AbstractIn vitro human tissue engineered human blood vessels (TEBV) that exhibit vasoactivity can be used to test human toxicity of pharmaceutical drug candidates prior to pre-clinical animal studies. TEBVs with 400–800 μM diameters were made by embedding human neonatal dermal fibroblasts or human bone marrow-derived mesenchymal stem cells in dense collagen gel. TEBVs were mechanically strong enough to allow endothelialization and perfusion at physiological shear stresses within 3 hours after fabrication. After 1 week of perfusion, TEBVs exhibited endothelial release of nitric oxide, phenylephrine-induced vasoconstriction and acetylcholine-induced vasodilation, all of which were maintained up to 5 weeks in culture. Vasodilation was blocked with the addition of the nitric oxide synthase inhibitor L-N^G-Nitroarginine methyl ester (L-NAME). TEBVs elicited reversible activation to acute inflammatory stimulation by TNF-α which had a transient effect upon acetylcholine-induced relaxation and exhibited dose-dependent vasodilation in response to caffeine and theophylline. Treatment of TEBVs with 1 μM lovastatin for three days prior to addition of Tumor necrosis factor – α (TNF-α) blocked the injury response and maintained vasodilation. These results indicate the potential to develop a rapidly-producible, endothelialized TEBV for microphysiological systems capable of producing physiological responses to both pharmaceutical and immunological stimuli.