Department of Pharmacy, 302 Hospital of People's Liberation Army, Beijing, People's Republic of China; Chengde Medical University Chengde, Hebei, People's Republic of China.
Department of Health Statistics, Taishan Medical University, Taian, Shandong, People's Republic of China.
J Ethnopharmacol. 2018 Jan 10;210:232-241. doi: 10.1016/j.jep.2017.08.029. Epub 2017 Aug 31.
Liuweiwuling (LWWL) tablets contain a six-herb Chinese formula and are commonly prescribed to facilitate nourishment of the liver and kidneys, clear away toxic materials and activate blood circulation. Administration of LWWL is well known for its protective effects on the liver and its capacity to confer long-term stability in patients exhibiting reduced transaminase levels. Clinical studies have reported that LWWL can also be used for the treatment of liver fibrosis with associated treatment regimens resulting in a concomitant reduction in transforming growth factor β1 (TGF-β1) levels in the serum of patients with hepatic fibrosis. TGF-β1 plays a prominent role in stimulating liver fibrogenesis and this effect is mediated by myofibroblasts (MFB) derived from hepatic stellate cells (HSCs). It is likely that this phenomenon underpins the antifibrotic effects associated with LWWL.
The purpose of this study was to investigate the antifibrotic effects and mechanisms pertaining to LWWL.
Hepatic fibrosis was induced in rats following bile duct ligation (BDL). Rats that underwent BDL received daily gavage administration of colchicine (0.2mg/kg per day), LWWL (0.4, 1.6, 6.4g/kg per day) or PBS (for the control group). Pathological changes in hepatic tissue were examined using hematoxylin and eosin (HE) and sirius red staining. Immunohistochemical analysis was performed to monitor α-SMA and type I collagen (Collagen I) protein expression. Real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) and Western blot analyses were used to monitor the expression of genes and proteins in the TGF-β/Smad signaling pathway, including TGF-β1, bone morphogenic protein and activin membrane-bound inhibitor (Bambi), Smad3, phosphorylated Smad3 (p-Smad3) and Smad7. We also monitored the expression of genes and proteins in the nuclear factor-κB (NF-κB) signaling pathway, including NF-κB p65, IκBα and phosphorylation of IκBα (p-IκBα), tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6) and interleukin 1β (IL-1β).
LWWL dose-dependently inhibited BDL-induced liver injury and hepatic fibrosis in rats. Furthermore, LWWL reduced liver tissue collagen deposition, hydroxyproline content, liver dysfunction and α-SMA expression in BDL-induced hepatic fibrosis rats. Moreover, LWWL markedly prevented activation of the TGF-β/Smad signaling pathway by inhibiting expression of Smad2/3 and phosphorylation of Smad3, and upregulating the expression of Bambi and Smad7. In addition, LWWL regulated the expression of the inflammatory cytokines IL-1β, TNF-α and IL-6 by inhibiting the activation of NF-κB p65 and the phosphorylation of IκBα, and increasing the expression of IκBα.
LWWL can attenuate BDL-induced hepatic fibrosis in rats, and this effect may be due to modulation of the NF-κB-dependent inflammatory response and activation of HSC and TGF-β/Smad-mediated synthesis and degradation of Collagen I.
六味五灵片（LWWL）含有一种六草药配方，常用于滋养肝脏和肾脏，清除毒素并促进血液循环。LWWL 的给药以其对肝脏的保护作用以及在降低转氨酶水平的患者中实现长期稳定性的能力而闻名。临床研究表明，LWWL 还可用于治疗肝纤维化，相关治疗方案可导致肝纤维化患者血清中转化生长因子-β1（TGF-β1）水平同时降低。TGF-β1 在刺激肝纤维化中起重要作用，这种作用是由来自肝星状细胞（HSCs）的肌成纤维细胞（MFB）介导的。这可能是 LWWL 相关抗纤维化作用的基础。
本研究旨在探讨 LWWL 的抗纤维化作用和机制。
胆总管结扎（BDL）后诱导大鼠肝纤维化。接受 BDL 的大鼠每天给予秋水仙碱（0.2mg/kg/天）、LWWL（0.4、1.6、6.4g/kg/天）或 PBS（对照组）灌胃。使用苏木精和伊红（HE）和天狼猩红染色检查肝组织的病理变化。通过免疫组织化学分析监测α-SMA 和 I 型胶原（Collagen I）蛋白表达。实时定量逆转录聚合酶链反应（RT-qPCR）和 Western blot 分析用于监测 TGF-β/Smad 信号通路中基因和蛋白的表达，包括 TGF-β1、骨形态发生蛋白和激活素膜结合抑制剂（Bambi）、Smad3、磷酸化 Smad3（p-Smad3）和 Smad7。我们还监测了核因子-κB（NF-κB）信号通路中基因和蛋白的表达，包括 NF-κB p65、IκBα 和磷酸化 IκBα（p-IκBα）、肿瘤坏死因子-α（TNF-α）、白细胞介素 6（IL-6）和白细胞介素 1β（IL-1β）。
LWWL 剂量依赖性地抑制 BDL 诱导的大鼠肝损伤和肝纤维化。此外，LWWL 减少了 BDL 诱导的肝纤维化大鼠肝组织胶原沉积、羟脯氨酸含量、肝功能和α-SMA 表达。此外，LWWL 通过抑制 Smad2/3 的表达和 Smad3 的磷酸化，上调 Bambi 和 Smad7 的表达，显著阻止了 TGF-β/Smad 信号通路的激活。此外，LWWL 通过抑制 NF-κB p65 的激活和 IκBα 的磷酸化，增加 IκBα 的表达，调节炎性细胞因子 IL-1β、TNF-α 和 IL-6 的表达。
LWWL 可减轻 BDL 诱导的大鼠肝纤维化，其作用可能是通过调节 NF-κB 依赖性炎症反应以及 HSC 的激活和 TGF-β/Smad 介导的 Collagen I 的合成和降解来实现的。
