RESUMEN
The present study aimed to investigate the inhibitory effect and mechanism of Isodon terricolous-medicated serum on lipopolysaccharide(LPS)-induced hepatic stellate cell(HSC) activation. LPS-induced HSCs were divided into a blank control group, an LPS model group, a colchicine-medicated serum group, an LPS + blank serum group, an I. terricolous-medicated serum group, a Toll-like receptor 4(TLR4) blocker group, and a TLR4 blocker + I. terricolous-medicated serum group. HSC proliferation was detected by methyl thiazolyl tetrazolium(MTT) assay. Enzyme-linked immunosorbent assay(ELISA) was used to measure type â collagen(COL â ), COL â ¢, transforming growth factor-ß1(TGF-ß1), intercellular adhesion molecule-1(ICAM-1), α-smooth muscle actin(α-SMA), vascular cell adhesion molecule-1(VCAM-1), cysteinyl aspartate-specific proteinase-1(caspase-1), and monocyte chemotactic protein-1(MCP-1). Real-time PCR(RT-PCR) was used to detect mRNA expression of TLR4, IκBα, and NOD-like receptor thermal protein domain associated protein 3(NLRP3), nuclear factor-κB(NF-κB) p65, gasdermin D(GSDMD), and apoptosis-associated speck-like protein containing a CARD(ASC) in HSCs. Western blot(WB) was used to detect the protein levels of TLR4, p-IκBα, NF-κB p65, NLRP3, ASC, and GSDMD in HSCs. The results showed that I. terricolous-medicated serum could inhibit the proliferation activity of HSCs and inhibit the secretion of COL â , COL â ¢, α-SMA, TGF-ß1, caspase-1, MCP-1, VCAM-1, and ICAM-1 in HSCs. Compared with the LPS model group, the I. terricolous-medicated serum group, the colchicine-medicated serum group, and the TLR4 blocker group showed down-regulated expression of p-IκBα, NLRP3, NF-κB p65, GSDMD, and ASC, and up-regulated expression of IκBα. Compared with the TLR4 blocker group, the TLR4 blocker + I. terricolous-medicated serum group showed decreased expression of TLR4, p-IκBα, NLRP3, NF-κB p65, GSDMD, and ASC, and increased expression of IκBα. In conclusion, I. terricolous-medicated serum down-regulates HSC activation by inhibiting the TLR4/NF-κB/NLRP3 signaling pathway.
Asunto(s)
Isodon , FN-kappa B , FN-kappa B/genética , FN-kappa B/metabolismo , Células Estrelladas Hepáticas , Factor de Crecimiento Transformador beta1/metabolismo , Inhibidor NF-kappaB alfa/metabolismo , Molécula 1 de Adhesión Intercelular/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismo , Molécula 1 de Adhesión Celular Vascular/metabolismo , Lipopolisacáridos/farmacología , Transducción de Señal , Colchicina/metabolismo , Colchicina/farmacología , CaspasasRESUMEN
A simple high-performance liquid chromatography (HPLC) method for the simultaneous separation of the highly polar and weakly polar components of traditional Chinese medicines was developed via a strategic combination of isocratic and gradient elution methods. Liu-Shen-Wan and Liu-Wei-Di-Huang-Wan were used as representative examples of traditional Chinese medicines. This is the first time that 6 components of varying degrees of polarity in Liu-Shen-Wan had been successfully resolved in a single chromatographic run using an ultraviolet-visible detector with a fixed wavelength of 296 nm. In contrast to conventional gradient separation methods, this novel method offered a viable route for separation of the highly and weakly polar fractions simultaneously, thus greatly reducing the time and cost of analysis. This method therefore provides a more efficient way to determine the polar components present in traditional Chinese medicines. It would find potential application in drug screening, drug authentication, and product quality control.