Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Expression Was Significantly Increased After MI (A, B) Quantification and representative images of Western blot analyses showing Trim55 protein expression in the border zone of myocardium at 3, 7, and 28 days after MI (n = 3). (C) Quantification of Trim55 mRNA levels in the border zone of myocardium at 3, 7, and 28 days after MI (n = 3). (D, E) Quantification and representative images of Trim55 immunohistochemistry in the border zone of myocardium at 3 and 28 days after MI (n = 3; scale bar: 50 μm). (F, G) Quantification and representative images of Western blot analyses showing Trim55 protein expression in NRCM cells after 400 μmol/L stimulation at 24 hours (n = 3). (H) Quantification of Trim55 mRNA levels in NRCM cells after CoCl 2 stimulation (400 μmol/L; 24 hours; n = 3). All data are expressed as mean ± SEM. Data in B, C, E, G, and H were analyzed using Student’s t -test. ∗ P < 0.05, ∗∗ P < 0.01 vs the control or sham group. CoCl 2 = cobalt chloride; CON = control; GADPH = glyceraldehyde-3-phosphate dehydrogenase; IHC = immunohistochemistry; MI = myocardial infarction; mRNA = messenger RNA; NRCM = neonatal rat cardiomyocyte; Trim55 = tripartite motif-containing 55.

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Expressing, Western Blot, Immunohistochemistry, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Knockout Alleviated Myocardial Injury and Cardiomyocyte Apoptosis After MI (A) Echocardiographic analysis of the LVEF and FS at 28 days after MI in the C57-sham (n = 10), Trim55 –/– -sham (n = 10), C57-MI (n = 10), and Trim55 –/– -MI (n = 10) groups. (B) Quantitative analysis of the ratio of heart weight to body weight at 28 days after MI in the C57-sham (n = 10), Trim55 –/– -sham (n = 10), C57-MI (n = 8), and Trim55 –/– -MI (n = 10) groups. (C, D) Representative HE and Masson staining pictures and quantification of the infarction area at 28 days after MI in the C57-sham, Trim55 –/– -sham, C57-MI, and Trim55 –/– -MI groups (n = 3). (E, F) Quantification and representative images of Western blot analyses showing the Trim55, collagen I, and TGF-β protein expression in the C57-sham, Trim55 –/– -sham, C57-MI, and Trim55 –/– -MI groups (n = 3). (G, H) Quantification and representative images of Western blot analyses showing the Trim55, Bax, Bcl2, cleaved caspase-3, and cleaved caspase-8 protein expression in the C57-sham (n = 3), Trim55 –/– -sham (n = 3), C57-MI (n = 4), and Trim55 –/– -MI (n = 3) groups. (I, J) Representative TUNEL staining images and quantification of apoptotic positive cells at 3 days after MI in the C57-sham (n = 3), Trim55 –/– -sham (n = 3), C57-MI (n = 3), and Trim55 –/– -MI (n = 4) groups. All data in A, B, D, F, H, and J are expressed as mean ± SEM. Data in A, B, D, F, H, and J were analyzed using 2-way analysis of variance followed by the Bonferroni post hoc test. ∗∗ P < 0.01 vs C57-sham; † P < 0.05 vs Trim55 –/– -sham; ‡ P < 0.05 vs C57-MI. Bax = BCL2 associated X apoptosis regulator; Bcl-2 = BCL2 apoptosis regulator; BW = body weight; FS = fraction shortening; HE = hematoxylin and eosin; HW = heart weight; LVEF = left ventricular ejection fraction; TGF = transforming growth factor; TUNEL = terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling; other abbreviations as in Figure 1 .

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Knock-Out, Staining, Western Blot, Expressing, TUNEL Assay, End Labeling

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Overexpression Promoted Cardiomyocyte Apoptosis Induced by Hypoxia (A, B) Quantification and representative images of Western blots showing the expression of apoptosis-related proteins after Trim55 overexpression (n = 3). (C, D) Effects of Trim55 overexpression and CoCl treatment on the expression of apoptosis-related proteins by Western blot (n = 3). (E, F) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by flow cytometry analysis (n = 4). (G, H) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by TUNEL staining (n = 3). CoCl : 400 μmol/L, 24 hours. All data in B, D, F, and H are expressed as mean ± SEM. Data in B were analyzed using an unpaired Student’s t -test. Data in D, F, and H were analyzed using 2-way analysis of variance followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon; † P < 0.05 vs adTrim55; ‡ P < 0.05 vs adcon + CoCl . adcon = adenovirus control; adTrim55 = Trim55-overexpressing adenovirus; DAPI = 4′,6-diamidino-2-phenylindole; ns = not significant; other abbreviations as in Figures 1 and .

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Over Expression, Western Blot, Expressing, Flow Cytometry, TUNEL Assay, Staining, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Knockdown Alleviated Cardiomyocyte Apoptosis Induced by Hypoxia (A, B) Quantification and representative images of Western blot analyses showing the expression of apoptosis-related proteins after Trim55 knockdown (n = 3). (C, D) Effects of Trim55 knockdown and CoCl 2 treatment on the expression of apoptosis-related proteins by Western blot (n = 3). (E, F) Effects of Trim55 knockout and CoCl 2 treatment on cardiomyocyte apoptosis by flow cytometry analysis (n = 4). (G, H) Effects of Trim55 knockdown and CoCl 2 treatment on cardiomyocyte apoptosis by TUNEL staining (n = 3). All data in B, D, F, and H are expressed as mean ± SEM. Data in B were analyzed using an unpaired Student’s t -test. Data in D, F, and H were analyzed using 2-way analysis of variance, followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs si-nc; ‡ P < 0.05 vs si-Trim55; † P < 0.05 vs si-nc + CoCl 2 . si-nc = siRNA-normal control; si-Trim55 = Trim55-interfering RNA; other abbreviations as in Figure 1 , Figure 2 , Figure 3 .

