Role In Endothelial Barrier Function Research Articles

Functional roles of CD26/DPP4 in lipopolysaccharide-induced lung injury.

Acute respiratory distress syndrome (ARDS) is characterized by dysregulated inflammation and increased permeability of lung microvascular cells. CD26/dipeptidyl peptidase-4 (DPP4) is a type II membrane protein that is expressed in several cell types and mediates multiple pleiotropic effects. We previously reported that DPP4 inhibition by sitagliptin attenuates lipopolysaccharide (LPS)-induced lung injury in mice. The current study characterized the functional role of CD26/DPP4 expression in LPS-induced lung injury in mice, isolated alveolar macrophages, and cultured lung endothelial cells. In LPS-induced lung injury, inflammatory responses [bronchoalveolar lavage fluid (BALF) neutrophil numbers and several proinflammatory cytokine levels] were attenuated in Dpp4 knockout (Dpp4 KO) mice. However, multiple assays of alveolar capillary permeability were similar between the Dpp4 KO and wild-type mice. TNF-α and IL-6 production was suppressed in alveolar macrophages isolated from Dpp4 KO mice. In contrast, in cultured mouse lung microvascular endothelial cells (MLMVECs), reduction in CD26/DPP4 expression by siRNA resulted in greater ICAM-1 and IL-6 expression after LPS stimulation. Moreover, the LPS-induced vascular monolayer permeability in vitro was higher in MLMVECs treated with Dpp4 siRNA, suggesting that CD26/DPP4 plays a protective role in endothelial barrier function. In summary, this study demonstrated that genetic deficiency of Dpp4 attenuates inflammatory responses but not permeability in LPS-induced lung injury in mice, potentially through differential functional roles of CD26/DPP4 expression in resident cellular components of the lung. CD26/DPP4 may be a potential therapeutic target for ARDS and warrants further exploration to precisely identify the multiple functional effects of CD26/DPP4 in ARDS pathophysiology.NEW & NOTEWORTHY We aimed to clarify the functional roles of CD26/DPP4 in ARDS pathophysiology using Dpp4-deficient mice and siRNA reduction techniques in cultured lung cells. Our results suggest that CD26/DPP4 expression plays a proinflammatory role in alveolar macrophages while also playing a protective role in the endothelial barrier. Dpp4 genetic deficiency attenuates inflammatory responses but not permeability in LPS-induced lung injury in mice, potentially through differential roles of CD26/DPP4 expression in the resident cellular components of the lung.

Regulation of Rho GTPases in the Vasculature by Cullin3-Based E3 Ligase Complexes.

Cullin3-based ubiquitin E3 ligases induce ubiquitination of substrates leading to their proteasomal or lysosomal degradation. BTB proteins serve as adaptors by binding to Cullin3 and recruiting substrate proteins, which enables specific recognition of a broad spectrum of targets. Hence, Cullin3 and its adaptors are involved in myriad cellular processes and organ functions. Cullin3-based ubiquitin E3 ligase complexes target small GTPases of the Rho subfamily, which are key regulators of cytoskeletal dynamics and cell adhesion. In this mini review, we discuss recent insights in Cullin3-mediated regulation of Rho GTPases and their impact on cellular function and disease. Intriguingly, upstream regulators of Rho GTPases are targeted by Cullin3 complexes as well. Thus, Rho GTPase signaling is regulated by Cullin3 on multiple levels. In addition, we address current knowledge of Cullin3 in regulating vascular function, focusing on its prominent role in endothelial barrier function, angiogenesis and the regulation of blood pressure.

The Role of Cytoplasmic Deacetylase, SIRT2, in Stabilizing Lung Endothelial Barrier Function

