Alloantigenicity of human endothelial cells: 1. Frequency and phenotype of human T helper lymphocytes that can react to allogeneic endothelial cells

To determine the relative ability of allogeneic endothelial cells to stimulate helper T lymphocytes (HTL), human PBMC or purified T cells were incubated in conventional lymphocyte microcultures or in limiting dilution microcultures with allogeneic human umbilical vein endothelia (HUVE), with cytokine-treated allogeneic HUVE, or with allogeneic peripheral blood monocytes. These cultures were tested for IL-2 production as an index of HTL stimulation. Dose-response studies in conventional lymphocyte cultures indicated that allogeneic monocytes were better than allogeneic HUVE at stimulating IL-2 production. Limiting dilution analyses revealed that untreated HUVE and TNF-treated HUVE stimulated small numbers of HTL (approximately 1 HTL/30,000 PBMC), whereas 5 to 10 times more HTL were stimulated by IFN-γ-treated HUVE and 10 to 20 times more HTL were stimulated by allogeneic monocytes. SerologiC deletion studies revealed that most of the high frequency HTL responding to IFN-γ-treated HUVE were CD4⁺, whereas most of the low frequency HTL responding to nontreated HUVE or to TNF-treated HUVE were CD8⁺. Interestingly, mAb to MHC class I and class II molecules, which significantly impaired HUVE-induced proliferation, caused little interference with HUVE-induced IL-2 production. Finally, polymerase chain reaction analysis demonstrated that untreated allogeneic HUVE cells could stimulate PBMC to produce mRNA for IFN-γ, as well as for IL-2. These data demonstrate the following hierarchy of allogeneic stimulatory capacity for human HTL: monocytes > IFN-γ-treated HUVE ≫ TNF-treated HUVE = non-treated HUVE. Further, these data suggest that non-activated allogeneic endothelial cells can initiate immune responses by inducing IL-2 and IFN-γ. Because IFN-γ can induce MHC class II expression by the endothelial cells, this could recruit large numbers of CD4⁺ T cells for IL-2 production.