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Knockdown, Western Blot, Expressing, Knock-Out, Flow Cytometry, TUNEL Assay, Staining, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Promoted Oxidative Stress by Inhibiting the Nrf2/HO-1 Pathway in Cardiomyocytes (A) Quantification and representative images of Western blot analyses showing the expression of antioxidant proteins (P62, Nrf2, HO-1) after overexpression of Trim55 in cardiomyocytes (n = 3). (B) Quantification and representative images of Western blot analyses showing the expression of antioxidant proteins (P62, Nrf2, HO-1) after Trim55 knockdown (n = 3). (C) Effects of Trim55 overexpression on the intracellular ROS levels by flow cytometry (n = 5). (D) Effects of Trim55 silencing on the intracellular ROS levels by flow cytometry (n = 5). (E) Quantification and representative images of CellROX staining after Trim55 overexpression (n = 3). (F) Quantification and representative images of CellROX staining after Trim55 knockdown (n = 3). All data in A to F are expressed as mean ±SEM. Data in A to F were analyzed by Student’s t -test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon or si-nc. APC = allophycocyanin; HO-1 = heme oxygenase 1; ISO = isotype control; Nrf2 = nuclear factor, erythroid derived 2; ROS = reactive oxygen species; other abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 .

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Western Blot, Expressing, Over Expression, Knockdown, Flow Cytometry, Staining, Control, Derivative Assay

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Promoted Cardiomyocyte Apoptosis Through Inhibiting the Nrf2/HO-1 Signaling Pathway (A) Effects of Trim55 overexpression and HO-1 overexpression on the expression of Trim55, HO-1, cleaved caspase-3, and cleaved caspase-8 by Western blot (n = 3). (B) Effects of Trim55 overexpression and HO-1 overexpression on cardiomyocyte apoptosis by flow cytometry (n = 4). (C) Effects of Trim55 overexpression and HO-1 overexpression on cardiomyocyte apoptosis by TUNEL staining (n = 3). (D) Effects of Trim55 knockdown and HO-1 knockdown on the expressions of cleaved caspase-3 and cleaved caspase-8 by Western blot (n = 3). (E) Effects of Trim55 knockdown and HO-1 knockdown on cardiomyocyte apoptosis by flow cytometry (n = 4). (F) Effects of Trim55 knockdown and HO-1 knockdown on cardiomyocyte apoptosis by TUNEL staining (n = 3). All data in A to F are expressed as mean ± SEM. Data in A to F were analyzed by 2-way analysis of variance followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon or si-nc; † P < 0.05 vs adTrim55 or si-nc + si HO-1; ‡ P < 0.05 vs adcon + pcDNA3.1 HO-1 or si-Trim55. Abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5 .

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Over Expression, Expressing, Western Blot, Flow Cytometry, TUNEL Assay, Staining, Knockdown

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Foxo3 Was a Direct Upstream Transcription Factor of Trim55 (A, B) Quantification and representative images of Western blots showing the expression of Foxo3, Gata3, Hoxa5, and Stat3 after CoCl 2 hypoxia in NRCMs (n = 3). ∗ P < 0.05 vs CON. (C, D) Foxo3 expression in the myocardium post-MI. (n = 3). ∗ P < 0.05, ∗∗ P < 0.01 vs sham (E, F) Quantification and representative images of Western blot analyses showing the expression of Trim55 after Foxo3 overexpression in NRCMs (n = 3). ∗∗ P < 0.01 vs CON. (G) Quantification of Trim55 mRNA levels in NRCMs after Foxo3 overexpression (n = 3). ∗∗ P < 0.01 vs CON. (H, I) Quantification and representative images of Western blot analyses showing the expression of Trim55 after knockdown in NRCMs (n = 3). † P < 0.05 vs si-nc. (J) Quantification of Trim55 mRNA levels in NRCMs after Foxo3 knockdown (n = 3). † P < 0.05 vs si-nc. (K) Possible binding sites of Foxo3 and Trim55 were predicted using the JASPAR website. (L) Relative luciferase activity in HEK293T cells with cotransfection with Foxo3-overexpressing plasmid and with WT-Trim55 plasmid or Mut-Trim55 plasmid. ∗∗ P < 0.01 vs NC Foxo3 + WT-Trim55; ‡ P < 0.05 vs NC Foxo3 + Mut-Trim55 (n = 12). (M) The effect of Trim55 on cardiomyocyte apoptosis after MI (by Figdraw). All data in B, D, F, G, H, and J are expressed as mean ± SEM. Data in B, D, F, G, and H were analyzed using Student’s t -test. Data in J were analyzed using 2-way analysis of variance, followed by the Bonferroni post hoc test. Foxo3 = forkhead box transcription factor 3; Mut = mutated; NC = normal control; WT = wild type; other abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5 .

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Western Blot, Expressing, Over Expression, Knockdown, Binding Assay, Luciferase, Activity Assay, Cotransfection, Plasmid Preparation, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Overexpression Promoted Cardiomyocyte Apoptosis Induced by Hypoxia (A, B) Quantification and representative images of Western blots showing the expression of apoptosis-related proteins after Trim55 overexpression (n = 3). (C, D) Effects of Trim55 overexpression and CoCl treatment on the expression of apoptosis-related proteins by Western blot (n = 3). (E, F) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by flow cytometry analysis (n = 4). (G, H) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by TUNEL staining (n = 3). CoCl : 400 μmol/L, 24 hours. All data in B, D, F, and H are expressed as mean ± SEM. Data in B were analyzed using an unpaired Student’s t -test. Data in D, F, and H were analyzed using 2-way analysis of variance followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon; † P < 0.05 vs adTrim55; ‡ P < 0.05 vs adcon + CoCl . adcon = adenovirus control; adTrim55 = Trim55-overexpressing adenovirus; DAPI = 4′,6-diamidino-2-phenylindole; ns = not significant; other abbreviations as in Figures 1 and .

Article Snippet: To establish Trim55-overexpressing or -knockdown cells, NRCMs were transfected with Trim55-overexpressing adenovirus (adTrim55, OBiO Technology) or Trim55-interfering RNA (si-Trim55, RiboBio).

