Li Xiu-Qin, Peng Wen-Ting, Shan Shan, Wu Jing-Jing, Li Nan, Du Jia-Jia, Sun Jia-Chang, Chen Ting-Ting, Wei Wei, Sun Wu-Yi
Institute of Clinical Pharmacology, Anhui Medical University, Key Laboratory of Anti-inflammatory and Immune Medicine, Ministry of Education, Anhui Collaborative Innovation Center of Anti-inflammatory and Immune Medicine, Hefei 230032, China.
J Cancer. 2021 Oct 28;12(24):7287-7299. doi: 10.7150/jca.59291. eCollection 2021.
β-arrestin2 and β2-adrenergic receptor (β2-AR) have important roles in malignant tumors, the present study aims to investigate the role of activated β2-AR in hepatic stellate cells (HSCs) during hepatocellular carcinoma (HCC) progression and the regulatory effect of β-arrestin2. Immunofluorescence and Western blot were used to detect the expression of β-arrestin2 and β2-AR in HSCs of liver tissues from human HCC samples and diethylnitrosamine (DEN)-induced HCC model mice. We next used β-arrestin2 mice to demonstrate the regulatory role of β-arrestin2 in DEN mice. The subsets of T cells were quantified by flow cytometry. MTT and wound healing assay were applied to detect the proliferation and migration of cells. Co-immunoprecipitation assay was used to detect the link of β-arrestin2 and β2-AR in HSCs. Effect of β-arrestin2 overexpression on β2-AR downstream signaling pathway was verified by Western blot. The secretion of CCL2 was detected by ELISA. The expression of β2-AR was significantly increased, while β-arrestin2 was decreased in HSCs of HCC tissues. And β-arrestin2 deficiency exacerbates DEN-induced HCC accompanied with increased β2-AR expression. The results of flow cytometry showed that the percentage of activated T cells decreased gradually after DEN injection. β-arrestin2 knockout down-regulated the ratio of activated T cells. , selective activation of β2-AR in HSCs promoted the proliferation and migration of HCC cells. β-arrestin2 overexpression enhanced co-immunoprecipitation of β-arrestin2 and β2-AR in activated HSCs, and decreased its downstream Akt phosphorylation. Akt inhibitor decreased secretion of CCL2 in activated HSCs. Our study demonstrated that β2-AR activation in HSCs induces the proliferation and migration of HCC cells may be through Akt signaling, and this effect appears to be regulated by β-arrestin2.
β-抑制蛋白2和β2-肾上腺素能受体（β2-AR）在恶性肿瘤中具有重要作用，本研究旨在探讨活化的β2-AR在肝细胞癌（HCC）进展过程中对肝星状细胞（HSCs）的作用以及β-抑制蛋白2的调节作用。采用免疫荧光和蛋白质印迹法检测人HCC样本和二乙基亚硝胺（DEN）诱导的HCC模型小鼠肝组织HSCs中β-抑制蛋白2和β2-AR的表达。接下来，我们使用β-抑制蛋白2基因敲除小鼠来证明β-抑制蛋白2在DEN小鼠中的调节作用。通过流式细胞术对T细胞亚群进行定量分析。采用MTT法和伤口愈合实验检测细胞的增殖和迁移能力。采用免疫共沉淀实验检测HSCs中β-抑制蛋白2与β2-AR的相互作用。通过蛋白质印迹法验证β-抑制蛋白2过表达对β2-AR下游信号通路的影响。采用酶联免疫吸附测定（ELISA）法检测CCL2的分泌水平。HCC组织的HSCs中β2-AR表达显著增加，而β-抑制蛋白2表达降低。并且β-抑制蛋白2缺乏会加剧DEN诱导的HCC，同时伴有β2-AR表达增加。流式细胞术结果显示，注射DEN后活化T细胞百分比逐渐降低。β-抑制蛋白2基因敲除下调了活化T细胞的比例。HSCs中β2-AR的选择性激活促进了HCC细胞的增殖和迁移。β-抑制蛋白2过表达增强了活化HSCs中β-抑制蛋白2与β2-AR的免疫共沉淀，并降低了其下游Akt磷酸化水平。Akt抑制剂降低了活化HSCs中CCL2的分泌。我们的研究表明，HSCs中β2-AR的激活可能通过Akt信号通路诱导HCC细胞的增殖和迁移，并且这种作用似乎受β-抑制蛋白2的调节。
