School of Medical Laboratory Science and Biotechnology, College of Medical Science and Technology, Taipei Medical University, Taipei 110, Taiwan.
Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei 112, Taiwan.
Int J Mol Sci. 2018 Jun 5;19(6):1678. doi: 10.3390/ijms19061678.
Liver fibrosis is the first step toward the progression to cirrhosis, portal hypertension, and hepatocellular carcinoma. A high-cholesterol diet is associated with liver fibrosis via the accumulation of free cholesterol in hepatic stellate cells (HSCs). Niemann-Pick type C2 (NPC2) plays an important role in the regulation of intracellular free cholesterol homeostasis via direct binding with free cholesterol. Previously, we reported that NPC2 was downregulated in liver cirrhosis tissues. Loss of NPC2 enhanced the accumulation of free cholesterol in HSCs and made them more susceptible to transforming growth factor (TGF)-β1. In this study, we showed that knockdown of NPC2 resulted in marked increases in platelet-derived growth factor BB (PDGF-BB)-induced HSC proliferation through enhanced extracellular signal-regulated kinases (ERK), p38, c-Jun N-terminal kinases (JNK), and protein kinase B (AKT) phosphorylation. In contrast, NPC2 overexpression decreased PDGF-BB-induced cell proliferation by inhibiting p38, JNK, and AKT phosphorylation. Although NPC2 expression did not affect caspase-related apoptosis, the autophagy marker light chain 3β (LC3B) was decreased in NPC2 knockdown, and free cholesterol accumulated in the HSCs. The mitochondrial respiration functions (such as oxygen consumption rate, ATP production, and maximal respiratory capacity) were decreased in NPC2 knockdown, and free cholesterol accumulated in the HSCs, while NPC2-overexpressed cells remained normal. In addition, NPC2 expression did not affect the susceptibility of HSCs to lipopolysaccharides (LPS), and U18666A treatment induced free cholesterol accumulation, which enhanced LPS-induced Toll-like receptor 4 (TLR4), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 phosphorylation, interleukin (IL)-1 and IL-6 expression. Our study demonstrated that NPC2-mediated free cholesterol homeostasis controls HSC proliferation and mitochondrial function.
肝纤维化是向肝硬化、门静脉高压和肝细胞癌进展的第一步。高胆固醇饮食通过在肝星状细胞（HSCs）中积累游离胆固醇与肝纤维化有关。尼曼-匹克 C2 型（NPC2）通过与游离胆固醇的直接结合，在调节细胞内游离胆固醇稳态方面发挥重要作用。以前，我们报道 NPC2 在肝硬化组织中下调。NPC2 的缺失增强了 HSCs 中游离胆固醇的积累，使它们更容易受到转化生长因子（TGF）-β1 的影响。在这项研究中，我们表明 NPC2 的敲低导致血小板衍生生长因子 BB（PDGF-BB）诱导的 HSC 增殖明显增加，这是通过增强细胞外信号调节激酶（ERK）、p38、c-Jun N-末端激酶（JNK）和蛋白激酶 B（AKT）磷酸化实现的。相反，NPC2 的过表达通过抑制 p38、JNK 和 AKT 磷酸化降低了 PDGF-BB 诱导的细胞增殖。虽然 NPC2 表达不影响与 Caspase 相关的细胞凋亡，但 NPC2 敲低时自噬标志物微管相关蛋白轻链 3β（LC3B）减少，HSCs 中游离胆固醇积累。NPC2 敲低时，HSCs 中的线粒体呼吸功能（如耗氧量、ATP 产生和最大呼吸能力）降低，游离胆固醇积累，而过表达 NPC2 的细胞保持正常。此外，NPC2 表达不影响 HSCs 对脂多糖（LPS）的敏感性，U18666A 处理诱导游离胆固醇积累，增强 LPS 诱导的 Toll 样受体 4（TLR4）、核因子 kappa-轻链增强子的激活 B 细胞（NF-κB）p65 磷酸化、白细胞介素（IL）-1 和 IL-6 的表达。我们的研究表明，NPC2 介导的游离胆固醇稳态控制 HSC 增殖和线粒体功能。