Techniques: Over Expression, Western Blot, Expressing, Flow Cytometry, TUNEL Assay, Staining, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Expression Was Significantly Increased After MI (A, B) Quantification and representative images of Western blot analyses showing Trim55 protein expression in the border zone of myocardium at 3, 7, and 28 days after MI (n = 3). (C) Quantification of Trim55 mRNA levels in the border zone of myocardium at 3, 7, and 28 days after MI (n = 3). (D, E) Quantification and representative images of Trim55 immunohistochemistry in the border zone of myocardium at 3 and 28 days after MI (n = 3; scale bar: 50 μm). (F, G) Quantification and representative images of Western blot analyses showing Trim55 protein expression in NRCM cells after 400 μmol/L stimulation at 24 hours (n = 3). (H) Quantification of Trim55 mRNA levels in NRCM cells after CoCl 2 stimulation (400 μmol/L; 24 hours; n = 3). All data are expressed as mean ± SEM. Data in B, C, E, G, and H were analyzed using Student’s t -test. ∗ P < 0.05, ∗∗ P < 0.01 vs the control or sham group. CoCl 2 = cobalt chloride; CON = control; GADPH = glyceraldehyde-3-phosphate dehydrogenase; IHC = immunohistochemistry; MI = myocardial infarction; mRNA = messenger RNA; NRCM = neonatal rat cardiomyocyte; Trim55 = tripartite motif-containing 55.

Article Snippet: To generate Trim55-overexpressing mice, adeno-associated virus (AAV) overexpressing Trim55 (AAV2/9-CMV-Trim55, 2 × 10 11 genome copies per mouse in 200 μL of saline [OBiO Technology]) was injected into C57 mice by tail vein injection.

Techniques: Expressing, Western Blot, Immunohistochemistry, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Knockout Alleviated Myocardial Injury and Cardiomyocyte Apoptosis After MI (A) Echocardiographic analysis of the LVEF and FS at 28 days after MI in the C57-sham (n = 10), Trim55 –/– -sham (n = 10), C57-MI (n = 10), and Trim55 –/– -MI (n = 10) groups. (B) Quantitative analysis of the ratio of heart weight to body weight at 28 days after MI in the C57-sham (n = 10), Trim55 –/– -sham (n = 10), C57-MI (n = 8), and Trim55 –/– -MI (n = 10) groups. (C, D) Representative HE and Masson staining pictures and quantification of the infarction area at 28 days after MI in the C57-sham, Trim55 –/– -sham, C57-MI, and Trim55 –/– -MI groups (n = 3). (E, F) Quantification and representative images of Western blot analyses showing the Trim55, collagen I, and TGF-β protein expression in the C57-sham, Trim55 –/– -sham, C57-MI, and Trim55 –/– -MI groups (n = 3). (G, H) Quantification and representative images of Western blot analyses showing the Trim55, Bax, Bcl2, cleaved caspase-3, and cleaved caspase-8 protein expression in the C57-sham (n = 3), Trim55 –/– -sham (n = 3), C57-MI (n = 4), and Trim55 –/– -MI (n = 3) groups. (I, J) Representative TUNEL staining images and quantification of apoptotic positive cells at 3 days after MI in the C57-sham (n = 3), Trim55 –/– -sham (n = 3), C57-MI (n = 3), and Trim55 –/– -MI (n = 4) groups. All data in A, B, D, F, H, and J are expressed as mean ± SEM. Data in A, B, D, F, H, and J were analyzed using 2-way analysis of variance followed by the Bonferroni post hoc test. ∗∗ P < 0.01 vs C57-sham; † P < 0.05 vs Trim55 –/– -sham; ‡ P < 0.05 vs C57-MI. Bax = BCL2 associated X apoptosis regulator; Bcl-2 = BCL2 apoptosis regulator; BW = body weight; FS = fraction shortening; HE = hematoxylin and eosin; HW = heart weight; LVEF = left ventricular ejection fraction; TGF = transforming growth factor; TUNEL = terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling; other abbreviations as in Figure 1 .

Article Snippet: To generate Trim55-overexpressing mice, adeno-associated virus (AAV) overexpressing Trim55 (AAV2/9-CMV-Trim55, 2 × 10 11 genome copies per mouse in 200 μL of saline [OBiO Technology]) was injected into C57 mice by tail vein injection.

Techniques: Knock-Out, Staining, Western Blot, Expressing, TUNEL Assay, End Labeling

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Overexpression Promoted Cardiomyocyte Apoptosis Induced by Hypoxia (A, B) Quantification and representative images of Western blots showing the expression of apoptosis-related proteins after Trim55 overexpression (n = 3). (C, D) Effects of Trim55 overexpression and CoCl treatment on the expression of apoptosis-related proteins by Western blot (n = 3). (E, F) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by flow cytometry analysis (n = 4). (G, H) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by TUNEL staining (n = 3). CoCl : 400 μmol/L, 24 hours. All data in B, D, F, and H are expressed as mean ± SEM. Data in B were analyzed using an unpaired Student’s t -test. Data in D, F, and H were analyzed using 2-way analysis of variance followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon; † P < 0.05 vs adTrim55; ‡ P < 0.05 vs adcon + CoCl . adcon = adenovirus control; adTrim55 = Trim55-overexpressing adenovirus; DAPI = 4′,6-diamidino-2-phenylindole; ns = not significant; other abbreviations as in Figures 1 and .

Article Snippet: To generate Trim55-overexpressing mice, adeno-associated virus (AAV) overexpressing Trim55 (AAV2/9-CMV-Trim55, 2 × 10 11 genome copies per mouse in 200 μL of saline [OBiO Technology]) was injected into C57 mice by tail vein injection.

