BACKGROUND MicroRNA-612 (miR-612) has been proven to suppress EMT, stemness, and tumor metastasis of hepatocellular carcinoma (HCC) via PI3K/AKT2 and Sp1/Nanog signaling. However, its biological roles on HCC progression are far from elucidated. METHODS We found direct downstream target of miR-612, hadha by RNA immunoprecipitation and sequencing. To explore its biological characteristic, potential molecular mechanism, and clinical relevance in HCC patients, we performed several in-vitro and in-vivo models, as well as human tissue chip. RESULTS Ectopic expression of miR-612 could partially reverse the level of HADHA, then suppress function of pseudopods, and diminish metastatic and invasive potential of HCC by lipid reprogramming. In detail, miR-612 might reduce invadopodia formation via HADHA-mediated cell membrane cholesterol alteration and accompanied with the inhibition of Wnt/β-catenin regulated EMT occurrence. Our results showed that the maximum oxygen consumption rates (OCR) of HCCLM3miR-612-OE and HCCLM3hadha-KD cells were decreased nearly by 40% and 60% of their counterparts (p < 0.05). The levels of acetyl CoA were significantly decreased, about 1/3 (p > 0.05) or 1/2 (p < 0.05) of their controls, in exogenous miR-612 or hadha-shRNA transfected HCCLM3 cell lines. Besides, overexpression of hadha cell lines had a high expression level of total cholesterol, especially 27-hydroxycholesterol (p < 0.005). SREBP2 protein expression level as well as its downstream targets, HMGCS1, HMGCR, MVD, SQLE were all deregulated by HADHA. Meanwhile, the ATP levels were reduced to 1/2 and 1/4 in HCCLM3miR-612-OE (p < 0.05) and HCCLM3hadha-KD (p < 0.01) respectively. Moreover, patients with low miR-612 levels and high HADHA levels had a poor prognosis with shorter overall survival. CONCLUSION miR-612 can suppress the formation of invadopodia, EMT, and HCC metastasis and by HADHA-mediated lipid programming, which may provide a new insight of miR-612 on tumor metastasis and progression.

中文翻译：

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MiR-612通过HADHA介导的脂质重编程调节肝细胞癌的侵袭。

背景技术已证明MicroRNA-612（miR-612）可通过PI3K / AKT2和Sp1 / Nanog信号传导抑制肝细胞癌（HCC）的EMT，干性和肿瘤转移。但是，其在肝癌进展中的生物学作用尚不清楚。方法我们通过RNA免疫沉淀和测序发现了miR-612的直接下游靶标hadha。为了探索其在肝癌患者中的生物学特性，潜在的分子机制和临床相关性，我们进行了几种体外和体内模型以及人体组织芯片的研究。结果miR-612的异位表达可以部分逆转HADHA的水平，然后抑制拟足细胞的功能，并通过脂质重编程降低HCC的转移和侵袭潜能。详细，miR-612可能通过HADHA介导的细胞膜胆固醇改变减少内脏足形成，并伴随Wnt /β-catenin调控的EMT发生的抑制作用。我们的结果表明，HCCLM3miR-612-OE和HCCLM3hadha-KD细胞的最大耗氧率（OCR）分别降低了40％和60％（p <0.05）。在外源性miR-612或hadha-shRNA转染的HCCLM3细胞系中，乙酰辅酶A的水平显着降低，约为其对照的1/3（p> 0.05）或1/2（p <0.05）。此外，hadha细胞系的过表达具有较高的总胆固醇表达水平，尤其是27-羟基胆固醇（p <0.005）。SREBP2蛋白表达水平及其下游靶标HMGCS1，HMGCR，MVD，SQLE均已被HADHA调节。与此同时，HCCLM3miR-612-OE（p <0.05）和HCCLM3hadha-KD（p <0.01）的ATP水平分别降低到1/2和1/4。而且，miR-612水平低和HADHA水平高的患者预后较差，总生存期较短。结论miR-612可以抑制恶性足病，EMT和HCC转移的形成，并且可以通过HADHA介导的脂质编程来抑制，这可能为miR-612提供关于肿瘤转移和进展的新见识。