Liu Shiying, Yin Yue, Yu Ruili, Li Yin, Zhang Weizhen
Department of Physiology and Pathophysiology, Peking University Health Science Center, Beijing 100191, China.
Department of Physiology and Pathophysiology, Peking University Health Science Center, Beijing 100191, China; Department of Surgery, University of Michigan Medical Center, Ann Arbor, MI 48109-0346, USA.
Biochem Biophys Res Commun. 2018 Apr 30;499(1):59-65. doi: 10.1016/j.bbrc.2018.03.126. Epub 2018 Mar 19.
BACKGROUND & AIMS: Leucine-rich repeat G-protein-coupled receptor 4 (LGR4) and its ligands R-spondin1-4 (Rspos) have been vastly investigated in embryonic development. The biological functions of Rspos-LGR4 system in liver remains largely unknown. Here, we explored whether it protects hepatocytes against hypoxia/reoxygenation (H/R) induced damage.
H/R injury was induced by dimethyloxalylglycine (DMOG) in AML12 cells and the effects of Rspo3 on cell proliferation and apoptosis were assessed. Specific shRNAs were used to interfere LGR4 or β-catenin.
DMOG caused hepatocytes damage evidenced by increase in HIF-1α, cell death and apoptosis genes p27 and Bax, with concurrent decrease of cell proliferation genes PCNA and CyclinD1. Of all the Rspos, Rspo3 is predominantly expressed in AML12 hepatocytes. Importantly, Rspo3 demonstrated an alteration in a manner similar to proliferation-related genes during H/R injury. Rspo3 pretreatment rendered hepatocytes less vulnerable to DMOG induced H/R injury. Ablation of LGR4 using shRNA attenuated the protective effects of Rspo3. Wnt3a also protected AML12 cells from damages caused by H/R, showing enhanced proliferation activity. Notably, knockdown of β-catenin in hepatocytes completely abolished the effect of Rspo3 pretreatment on the expression levels of PCNA and CyclinD1.
Rspo3-LGR4 axis protects hepatocytes from H/R injury via activating β-catenin.
富含亮氨酸重复序列的G蛋白偶联受体4（LGR4）及其配体R-spondin1-4（Rspos）在胚胎发育中已得到广泛研究。Rspos-LGR4系统在肝脏中的生物学功能仍 largely未知。在此，我们探讨了它是否能保护肝细胞免受缺氧/复氧（H/R）诱导的损伤。
用二甲基草酰甘氨酸（DMOG）诱导AML12细胞发生H/R损伤，并评估Rspo3对细胞增殖和凋亡的影响。使用特异性短发夹RNA（shRNAs）干扰LGR4或β-连环蛋白。
DMOG导致肝细胞损伤，表现为缺氧诱导因子-1α（HIF-1α）增加、细胞死亡以及凋亡相关基因p27和Bax升高，同时细胞增殖相关基因增殖细胞核抗原（PCNA）和细胞周期蛋白D1（CyclinD1）降低。在所有的Rspos中，Rspo3在AML12肝细胞中表达最为显著。重要的是，在H/R损伤期间，Rspo3的变化方式与增殖相关基因相似。Rspo3预处理使肝细胞对DMOG诱导的H/R损伤更具抵抗力。用shRNA敲除LGR4减弱了Rspo3的保护作用。Wnt3a也能保护AML12细胞免受H/R引起的损伤，表现出增强的增殖活性。值得注意的是，敲低肝细胞中的β-连环蛋白完全消除了Rspo3预处理对PCNA和CyclinD1表达水平的影响。
Rspo3-LGR4轴通过激活β-连环蛋白保护肝细胞免受H/R损伤。
