Journal: Cell death & disease

Article Title: Investigation into the role of the MITA-TRIM38 interaction in regulating pyroptosis and maintaining immune tolerance at the maternal-fetal interface.

doi: 10.1038/s41419-023-06314-w

Figure Lengend Snippet: Fig. 1 Differential expression of pyroptosis-associated markers and macrophage subpopulations in decidual tissues between patients with URSA and control patients. A Differences in the protein expression levels of GSDMD, GSDMD-N, pro-caspase-1 and mature caspase-1 in decidual tissues between patients in the URSA group and the control group. B FCM analysis shows the content of CD14-labeled macrophages, FVS-labeled live/dead cells, and the contents of M1 macrophages labeled by CD86 and M2 macrophages labeled in CD209 in viable and dead cells in the decidual tissues in one patient with URSA and one control patient. a shows that the percentages of all the macrophages with CD14 (+) in the URSA and control groups are 5.02% and 1.32%, respectively; b shows that the percentages of CD14 (+) in FVS (−) and FVS (+) in the URSA and control groups are 53.6%, 42.1%, 28.4% and 38.2%; C shows that the percentages of macrophages showing the FVS (−) survival status represented by CD86 (+) M1 macrophages in the URSA group and control group are 38.5% and 33.3%, respectively; d shows that the percentages of macrophages showing the FVS (−) survival status represented by CD209 ( + ) M2 macrophages in the URSA and control groups are 31.2% and 71.5%, respectively; e shows that the percentages of macrophages showing the FVS (+) dead status represented by CD86 (+) M1 macrophages in the URSA group and control group are 45.4% and 30.4%; f shows that the percentages of macrophages showing the FVS (+) dead status represented by CD209 (+) M2 macrophages in the URSA group and control group are 22.5% and 55.2%. C FCM analysis of differences in total macrophage content in decidual tissues between patients in the URSA group and the control group. D FCM analysis of the proportion of macrophage survival and death in the uterine decidual tissues of patients in the URSA group and the control group. E FCM analysis of the difference in the proportion of M1 and M2 macrophages in the surviving and dead macrophages of the URSA group and the control group. *P < 0.05, **P < 0.01, ns = nonsignificant.

Article Snippet: For triple staining of CD86+ CD206 and TRIM38, the sections were treated with 3% bovine serum albumin at room temperature for 30min, followed by overnight incubation at 4 °C with the primary antihuman antibody CD206 (#60143-1-Ig, Proteintech, dilution 1:400).

Techniques: Quantitative Proteomics, Control, Expressing, Labeling

Journal: Cell death & disease

Article Title: Investigation into the role of the MITA-TRIM38 interaction in regulating pyroptosis and maintaining immune tolerance at the maternal-fetal interface.

doi: 10.1038/s41419-023-06314-w

Figure Lengend Snippet: Fig. 2 Differential expression of TRIM38 and MITA in decidual tissues between patients in the URSA group and the control group as determined by immunohistochemistry and immunofluorescence staining. A Immunohistochemistry shows the differences in MITA and TRIM38 expression in the uterine decidual tissue of patients with URSA and controls (brown part). B Triple immunofluorescence staining shows the distribution and expression differences of M1 macrophages (CD86 marker, red), M2 macrophages (CD206 marker, green) and TRIM38 (pink) in the uterine decidual tissue of patients with URSA and controls. C Immunofluorescence staining shows the differences in the distribution and expression differences of M1 macrophages (CD86 marker, red), M2 macrophages (CD206 marker, green) and MITA (pink) in the uterine decidual tissue of patients with URSA and controls. D Local magnification plot corresponding to B. E Local magnification plot corresponding to C.

Article Snippet: For triple staining of CD86+ CD206 and TRIM38, the sections were treated with 3% bovine serum albumin at room temperature for 30min, followed by overnight incubation at 4 °C with the primary antihuman antibody CD206 (#60143-1-Ig, Proteintech, dilution 1:400).

Techniques: Quantitative Proteomics, Control, Immunohistochemistry, Staining, Expressing, Marker

Journal: Cell death & disease

Article Title: Investigation into the role of the MITA-TRIM38 interaction in regulating pyroptosis and maintaining immune tolerance at the maternal-fetal interface.

