Department of General Surgery, The First Affiliated Hospital of Soochow University, Suzhou, China.
Department of Orthopedics, The First Affiliated Hospital of Soochow University, Suzhou, China.
Cancer Sci. 2022 Feb;113(2):634-647. doi: 10.1111/cas.15238. Epub 2021 Dec 22.
Noncoding RNAs have been verified to regulate the infiltration of macrophages to accelerate tumor biological progression, however the regulation of macrophages by circular RNAs in hepatocellular carcinoma (HCC) remains unresolved. Using high-throughput RNA sequencing, we demonstrated that hsa_circ_0003410 was clearly upregulated in HCC. 5-Ethynyl-2'-deoxyuridine and transwell assays showed that hsa_circ_0003410 facilitated the proliferation and migration of HCC cells in vitro. We knocked down the expression of hsa_circ_0003410 in HepG2 cells and performed next-generation sequencing to determine possible target genes of hsa_circ_0003410. Kyoto Encyclopedia of Genes and Genomes analysis revealed that different genes were mainly enriched in immune-related pathways. Mechanistically, we identified CCL5 as the target gene of hsa_circ_0003410. RNA-FISH showed the co-expression of hsa_circ_0003410 and CCL5. Western blot and ELISA also verified that hsa_circ_0003410 could upregulate the expression of CCL5 protein. Flow cytometry and immunofluorescence assays indicated that CCL5 activated and recruited M2 macrophages and increased the ratio of M2/M1 macrophages to promote the progression of HCC. Animal experiments in vitro also confirmed our results. Taken together, our experiments revealed that noncoding RNAs play a critical role in the HCC microenvironment and can be considered as markers for the diagnosis and prognosis of HCC.
非编码 RNA 已被证实可调节巨噬细胞浸润，从而加速肿瘤的生物学进展，但环状 RNA 对肝癌（HCC）中巨噬细胞的调节作用仍未得到解决。通过高通量 RNA 测序，我们证实 hsa_circ_0003410 在 HCC 中明显上调。5-乙炔基-2'-脱氧尿苷和 Transwell 分析表明，hsa_circ_0003410 促进了 HCC 细胞在体外的增殖和迁移。我们敲低了 HepG2 细胞中 hsa_circ_0003410 的表达，并进行了下一代测序，以确定 hsa_circ_0003410 的可能靶基因。京都基因与基因组百科全书分析显示，不同的基因主要富集在免疫相关途径中。从机制上讲，我们确定 CCL5 是 hsa_circ_0003410 的靶基因。RNA-FISH 显示 hsa_circ_0003410 和 CCL5 的共表达。Western blot 和 ELISA 也验证了 hsa_circ_0003410 可以上调 CCL5 蛋白的表达。流式细胞术和免疫荧光分析表明，CCL5 激活并募集 M2 巨噬细胞，增加 M2/M1 巨噬细胞的比例，从而促进 HCC 的进展。体外动物实验也证实了我们的结果。总之，我们的实验揭示了非编码 RNA 在 HCC 微环境中发挥着关键作用，可以作为 HCC 诊断和预后的标志物。
