Department of Gastroenterology, The First Affiliated Hospital of Jiaxing College, 1882 Central-South Road, Jiaxing, 314001, Zhejiang Province, People's Republic of China,
Dig Dis Sci. 2014 Oct;59(10):2452-61. doi: 10.1007/s10620-014-3208-1. Epub 2014 May 23.
The development of liver fibrosis is the fundamental stage toward a number of mortal complications of liver diseases, including cirrhosis and hepatocellular carcinoma. Canonical Wnt pathway is crucial in diverse biological processes and mediates the progression and regression of liver fibrosis. As a potent Wnt pathway agonist, the role of roof plate-specific spondin-2 (R-Spondin2) in the hepatic fibrosis has not been well elucidated.
The purpose of this study was to investigate whether R-Spondin2 contributes to hepatic stellate cells (HSCs) activation, the key event in liver fibrogenesis.
Human liver tissues, hepatic fibrosis mouse model, and freshly isolated mice HSCs were used. Protein expression and transcriptional level were analyzed by Western-blot assays and real-time PCR, respectively. Exogenous stimulation with recombinant R-Spondin2 and knockdown of R-Spondin2 were performed to investigate functionality. Nuclear β-catenin level and T cell-specific transcription factors activity were analyzed, and HSC proliferation was tested by MTT assay.
Overexpression of R-Spondin2 was observed in both human fibrotic liver tissues and hepatic fibrosis mouse model. Exogenous stimulation with R-Spondin2 in the freshly isolated mice HSCs induced a dose-dependent increase in Wnt pathway activities, HSC proliferation, and the expression of α-smooth muscle actin (α-SMA) and Collagen I. Additionally, Wnt pathway activities, HSC proliferation, and the expressions of α-SMA and Collagen I decreased in the R-Spondin2 knockdown HSCs.
These findings suggest that R-Spondin2 may promote HSC activation by enhancing the canonical Wnt pathway.
肝纤维化的发展是许多肝脏疾病致命并发症的根本阶段，包括肝硬化和肝细胞癌。经典 Wnt 通路在多种生物过程中至关重要，介导肝纤维化的进展和消退。作为一种有效的 Wnt 通路激动剂，屋顶板特异性 Spondin-2（R-Spondin2）在肝纤维化中的作用尚未得到充分阐明。
本研究旨在探讨 R-Spondin2 是否有助于肝星状细胞（HSCs）激活，这是肝纤维化发生的关键事件。
使用人肝组织、肝纤维化小鼠模型和新鲜分离的小鼠 HSCs。通过 Western-blot 分析和实时 PCR 分别分析蛋白表达和转录水平。通过重组 R-Spondin2 的外源性刺激和 R-Spondin2 的敲低来研究其功能。分析核 β-连环蛋白水平和 T 细胞特异性转录因子活性，并通过 MTT 测定检测 HSC 增殖。
在人纤维性肝组织和肝纤维化小鼠模型中均观察到 R-Spondin2 的过表达。在新鲜分离的小鼠 HSCs 中，R-Spondin2 的外源性刺激诱导 Wnt 通路活性、HSC 增殖以及α-平滑肌肌动蛋白（α-SMA）和胶原 I 的表达呈剂量依赖性增加。此外，在 R-Spondin2 敲低的 HSCs 中，Wnt 通路活性、HSC 增殖以及α-SMA 和胶原 I 的表达降低。
这些发现表明，R-Spondin2 可能通过增强经典 Wnt 通路促进 HSC 激活。
