Targeting NOX4 alleviates sepsis-induced acute lung injury via attenuation of redox-sensitive activation of CaMKII/ERK1/2/MLCK and endothelial cell barrier dysfunction

Abstract

Increased pulmonary vascular permeability due to endothelial cell (EC) barrier dysfunction is a major pathological feature of acute respiratory distress syndrome/acute lung injury (ARDS/ALI), which is a devastating critical illness with high incidence and excessive mortality. Activation of NADPH oxidase (NOX) induces EC dysfunction via production of reactive oxygen species (ROS). However, the role(s) of NOX isoform(s), and their downstream signaling events, in the development of ARDS/ALI have remained unclear. Cecal Ligation Puncture (CLP) was used to induce preclinical septic ALI in wild-type mice and mice deficient in NOX2 or p47phox, or mice transfected of control siRNA, NOX1 or NOX4 siRNA in vivo. The survival rate of the CLP group at 24 h (26.6%, control siRNA treated) was substantially improved by NOX4 knockdown (52.9%). Mice lacking NOX2 or p47phox, however, had worse outcomes after CLP (survival rates at 0% and 8.3% respectively), whereas NOX1-silenced mice had similar survival rate (30%). NOX4 knockdown attenuated lung ROS production in septic mice, whereas NOX1 knockdown, NOX2 knockout, or p47phox knockout in mice had no effects. In addition, NOX4 knockdown attenuated redox-sensitive activation of the CaMKII/ERK1/2/MLCK pathway, and restored expression of EC tight junction proteins ZO-1 and Occludin to maintain EC barrier integrity. Correspondingly, NOX4 knockdown in cultured human lung microvascular ECs also reduced LPS-induced ROS production, CaMKII/ERK1/2/MLCK activation and EC barrier dysfunction. Scavenging superoxide in vitro and in vivo with TEMPO, or inhibiting CaMKII activation with KN93, had similar effects as NOX4 knockdown in preserving EC barrier dysfunction. In summary, we have identified a novel, selective and causal role of NOX4 (versus other NOX isoforms) in inducing lung EC barrier dysfunction and injury/mortality in a preclinical CLP-induced septic model, which involves redox-sensitive activation of CaMKII/ERK1/2/MLCK pathway. Targeting NOX4 may therefore prove to an innovative therapeutic option that is markedly effective in treating ALI/ARDS.

Inorganic polyphosphate modulates leukocyte accumulation and vascular endothelial cell permeability and ameliorates cecal ligation and puncture-induced lethality

“…Our previous report showed that polyP 150 protects against LPS-induced death and organ damage by modulating the recruitment of macrophages to tissues such as lung and liver (Terashima-Hasegawa et al, 2019). It has been suggested that activated inflammatory cells accumulate in the lungs under septic conditions and release cytokines and reactive oxygen species, disrupting pulmonary vascular endothelial and epithelial cells and increasing pulmonary vascular permeability (Gill et al, 2015;Jiang et al, 2020;Marcus et al, 1997). Excessive leukocyte accumulation may destabilize alveolar barrier function and increase vascular permeability, resulting in further disruption of alveolar barrier function.…”

“…Increased vascular permeability and disruption of its barrier function are largely responsible for the pathogenesis of CLP-induced septic shock ( Jiang et al, 2020 ;Joff re et al, 2020 ). Therefore, the eff ects of polyP 150 on the increased vascular permeability in the CLP model, which has been known to occur in this animal model, was examined.…”

“…In general, the increased vascular permeability in sepsis is thought to be closely related to the fact that overproduced inflammatory mediators reduce the function of endothelial cell adhesion (Joffre et al, 2020). Meanwhile, it has also been shown that adhesion and infiltration of neutrophils to the pulmonary vascular endothelium play an important role in vascular permeability in the acute lung injury secondary to sepsis (Gill et al, 2015;Jiang et al, 2020;Marcus et al, 1997). Therefore, we investigated monocyte-endothelial cell interactions in vitro using monocytic cells that are recruited by a mechanism similar to neutrophil adsorption to the vascular endothelium (Ley et al, 2007;Nourshargh and Alon, 2014).…”

Nanotherapies for sepsis by regulating inflammatory signals and reactive oxygen and nitrogen species: New insight for treating COVID-19

“…The pathogenesis of sepsis has not been well investigated, but uncontrollable inflammation and excessive RONS are thought to be two key factors that promote the progression of sepsis [ 24 ]. Therefore, an in-depth understanding of the mechanisms involved in immune response-induced inflammation and inflammation-induced RONS production is crucial for exploring sepsis pathogenesis and effective means of therapy [ 25 ]. In sepsis patients, the activated innate immune system is mediated by PRRs expressed on immune cells (e.g., macrophages and neutrophils) and non-immune cells (e.g., vascular endothelial cells) [ 9 , 26 ].…”

Nanomedicine-Based Therapeutics to Combat Acute Lung Injury

“…Restoring the integrity of the endothelial–epithelial barrier is critical in ALI therapy. 83 , 84 As the primary injured structure, the pulmonary epithelium is subject to dissociation of intercellular junctions 85 and cell death, 31 but is more resistant to injury than the endothelium. 86 The endothelial junctions’ breakdown or the endothelial cells’ death will increase the lung’s vascular permeability, thus resulting in excessive fluid and protein leakage into the alveoli.…”