ABSTRACT
The Chinese herb Salvia miltiorrhiza (SM) has been found to have beneficial effects on the circulatory system. In the present study, we investigated the effects of cryptotanshinone (derived from SM) on endothelin-1 (ET-1) expression in human umbilical vein endothelial cells (HUVECs). The effect of cryptotanshinone on nitric oxide (NO) in HUVECs was also examined. We found that cryptotanshinone inhibited basal and tumor necrosis factor-alpha (TNF-alpha) stimulated ET-1 secretion in a concentration-dependent manner. Cryptotanshinone also induced a concentration-dependent decrease in ET-1 mRNA expression. Cryptotanshinone increased basal and TNF-alpha-attenuated NO production in a dose-dependent fashion. Cryptotanshinone induced a concentration-dependent increase in endothelial nitric oxide synthase (eNOS) expression without significantly changing neuronal nitric oxide synthase (nNOS) expression in HUVECs in the presence or absence of TNF-alpha. NOS activities in the HUVECs were also induced by cryptotanshinone. Furthermore, decreased ET-1 expression in response to cryptotanshinone was not antagonized by the NOS inhibitor l-NAME. A gel shift assay further showed that TNF-alpha-induced Nuclear Factor-kappaB (NF-kappaB) activity was significantly reduced by cryptotanshinone. These data suggest that cryptotanshinone inhibits ET-1 production, at least in part, through a mechanism that involves NF-kappaB but not NO production.
Subject(s)
Endothelin-1/antagonists & inhibitors , Endothelium, Vascular/cytology , Gene Expression Regulation/drug effects , Nitric Oxide/biosynthesis , Phenanthrenes/pharmacology , Cells, Cultured , Drugs, Chinese Herbal/pharmacology , Endothelial Cells/drug effects , Endothelin-1/genetics , Endothelium, Vascular/drug effects , Humans , NF-kappa B/metabolism , Nitric Oxide Synthase Type I/genetics , Nitric Oxide Synthase Type III/genetics , RNA, Messenger/analysis , Salvia miltiorrhiza/chemistry , Tumor Necrosis Factor-alpha/pharmacology , Umbilical VeinsABSTRACT
Plasminogen activator inhibitor type 1 (PAI-1), which plays a role in the development of atherosclerosis, is produced by endothelial cells following stimulation with various inflammatory cytokines such as tumor necrosis factor (TNF-alpha). In the present study, we investigated the effects of a potent water-soluble antioxidant, salvianolic acid B (SalB; derived from the Chinese herb, Salvia miltiorrhiza), on the expression of PAI-1 in TNF-alpha-treated human umbilical vein endothelial cells (HUVECs). We found that SalB inhibited TNF-alpha-induced PAI-1 mRNA production and protein secretion in HUVECs. Treatment with SalB (0.05 and 0.15 microM) notably attenuated TNF-alpha induced expression of PAI-1 to 90.5% and 74.6%, respectively, after 12 h, and to 75.1% and 64.2%, respectively, after 18 h. We also observed a dose-dependent decrease in PAI-1 protein production in the presence of SalB. We then used pathway inhibitors to investigate which step of the TNF-alpha induced signaling pathway was targeted by SalB. We found that the c-Jun N-terminal kinase (JNK) inhibitor, SP600125, increased the inhibitory effects of SalB on TNF-alpha-induced PAI-1 secretion, whereas the nuclear factor-kappaB (NF-kappaB) inhibitor, emodin, and the extracellular signal-regulated kinase (ERK) inhibitor, PD98059, did not. A gel shift assay further showed that SalB inhibited the TNF-alpha-activated NF-kappaB and AP-1 DNA binding activities in a dose-dependent manner. Collectively, these results indicate that the NF-kappaB and ERK-AP-1 pathways are possible targets of SalB in the regulation of TNF-alpha-stimulated PAI-1 production in HUVECs.
Subject(s)
Benzofurans/pharmacology , Drugs, Chinese Herbal/pharmacology , Endothelium, Vascular/drug effects , Plasminogen Activator Inhibitor 1/biosynthesis , Tumor Necrosis Factor-alpha/physiology , Cells, Cultured , Endothelial Cells/drug effects , Endothelial Cells/metabolism , Endothelium, Vascular/metabolism , Humans , NF-kappa B/metabolism , Plasminogen Activator Inhibitor 1/genetics , Plasminogen Activator Inhibitor 1/metabolism , RNA, Messenger/metabolism , Signal Transduction/drug effects , Tissue Plasminogen Activator/metabolism , Transcription Factor AP-1/metabolism , Umbilical Veins/drug effects , Umbilical Veins/metabolismABSTRACT
Adhesion molecules, which play a crucial role in the development of atherogenesis, are produced by endothelial cells following stimulation with various inflammatory cytokines. The current studies examined the effect of a potent water-soluble antioxidant, protocatechuic aldehyde (derived from the Chinese herb, Salvia miltiorrhiza), on the expression of adhesion molecules in human umbilical vein endothelial cells (HUVECs) stimulated with tumor necrosis factor-alpha (TNF-alpha). Protocatechuic aldehyde appeared to specifically downregulate the TNF-alpha-induced cell surface expression of vascular adhesion molecule-1 (VCAM-1) and intercellular cell adhesion molecule-1 (ICAM-1) on HUVECs as well as the release of soluble VCAM-1and ICAM-1 from HUVECs in a dose-response manner at pharmacologically relevant concentrations (0.15-1.35 mM). We also observed a dose-dependent lowering of mRNA expression of VCAM-1 and ICAM-1 in the presence of protocatechuic aldehyde. Furthermore, protocatechuic aldehyde (0.15, 0.45, and 1.35 mM) notably inhibited TNF-alpha-induced upregulation of U937 cell adhesion to HUVECs to 83.7%, 60.9%, and 40.8%, respectively. A gel shift assay further showed that protocatechuic aldehyde inhibited the TNF-alpha-activated NF-kappaB and AP-1 DNA binding activities in a dose-dependent manner. Collectively, these results indicate that protocatechuic aldehyde inhibits TNF-alpha-stimulated VCAM-1 and ICAM-1expression in HUVECs through a mechanism that involves NF-kappaB and AP-1.
Subject(s)
Benzaldehydes/pharmacology , Catechols/pharmacology , Endothelial Cells/drug effects , Intercellular Adhesion Molecule-1/genetics , Tumor Necrosis Factor-alpha/pharmacology , Vascular Cell Adhesion Molecule-1/genetics , Blotting, Western , Cell Adhesion/drug effects , Cell Line , Dose-Response Relationship, Drug , E-Selectin/genetics , E-Selectin/metabolism , Endothelial Cells/cytology , Endothelial Cells/metabolism , Gene Expression/drug effects , Humans , Intercellular Adhesion Molecule-1/metabolism , NF-kappa B/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Transcription Factor AP-1/metabolism , U937 Cells , Umbilical Veins/cytology , Vascular Cell Adhesion Molecule-1/metabolismABSTRACT
OBJECTIVE: To establish the HPLC fingerprint of Xuelian injection. METHOD: HPLC with Diamonsil C18 (4.6 mm x 250 mm, 5 microm) column was used, the methanol-1% acetic acid as a mobile phase and detection wavelength at 270 nm. RESULT: Indicating 11 peaks on HPLC fingerprint. CONCLUSION: This method is simple and acurate with a good reproducibility and can be used as a quality control method for Xuelian injection.