Lung microvascular endothelial functional and structural integrity is essential for maintenance of tissue‐fluid homeostasis and normal function of the respiratory system. Disruption of the pulmonary endothelial barrier leads to pulmonary edema, acute lung injury, and respiratory failure. Adherens junction (AJ) proteins, actin cytoskeleton, and microtubules are major components required for maintenance of barrier integrity. Post‐translational protein modifications of AJ constituents, such as phosphorylation, nitration, and nitrosylation, have been well known to regulate endothelial barrier function. More recently, lysine acetylation/deacetylation has emerged as an important and physiologically significant posttranslational protein modification in physiological and pathological conditions. However, its role in endothelial barrier function remains largely unknown. Sirtuins (SIRTs) are NAD+‐dependent deacetylases, and critical regulators of energy metabolism and oxidative stress response in different cell types. We studied the function of SIRT2, the only cytosolic sirtuin family member, in regulating pulmonary endothelial barrier function during inflammatory response. We showed that SIRT2 is the most abundant sirtuin in human lung endothelial cells. Importantly, SIRT2 protein level is significantly increased in lung endothelial cells upon challenging mice with a sublethal dose of lipopolysaccharides, a gram‐negative bacteria endotoxin which causes systemic inflammatory responses resulting in acute lung injury. Increase in SIRT2 protein level was also observed in human lung microvascular endothelial cells (HLMVEC) upon challenge with permeability‐increasing agents such as human α‐thrombin and TNF‐α, further emphasizing the potentially important role of SIRT2 in regulating adaptation of endothelial cells to inflammatory responses. To determine the role of SIRT2 activity in the maintenance of lung endothelial barrier function, we knocked down SIRT2 in HLMVECs and assessed changes in endothelial permeability upon challenge with α‐thrombin. We found that SIRT2 knock down significantly augmented α‐thrombin‐induced permeability increases as compared to control siRNA‐treated cells, suggesting that SIRT2 activity may play a critical role in regulating endothelial barrier integrity. Furthermore, treatment of endothelial cells with AGK2, a SIRT2‐selective inhibitor, significantly decreased assembly of endothelial junctions due to increased accumulation of VE‐cadherin in the cytoplasm. Our data suggest that SIRT2 plays a critical role in the maintenance of endothelial barrier function under basal conditions and upon inflammatory response.Support or Funding InformationXiaoyan Yang : T32HL007829‐23, AHA 17SDG33410608.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Terminal sialic acids are an important determinant of pulmonary endothelial barrier integrity.

The surface of vascular endothelium bears a glycocalyx comprised, in part, of a complex mixture of oligosaccharide chains attached to cell-surface proteins and membrane lipids. Importantly, understanding of the structure and function of the endothelial glycocalyx is poorly understood. Preliminary studies have demonstrated structural differences in the glycocalyx of pulmonary artery endothelial cells compared with pulmonary microvascular endothelial cells. Herein we begin to probe in more detail structural and functional attributes of endothelial cell-surface carbohydrates. In this study we focus on the expression and function of sialic acids in pulmonary endothelium. We observed that, although pulmonary microvascular endothelial cells express similar amounts of total sialic acids as pulmonary artery endothelial cells, the nature of the sialic acid linkages differs between the two cell types such that pulmonary artery endothelial cells express both α(2,3)- and α(2,6)-linked sialic acids on the surface (i.e., surficially), whereas microvascular endothelial cells principally express α(2,3)-linked sialic acids. To determine whether sialic acids play a role in endothelial barrier function, cells were treated with neuraminidases to hydrolyze sialic acid moieties. Disruption of cell-cell and cell-matrix adhesions was observed following neuraminidase treatment, suggesting that terminal sialic acids promote endothelial barrier integrity. When we measured transendothelial resistance, differential responses of pulmonary artery and microvascular endothelial cells to neuraminidase from Clostridium perfringens suggest that the molecular architecture of the sialic acid glycomes differs between these two cell types. Collectively our observations reveal critical structural and functional differences of terminally linked sialic acids on the pulmonary endothelium.

The pulmonary microvascular endothelial cell glycocalyx includes sialic acid important for endothelial barrier function

The endothelium forms a semi‐permeable barrier between constituents of the blood and underlying tissue. The endothelial glycocalyx consists of glycoproteins, glycolipids and proteoglycans which coat the cell surface. The carbohydrate network that contributes to the glycocalyx is very complex, and to date, the structure of these complex carbohydrates and their role in endothelial barrier function are unknown. We have begun to probe the molecular identities of endothelial cell surface carbohydrates in a systematic way. Initial studies examined the presence of terminal sialic acid residues. Neuraminidase is an enzyme that cleaves sialic acids from carbohydrate chains. Cultured rat pulmonary microvascular endothelial cells (PMVECs) were treated with neuraminidase from Clostridium perfringe (1 unit) for two hours. Neuraminidase treated cells exhibited loss of cell‐cell adhesion as noted by the formation of interendothelial cell gaps, and in a scratch‐wound assay neuraminidase treated cells were slower to reseal as compared to control. These observations suggest that the PMVEC glycocalyx contains terminal sialic acid residues that play a role in endothelial barrier function. Supported by HL60024.