Cross-Talk between HLA Class I and TLR4 Mediates P-Selectin Surface Expression and Monocyte Capture to Human Endothelial Cells

Abstract

Donor-specific HLA Abs contribute to Ab-mediated rejection (AMR) by binding to HLA molecules on endothelial cells (ECs) and triggering intracellular signaling, leading to EC activation and leukocyte recruitment. The molecular mechanisms involving donor-specific HLA Ab–mediated EC activation and leukocyte recruitment remain incompletely understood. In this study, we determined whether TLRs act as coreceptors for HLA class I (HLA I) in ECs. We found that human aortic ECs express TLR3, TLR4, TLR6, and TLR10, but only TLR4 was detected on the EC surface. Consequently, we performed coimmunoprecipitation experiments to examine complex formation between HLA I and TLR4. Stimulation of human ECs with HLA Ab increased the amount of complex formation between HLA I and TLR4. Reciprocal coimmunoprecipitation with a TLR4 Ab confirmed that the crosslinking of HLA I increased complex formation between TLR4 and HLA I. Knockdown of TLR4 or MyD88 with small interfering RNAs inhibited HLA I Ab–stimulated P-selectin expression, von Willebrand factor release, and monocyte recruitment on ECs. Our results show that TLR4 is a novel coreceptor for HLA I to stimulate monocyte recruitment on activated ECs. Taken together with our previous published results, we propose that HLA I molecules form two separate signaling complexes at the EC surface, that is, with TLR4 to upregulate P-selectin surface expression and capture of monocytes to human ECs and integrin β4 to induce mTOR-dependent firm monocyte adhesion via ICAM-1 clustering on ECs, two processes implicated in Ab-mediated rejection.

Spatial Multi-omics of Arterial Regions from Cardiac Allograft Vasculopathy Rejected Grafts

“…Monocyte recruitment may occur in the early phases of CAV as we observed significantly higher protein expression of CD14 and classical monocytes counts in arteries with low neointima. HLA DSA can rapidly increase intracellular calcium and endothelial presentation of P-selectin, which supports monocyte binding 7,49 . Notably, CD68 protein expression also significantly correlated with coexpressing module ME206 in AOIs with high neointima.…”

“…Episodes of antibody-mediated rejection (AMR) in which donor specific antibodies (DSA) target human leukocyte antigens (HLA) present on vascular endothelial cells (ECs) have been increasingly recognized as a major risk factor contributing to CAV 3 . Specifically, DSA can activate EC intracellular signaling cascades inducing EC proliferation, migration, and increased surface expression of adhesion molecules promoting monocyte and NK cell recruitment [4][5][6][7][8] . Moreover, DSA can directly mediate EC-injury by triggering activation of the classical complement cascade 8 .…”

Lack of NPR1 Increases Vascular Endothelial Adhesion through Induction of Integrin Beta 4

“…Through Western analysis, it was observed that ITGB4 expression was increased in NPR1 -kncokdown HUVECs ( Figure 3 E). Furthermore, ITGB4 binds to HLA class I (HLAI) as a complex that strengthens monocyte adhesion via ICAM-1 clustering on endothelial cells by the mTOR signaling pathway [ 27 ]. All of the data suggest that NPR1 deficiency promotes the ITGB4 expression level.…”

“…A mouse model of ITGB4 with deletion of cytoplasmic signaling domain reduces the endothelial inflammatory response induced by mechanical stress [ 48 ], while mice with atherosclerotic plaques display an increased ITGB4 [ 9 ]. Recently, a study found that the HLA I/ITGB4 complex stabilizes monocyte adhesion via ICAM-1 aggregation on endothelial cells in an mTOR-dependent manner [ 27 ]. This evidence indicates that ITGB4 participates in endothelial cell adhesion, inflammation, and atherogenesis.…”

Posttransplant de novo DSA and NDSA affect GvHD, OS, and DFS after haplo-HSCT in patients without pre-existing HLA antibodies of hematological malignancies

“…HLA antibodies crosslinking of HLA molecules on endothelial cells stimulates intracellular signaling, including FAK/Src, ERK, PIK, and mTORC1. HLA molecules trigger these signaling pathways by physically associating with integrin β4 or TLR4, thereby leading to activation of endothelial cells and recruitment of leukocytes, which manifests as transplant vasculopathy ( 26 , 27 ).…”