Geniposide alleviates VEGF-induced angiogenesis by inhibiting VEGFR2/PKC/ERK1/2-mediated SphK1 translocation.

BACKGROUND: Rheumatoid arthritis (RA) is an angiogenesis-dependent disease caused by the imbalance of pro- and anti-angiogenic factors. More effective strategies to block synovial angiogenesis in RA should be studied. Geniposide (GE), a natural product isolated from the fruit of Gardenia jasminoides Ellis (GJ), is reported to have anti-inflammatory, anti-angiogenic and other pharmacological effects. However, the underlying mechanism through which GE affects synovial angiogenesis in RA remains unclear. PURPOSE: In this research, we aimed to elucidate the effect and potential mechanisms of GE on angiogenesis in RA. MATERIALS AND METHODS: Synovial angiogenesis in patients with RA and a rat model of adjuvant arthritis (AA) was detected by hematoxylin and eosin (HE) staining, immunohistochemistry (IHC), and western blottiing. The biological functions of vascular endothelial cells (VECs) and sphingosine kinase 1 (SphK1) translocation were checked by CCK-8, EdU, Transwell, tube formation, co-immunoprecipitation assays, and laser scanning confocal microscopy. The effect of the SphK1 gene on angiogenesis was assessed by transfection of SphK1-siRNA in cells and mices. The effect of GE on VEGF-induced angiogenesis was measured by Matrigel plug assay in a mouse model of AA. RESULTS: GE effectively inhibited synovial angiogenesis and alleviated the disease process. SphK1, as a new regulatory molecule, has a potentially important relationship in regulating VEGF/VEGFR2 and S1P/S1PR1 signals. SphK1 translocation was activated via the VEGFR2/PKC/ERK1/2 pathway and was closely linked to the biological function of VECs. GE significantly reduced SphK1 translocation, thereby ameliorating the abnormal biological function of VECs. Furthermore, after transfection of SphK1 siRNA in VECs and C57BL/6 mice, silencing SphK1 caused effectively attenuation of VEGF-induced VEC biological functions and angiogenesis. In vivo, the Matrigel plug experiment indicated that GE significantly inhibited pericyte coverage, basement membrane formation, vascular permeability, and fibrinogen deposition. CONCLUSIONS: Our findings suggest that GE inhibited VEGF-induced VEC biological functions and angiogenesis by reducing SphK1 translocation. Generally, studies have revealed that GE down-regulated VEGFR2/PKC/ERK1/2-mediated SphK1 translocation and inhibited S1P/S1PR1 signaling activation, thereby alleviating VEGF-stimulated angiogenesis. The above evidences indicated that angiogenesis inhibition may provide a new direction for RA treatment.

Comparison of tranexamic acid pharmacokinetics after intra-articular and intravenous administration in rabbits.

Tranexamic Acid (TXA) is commonly administered in total knee arthroplasty for reducing blood loss. There has been a growing interest in the topical use of TXA except intravenous use for prevention of bleeding in TKA. The aim of this study was to develop and validate a HPLC-MS method to detect TXA and apply to compare the pharmacokinetic profile of TXA after intravenous (IV) and topical intra-articular (IA) application of TXA at a dose of 20 mg/kg in rabbits. In order to prove intra-articular administration is better than that of intravenous administration from the point of rabbit pharmacokinetic. Two groups of rabbits (n=6/group) respectively received TXA intra-articularly or intravenously. Blood samples were collected at scheduled time. The concentration of TXA in plasma was determined by a validated HPLC-MS method. Excellent linearity was found between 0.015 and 70.0µg/ml with a lower limit of quantitation (LLOQ) of 0.015µg/ml (r>0.99); moreover, all the validation data including accuracy and precision (intra- and inter-day) were all within the required limits. The pharmacokinetic parameters in IA and IV group were: Cmax: 30.65±3.31 VS 54.05± 6.21µg/ml (p<0.01); t1/2: 1.26±0.05 VS 0.68±0.13h (p<0.05); AUC0-t: 42.98±7.73 VS 23.39±4.14µg/ml• h (p<0.01), time above the minimum effective concentration (%T > MEC): 1.5-2.2 VS 0.7-1.2h (p<0.05). HPLC-MS method is suitable for TXA pharmacokinetic studies. The results demonstrated that topical intra-articular application of TXA showed a reduced peak plasma concentration and prolonged therapeutic drug level compared with intravenous TXA from the point of rabbit pharmacokinetic.