Abstract
BackgroundApart from the platelet/endothelial cell adhesion molecule 1 (PECAM-1, CD31), endoglin (CD105) and a positive factor VIII-related antigen staining, human primary and immortalized macro- and microvascular endothelial cells (ECs) differ in their cell surface expression of activating and inhibitory ligands for natural killer (NK) cells. Here we comparatively study the effects of irradiation on the phenotype of ECs and their interaction with resting and activated NK cells.Methodology/Principal FindingsPrimary macrovascular human umbilical vein endothelial cells (HUVECs) only express UL16 binding protein 2 (ULBP2) and the major histocompatibility complex (MHC) class I chain-related protein MIC-A (MIC-A) as activating signals for NK cells, whereas the corresponding immortalized EA.hy926 EC cell line additionally present ULBP3, membrane heat shock protein 70 (Hsp70), intercellular adhesion molecule ICAM-1 (CD54) and HLA-E. Apart from MIC-B, the immortalized human microvascular endothelial cell line HMEC, resembles the phenotype of EA.hy926. Surprisingly, primary HUVECs are more sensitive to Hsp70 peptide (TKD) plus IL-2 (TKD/IL-2)-activated NK cells than their immortalized EC counterpatrs. This finding is most likely due to the absence of the inhibitory ligand HLA-E, since the activating ligands are shared among the ECs. The co-culture of HUVECs with activated NK cells induces ICAM-1 (CD54) and HLA-E expression on the former which drops to the initial low levels (below 5%) when NK cells are removed. Sublethal irradiation of HUVECs induces similar but less pronounced effects on HUVECs. Along with these findings, irradiation also induces HLA-E expression on macrovascular ECs and this correlates with an increased resistance to killing by activated NK cells. Irradiation had no effect on HLA-E expression on microvascular ECs and the sensitivity of these cells to NK cells remained unaffected.Conclusion/SignificanceThese data emphasize that an irradiation-induced, transient up-regulation of HLA-E on macrovascular ECs might confer protection against NK cell-mediated vascular injury.
Highlights
The endothelial cell (EC) monolayer which lines blood vessels performs multiple tasks including the regulation of tissue fluid homeostasis and blood cell transmigration
All three endothelial cell types show a positive staining for the factor VIII-related antigen (Figure 1A–C) and strongly express the endothelial cell associated markers CD31, [35] and CD105 [36], a component of the TGF-beta 1 receptor complex, which is involved in vascular remodelling (Table 1)
A translocation of the major stress-inducible heat shock protein 70 (Hsp70) to the plasma membrane [36,37] is typically associated with a tumorigenic phenotype, since normal cells are membrane Hsp702, a membrane Hsp70+ phenotype was selectively detectable on immortalized EA.hy926 cells and HMECs using the cmHsp70.1 monoclonal antibodies (mAbs), but not on primary human umbilical vein endothelial cells (HUVECs)
Summary
The endothelial cell (EC) monolayer which lines blood vessels performs multiple tasks including the regulation of tissue fluid homeostasis and blood cell transmigration. The interaction of ECs with leukocytes, such as activated NK cells, involves attachment and rolling which are predominantly mediated by selectins and carbohydrate-rich ligands, such as mucins [1,2,3]. The functionality of NK cells is regulated by interactions of inhibitory and activating receptors [11,12] which belong either to the immunoglobulin (Ig)-like (KIR), Ig-like transcript (ILT), C-type lectin or natural cytotoxicity group, with the signals being derived from the target cell ligands [13]. Inhibitory receptors with specificity for classical and non-classical MHC class I molecules mediate protection for the target cells. HLA-E, a prominent member of the non-classical MHC (MHC class Ib), which is recognized by CD94/NKG2A (inhibitory) and CD94/NKG2C (activating) expressing NK cells, is characterized by a limited polymorphism and a tissue-specific and inducible expression pattern [14,15]. We comparatively study the effects of irradiation on the phenotype of ECs and their interaction with resting and activated NK cells
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