Techniques: Over Expression, Western Blot, Expressing, Flow Cytometry, TUNEL Assay, Staining, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Knockdown Alleviated Cardiomyocyte Apoptosis Induced by Hypoxia (A, B) Quantification and representative images of Western blot analyses showing the expression of apoptosis-related proteins after Trim55 knockdown (n = 3). (C, D) Effects of Trim55 knockdown and CoCl 2 treatment on the expression of apoptosis-related proteins by Western blot (n = 3). (E, F) Effects of Trim55 knockout and CoCl 2 treatment on cardiomyocyte apoptosis by flow cytometry analysis (n = 4). (G, H) Effects of Trim55 knockdown and CoCl 2 treatment on cardiomyocyte apoptosis by TUNEL staining (n = 3). All data in B, D, F, and H are expressed as mean ± SEM. Data in B were analyzed using an unpaired Student’s t -test. Data in D, F, and H were analyzed using 2-way analysis of variance, followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs si-nc; ‡ P < 0.05 vs si-Trim55; † P < 0.05 vs si-nc + CoCl 2 . si-nc = siRNA-normal control; si-Trim55 = Trim55-interfering RNA; other abbreviations as in Figure 1 , Figure 2 , Figure 3 .

Article Snippet: To generate Trim55-overexpressing mice, adeno-associated virus (AAV) overexpressing Trim55 (AAV2/9-CMV-Trim55, 2 × 10 11 genome copies per mouse in 200 μL of saline [OBiO Technology]) was injected into C57 mice by tail vein injection.

Techniques: Knockdown, Western Blot, Expressing, Knock-Out, Flow Cytometry, TUNEL Assay, Staining, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Promoted Oxidative Stress by Inhibiting the Nrf2/HO-1 Pathway in Cardiomyocytes (A) Quantification and representative images of Western blot analyses showing the expression of antioxidant proteins (P62, Nrf2, HO-1) after overexpression of Trim55 in cardiomyocytes (n = 3). (B) Quantification and representative images of Western blot analyses showing the expression of antioxidant proteins (P62, Nrf2, HO-1) after Trim55 knockdown (n = 3). (C) Effects of Trim55 overexpression on the intracellular ROS levels by flow cytometry (n = 5). (D) Effects of Trim55 silencing on the intracellular ROS levels by flow cytometry (n = 5). (E) Quantification and representative images of CellROX staining after Trim55 overexpression (n = 3). (F) Quantification and representative images of CellROX staining after Trim55 knockdown (n = 3). All data in A to F are expressed as mean ±SEM. Data in A to F were analyzed by Student’s t -test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon or si-nc. APC = allophycocyanin; HO-1 = heme oxygenase 1; ISO = isotype control; Nrf2 = nuclear factor, erythroid derived 2; ROS = reactive oxygen species; other abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 .

Article Snippet: To generate Trim55-overexpressing mice, adeno-associated virus (AAV) overexpressing Trim55 (AAV2/9-CMV-Trim55, 2 × 10 11 genome copies per mouse in 200 μL of saline [OBiO Technology]) was injected into C57 mice by tail vein injection.

Techniques: Western Blot, Expressing, Over Expression, Knockdown, Flow Cytometry, Staining, Control, Derivative Assay

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Promoted Cardiomyocyte Apoptosis Through Inhibiting the Nrf2/HO-1 Signaling Pathway (A) Effects of Trim55 overexpression and HO-1 overexpression on the expression of Trim55, HO-1, cleaved caspase-3, and cleaved caspase-8 by Western blot (n = 3). (B) Effects of Trim55 overexpression and HO-1 overexpression on cardiomyocyte apoptosis by flow cytometry (n = 4). (C) Effects of Trim55 overexpression and HO-1 overexpression on cardiomyocyte apoptosis by TUNEL staining (n = 3). (D) Effects of Trim55 knockdown and HO-1 knockdown on the expressions of cleaved caspase-3 and cleaved caspase-8 by Western blot (n = 3). (E) Effects of Trim55 knockdown and HO-1 knockdown on cardiomyocyte apoptosis by flow cytometry (n = 4). (F) Effects of Trim55 knockdown and HO-1 knockdown on cardiomyocyte apoptosis by TUNEL staining (n = 3). All data in A to F are expressed as mean ± SEM. Data in A to F were analyzed by 2-way analysis of variance followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon or si-nc; † P < 0.05 vs adTrim55 or si-nc + si HO-1; ‡ P < 0.05 vs adcon + pcDNA3.1 HO-1 or si-Trim55. Abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5 .

Article Snippet: To generate Trim55-overexpressing mice, adeno-associated virus (AAV) overexpressing Trim55 (AAV2/9-CMV-Trim55, 2 × 10 11 genome copies per mouse in 200 μL of saline [OBiO Technology]) was injected into C57 mice by tail vein injection.

Techniques: Over Expression, Expressing, Western Blot, Flow Cytometry, TUNEL Assay, Staining, Knockdown

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Foxo3 Was a Direct Upstream Transcription Factor of Trim55 (A, B) Quantification and representative images of Western blots showing the expression of Foxo3, Gata3, Hoxa5, and Stat3 after CoCl 2 hypoxia in NRCMs (n = 3). ∗ P < 0.05 vs CON. (C, D) Foxo3 expression in the myocardium post-MI. (n = 3). ∗ P < 0.05, ∗∗ P < 0.01 vs sham (E, F) Quantification and representative images of Western blot analyses showing the expression of Trim55 after Foxo3 overexpression in NRCMs (n = 3). ∗∗ P < 0.01 vs CON. (G) Quantification of Trim55 mRNA levels in NRCMs after Foxo3 overexpression (n = 3). ∗∗ P < 0.01 vs CON. (H, I) Quantification and representative images of Western blot analyses showing the expression of Trim55 after knockdown in NRCMs (n = 3). † P < 0.05 vs si-nc. (J) Quantification of Trim55 mRNA levels in NRCMs after Foxo3 knockdown (n = 3). † P < 0.05 vs si-nc. (K) Possible binding sites of Foxo3 and Trim55 were predicted using the JASPAR website. (L) Relative luciferase activity in HEK293T cells with cotransfection with Foxo3-overexpressing plasmid and with WT-Trim55 plasmid or Mut-Trim55 plasmid. ∗∗ P < 0.01 vs NC Foxo3 + WT-Trim55; ‡ P < 0.05 vs NC Foxo3 + Mut-Trim55 (n = 12). (M) The effect of Trim55 on cardiomyocyte apoptosis after MI (by Figdraw). All data in B, D, F, G, H, and J are expressed as mean ± SEM. Data in B, D, F, G, and H were analyzed using Student’s t -test. Data in J were analyzed using 2-way analysis of variance, followed by the Bonferroni post hoc test. Foxo3 = forkhead box transcription factor 3; Mut = mutated; NC = normal control; WT = wild type; other abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5 .

