p < 0.01) and PMVEC apoptosis (FLIVO+; r = 0.991, p < 0.01). Treatment with the pan-caspase inhibitor Q-VD prior to CLP reduced PMVEC death/apoptosis and attenuated septic pulmonary microvascular dysfunction, including both albumin-permeability barrier dysfunction and pulmonary microvascular PMN sequestration (p < 0.05). Septic PMVEC apoptosis and pulmonary microvascular dysfunction were also abrogated following CLP-sepsis in mice deficient in iNOS (Nos2−/−) or NADPH oxidase (p47phox−/− or gp91phox−/−) and in wild-type mice treated with the NADPH oxidase inhibitor, apocynin.ConclusionsSeptic murine pulmonary microvascular dysfunction in vivo is due to PMVEC death, which is mediated through caspase-dependent apoptosis and iNOS/NADPH-oxidase dependent signaling.

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Section: Resultsmentioning

confidence: 99%

Role of pulmonary microvascular endothelial cell apoptosis in murine sepsis-induced lung injury in vivo

2015

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“…We have previously demonstrated that iNOS plays an important role in multiple features of septic lung injury in the murine CLP/sepsis model, including pulmonary microvascular endothelial barrier dysfunction and the resulting hyper-permeability [9] , [10] , [30] , [36] . Thus, we next examined whether iNOS was required for septic pulmonary cell death in CLP vs. sham-treated mice.…”

Section: Resultsmentioning

confidence: 99%

Pulmonary Microvascular Albumin Leak Is Associated with Endothelial Cell Death in Murine Sepsis-Induced Lung Injury In Vivo

et al. 2014

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Sepsis is a systemic inflammatory response that targets multiple components of the cardiovascular system including the microvasculature. Microvascular endothelial cells (MVEC) are central to normal microvascular function, including maintenance of the microvascular permeability barrier. Microvascular/MVEC dysfunction during sepsis is associated with barrier dysfunction, resulting in the leak of protein-rich edema fluid into organs, especially the lung. The specific role of MVEC apoptosis in septic microvascular/MVEC dysfunction in vivo remains to be determined. To examine pulmonary MVEC death in vivo under septic conditions, we used a murine cecal ligation/perforation (CLP) model of sepsis and identified non-viable pulmonary cells with propidium iodide (PI) by intravital videomicroscopy (IVVM), and confirmed this by histology. Septic pulmonary microvascular Evans blue (EB)-labeled albumin leak was associated with an increased number of PI-positive cells, which were confirmed to be predominantly MVEC based on specific labeling with three markers, anti-CD31 (PECAM), anti-CD34, and lectin binding. Furthermore, this septic death of pulmonary MVEC was markedly attenuated by cyclophosphamide-mediated depletion of neutrophils (PMN) or use of an anti-CD18 antibody developed for immunohistochemistry but shown to block CD18-dependent signaling. Additionally, septic pulmonary MVEC death was iNOS-dependent as mice lacking iNOS had markedly fewer PI-positive MVEC. Septic PI-positive pulmonary cell death was confirmed to be due to apoptosis by three independent markers: caspase activation by FLIVO, translocation of phosphatidylserine to the cell surface by Annexin V binding, and DNA fragmentation by TUNEL. Collectively, these findings indicate that septic pulmonary MVEC death, putatively apoptosis, is a result of leukocyte activation and iNOS-dependent signaling, and in turn, may contribute to pulmonary microvascular barrier dysfunction and albumin hyper-permeability during sepsis.

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“…Pulmonary MVEC isolation. Pulmonary MVEC were isolated from the lungs of WT and Timp3 Ϫ/Ϫ mice, as previously described (27,28,82). In brief, lungs were isolated, finely minced, and then digested with collagenase before incubation with magnetic microbeads (Dynal Biotech, Lake Success, NY) coupled to anti-CD31 (platelet endothelial cell adhesion molecule, PECAM) antibodies (BD Pharmingen, Franklin Lakes, NJ).…”

Section: Methodsmentioning

confidence: 99%

Tissue inhibitor of metalloproteinases 3-dependent microvascular endothelial cell barrier function is disrupted under septic conditions

Arpino

Mehta

Wang

et al. 2016

American Journal of Physiology-Heart and Circulatory Physiology

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Sepsis is associated with dysfunction of microvascular endothelial cells (MVEC) leading to tissue edema and multiple organ dysfunction. Metalloproteinases can regulate MVEC function through processing of cell surface proteins, and tissue inhibitor of metalloproteinases 3 (TIMP3) regulates metalloproteinase activity in the lung following injury. We hypothesize that TIMP3 promotes normal pulmonary MVEC barrier function through inhibition of metalloproteinase activity. Naive Timp3(-/-) mice had significantly higher basal pulmonary microvascular Evans blue (EB) dye-labeled albumin leak vs. wild-type (WT) mice. Additionally, cecal-ligation/perforation (CLP)-induced sepsis significantly increased pulmonary microvascular EB-labeled albumin leak in WT but not Timp3(-/-) mice. Similarly, PBS-treated isolated MVEC monolayers from Timp3(-/-) mice displayed permeability barrier dysfunction vs. WT MVEC, evidenced by lower transendothelial electrical resistance and greater trans-MVEC flux of fluorescein-dextran and EB-albumin. Cytomix (equimolar interferon γ, tumor necrosis factor α, and interleukin 1β) treatment of WT MVEC induced significant barrier dysfunction (by all three methods), and was associated with a time-dependent decrease in TIMP3 mRNA and protein levels. Additionally, basal Timp3(-/-) MVEC barrier dysfunction was associated with disrupted MVEC surface VE-cadherin localization, and both barrier dysfunction and VE-cadherin localization were rescued by treatment with GM6001, a synthetic metalloproteinase inhibitor. TIMP3 promotes normal MVEC barrier function, at least partially, through inhibition of metalloproteinase-dependent disruption of adherens junctions, and septic downregulation of TIMP3 may contribute to septic MVEC barrier dysfunction.

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Modulation of neutrophil apoptosis by murine pulmonary microvascular endothelial cell inducible nitric oxide synthase

Cited by 7 publications

References 45 publications

Role of pulmonary microvascular endothelial cell apoptosis in murine sepsis-induced lung injury in vivo

Role of pulmonary microvascular endothelial cell apoptosis in murine sepsis-induced lung injury in vivo

Pulmonary Microvascular Albumin Leak Is Associated with Endothelial Cell Death in Murine Sepsis-Induced Lung Injury In Vivo

Tissue inhibitor of metalloproteinases 3-dependent microvascular endothelial cell barrier function is disrupted under septic conditions

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