Ma Dong-Bo, Zhang Hui, Wang Xi-Ling, Wu Qiu-Ge
Department of Respiratory and Critical Care Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou City, China.
Kaohsiung J Med Sci. 2024 Aug;40(8):722-731. doi: 10.1002/kjm2.12843. Epub 2024 May 16.
Disruption of the alveolar barrier can trigger acute lung injury. This study elucidated the association of methyltransferase-like 3 (METTL3) with Streptococcus pneumoniae (SP)-induced apoptosis and inflammatory injury of alveolar epithelial cells (AECs). AECs were cultured and then infected with SP. Furthermore, the expression of METTL3, interleukin (IL)-10, IL-6, tumor necrosis factor-alpha (TNF-α), monocyte chemoattractant protein-1 (MCP-1), long noncoding RNA nuclear paraspeckle assembly transcript 1 (NEAT1), mucin 19 (MUC19), N6-methyladenosine (m6A), and NEAT1 after m6A modification were detected by qRT-PCR, Western blot, and enzyme-linked immunosorbent, m6A quantification, and methylated RNA immunoprecipitation-qPCR analyses, respectively. Moreover, the subcellular localization of NEAT1 was analyzed by nuclear/cytosol fractionation assay, and the binding between NEAT1 and CCCTC-binding factor (CTCF) was also analyzed. The results of this investigation revealed that SP-induced apoptosis and inflammatory injury in AECs and upregulated METTL3 expression. In addition, the downregulation of METTL3 alleviated apoptosis and inflammatory injury in AECs. METTL3-mediated m6A modification increased NEAT1 and promoted its binding with CTCF to facilitate MUC19 transcription. NEAT1 or MUC19 overexpression disrupted their protective role of silencing METTL3 in AECs, thereby increasing apoptosis and inflammatory injury. In conclusion, this is the first study to suggest that METTL3 aggravates SP-induced cell damage via the NEAT1/CTCF/MUC19 axis.
肺泡屏障的破坏可引发急性肺损伤。本研究阐明了甲基转移酶样3(METTL3)与肺炎链球菌(SP)诱导的肺泡上皮细胞(AECs)凋亡及炎性损伤之间的关联。培养AECs,然后用SP感染。此外,分别通过qRT-PCR、蛋白质免疫印迹法、酶联免疫吸附测定、m6A定量分析以及甲基化RNA免疫沉淀-qPCR分析检测METTL3、白细胞介素(IL)-10、IL-6、肿瘤坏死因子-α(TNF-α)、单核细胞趋化蛋白-1(MCP-1)、长链非编码RNA核旁斑组装转录本1(NEAT1)、粘蛋白19(MUC19)、N6-甲基腺苷(m6A)以及m6A修饰后的NEAT1的表达。此外,通过核/细胞质分级分离测定分析NEAT1的亚细胞定位,并分析NEAT1与CCCTC结合因子(CTCF)之间的结合情况。本研究结果显示,SP诱导AECs凋亡和炎性损伤,并上调METTL3表达。此外,METTL3的下调减轻了AECs的凋亡和炎性损伤。METTL3介导的m6A修饰增加了NEAT1的表达,并促进其与CTCF的结合以促进MUC19转录。NEAT1或MUC19的过表达破坏了它们在AECs中沉默METTL3的保护作用,从而增加了凋亡和炎性损伤。总之,这是第一项表明METTL3通过NEAT1/CTCF/MUC19轴加重SP诱导的细胞损伤的研究。