Journal: Journal of Cardiothoracic Surgery

Article Title: NSUN2 inhibits NCOA4 expression to alleviate ferroptosis and inflammation in sepsis-induced myocardial injury in a m 5 C manner

doi: 10.1186/s13019-025-03554-z

Figure Lengend Snippet: NSUN2 suppressed ferroptosis in LPS-induced SIMI. Cellular A , Fe 2 + level, B , GSH content, and C , relative ROS production in each group were determined by commercial kits; D , Western blot was performed to assess the protein levels of NCOA4, FTH1, and GPX4 in each group Fe 2+ , ferrous iron; GSH, glutathione; ROS, reactive oxygen species; NCOA, nuclear receptor coactivator; FTH1, ferritin heavy chain; GPX, glutathione peroxidase

Article Snippet: The primary antibodies used were rabbit antibodies specific for NCOA4 (1:1000; ab314553; Abcam, Cambridge, MA, USA), FTH1 (1:1000; CSB- PA008485 ; Cusabio Engineering Co., LTD, Wuhan, China), GPX4 (1:1000; ab125066; Abcam), and GAPDH (1:1000; ab8245; Abcam).

Techniques: Western Blot

Journal: Journal of Cardiothoracic Surgery

Article Title: NSUN2 inhibits NCOA4 expression to alleviate ferroptosis and inflammation in sepsis-induced myocardial injury in a m 5 C manner

doi: 10.1186/s13019-025-03554-z

Figure Lengend Snippet: NSUN2 inhibited NCOA4 expression in a m 5 C-dependent manner. A , The expression of NCOA4, FTH1, and GPX4 after NSUN2 overexpression in H9c2 cells was detected by RT-qPCR; B , MeRIP-qPCR assay was performed to detect m 5 C levels of NCOA4, FTH1, and GPX4 in vector and NSUN2 groups; C , RIP assay evaluated the interaction of NSUN2 and NCOA4 in H9c2 cells; D , Random Forest method was used to predict possible m 5 C sites of NCOA4; Dual-luciferase gene reporter assay evaluated the binding of NSUN2 and NCOA4 at sites E , 501, F , 1138, and G , 2917; H , RNA stability assay was used to detect the NCOA4 expression when actinomycin D treated at different time points (0, 4, 8, and 12 h) after NSUN2 overexpression NCOA, nuclear receptor coactivator; FTH1, ferritin heavy chain; GPX, glutathione peroxidase; NSUN, NOL1/NOP2/SUN domain; RT-qPCR, reverse transcription-polymerase chain reaction; MeRIP, Methylated RNA immunoprecipitation; RIP, RNA immunoprecipitation; m 5 C, 5-methylcytosine

Article Snippet: The primary antibodies used were rabbit antibodies specific for NCOA4 (1:1000; ab314553; Abcam, Cambridge, MA, USA), FTH1 (1:1000; CSB- PA008485 ; Cusabio Engineering Co., LTD, Wuhan, China), GPX4 (1:1000; ab125066; Abcam), and GAPDH (1:1000; ab8245; Abcam).

Techniques: Expressing, Over Expression, Quantitative RT-PCR, Plasmid Preparation, Luciferase, Reporter Assay, Binding Assay, Stability Assay, Reverse Transcription, Polymerase Chain Reaction, Methylation, RNA Immunoprecipitation

Journal: Journal of Cardiothoracic Surgery

Article Title: NSUN2 inhibits NCOA4 expression to alleviate ferroptosis and inflammation in sepsis-induced myocardial injury in a m 5 C manner

doi: 10.1186/s13019-025-03554-z

Figure Lengend Snippet: Overexpressing NCOA4 downregulated cell viability and upregulated LDH activity, inflammation, and ferroptosis in LPS-induced SIMI. A , NCOA4 mRNA level was detected by RT-qPCR; B , The cell viability of H9c2 cells in each group was analyzed by CCK-8 assay; C , Determination of LDH enzyme activity in each group; D , The contents of TNF-α, IL-6, and IL-8 in each group was evaluated by ELISA; Cellular E , Fe 2 + level, F , GSH content, and G , relative ROS production in each group were determined by commercial kits; H , The protein levels of NCOA4, FTH1, and GPX4 in each group was analyzed by Western blot RT-qPCR, reverse transcription-polymerase chain reaction; CCK-8, cell counting kit-8; LDH, lactate dehydrogenase; ELISA, enzyme-linked immunosorbent assay; TNF-α, tumor necrosis factor-α; IL, interleukin; Fe 2+ , ferrous iron; GSH, glutathione; ROS, reactive oxygen species; NCOA, nuclear receptor coactivator; FTH1, ferritin heavy chain; GPX, glutathione peroxidase

Article Snippet: The primary antibodies used were rabbit antibodies specific for NCOA4 (1:1000; ab314553; Abcam, Cambridge, MA, USA), FTH1 (1:1000; CSB- PA008485 ; Cusabio Engineering Co., LTD, Wuhan, China), GPX4 (1:1000; ab125066; Abcam), and GAPDH (1:1000; ab8245; Abcam).

Techniques: Activity Assay, Quantitative RT-PCR, CCK-8 Assay, Enzyme-linked Immunosorbent Assay, Western Blot, Reverse Transcription, Polymerase Chain Reaction, Cell Counting