Article Snippet: To generate Trim55-overexpressing mice, adeno-associated virus (AAV) overexpressing Trim55 (AAV2/9-CMV-Trim55, 2 × 10 11 genome copies per mouse in 200 μL of saline [OBiO Technology]) was injected into C57 mice by tail vein injection.

Techniques: Western Blot, Expressing, Over Expression, Knockdown, Binding Assay, Luciferase, Activity Assay, Cotransfection, Plasmid Preparation, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Expression Was Significantly Increased After MI (A, B) Quantification and representative images of Western blot analyses showing Trim55 protein expression in the border zone of myocardium at 3, 7, and 28 days after MI (n = 3). (C) Quantification of Trim55 mRNA levels in the border zone of myocardium at 3, 7, and 28 days after MI (n = 3). (D, E) Quantification and representative images of Trim55 immunohistochemistry in the border zone of myocardium at 3 and 28 days after MI (n = 3; scale bar: 50 μm). (F, G) Quantification and representative images of Western blot analyses showing Trim55 protein expression in NRCM cells after 400 μmol/L stimulation at 24 hours (n = 3). (H) Quantification of Trim55 mRNA levels in NRCM cells after CoCl 2 stimulation (400 μmol/L; 24 hours; n = 3). All data are expressed as mean ± SEM. Data in B, C, E, G, and H were analyzed using Student’s t -test. ∗ P < 0.05, ∗∗ P < 0.01 vs the control or sham group. CoCl 2 = cobalt chloride; CON = control; GADPH = glyceraldehyde-3-phosphate dehydrogenase; IHC = immunohistochemistry; MI = myocardial infarction; mRNA = messenger RNA; NRCM = neonatal rat cardiomyocyte; Trim55 = tripartite motif-containing 55.

Article Snippet: After rehydration in citrate buffer solution and a 40-minute antigen recovery process, mouse heart slices were permeabilized with 0.2% Triton X-100 in PBS for 15 minutes, sealed with 5% bovine serum albumin for 1 hour, and stained overnight with Trim55 (1:50, Thermo Fisher Scientific) antibody.

Techniques: Expressing, Western Blot, Immunohistochemistry, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Knockout Alleviated Myocardial Injury and Cardiomyocyte Apoptosis After MI (A) Echocardiographic analysis of the LVEF and FS at 28 days after MI in the C57-sham (n = 10), Trim55 –/– -sham (n = 10), C57-MI (n = 10), and Trim55 –/– -MI (n = 10) groups. (B) Quantitative analysis of the ratio of heart weight to body weight at 28 days after MI in the C57-sham (n = 10), Trim55 –/– -sham (n = 10), C57-MI (n = 8), and Trim55 –/– -MI (n = 10) groups. (C, D) Representative HE and Masson staining pictures and quantification of the infarction area at 28 days after MI in the C57-sham, Trim55 –/– -sham, C57-MI, and Trim55 –/– -MI groups (n = 3). (E, F) Quantification and representative images of Western blot analyses showing the Trim55, collagen I, and TGF-β protein expression in the C57-sham, Trim55 –/– -sham, C57-MI, and Trim55 –/– -MI groups (n = 3). (G, H) Quantification and representative images of Western blot analyses showing the Trim55, Bax, Bcl2, cleaved caspase-3, and cleaved caspase-8 protein expression in the C57-sham (n = 3), Trim55 –/– -sham (n = 3), C57-MI (n = 4), and Trim55 –/– -MI (n = 3) groups. (I, J) Representative TUNEL staining images and quantification of apoptotic positive cells at 3 days after MI in the C57-sham (n = 3), Trim55 –/– -sham (n = 3), C57-MI (n = 3), and Trim55 –/– -MI (n = 4) groups. All data in A, B, D, F, H, and J are expressed as mean ± SEM. Data in A, B, D, F, H, and J were analyzed using 2-way analysis of variance followed by the Bonferroni post hoc test. ∗∗ P < 0.01 vs C57-sham; † P < 0.05 vs Trim55 –/– -sham; ‡ P < 0.05 vs C57-MI. Bax = BCL2 associated X apoptosis regulator; Bcl-2 = BCL2 apoptosis regulator; BW = body weight; FS = fraction shortening; HE = hematoxylin and eosin; HW = heart weight; LVEF = left ventricular ejection fraction; TGF = transforming growth factor; TUNEL = terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling; other abbreviations as in Figure 1 .

Article Snippet: After rehydration in citrate buffer solution and a 40-minute antigen recovery process, mouse heart slices were permeabilized with 0.2% Triton X-100 in PBS for 15 minutes, sealed with 5% bovine serum albumin for 1 hour, and stained overnight with Trim55 (1:50, Thermo Fisher Scientific) antibody.

