anti tr4  (R&D Systems)

Journal: European Journal of Medical Research

Article Title: TR4 worsen urosepsis by regulating GSDMD

doi: 10.1186/s40001-024-01742-6

Figure Lengend Snippet: TR4 is associated with the sepsis prognosis. A Normalize the gene expression values of different samples by log2 transformation. B Principal component analysis (PCA) was performed on different samples. C Volcano plot of differentially expressed genes; red dots represent upregulated genes and blue dots represent downregulated genes. The fold change was set as ≥ 2 or ≤ 0.5, p < 0.05. D Heatmap of the top 50 differentially expressed genes. E Expression of TR4 in the sepsis survival and death groups. N = 10, * p < 0.05. F Relative mRNA expression of TR4 in the sepsis and nonsepsis groups. N = 20, * p < 0.05

Article Snippet: After blocking with nonfat milk on the incubator shaker for 1 h at room temperature, the membranes were incubated with primary antibodies at 4 °C overnight, which included GAPDH (1:1000, sc-202525, Santa Cruz, CA, USA), GSDMD (1:1000, A18281, ABclonal, Wuhan, China), and TR4 (1:1000, ab109301, Abcam, USA).

Techniques: Expressing, Transformation Assay

Journal: European Journal of Medical Research

Article Title: TR4 worsen urosepsis by regulating GSDMD

doi: 10.1186/s40001-024-01742-6

Figure Lengend Snippet: TR4 worsens the outcomes of urosepsis. A HE staining of the kidney, liver, lung, and intestine of TR4 knockdown mice and TR4 WT mice induced by E. coli bacteria. B Quantification of the injury score based on H&E-stained sections of WT mice and TR4 ± mice; N = 5; * p < 0.05, ** p < 0.01, *** p < 0.001. C Overall survival time of the WT mice and TR4 ± mice; N = 10

Article Snippet: After blocking with nonfat milk on the incubator shaker for 1 h at room temperature, the membranes were incubated with primary antibodies at 4 °C overnight, which included GAPDH (1:1000, sc-202525, Santa Cruz, CA, USA), GSDMD (1:1000, A18281, ABclonal, Wuhan, China), and TR4 (1:1000, ab109301, Abcam, USA).

Techniques: Staining, Bacteria

Journal: European Journal of Medical Research

Article Title: TR4 worsen urosepsis by regulating GSDMD

doi: 10.1186/s40001-024-01742-6

Figure Lengend Snippet: TR4 promotes inflammatory factor secretion during urosepsis. A THP-1 cells can be induced to differentiate into M0 macrophages after treatment with 100 ng/ml PMA for 48 h, and M0 macrophages, which are round or oval in shape, can be observed under a microscope. The M0 cells were then treated with 500 ng/ml LPS for 48 h to induce M1 macrophages. Under the microscope, irregular cell morphology and obvious cell protrusions could be observed. B After LPS stimulation, the expression levels of TR4, TNFα, and IL-6 in macrophages were significantly increased. When TR4 was knocked down, the expression levels of TR4, TNFα, and IL-6 in macrophages decreased. * p < 0.05, ** p < 0.01, *** p < 0.001. C After knocking down TR4, the expression levels of MCP1, IL-6, TNFα, and IL-1β were significantly decreased; ** p < 0.01. D After overexpressing TR4, the expression levels of MCP1, IL-6, TNFα, and IL-1β were significantly increased; * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: After blocking with nonfat milk on the incubator shaker for 1 h at room temperature, the membranes were incubated with primary antibodies at 4 °C overnight, which included GAPDH (1:1000, sc-202525, Santa Cruz, CA, USA), GSDMD (1:1000, A18281, ABclonal, Wuhan, China), and TR4 (1:1000, ab109301, Abcam, USA).

Techniques: Microscopy, Expressing

Journal: European Journal of Medical Research

Article Title: TR4 worsen urosepsis by regulating GSDMD

doi: 10.1186/s40001-024-01742-6

Figure Lengend Snippet: TR4 promotes pyroptosis during urosepsis. A Propidium iodide (PI; red indicates dead cells) and Hoechst (blue indicates all nuclei) double staining were used to observe the morphology of pyroptotic cell, and the numbers of pyroptotic cells were compared; * p < 0.05, ** p < 0.01, *** p < 0.001. B Comparison of the IL-1β concentration in the culture supernatant of control and TR4-overexpressing macrophages; * p < 0.05, ** p < 0.01, *** p < 0.001. C Comparison of 24-h pyroptotic cell numbers in control and TR4-overexpressing macrophages

Article Snippet: After blocking with nonfat milk on the incubator shaker for 1 h at room temperature, the membranes were incubated with primary antibodies at 4 °C overnight, which included GAPDH (1:1000, sc-202525, Santa Cruz, CA, USA), GSDMD (1:1000, A18281, ABclonal, Wuhan, China), and TR4 (1:1000, ab109301, Abcam, USA).

Techniques: Double Staining, Comparison, Concentration Assay

Journal: European Journal of Medical Research

Article Title: TR4 worsen urosepsis by regulating GSDMD

doi: 10.1186/s40001-024-01742-6

Figure Lengend Snippet: TR4 regulates the expression of GSDMD. A The expression of GSDMD mRNA was changed when TR4 was overexpressed or knocked down, * p < 0.05, ** p < 0.01, *** p < 0.001. B Changes in the protein level of GSDMD when TR4 was overexpressed or knocked down. The quantification of TR4 and GSDMD expression based on band density is shown in the right panel. C Correlation between TR4 and GSDMD expression in blood cells. D The upper picture shows the motif structure of TR4 predicted by the website, and the lower picture shows the promoter region of GSDMD to which TR4 may bind, which was predicted by the hTFtarget website. E ChIP experiments verified that TR4 binds to the GSDMD promoter region (chr8:143553465–143,553867); ** p < 0.01

Article Snippet: After blocking with nonfat milk on the incubator shaker for 1 h at room temperature, the membranes were incubated with primary antibodies at 4 °C overnight, which included GAPDH (1:1000, sc-202525, Santa Cruz, CA, USA), GSDMD (1:1000, A18281, ABclonal, Wuhan, China), and TR4 (1:1000, ab109301, Abcam, USA).

Techniques: Expressing

Journal: European Journal of Medical Research

Article Title: TR4 worsen urosepsis by regulating GSDMD

doi: 10.1186/s40001-024-01742-6

Figure Lengend Snippet: Hypothetical model by which TR4-regulated macrophage pyroptosis promotes urosepsis via GSDMD. The mechanism map of the TR4 acts as a key regulator of GSDMD gene expression and pyroptosis in macrophages

Article Snippet: After blocking with nonfat milk on the incubator shaker for 1 h at room temperature, the membranes were incubated with primary antibodies at 4 °C overnight, which included GAPDH (1:1000, sc-202525, Santa Cruz, CA, USA), GSDMD (1:1000, A18281, ABclonal, Wuhan, China), and TR4 (1:1000, ab109301, Abcam, USA).

Techniques: Expressing