AIM Urtica dentata hand (UDH) are traditionally used in the alpine region as a herbal medicine.Our goal was to elucidate the cellular and molecular pathway and explore a method to quantify the total coumarins (TC) in UDH.METHODS We established a specific HPLC method to quantify TC in UDH.The mechanism of TC in promoting immune tolerance was studied using dendritic cell (DC) and regulatory cell (Treg).RESULTS We described the ability of TC-conditioned DC, reduced capacity to stimulate effector T cell responses.We also found that PD-LI (programmed death ligand-1) that plays an inhibitory role in regulating T cell activation in the periphery, were up-regulated on DC by TC.Consequently, TC-DC-stirsulated Treg were superior alloantigenspecific suppressor of T effector response compared with those stimulated by control (CTR)-DC.Furthermore, TC-conditioned DC increased levels of Foxp3 and CTLA-4 in the CD25 T cell population.TC-DC did not down-regulate TLR4 protein expression in response to LPS.This shows that down-regulation of TLR4 in response to TC on DC is a critical signaling pathway that regulate DCs phenotype and function.We also established a sensitive and specific high-performance liquid chromatography-diodearray detection-mass spectrometry (HPLC-DAD-MS) for simultaneous identification of its main coumarins ,6,6 ,7,7-tetramethoxyl-8,8-biscoumaxin(1),7,7 -dihydroxy-6,6-dimethoxy-8,80-biscoumarin (2), 7,7 -dimethoxy-6,6-biscoumarin (3), scoparone (4).CONCLUSION A demonstration of this mechanism and method for identification and quantification of TC in UDH endorses their potential as tolerancepromoting herbal medicine to prevent or treat transplantation rejection and autoimmune diseases.