Techniques: Knock-Out, Staining, Western Blot, Expressing, TUNEL Assay, End Labeling

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Overexpression Promoted Cardiomyocyte Apoptosis Induced by Hypoxia (A, B) Quantification and representative images of Western blots showing the expression of apoptosis-related proteins after Trim55 overexpression (n = 3). (C, D) Effects of Trim55 overexpression and CoCl treatment on the expression of apoptosis-related proteins by Western blot (n = 3). (E, F) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by flow cytometry analysis (n = 4). (G, H) Effects of Trim55 overexpression and CoCl treatment on cardiomyocyte apoptosis by TUNEL staining (n = 3). CoCl : 400 μmol/L, 24 hours. All data in B, D, F, and H are expressed as mean ± SEM. Data in B were analyzed using an unpaired Student’s t -test. Data in D, F, and H were analyzed using 2-way analysis of variance followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon; † P < 0.05 vs adTrim55; ‡ P < 0.05 vs adcon + CoCl . adcon = adenovirus control; adTrim55 = Trim55-overexpressing adenovirus; DAPI = 4′,6-diamidino-2-phenylindole; ns = not significant; other abbreviations as in Figures 1 and .

Article Snippet: After rehydration in citrate buffer solution and a 40-minute antigen recovery process, mouse heart slices were permeabilized with 0.2% Triton X-100 in PBS for 15 minutes, sealed with 5% bovine serum albumin for 1 hour, and stained overnight with Trim55 (1:50, Thermo Fisher Scientific) antibody.

Techniques: Over Expression, Western Blot, Expressing, Flow Cytometry, TUNEL Assay, Staining, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Knockdown Alleviated Cardiomyocyte Apoptosis Induced by Hypoxia (A, B) Quantification and representative images of Western blot analyses showing the expression of apoptosis-related proteins after Trim55 knockdown (n = 3). (C, D) Effects of Trim55 knockdown and CoCl 2 treatment on the expression of apoptosis-related proteins by Western blot (n = 3). (E, F) Effects of Trim55 knockout and CoCl 2 treatment on cardiomyocyte apoptosis by flow cytometry analysis (n = 4). (G, H) Effects of Trim55 knockdown and CoCl 2 treatment on cardiomyocyte apoptosis by TUNEL staining (n = 3). All data in B, D, F, and H are expressed as mean ± SEM. Data in B were analyzed using an unpaired Student’s t -test. Data in D, F, and H were analyzed using 2-way analysis of variance, followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs si-nc; ‡ P < 0.05 vs si-Trim55; † P < 0.05 vs si-nc + CoCl 2 . si-nc = siRNA-normal control; si-Trim55 = Trim55-interfering RNA; other abbreviations as in Figure 1 , Figure 2 , Figure 3 .

Article Snippet: After rehydration in citrate buffer solution and a 40-minute antigen recovery process, mouse heart slices were permeabilized with 0.2% Triton X-100 in PBS for 15 minutes, sealed with 5% bovine serum albumin for 1 hour, and stained overnight with Trim55 (1:50, Thermo Fisher Scientific) antibody.

Techniques: Knockdown, Western Blot, Expressing, Knock-Out, Flow Cytometry, TUNEL Assay, Staining, Control

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Promoted Oxidative Stress by Inhibiting the Nrf2/HO-1 Pathway in Cardiomyocytes (A) Quantification and representative images of Western blot analyses showing the expression of antioxidant proteins (P62, Nrf2, HO-1) after overexpression of Trim55 in cardiomyocytes (n = 3). (B) Quantification and representative images of Western blot analyses showing the expression of antioxidant proteins (P62, Nrf2, HO-1) after Trim55 knockdown (n = 3). (C) Effects of Trim55 overexpression on the intracellular ROS levels by flow cytometry (n = 5). (D) Effects of Trim55 silencing on the intracellular ROS levels by flow cytometry (n = 5). (E) Quantification and representative images of CellROX staining after Trim55 overexpression (n = 3). (F) Quantification and representative images of CellROX staining after Trim55 knockdown (n = 3). All data in A to F are expressed as mean ±SEM. Data in A to F were analyzed by Student’s t -test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon or si-nc. APC = allophycocyanin; HO-1 = heme oxygenase 1; ISO = isotype control; Nrf2 = nuclear factor, erythroid derived 2; ROS = reactive oxygen species; other abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 .

Article Snippet: After rehydration in citrate buffer solution and a 40-minute antigen recovery process, mouse heart slices were permeabilized with 0.2% Triton X-100 in PBS for 15 minutes, sealed with 5% bovine serum albumin for 1 hour, and stained overnight with Trim55 (1:50, Thermo Fisher Scientific) antibody.

Techniques: Western Blot, Expressing, Over Expression, Knockdown, Flow Cytometry, Staining, Control, Derivative Assay

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Trim55 Promoted Cardiomyocyte Apoptosis Through Inhibiting the Nrf2/HO-1 Signaling Pathway (A) Effects of Trim55 overexpression and HO-1 overexpression on the expression of Trim55, HO-1, cleaved caspase-3, and cleaved caspase-8 by Western blot (n = 3). (B) Effects of Trim55 overexpression and HO-1 overexpression on cardiomyocyte apoptosis by flow cytometry (n = 4). (C) Effects of Trim55 overexpression and HO-1 overexpression on cardiomyocyte apoptosis by TUNEL staining (n = 3). (D) Effects of Trim55 knockdown and HO-1 knockdown on the expressions of cleaved caspase-3 and cleaved caspase-8 by Western blot (n = 3). (E) Effects of Trim55 knockdown and HO-1 knockdown on cardiomyocyte apoptosis by flow cytometry (n = 4). (F) Effects of Trim55 knockdown and HO-1 knockdown on cardiomyocyte apoptosis by TUNEL staining (n = 3). All data in A to F are expressed as mean ± SEM. Data in A to F were analyzed by 2-way analysis of variance followed by the Bonferroni post hoc test. ∗ P < 0.05, ∗∗ P < 0.01 vs adcon or si-nc; † P < 0.05 vs adTrim55 or si-nc + si HO-1; ‡ P < 0.05 vs adcon + pcDNA3.1 HO-1 or si-Trim55. Abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5 .

Article Snippet: After rehydration in citrate buffer solution and a 40-minute antigen recovery process, mouse heart slices were permeabilized with 0.2% Triton X-100 in PBS for 15 minutes, sealed with 5% bovine serum albumin for 1 hour, and stained overnight with Trim55 (1:50, Thermo Fisher Scientific) antibody.

