Journal: Journal of Cardiothoracic Surgery

Article Title: Study on the mechanism and clinical value of miR-210-3p and miR-582-5p in acute myocardial infarction by targeting MXD1

doi: 10.1186/s13019-025-03518-3

Figure Lengend Snippet: Expressions of miR-210-3p and miR-582-5p in AMI. (A) Intersection of miRNAs differentially expressed in STEMI and non-STEMI patients, respectively, compared with healthy volunteers. Expressions of miR-210-3p (B and C) and miR-582-5p (D and E) in the GSE168856 dataset. (F) The secretion level of cTnI in cardiomyocytes. (G) Expressions of miR-210-3p and miR-582-5p in the hypoxic cardiomyocytes. Expressions of miR-210-3p (H) and miR-582-5p (I) in AMI patients. ** p < 0.01; *** p < 0.001

Article Snippet: The secretion level of cardiac troponin I (cTnI) and the intracellular levels of MDA and Fe 2+ were detected using the Human cTnI ELISA Kit (MULTI SCIENCES, CHN), lipid oxidation (MDA) Assay Kit (Beyotime) and Iron Assay Kit (Abcam, UK), respectively.

Techniques:

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Platelet Internalization Mediates Ferroptosis in Myocardial Infarction

doi: 10.1161/atvbaha.122.318161

Figure Lengend Snippet: Figure 2. Platelets are internalized by cardiomyocytes. A, Distribution of platelets in myocardial tissue (n=4). Platelets were labeled with CD41 (green), cardiomyocytes were labeled with cTnI (red), and nuclei visualized with DAPI (blue). Scale bar, 500 μm. B, Neonatal mouse cardiomyocytes (CMs) internalize platelets (n=3). CMs were labeled by CM-Dil (red), platelets were labeled by CFSE (green). Scale bar, 100 µm. C, Three-dimensional reconstruction of confocal Z-stack images of CMs co-cultured with platelets to demonstrate uptake and perinuclear localization (n=3; view angle: 30, z-Scaling: 2).

Article Snippet: Primary antibodies included cTnI antibody (1:100, Proteintech, catalog 21652-1- AP) and CD41 antibody (1:200, Santa cruz, sc-365938).

Techniques: Labeling, Cell Culture

Journal: Scientific Reports

Article Title: Human platelet lysate combined with mesenchymal stem cells pretreated with platelet lysate improved cardiac function in rats with myocardial infarction

doi: 10.1038/s41598-024-79050-6

Figure Lengend Snippet: Immunofluorescent analysis indicates the expression of NKX2.5 and cTnI (green) in the hearts of rats with MI two and four weeks after PMSCs and HPL + PMSCs injection: ( A , B ): Sample micrographs of NKX2.5 and cTnI in an animal of each group. Quantification of the levels of NKX2.5 + and cTnI+ ( C and E ) and the number of Dil+/NKX2.5 + and Dil+/ cTnI+ ( D and F ) in the study groups. Stem cells stained with Dil dye (red) and nuclei with DAPI (blue). *** P < 0.001 vs. PMSCs 2 weeks; # < P < 0.05, ### P < 0.001 vs. HPL + PMSCs 2 weeks; &&& P < 0.01 vs. PMSCs; $$ P < 0.001, $$$ P < 0.001 vs. HPL + PMSCs. n = 3 in each group.

Article Snippet: Rabbit polyclonal primary antibody for NKX2 (No: orb540657), rabbit polyclonal antibody for cTnI (No: orb304638), and goat anti-rabbit IgG (H + L) antibody (FITC) (No orb688925) were purchased from Biorbyt, United Kingdom.

Techniques: Expressing, Injection, Staining

Journal: International Journal of Molecular and Cellular Medicine

Article Title: Innovative Aptamer-HRP Conjugation for Cardiac Troponin I Detection: A Novel ELASA Approach for AMI Diagnostics

doi: 10.22088/IJMCM.BUMS.14.2.705

Figure Lengend Snippet: Four modes of Tro4 and Tro6 molecular docking with Docking score and confidence score from HDOCK web server. The table summarizes the information about the Docking Score and Confidence Score of the 1 to 4 molecular docking models of the cTnI protein and Tro4 and Tro6 aptamer, based on data retrieved from the HDOCK web server. It seems that Model 1 indicates the highest affinity and binding probability between both aptamers and cTnI protein.

Article Snippet: A cTnI ELISA kit was sourced from Monobind (USA).

Techniques: Binding Assay

Journal: International Journal of Molecular and Cellular Medicine

Article Title: Innovative Aptamer-HRP Conjugation for Cardiac Troponin I Detection: A Novel ELASA Approach for AMI Diagnostics

doi: 10.22088/IJMCM.BUMS.14.2.705

Figure Lengend Snippet: Screening of two aptamers binding ability against cTnI. Direct ELASA was used to screen two aptamer sequences for the detection of cTnI. Different concentrations of both aptamers were used to find the optimum concentration and it was shown that concentrations of 10 and 5μM were considered as the optimum.

