Endothelial nuclear factor-kappa B translocation and vascular cell adhesion molecule-1 induction by complement inhibition with anti-human C5 therapy or cGMP analogues

We have previously shown that reoxygenation of hypoxic human umbilical vein endothelial cells (HUVECs) leads to the activation and deposition of compl ement. In the present study, we investigated whether the terminal complemen t complex (C5b-9) influences HUVEC nuclear factor-kappa B (NF-kappa B) tran slocation and vascular cell adhesion molecule-1 (VCAM-1) protein expression after hypoxia/reoxygenation by decreasing endothelial cGMP. Additionally, we investigated the action of anti-human C5 therapy on endothelial cGMP, NF -kappa B translocation, and VCAM-1 protein expression. Reoxygenation (0.5 t o 3 hours, 21% O-2) of hypoxic (12 hours, 1% O-2) HUVECs in human serum (HS ) significantly increased C5b-9 deposition, VCAM-1 expression, and NF-kappa B translocation compared with hypoxic/reoxygenated HUVECs treated with the recombinant human C5 inhibitor h5G1.1scFv. Acetylcholine (ACh)-induced cGM P synthesis was significantly higher in normoxic HUVECs compared with hypox ic HUVECs reoxygenated in HS but did not differ from hypoxic HUVECs reoxyge nated in buffer or HS treated with h5G1.1-scFv. Treatment of hypoxic/reoxyg enated HUVECs with h5G1.1-scFv or cGMP analogues significantly attenuated N F-kappa B translocation and VCAM-1 protein expression, Treatment with NO an alogues, but not a cAMP analogue, cGMP antagonists, or an NO antagonist, al so significantly attenuated VCAM-1 expression. We conclude that (1) C5b-9 d eposition, NF-kappa B translocation, and VCAM-1 protein expression are incr eased in hypoxic HUVECs reoxygenated in HS; (2) reoxygenation of hypoxic HU VECs in HS, but not buffer alone, attenuates ACh-induced cGMP synthesis; an d (3) treatment of hypoxic/reoxygenated HUVECs with h5G1.1-scFv attenuates C5b-9 deposition, NF-kappa B translocation, and VCAM-1 expression while pre serving ACh-induced cGMP synthesis, C5b-9-induced VCAM-1 expression may thu s involve an NO/cGMP-regulated NF-kappa B translocation mechanism.