Techniques: Over Expression, Expressing, Western Blot, Flow Cytometry, TUNEL Assay, Staining, Knockdown

Journal: JACC: Basic to Translational Science

Article Title: TRIM55 Aggravates Cardiomyocyte Apoptosis After Myocardial Infarction via Modulation of the Nrf2/HO-1 Pathway

doi: 10.1016/j.jacbts.2024.05.006

Figure Lengend Snippet: Foxo3 Was a Direct Upstream Transcription Factor of Trim55 (A, B) Quantification and representative images of Western blots showing the expression of Foxo3, Gata3, Hoxa5, and Stat3 after CoCl 2 hypoxia in NRCMs (n = 3). ∗ P < 0.05 vs CON. (C, D) Foxo3 expression in the myocardium post-MI. (n = 3). ∗ P < 0.05, ∗∗ P < 0.01 vs sham (E, F) Quantification and representative images of Western blot analyses showing the expression of Trim55 after Foxo3 overexpression in NRCMs (n = 3). ∗∗ P < 0.01 vs CON. (G) Quantification of Trim55 mRNA levels in NRCMs after Foxo3 overexpression (n = 3). ∗∗ P < 0.01 vs CON. (H, I) Quantification and representative images of Western blot analyses showing the expression of Trim55 after knockdown in NRCMs (n = 3). † P < 0.05 vs si-nc. (J) Quantification of Trim55 mRNA levels in NRCMs after Foxo3 knockdown (n = 3). † P < 0.05 vs si-nc. (K) Possible binding sites of Foxo3 and Trim55 were predicted using the JASPAR website. (L) Relative luciferase activity in HEK293T cells with cotransfection with Foxo3-overexpressing plasmid and with WT-Trim55 plasmid or Mut-Trim55 plasmid. ∗∗ P < 0.01 vs NC Foxo3 + WT-Trim55; ‡ P < 0.05 vs NC Foxo3 + Mut-Trim55 (n = 12). (M) The effect of Trim55 on cardiomyocyte apoptosis after MI (by Figdraw). All data in B, D, F, G, H, and J are expressed as mean ± SEM. Data in B, D, F, G, and H were analyzed using Student’s t -test. Data in J were analyzed using 2-way analysis of variance, followed by the Bonferroni post hoc test. Foxo3 = forkhead box transcription factor 3; Mut = mutated; NC = normal control; WT = wild type; other abbreviations as in Figure 1 , Figure 2 , Figure 3 , Figure 4 , Figure 5 .

Article Snippet: After rehydration in citrate buffer solution and a 40-minute antigen recovery process, mouse heart slices were permeabilized with 0.2% Triton X-100 in PBS for 15 minutes, sealed with 5% bovine serum albumin for 1 hour, and stained overnight with Trim55 (1:50, Thermo Fisher Scientific) antibody.

Techniques: Western Blot, Expressing, Over Expression, Knockdown, Binding Assay, Luciferase, Activity Assay, Cotransfection, Plasmid Preparation, Control

Journal: Journal of Hepatocellular Carcinoma

Article Title: TRIM55 Promotes Proliferation of Hepatocellular Carcinoma Through Stabilizing TRIP6 to Activate Wnt/β-Catenin Signaling

doi: 10.2147/JHC.S418049

Figure Lengend Snippet: TRIM55 expression was upregulated in HCC and associated with a poor prognosis in HCC patients. ( A ) TRIM55 mRNA expression in HCC tumor tissues and adjacent non-tumor tissues extracted from GSE84005, GSE64041, GSE39791 and GSE36376. ( B ) TRIM55 expression in HCC tumor tissues and adjacent non-tumor tissues in the TCGA database. ( C ) Representative immunohistochemical staining images of TRIM55 in 110 paired HCC tumor tissues and adjacent non-tumor tissues (Scale bars: 50 μm, 20 μm). ( D ) Statistical analysis of TRIM55 expression according to the immunohistochemistry results. ( E ) Kaplan-Meier overall survival curve of two HCC groups: high TRIM55 group: n = 67; low TRIM55 group: n = 43. ( F ) Kaplan-Meier disease-free survival curve of two HCC groups: high TRIM55 group: n = 67; low TRIM55 group: n = 43. (*P < 0.05, **P < 0.01, ****P < 0.0001).

Article Snippet: To construct TRIM55 overexpressing cell lines, the human TRIM55 cDNA and negative control were cloned into the pLenti-CMV-Puro plasmid (Addgene #17448).

Techniques: Expressing, Immunohistochemical staining, Staining, Immunohistochemistry

Journal: Journal of Hepatocellular Carcinoma

Article Title: TRIM55 Promotes Proliferation of Hepatocellular Carcinoma Through Stabilizing TRIP6 to Activate Wnt/β-Catenin Signaling

doi: 10.2147/JHC.S418049

Figure Lengend Snippet: TRIM55 promotes HCC cell proliferation in vitro and in vivo. ( A ) The overexpression of TRIM55 in HLF and HepG2 cells were verified by Western blot analysis. ( B ) Knockdown of TRIM55 in 97H and Alex cells were verified by Western blot analysis. ( C and D ) The effect of TRIM55 overexpression or knockdown on cell proliferation evaluated by CCK8 assay. ( E and F ) The effect of TRIM55 overexpression or knockdown on cell proliferation evaluated by colony formation assay. ( G and H ) The effect of TRIM55 overexpression or knockdown on growth of xenograft tumors. Tumor growth was evaluated by measuring tumor volume. ( I and J ) Representative images of the effect of TRIM55 overexpression or knockdown on growth of intrahepatic tumors. Tumor growth was evaluated by measuring tumor volume. ( K and L ) Representative immunohistochemical images of expression level of Ki-67 in xenograft tumor tissues. (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.).

Article Snippet: To construct TRIM55 overexpressing cell lines, the human TRIM55 cDNA and negative control were cloned into the pLenti-CMV-Puro plasmid (Addgene #17448).