Article Snippet: A cTnI ELISA kit was sourced from Monobind (USA).

Techniques: Binding Assay, Concentration Assay

Journal: International Journal of Molecular and Cellular Medicine

Article Title: Innovative Aptamer-HRP Conjugation for Cardiac Troponin I Detection: A Novel ELASA Approach for AMI Diagnostics

doi: 10.22088/IJMCM.BUMS.14.2.705

Figure Lengend Snippet: A calibration curve between absorbance and cTnI concentration from 0.1 to 22ng/Ml. To determine the linearity of the aptamer-based ELISA method, a sample with a specific concentration was diluted as 1.5, 1.10, 1.25, 1.175, 1.875, and 1.1000. The fitted linear equation is y=0/0842ln (cTnI concentration)+0/3585 (R²=0.94).

Article Snippet: A cTnI ELISA kit was sourced from Monobind (USA).

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Biochemical Journal

Article Title: Calpain-2 specifically cleaves Junctophilin-2 at the same site as Calpain-1 but with less efficacy

doi: 10.1042/BCJ20210629

Figure Lengend Snippet: Representative Western blots and quantification of in vitro β2-Spectrin ( A , B ), cardiac troponin T (cTnT) ( C , D ), and cardiac troponin I (cTnI) ( E , F ) cleavage assays in response to CAPN1 and CAPN2 expression and different Ca 2+ concentrations. Assays were performed as indicated and as described in . n = 3 for each group. Data are mean ± SEM. One-way ANOVA followed by the Tukey multiple-comparisons test. * P < 0.05.

Article Snippet: The pCMV6-XL5-CAPN1 and pCMV6-entry-CAPN2 plasmids expressing human CAPN1 and CAPN2 were provided by Dr. Tianqing Peng (Western University, Canada). cDNA of human cTnT (Origene, RC201218), human cTnI (Origene, RC214740) were fused with C-terminal Flag tag and human β2-Spectrin (Addgene, #31070) were fused with C-terminal HA tag.

Techniques: Western Blot, In Vitro, Expressing

Journal: Nature Communications

Article Title: The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

doi: 10.1038/s41467-022-29733-3

Figure Lengend Snippet: a Real-time quantitative polymerase chain reaction (RT-qPCR)-determined levels of the designated mRNAs during human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) differentiation. Levels of indicated genes are shown normalized to RPLP0 levels at each timepoint. Day 0 and 2, n = 9; Day 4, 6, 8, and 15, n = 8. * p < 0.05, *** p < 0.001, **** p < 0.0001 vs Day 0; one-way ANOVA followed by Dunnett’s multiple comparison test. b Representative immunoblots of estrogen-related receptor (ERR) α and γ protein levels during hiPSC-CM differentiation. α-Tubulin was used as a loading control. c Heatmap representing log 2 fold change [ERRα/γ knockout (KO)/wild-type control (WT)] of mRNA levels of genes involved in cardiac structural component, Ca 2+ handling function, and ion transport in two different KO cell lines (KO1, KO6). All changes were significant (Benjamini-Hochberg false discovery rate <0.05) except for SCN5A in KO1 ( p = 0.057). d Representative immunoblots of indicated protein levels in WT Control and ERRα/γ KO hiPSC-CMs. Total protein staining was shown as a loading control. e The mRNA levels of indicated genes determined by RT-qPCR in hiPSC-CMs following the infection of adenovirus expressing GFP or ERRγ (Ad-GFP or Ad-ERRγ). ** p < 0.01, *** p < 0.001, **** p < 0.0001, Ad-ERRγ vs Ad-GFP, two-tailed student’s t-test. PDK4 (Ad-GFP, n = 7 and Ad-ERRγ, n = 8); COX6A2 (Ad-GFP, n = 14 and Ad-ERRγ, n = 15); MDH1 and CKMT2 , (Ad-GFP, n = 7 and Ad-ERRγ, n = 8); TNNI3 (Ad-GFP, n = 14 and Ad-ERRγ, n = 15); TNNI1 (Ad-GFP, n = 10 and Ad-ERRγ, n = 11). f The mRNA levels of indicated hiPSC-CM maturation markers determined by RT-qPCR in WT and ERRα/γ KO hiPSC-CMs derived from KO1 and KO6 with ( n = 15) or without ( n = 9) maturation cocktail treatment for 7 days. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, WT vs ERRα/γ KO hiPSC-CMs, two-way ANOVA followed by Tukey’s multiple comparisons test. All graphs in a , e , and f represent the means ± SEM. n denotes independent biological replicates. g Representative immunoblot images to show ACSL1 and TNNI3 protein expression levels in WT and ERRα/γ KO hiPSC-CMs with or without maturation cocktail. Total protein staining was used as a loading control.

