Slit2–Robo4 Pathway Modulates Lipopolysaccharide-Induced Endothelial Inflammation and Its Expression Is Dysregulated during Endotoxemia

Abstract

The secretory protein Slit2 and its receptors Robo1 and Robo4 are considered to regulate mobility and permeability of endothelial cells and other cell types. However, the roles of Slit2 and its two receptors in endothelial inflammatory responses remain to be clarified. Here we show that, in primary Human Umbilical Vein Endothelial Cells (HUVECs), Slit2 represses lipopolysaccharide (LPS)-induced secretion of certain inflammatory cytokines/chemokines, cell adhesion molecule ICAM-1 upregulation and monocyte adhesion. Slit2 anti-inflammatory effect is mediated by its dominant endothelial specific receptor Robo4. However the minor receptor Robo1 has pro-inflammatory properties, and is downregulated by Slit2 via targeting of miR-218. Elucidation of molecular mechanism reveals that Slit2 represses inflammatory responses by inhibiting the Pyk2-NF-kB pathway downstream of LPS-TLR4. Further studies reveal that LPS enhances endothelial inflammation by downregulating the anti-inflammatory Slit2 and Robo4 in HUVECs in vitro as well as in arterial endothelial cells and liver in vivo during endotoxemia. These results suggest that Slit2-Robo4 signaling is important in regulating LPS-induced endothelial inflammation, and LPS, in turn, enhances inflammation by interfering with the expression of the anti-inflammatory Slit2-Robo4 during disease state. This implies that Slit2-Robo4 is a key regulator of endothelial inflammation and its dysregulation during endotoxemia is a novel mechanism for LPS-induced vascular pathogenesis.

The Neurorepellent Slit2 Inhibits Postadhesion Stabilization of Monocytes Tethered to Vascular Endothelial Cells

“…Our observations are consistent with work showing that Slit2 inhibited CXCL12-mediated adhesion of human breast cancer cells to fibronectin and collagen (15). Recently, Zhao et al (39) demonstrated that Slit2 prevented adhesion of monocytic cells to activated HUVEC by inhibiting LPS-induced upregulation of ICAM-1 in endothelial cells. It is possible that Slit2 similarly inhibits upregulation of ICAM-1 on the surface of endothelial cells exposed to TNF-a, contributing to the decreased adhesion of monocytic cells that we observed.…”

“…A recent study demonstrated that Robo-1 expression is downregulated in endothelial cells via targeting of miR-218 (39). Because Robo-1 is a single-pass transmembrane protein, cell surface levels could, in principle, also be determined by endocytosis and recycling back to the plasma membrane.…”

Chronic Administration of Δ 9 -Tetrahydrocannabinol Induces Intestinal Anti-Inflammatory MicroRNA Expression during Acute Simian Immunodeficiency Virus Infection of Rhesus Macaques

“…Further, among the seven, miR-29b, miR-101, miR-130a, and miR-362-3p have bioinformatically predicted binding sites on the 3= UTR of CXCL12 (see Tables S5 and S7 in the supplemental material). Similarly, miR- 218, an miRNA previously shown to exert anti-inflammatory effects by directly targeting Robo1 protein translation (47), showed markedly elevated expression in the THC/SIV group (8.6-fold increase) compared to the VEH/SIV (3.9-fold increase) and THConly (2.9-fold increase) groups (see Tables S1 to S3 in the supplemental material). Similar to the 30-day p.i.…”

Role of Fibrinogen in the Attenuation of the Ischemic Reperfusion Injury and in Remote Ischemic Preconditioning

“…The increased expression/ activity of Rap-1 leads to NF-κB induction, while Rap-1 depletion leads to NF-κB decreased activity as Rap-1 is also important for the phosphorylation of p65 subunit of NF-κB. Remarkably, similar to inhibiting NF-κB, knockdown of Rap-1 sensitizes some cells to apoptosis [35][36][37].…”