doi: 10.1038/s41419-023-06314-w

Figure Lengend Snippet: Fig. 7 The relationship between pyroptosis and K48. A Morphology and number of pyroptotic cells in the supernatants of wild-type M1, M1 with DMSO (M1-vehicle), M1 with VX765 (M1-VX765) and wild-type M2 macrophages under a ×40 microscope. B Differences in the protein expression levels of GSDMD-N, caspase-1 and cleaved caspase-1 in the supernatants of M1, M1 with DMSO (M1-vehicle), M1 with VX765 (M1- VX765) and M2 macrophages. C Differences in the protein expression levels of MITA and cGAS in M1 macrophages, M1 macrophages treated with DMSO (M1-vehicle), M1 macrophages treated with VX765 (M1-VX765) and M2 macrophages. D Differential expression of MITA by ubiquitination of type K48 in M1, M1 with DMSO (M1-vehicle) and M1 with VX765 (M1-VX765) macrophages. E FCM analysis shows wild-type M1, M1 with DMSO (M1-vehicle), M1 with VX765 (M1-VX765) of FVS-labeled live cells, and the content of macrophages labeled by CD86 or CD209 in viable cells in the above three kinds of cells. a–c shows that the percentages of FVS (−) in M1, M1-vehicle and M1-VX765 are 59.5%, 58.9% and 58.2%, respectively. d–f shows that the percentages of macrophages showing the FVS (−) survival status represented by CD86 (+) macrophages in M1, M1-vehicle and M1-VX765 are 49.1%, 48.2% and 42.5%, respectively. g–i shows that the percentages of macrophages showing the FVS (−) survival status represented by CD209 (+) macrophages in M1, M1-vehicle and M1-VX765 are 0.09%, 0.10% and 0.86%, respectively. F The phenotypes of CD86 and CD209 in wild-type M1, M1-vehicle, and M1-VX765 cells were analyzed by FCM. *P < 0.05, ****P < 0.0001, ns = nonsignificant.

Article Snippet: For triple staining of CD86+ CD206 and TRIM38, the sections were treated with 3% bovine serum albumin at room temperature for 30min, followed by overnight incubation at 4 °C with the primary antihuman antibody CD206 (#60143-1-Ig, Proteintech, dilution 1:400).

Techniques: Microscopy, Expressing, Quantitative Proteomics, Ubiquitin Proteomics, Labeling

Journal: Cell death & disease

Article Title: Investigation into the role of the MITA-TRIM38 interaction in regulating pyroptosis and maintaining immune tolerance at the maternal-fetal interface.

doi: 10.1038/s41419-023-06314-w

Figure Lengend Snippet: Fig. 9 Effect of lentiviral knockdown of TRIM38 or MITA on the polarization efficiency of macrophages. A FCM analysis shows the wild- type M1, shTRIM38-M1, shMITA-M1 of FVS-labeled live cells, and the content of macrophages labeled by CD86 in viable cells in the above three kinds of cells, respectively. a–c shows the percentage of FVS (−) in M1, shTRIM38-M1, and shMITA-M1 are 75.0%, 75.7% and 74.7%, respectively. d–f shows that the percentages of macrophages showing the FVS (−) survival status represented by CD86 (+) macrophages in M1, shTRIM38-M1, and shMITA-M1 are 76.5%, 86.4% and 65.3%, respectively. B The phenotypes of FVS(−) and surviving macrophages with CD86(+) in wild-type M1, shTRIM38-M1, and shMITA-M1 cells were analyzed by FCM. **P < 0.01, ns = nonsignificant. C FCM analysis shows the wild-type M2, shTRIM38-M2, and shMITA-M2 FVS-labeled live cells and the content of macrophages labeled by CD209 in viable cells in the above three kinds of cells; a–c shows that the percentages of FVS (−) in M2, shTRIM38-M2, and shMITA-M2 are 74.9%, 72.4% and 72.9%, respectively; d–f shows the percentage of macrophages showing the FVS (−) survival status represented by CD209 (+) macrophages in M2, shTRIM38-M2, and shMITA-M2 are 50.9%, 32.7% and 76.6%, respectively. D The phenotypes of FVS(−) and surviving macrophages with CD209(+) in wild-type M2, shTRIM38-M2, and shMITA-M2 cells were analyzed by FCM. ***P < 0.001, ns nonsignificant.

Article Snippet: For triple staining of CD86+ CD206 and TRIM38, the sections were treated with 3% bovine serum albumin at room temperature for 30min, followed by overnight incubation at 4 °C with the primary antihuman antibody CD206 (#60143-1-Ig, Proteintech, dilution 1:400).

Techniques: Knockdown, Labeling

Journal: bioRxiv

Article Title: GREM1 is epigenetically reprogrammed in muscle cells after exercise training and controls myogenesis and metabolism

doi: 10.1101/2020.02.20.956300

Figure Lengend Snippet: a , Schematic of experimental setup. TA muscles of adult mice were injured by cardiotoxin (CTX) injection, and recombinant GREM1 protein or anti-GREM1 blocking antibodies (GREM1Ab) were administrated 3 days post injury. b , Quantification of the number of PAX7+ cells on cryosections generated 7 days after injury in regenerating TA muscles. c , Immunolocalization of dystrophin (green) and TCF4 (red) proteins on regenerated TA muscles (14 days post injury). d , Immunolocalization of PAX7 (green) and laminin (red) proteins on 14 days post injury in regenerated TA muscles. e , Quantification of regenerated myofiber cross-sectional area. f , Quantification of area covered by Collagen. g , Quantification of muscle satellite cell (MuSCs) self-renewal. Nuclei are stained with Hoechst (blue). Scale bars, 100μm (C), 50μm (D). n = 3 or more for each experimental point. Error bars indicate ±SD. *p<0.05; **p< 0.01; ***p< 0.001. n.s.: not statistically significant.

Article Snippet: Primary antibodies used were against Dystrophin (Leica), embryonic MyHC (Santa Cruz), Grem1 (R&D Systems), Laminin (Santa Cruz), Pax7 (Santa Cruz), Tcf4 (Cell Signaling Tech.).

Techniques: Injection, Recombinant, Blocking Assay, Generated, Staining