Colorectal cancer (CRC) is a global healthcare problem. Radioresistance is a huge setback for CRC radiotherapy. In this text, the roles and molecular mechanisms of long non-coding RNA HOTAIR in CRC tumorigenesis and radioresistance were further investigated. ATG12 mRNA, HOTAIR, and microRNA-93 (miR-93) levels were measured by quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay. Protein levels of LC3 I, LC3 II, p62, ATG12, cleaved caspase 3, Bax, and Bcl-2 were detected by western blotting assay in cells and were examined by immunohistochemistry (IHC) assay in tissues. Cell survival fractions, viability, and apoptotic rates were determined by clonogenic survival assay, CCK-8 assay, and flow cytometry analysis, respectively. The relationships of HOTAIR, miR-93, and ATG12 were tested by bioinformatics analysis and luciferase reporter assay. Mouse xenograft tumor models were established to investigate the influence of HOTAIR knockdown on CRC radioresistance in vivo. We found that HOTAIR expression was markedly upregulated in plasma from CRC patients after radiotherapy and CRC cells after irradiation. HOTAIR knockdown, miR-93 overexpression, or ATG12 silencing weakened cell viability, induced cell apoptosis, inhibited cell autophagy, and enhanced cell radiosensitivity in CRC. HOTAIR exerted its functions by downregulating miR-93. Moreover, HOTAIR functioned as a molecular sponge of miR-93 to regulate ATG12 expression. ATG12 protein expression was markedly upregulated and associated with miR-93 and HOTAIR expression in CRC tissues. Furthermore, HOTAIR knockdown enhanced radiosensitivity of CRC xenograft tumors by regulating miR-93/ATG12 axis. In conclusion, HOTAIR knockdown potentiated radiosensitivity through regulating miR-93/ATG12 axis in CRC, further elucidating the roles and molecular basis of HOTAIR in CRC radioresistance.

中文翻译：

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长期的非编码RNA HOTAIR组合可通过调节microRNA-93 / ATG12轴增强结直肠癌的放射敏感性。

大肠癌（CRC）是全球性的医疗保健问题。放射抵抗是CRC放射治疗的巨大挫折。本文进一步研究了长非编码RNA HOTAIR在CRC肿瘤发生和放射抵抗中的作用和分子机制。通过定量逆转录聚合酶链反应（RT-qPCR）分析来测量ATG12 mRNA，HOTAIR和microRNA-93（miR-93）的水平。通过蛋白质印迹法在细胞中检测LC3 I，LC3 II，p62，ATG12，裂解的胱天蛋白酶3，Bax和Bcl-2的蛋白水平，并在组织中通过免疫组织化学（IHC）检测。细胞存活分数，存活率和凋亡率分别通过克隆形成存活测定，CCK-8测定和流式细胞术分析来确定。HOTAIR，miR-93，通过生物信息学分析和萤光素酶报告基因检测法检测ATG12和ATG12。建立小鼠异种移植肿瘤模型以研究HOTAIR组合对体内CRC放射抗性的影响。我们发现，放疗后CRC患者的血浆和放疗后CRC细胞的HOTAIR表达明显上调。HOTAIR敲低，miR-93过表达或ATG12沉默会削弱细胞活力，诱导细胞凋亡，抑制细胞自噬并增强CRC中的细胞放射敏感性。HOTAIR通过下调miR-93发挥其功能。此外，HOTAIR充当miR-93的分子海绵来调节ATG12的表达。ATG12蛋白表达明显上调，并与CRC组织中的miR-93和HOTAIR表达相关。此外，HOTAIR基因敲低通过调节miR-93 / ATG12轴增强了CRC异种移植肿瘤的放射敏感性。总之，HOTAIR基因敲低通过调节miR-93 / ATG12轴在CRC中增强了放射敏感性，进一步阐明了HOTAIR在CRC放射抗性中的作用和分子基础。