Hepatocellular carcinoma (HCC) is still the most common cause of cancer-related mortality worldwide and accumulating studies report that HCC is frequently linked to chronic inflammation. G-protein alpha-subunit (GNAS)-activating mutations have recently been reported to form a rare subgroup of inflammatory liver tumors. In this study, we investigated the roles of GNAS in inflammation-related HCC progression and its underlying mechanism. Lipopolysaccharides (LPS) and diethylnitrosamine were employed to stimulate HCC cells to an induced inflammatory response. qRT-PCR, immunohistochemistry and immunoblotting were performed to detect the expression of GNAS in HCC tissues and cell lines. Expression levels of proinflammatory cytokines were detected by qRT-PCR and ELISA. N6-methyladenosine (m6A) methylation of GNAS mRNA was detected by RNA-binding protein immunoprecipitation (RIP). Transcription factors activation profiling plate array was performed to investigate the underlying mechanism in GNAS promoting interleukin-6 (IL-6) expression in HCC cells. HCC cell invasion was determined by transwell assay in vitro, and tumorigenesis was assessed with a subcutaneous xenograft mouse model of HCC. We found that LPS stimulation promotes GNAS expression in HCC cells through increasing m6A methylation of GNAS mRNA. The high expression level of GNAS promotes LPS-induced HCC cell growth and invasion by interacting with signal transducer and activator of transcription 3 (STAT3). Furthermore, GNAS knockdown inhibits LPS induced-IL-6 expression in HCC cells by suppressing STAT3 activation. Moreover, we found that GNAS promotes LPS-induced STAT3 activation in HCC cells through inhibiting long non-coding RNA TPTEP1 interacting with STAT3. In addition, GNAS expression promotes HCC development in mice and is related to poor survival. Our findings for the first time indicate a tumor-promoting role of GNAS in inflammation-related HCC progression and provide a novel potential target for HCC therapy.

中文翻译：

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GNAS 通过促进 STAT3 激活来促进炎症相关的肝细胞癌进展。

肝细胞癌 (HCC) 仍然是全球癌症相关死亡的最常见原因，并且越来越多的研究报告称，HCC 经常与慢性炎症有关。最近有报道称 G 蛋白 α 亚基 (GNAS) 激活突变形成了一种罕见的炎症性肝肿瘤亚组。在这项研究中，我们研究了 GNAS 在炎症相关 HCC 进展中的作用及其潜在机制。脂多糖 (LPS) 和二乙基亚硝胺用于刺激 HCC 细胞诱导炎症反应。采用qRT-PCR、免疫组化和免疫印迹法检测GNAS在HCC组织和细胞系中的表达。通过qRT-PCR和ELISA检测促炎细胞因子的表达水平。通过 RNA 结合蛋白免疫沉淀 (RIP) 检测 GNAS mRNA 的 N6-甲基腺苷 (m6A) 甲基化。进行转录因子激活谱板阵列以研究 GNAS 促进 HCC 细胞中白细胞介素 6 (IL-6) 表达的潜在机制。HCC 细胞侵袭在体外通过 transwell 测定法确定，并用 HCC 的皮下异种移植小鼠模型评估肿瘤发生。我们发现 LPS 刺激通过增加 GNAS mRNA 的 m6A 甲基化促进 HCC 细胞中的 GNAS 表达。GNAS 的高表达水平通过与信号转导和转录激活因子 3 (STAT3) 相互作用促进 LPS 诱导的 HCC 细胞生长和侵袭。此外，GNAS 敲低通过抑制 STAT3 激活来抑制 LPS 诱导的 HCC 细胞中的 IL-6 表达。而且，我们发现 GNAS 通过抑制长链非编码 RNA TPTEP1 与 STAT3 相互作用来促进 LPS 诱导的 HCC 细胞中的 STAT3 活化。此外，GNAS 表达促进小鼠 HCC 的发展，并与较差的存活率有关。我们的研究结果首次表明 GNAS 在炎症相关 HCC 进展中的促肿瘤作用，并为 HCC 治疗提供了一个新的潜在靶点。