Techniques: In Vitro, In Vivo, Over Expression, Western Blot, CCK-8 Assay, Colony Assay, Immunohistochemical staining, Expressing

Journal: Journal of Hepatocellular Carcinoma

Article Title: TRIM55 Promotes Proliferation of Hepatocellular Carcinoma Through Stabilizing TRIP6 to Activate Wnt/β-Catenin Signaling

doi: 10.2147/JHC.S418049

Figure Lengend Snippet: TRIM55 interacts and colocalizes with TRIP6. ( A ) Proteins associated with TRIM55 were identified by mass spectrometry. ( B ) The TRIP6 peptides identified by mass spectrometry were shown. ( C ) The interaction between Flag-TRIM55 and HA-TRIP6 was analyzed by Co-IP in co-transfected HEK293T cells. ( D ) The interaction between endogenous TRIM55 and TRIP6 was analyzed by Co-IP in HLF and HepG2 cells with TRIM55-overexpression. ( E ) Confocal assays confirmed that TRIM55 and TRIP6 were mainly co-localized in the cytoplasm (Scale bar: 10 μm, 63 X objective).

Article Snippet: To construct TRIM55 overexpressing cell lines, the human TRIM55 cDNA and negative control were cloned into the pLenti-CMV-Puro plasmid (Addgene #17448).

Techniques: Mass Spectrometry, Co-Immunoprecipitation Assay, Transfection, Over Expression

Journal: Journal of Hepatocellular Carcinoma

Article Title: TRIM55 Promotes Proliferation of Hepatocellular Carcinoma Through Stabilizing TRIP6 to Activate Wnt/β-Catenin Signaling

doi: 10.2147/JHC.S418049

Figure Lengend Snippet: Tumor promoting effect of TRIM55 in HCC is partially dependent on TRIP6. ( A ) The effect of TRIM55 overexpression or knockdown on TRIP6, β-Catenin and Cyclin D1 protein levels by Western blot analysis. ( B ) The effect of TRIM55 overexpression or knockdown on Wnt activity by TOP/FOP luciferase reporter assay. ( C ) Knockdown of TRIP6 in HLF and HepG2 cells with TRIM55 overexpression was verified by Western blot analysis. ( D ) CCK8 assay was performed in HLF and HepG2 cells with TRIM55-overexpression or control cells transiently transfected with siNC, siTRIP6. ( E ) Colony formation assay was performed in HLF and HepG2 cells with TRIM55-overexpression or control cells transiently transfected with siNC, siTRIP6. ( F ) TRIP6 knockdown significantly decreased the TOP/FOP luciferase activity induced by TRIM55. ( G ) Western blot analysis of TRIP6, β-Catenin and Cyclin D1 protein levels in HLF and HepG2 cells with TRIM55-overexpression or control cells transiently transfected with siNC, siTRIP6. (*P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: To construct TRIM55 overexpressing cell lines, the human TRIM55 cDNA and negative control were cloned into the pLenti-CMV-Puro plasmid (Addgene #17448).

Techniques: Over Expression, Western Blot, Activity Assay, Luciferase, Reporter Assay, CCK-8 Assay, Transfection, Colony Assay

Journal: Journal of Hepatocellular Carcinoma

Article Title: TRIM55 Promotes Proliferation of Hepatocellular Carcinoma Through Stabilizing TRIP6 to Activate Wnt/β-Catenin Signaling

doi: 10.2147/JHC.S418049

Figure Lengend Snippet: TRIM55 stabilizes TRIP6 protein by inhibiting K48-linked ubiquitination and inducing K63-linked ubiquitination. ( A ) The effects of transfection of Flag-TRIM55 with different concentrations on HA-TRIP6 (0.5μg) were detected in HEK293T cells by Western blot assay. ( B and C ) HLF-TRIM55 and HepG2-TRIM55 or control cells were treated with CHX (10 μM) for 0, 2, 4 and 8 h, the levels of TRIP6 were detected by Western blot assay. Quantification of the TRIP6 levels relative to GAPDH expression is shown. ( D and E ) HLF-TRIM55 and HepG2-TRIM55 or control cells were treated with Mg132 (20 μM) for 6 h, and the ubiquitination of TRIP6 was detected by immunoprecipitation (IP). ( F ) Co-IP analysis of ubiquitination of TRIP6 in 293T cells co-transfected with HA-TRIP6, Flag-TRIM55 and Myc-Ub plasmids. ( G and H ) Co-IP analysis of ubiquitination of TRIP6 in 293T cells co-transfected with Flag-TRIM55, Myc -TRIP6, HA-Ub-K48 or HA-Ub-K63 plasmids.

Article Snippet: To construct TRIM55 overexpressing cell lines, the human TRIM55 cDNA and negative control were cloned into the pLenti-CMV-Puro plasmid (Addgene #17448).

Techniques: Transfection, Western Blot, Expressing, Immunoprecipitation, Co-Immunoprecipitation Assay

Journal: Journal of Hepatocellular Carcinoma

Article Title: TRIM55 Promotes Proliferation of Hepatocellular Carcinoma Through Stabilizing TRIP6 to Activate Wnt/β-Catenin Signaling

doi: 10.2147/JHC.S418049

Figure Lengend Snippet: Expression of TRIM55 and TRIP6 were positively correlated in human HCC tissues. ( A ) Representative IHC staining figures of TRIM55 and TRIP6 expression in serial sections of HCC tumor samples (Scale bar: 50 μm, 20 μm; 20 X, 40 X objective). ( B ) Distribution of TRIP6 expression level in high TRIM55 expression level group and low TRIM55 expression level group tissues. ( C ) Pearson correlation analysis between TRIM55 and TRIP6 expression, N = 110. (*P < 0.05).

Article Snippet: To construct TRIM55 overexpressing cell lines, the human TRIM55 cDNA and negative control were cloned into the pLenti-CMV-Puro plasmid (Addgene #17448).

Techniques: Expressing, Immunohistochemistry