Estrogen receptor‑β‑dependent effects of saikosaponin‑d on the suppression of oxidative stress‑induced rat hepatic stellate cell activation

Abstract

Saikosaponin-d (SSd) is one of the major triterpenoid saponins derived from Bupleurum falcatum L., which has been reported to possess antifibrotic activity. At present, there is little information regarding the potential target of SSd in hepatic stellate cells (HSCs), which serve an important role in excessive extracellular matrix (ECM) deposition during the pathogenesis of hepatic fibrosis. Our recent study indicated that SSd may be considered a novel type of phytoestrogen with estrogen-like actions. Therefore, the present study aimed to investigate the effects of SSd on the proliferation and activation of HSCs, and the underlying mechanisms associated with estrogen receptors. In the present study, a rat HSC line (HSC-T6) was used and cultured with dimethyl sulfoxide, SSd, or estradiol (E2; positive control), in the presence or absence of three estrogen receptor (ER) antagonists [ICI-182780, methylpiperidinopyrazole (MPP) or (R,R)-tetrahydrochrysene (THC)], for 24 h as pretreatment. Oxidative stress was induced by exposure to hydrogen peroxide for 4 h. Cell proliferation was assessed by MTT growth assay. Malondialdehyde (MDA), CuZn-superoxide dismutase (CuZn-SOD), tissue inhibitor of metalloproteinases-1 (TIMP- 1), matrix metalloproteinase-1 (MMP-1), transforming growth factor-β1 (TGF-β1), hydroxyproline (Hyp) and collagen-1 (COL1) levels in cell culture supernatants were determined by ELISA. Reactive oxygen species (ROS) was detected by flow cytometry. Total and phosphorylated mitogen-activated protein kinases (MAPKs) and α-smooth muscle actin (α-SMA) were examined by western blot analysis. TGF-β1 mRNA expression was determined by RT-quantitative (q)PCR. SSd and E2 were able to significantly suppress oxidative stress-induced proliferation and activation of HSC-T6 cells. Furthermore, SSd and E2 were able to reduce ECM deposition, as demonstrated by the decrease in transforming growth factor-β1, hydroxyproline, collagen-1 and tissue inhibitor of metalloproteinases-1, and by the increase in matrix metalloproteinase-1. These results suggested that the possible molecular mechanism could involve downregulation of the reactive oxygen species/mitogen-activated protein kinases signaling pathway. Finally, the effects of SSd and E2 could be blocked by co-incubation with ICI-182780 or THC, but not MPP, thus indicating that ERβ may be the potential target of SSd in HSC-T6 cells. In conclusion, these findings suggested that SSd may suppress oxidative stress-induced activation of HSCs, which relied on modulation of ERβ.

Inhibition of oxidative stress and NLRP3 inflammasome by Saikosaponin-d alleviates acute liver injury in carbon tetrachloride-induced hepatitis in mice

“…In addition, our previous study reported that SSd was a new kind of phytoestrogen, which could suppress the proliferation and activation of hepatic stellate cells through modulation of estrogen receptor β. 30,31 Therefore further studies are still needed to explore the specific action mechanisms of SSd.…”

IL‑22 alleviates the fibrosis of hepatic stellate cells via the inactivation of NLRP3 inflammasome signaling

“…Oxidative stress serves a crucial role in inducing HSC activation and fibrogenic potential ( 16 ). Superoxide dismutase (SOD), one of the most important protective enzymes, provides the first antioxidant defense system in various organs and tissues, including the liver ( 17 ).…”

Saikosaponin-d alleviates hepatic fibrosis through regulating GPER1/autophagy signaling

“…4A, the HSCs from CCl 4 mice exhibited a markedly decreased expression of ER-α, and ER-β, and GPER1, while SSd treatment rescued ER-β and GPER1 expression in primary HSCs. Here we focused on the correlation of SSd with GPER1 because the regulatory role of SSd in ER-β has been verified in our previous studies [30]. The results from qPCR and Western blot analysis showed that TGF-β treatment reduced the mRNA and protein expression of GPER1 in LX-2 cells, whereas SSd treatment reversed the effect (Fig.…”

“…Our previous studies have demonstrated that SSd represses oxidative stress-induced HSCs activation by regulating estrogen receptor (ER)-β [30]. Given the important role of Estrogens in repressing HSCs activation and liver fibrosis by interacting with its nuclear receptors (ER-α, and ER-β) and membrane receptor (G protein-coupled estrogen receptor, GPER1) [31], we next investigated whether SSd exerted its anti-liver fibrotic function by regulating estrogen receptors expression.…”