Monoclonal Anti-Endothelial Cell Antibodies From A Patient With Takayasu Arteritis Activate Endothelial Cells From Large Vessels

Objective. To create monoclonal anti-endothelial cell antibodies (mAECA) from a patient with Takayasu arteritis to evaluate their ability to activate human umbilical vein endothelial cells (HUVEC), and to characterize the mechanism of EC activation.Methods, A panel of mAECA was generated from peripheral blood lymphocytes of a patient with Takayasu arteritis, using Epstein-Barr virus transformation. Activity against macrovascular EC (HUVEC) and microvascular EC (human bone marrow EC immortalized by SV40) antigens was detected by enzyme-linked immunosorbent assay. Inhibition studies were used to select the monoclonal antibodies (mAECA) which share the same EC epitope binding specificity as the total IgG-AECA from the Takayasu arteritis patient. The binding of the mAECA to human aortic EC was studied by immunohistochemistry. The secretion levels of interleukin-6 (IL-6) and von Willebrand factor (vWF) were determined, to serve as markers for EC activation. The activated EC were examined for the adherence of a monocytic cell line (U937), as well as for expression of vascular cell adhesion molecule 1, intercellular adhesion molecule 1, and E-selectin, In addition, nuclear extracts of the mAECA-treated EC were analyzed for the induction of translocation of nuclear factor kappa B (NF-kappa B), using a specific NF-kappa B oligoprobe in an electrophoretic mobility shift assay.Results. Six mAECA were selected, the mixture of which produced 100% inhibition of binding of the original IgG (from the patient with Takayasu arteritis) to HUVEC. All mAECA possessed high activity against macrovascular EC, but none had significant anti-microvascular EC activity. The mAECA, but not normal human IgG, had anti-human aortic EC activity. Four of the 6 mAECA activated EC, manifested by increased IL-6 and vWF secretion. The 4 mAECA induced EC expression of adhesion molecules and increased adhesion of U937 monocytic cells to EC, In addition, these mAECA stimulated the nuclear translocation of the NF-kappa B transcription factor.Conclusion. Our findings suggest that AECA may directly stimulate EC in Takayasu arteritis through elevation of adhesion molecule expression associated with NF-kappa B activation and adhesion of monocytes, and may therefore play a pathogenic role in the development of the vasculopathy in Takayasu arteritis.