Article Snippet: The following antibodies were used: ERRα; SMYD1; VDAC; ACSL3; TNNI3 (Abcam catalog and clone #s: ab76228; EPR46Y, ab181372; EPR13574(B)-30, ab15895, ab151959, and ab47003 respectively); His tag; α-tubulin; ACSL1 (Cell Signaling Technology catalog and clone #s: 12698S; D3I1O, 3873; DM1A, and 9189; D2H5 respectively); HA tag and MYL2 (Proteintech catalog #s 51064-2-AP and 10906-1-AP); α-Actinin, FLAG M2, TNNI1, ACTB, and PGC-1α (Sigma-Aldrich catalog and clone #s: A7732-100UL; EA-53, F1804-50UG; M2, AV42117-100UL, A5316; AC-74, and ST1202; 4C1.3); GATA4 and MYBPC3 (SANTA CRUZ BIOTECHNOLOGY catalog and clone #s, sc-25310; G-4, and sc-137180; E-7).

Techniques: Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Derivative Assay, Comparison, Western Blot, Control, Knock-Out, Staining, Infection, Expressing, Two Tailed Test

Journal: Nature Communications

Article Title: The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

doi: 10.1038/s41467-022-29733-3

Figure Lengend Snippet: a Venn diagram indicates the overlapped peaks from published GATA4 chromatin immunoprecipitation sequencing (ChIP-seq; GSE85631) and ERRγ ChIP-seq (GSE113784) datasets generated from human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). GATA4 and ERRγ-shared peaks (ERRγ + GATA4) are presented by orange-colored region. p values were calculated using Fisher’s exact test. b Pie chart represents the overlapped sites from published GATA4 ChIP-seq, cardiac super-enhancers (SEs; GSE85631), and ERRγ ChIP-seq (GSE113784) datasets in hiPSC-CMs. The 45 SEs where ERRγ and GATA4 peaks are colocalized within 200 base pairs are highlighted with yellow. c Top 20 transcription factors that significantly overlapped with ERRγ + GATA4 and ERRγ-GATA4 are plotted. ESRRA was ranked in the top 30 in the ERRγ + GATA4 analysis. Y axis represents the similarity (GIGGLE score) between published datasets and each peak set. d Bars indicate enrichment score for Gene Ontology (GO) Biological Process and GO Cellular Component terms significantly enriched in ERRγ + GATA4 targets. e Representative genomic browser tracks of TNNI3 , MYBPC3 , and MYH6-7 regions. f Bar graphs represent mRNA expression levels of a subset of ERRγ + GATA4 and ERRγ targets in Negative Control (NC) ( n = 9) and siGATA4#1 ( n = 7) or siGATA4#2 ( n = 7) transfected hiPSC-CMs. * p < 0.05, ** p < 0.01, **** p < 0.0001, one-way ANOVA followed by Dunnett’s multiple comparisons test. All bars represent the means ± SEM. n denotes independent biological replicates.

Article Snippet: The following antibodies were used: ERRα; SMYD1; VDAC; ACSL3; TNNI3 (Abcam catalog and clone #s: ab76228; EPR46Y, ab181372; EPR13574(B)-30, ab15895, ab151959, and ab47003 respectively); His tag; α-tubulin; ACSL1 (Cell Signaling Technology catalog and clone #s: 12698S; D3I1O, 3873; DM1A, and 9189; D2H5 respectively); HA tag and MYL2 (Proteintech catalog #s 51064-2-AP and 10906-1-AP); α-Actinin, FLAG M2, TNNI1, ACTB, and PGC-1α (Sigma-Aldrich catalog and clone #s: A7732-100UL; EA-53, F1804-50UG; M2, AV42117-100UL, A5316; AC-74, and ST1202; 4C1.3); GATA4 and MYBPC3 (SANTA CRUZ BIOTECHNOLOGY catalog and clone #s, sc-25310; G-4, and sc-137180; E-7).

