Hsa_circ_0048674 通过调控 miR-223-3p/PDL1 促进肝癌进展和自然杀伤细胞耗竭。
Hsa_circ_0048674 facilitates hepatocellular carcinoma progression and natural killer cell exhaustion depending on the regulation of miR-223-3p/PDL1.
机构信息
Tumor Center, First Affiliated Hospital of Guangzhou University of Traditional Chinese Medicine, Guangzhou, Guangdong, China.
Institute of Tumor, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, China.
出版信息
Histol Histopathol. 2022 Dec;37(12):1185-1199. doi: 10.14670/HH-18-440. Epub 2022 Feb 21.
BACKGROUND
Circular RNAs (circRNAs) play vital regulatory roles in human cancers, including hepatocellular carcinoma (HCC). In this study, we aimed to explore the functions of hsa_circ_0048674 in HCC development.
METHODS
Quantitative real-time polymerase chain reaction (qRT-PCR) assay was used to detect hsa_circ_0048674, ubiquitin-like with PHD and RING finger domains 1 (UHRF1), microRNA-223-3p (miR-223-3p) and programmed death ligand 1 (PDL1). RNase R assay and Actinomycin D assay were employed to analyze the stability of hsa_circ_0048674. Cell Counting Kit-8 (CCK-8) assay, colony formation assay and 5-ethynyl-2'- deoxyuridine (EdU) assay were conducted to assess cell proliferation. Flow cytometry analysis, transwell assay and tube formation assay were carried out for cell apoptosis, migration, invasion and angiogenesis, respectively. Western blot assay was adopted for protein levels. Dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were used to analyze the relationship between miR-223-3p and hsa_circ_0048674 or PDL1. Murine xenograft model assay was conducted for the function of hsa_circ_0048674 in vivo. Immunohistochemistry (IHC) assay was used to detect Ki-67 level in tumor tissues. Enzyme linked immunosorbent assay (ELISA) kits were employed for the concentrations of inflammatory factors.
RESULTS
Hsa_circ_0048674 was highly expressed in HCC tissues and cells. Silencing of hsa_circ_0048674 repressed cell growth, migration, invasion and angiogenesis and promoted apoptosis in HCC cells in vitro and hampered tumor growth in vivo. Hsa_circ_0048674 served as an miR-223-3p sponge to alter PDL1 expression. MiR-223-3p inhibition or PDL1 overexpression restored the impacts of hsa_circ_0048674 silencing on HCC malignant behaviors. In addition, hsa_circ_0048674 knockdown promoted natural killer (NK) cell-mediated cytotoxicity to HCC cells.
CONCLUSION
Hsa_circ_0048674 knockdown decelerated HCC progression through the mediation of the miR-223-3p/PDL1 axis.
背景
环状 RNA（circRNA）在人类癌症中发挥着重要的调控作用，包括肝细胞癌（HCC）。在本研究中，我们旨在探索 hsa_circ_0048674 在 HCC 发展中的功能。
方法
采用实时定量聚合酶链反应（qRT-PCR）检测 hsa_circ_0048674、泛素样含 PH 和环指域 1（UHRF1）、微小 RNA-223-3p（miR-223-3p）和程序性死亡配体 1（PDL1）。采用 RNase R 试验和放线菌素 D 试验分析 hsa_circ_0048674 的稳定性。细胞计数试剂盒-8（CCK-8）试验、集落形成试验和 5-乙炔基-2'-脱氧尿苷（EdU）试验用于评估细胞增殖。采用流式细胞术分析、Transwell 试验和管形成试验分别检测细胞凋亡、迁移、侵袭和血管生成。采用蛋白质印迹法检测蛋白水平。采用双荧光素酶报告基因试验和 RNA 免疫沉淀（RIP）试验分析 miR-223-3p 与 hsa_circ_0048674 或 PDL1 之间的关系。采用小鼠异种移植模型试验研究 hsa_circ_0048674 在体内的功能。免疫组化（IHC）试验用于检测肿瘤组织中 Ki-67 水平。酶联免疫吸附试验（ELISA）试剂盒用于检测炎症因子的浓度。
结果
hsa_circ_0048674 在 HCC 组织和细胞中高表达。沉默 hsa_circ_0048674 可抑制 HCC 细胞的体外生长、迁移、侵袭和血管生成，并促进凋亡，同时阻碍体内肿瘤生长。hsa_circ_0048674 作为 miR-223-3p 的海绵，改变 PDL1 的表达。抑制 miR-223-3p 或过表达 PDL1 可恢复 hsa_circ_0048674 沉默对 HCC 恶性行为的影响。此外，hsa_circ_0048674 敲低可促进自然杀伤（NK）细胞对 HCC 细胞的细胞毒性。
结论
hsa_circ_0048674 敲低通过 miR-223-3p/PDL1 轴抑制 HCC 进展。
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