ErMiao San Inhibits Angiogenesis in Rheumatoid Arthritis by Suppressing JAK/STAT Signaling Pathways

Abstract

ErMiao San (EMS) is composed of the Cortex Phellodendri chinensis and Atractylodes lancea, and it has the function of eliminating heat and excreting dampness in terms of traditional Chinese medicine to damp heat syndrome. Previous reports indicate that EMS possesses anti-inflammatory activity; however, its action on angiogenesis of rheumatoid arthritis (RA) has not been clarified. The present study aims to determine the antiangiogenic activity of EMS in collagen-induced arthritis (CIA) mice and in various angiogenesis models. Our data showed that EMS (5 g/kg) markedly reduced the immature blood vessels in synovial membrane tissues of inflamed joints from CIA mice. It also inhibited vascular endothelial growth factor (VEGF)-induced microvessel sprout formation ex vivo. Meanwhile, EMS suppressed VEGF-induced migration, invasion, adhesion, and tube formation of human umbilical vein endothelial cells (HUVECs). Moreover, EMS significantly reduced the expression of angiogenic activators including interleukin (IL)-1β, IL-6, and tumor necrosis factor-alpha (TNF-α) in synovium of CIA mice. More interestingly, EMS blocked the autophosphorylation of VEGF-induced JAK1, STAT1, and STAT6 in CIA mice and VEGF-induced HUVECs. These findings suggest for the first time that EMS possesses the antiangiogenic effect in RA in vivo, ex vivo, and in vitro by interrupting the targeting of JAK/STAT activation.

Shikonin attenuates rheumatoid arthritis by targeting SOCS1/JAK/STAT signaling pathway of fibroblast like synoviocytes

“…The experiment was carried out as described previously [ 21 , 22 , 25 ]. FLS (5 × 10 4 cells per well) were cultured in a 48-well plate and incubated overnight yielding confluent monolayers for wounding.…”

“…The migration of FLS to TNF-α as mentioned earlier, Transwell migration analysis was used [ 21 , 22 , 25 ]. In the lower wells, 500 μL complete H-DMEM contained with 20 ng/mL TNF-α prepared was placed.…”

“…This experiment was carried out in Transwell room, of which the transwell insert upper surfaces were pre-coated with 1.25 mg/mL Matrigel (20 μL/well) at 37 °C for 30–60 min and the bottom chamber contained 600 μL of culture medium with or without 20 ng/mL TNF-α as mentioned previously [ 21 , 22 , 25 ]. 1 × 10 4 cells/well FLS were seeded on the upper chamber and incubated in normal H-DMEM medium with different concentrations of SKN (0, 0.05, 0.1 and 0.2 μM).…”

“…FLS (5 × 10 4 cells/mL) were cultured in the pore of 24-well plates as previously described [ 21 , 22 , 25 ], and then cultured with or without 20 ng/mL TNF-α, and different concentrations of SKN (0, 0.05, 0.1 and 0.2 μM) for 24 h. Treated/untreated cells were then seeded in 20 mg/L fibronectin (FN), or 10 mg/mL bovine serum albumin (BSA, used as negative control)-coated 96-well plates and incubated in sterile complete H-DMEM medium for 1.5 h. Then cells were washed twice with PBS, followed by sterile H-DMEM (200 μL) including 5% FBS and 10% (v/v) MTT reagent adding to the cells and measuring with a microplate reader at 490 nm. The results were expressed as cell adhesiveness.…”

“…The assay was measured as previously mentioned [ 21 , 22 , 25 ]. Synovium tissue of mice knee joints were collected.…”