Techniques: ChIP-sequencing, Generated, Derivative Assay, Expressing, Negative Control, Transfection

Journal: Nature Communications

Article Title: The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

doi: 10.1038/s41467-022-29733-3

Figure Lengend Snippet: a mRNA expression level of TNNI3 in wild-type (WT) control ( n = 6) and ERRα/γ knockout (KO) line 1 or 6 ( n = 3) human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) following the overexpression of PGC-1α and/or GATA4 (Ad-PGC-1α and/or Ad-GATA4). ** p < 0.01, **** p < 0.0001, two-way ANOVA followed by Tukey’s multiple comparisons test. b Representative immunoblot images to show hemagglutinin (HA)-tagged GATA4, His-tagged PGC-1α, TNNI3, and TNNI1. c Levels of HA-tagged GATA4 occupation on the indicated targets in WT ( n = 5 or n = 4 for TNNI3 and POU5F1 ) and ERRα/γ KO hiPSC-CMs ( n = 5). * p < 0.05 vs WT, two-tailed student’s t -test. MYH6 -7 denotes the enhancer site of MYH6-7 cluster. d Bar graphs represent relative light units (RLU) derived from TNNI3 promoter-luciferase reporter ( TNNI3 -luc) with overexpression of ERRγ, GATA4, and PGC-1α in H9c2 myoblast. of TNNI3 -luc. n = 12, **** p < 0.0001, one-way ANOVA followed by Tukey multiple comparisons test. e Bar graphs represent the RLU derived from TNNI3 -luc or COX6A2 luciferase reporter ( COX6A2 -luc) in H9c2 myoblasts transduced with CRISPR/Cas9 and non-targeting control (NTC) guide RNA (gRNA) or gRNA targeting ERRα and γ (ERRα/γ KD). n = 12 for NTC in COX6A2 -luc or n = 9 for others. *** p < 0.001, **** p < 0.0001, two-way ANOVA followed by Tukey’s multiple comparisons test. f Schematic of TNNI3 promoter with putative ERR response elements (ERRE) and GATA binding sites. g WT ( n = 25), ERRE-deleted ( n = 12) or GATA binding sites-deleted ( n = 16) - TNNI3 -luc were assessed with overexpression of ERRγ and/or GATA4 in H9c2 myoblasts. * p < 0.05, **** p < 0.0001, two-way ANOVA followed by Tukey’s multiple comparisons test. h Bar graphs represents RLU derived from TNNI3 -luc with overexpression of ERRγ ( n = 15), GATA4 WT ( n = 15), C-teriminal zinc finger deleted GATA4 (C-zincΔ, n = 12), N-terminal zinc finger deleted GATA4 (N-zincΔ, n = 6), the combination of ERRγ and WT (n = 15) or each GATA4 mutant (C-zincΔ, n = 12; N-zincΔ, n = 6) in H9c2 myoblasts. * p < 0.05, **** p < 0.0001, one-way ANOVA followed by Tukey’s multiple comparisons test. i Bar graphs represents RLU derived from control reporter (empty-luc; n = 8 for ERRγ + PGC-1α or n = 9 for others) and MYH6-7 enhancer-luc ( n = 9) with overexpressed indicated factors in AC16 cells. **** p < 0.0001, two-way ANOVA followed by Tukey’s multiple comparisons test. All bars represent the means ± SEM. n denotes independent biological replicates.

Article Snippet: The following antibodies were used: ERRα; SMYD1; VDAC; ACSL3; TNNI3 (Abcam catalog and clone #s: ab76228; EPR46Y, ab181372; EPR13574(B)-30, ab15895, ab151959, and ab47003 respectively); His tag; α-tubulin; ACSL1 (Cell Signaling Technology catalog and clone #s: 12698S; D3I1O, 3873; DM1A, and 9189; D2H5 respectively); HA tag and MYL2 (Proteintech catalog #s 51064-2-AP and 10906-1-AP); α-Actinin, FLAG M2, TNNI1, ACTB, and PGC-1α (Sigma-Aldrich catalog and clone #s: A7732-100UL; EA-53, F1804-50UG; M2, AV42117-100UL, A5316; AC-74, and ST1202; 4C1.3); GATA4 and MYBPC3 (SANTA CRUZ BIOTECHNOLOGY catalog and clone #s, sc-25310; G-4, and sc-137180; E-7).

Techniques: Expressing, Control, Knock-Out, Derivative Assay, Over Expression, Western Blot, Two Tailed Test, Luciferase, Transduction, CRISPR, Binding Assay, Mutagenesis

Journal: Nature Communications

Article Title: The nuclear receptor ERR cooperates with the cardiogenic factor GATA4 to orchestrate cardiomyocyte maturation

doi: 10.1038/s41467-022-29733-3

Figure Lengend Snippet: a Bar graphs represent the relative light unit (RLU) from pG5luc construct. ERRγ-Gal4, GATA4/PGC-1α/ERRγ-Gal4, GATA4/ERRγ-Gal4, and PGC-1α/ERRγ-Gal4, n = 18; Gal4, n = 17; PGC-1α/Gal4, n = 16; GATA4/Gal4, n = 15; GATA4/PGC-1α/Gal4, PGC-1α m/ERRγ-Gal4, PGC-1α m/ERRγ-Gal4, n = 9; PGC-1α m/Gal4, n = 6, **** p < 0.0001, one-way ANOVA followed by Tukey’s multiple comparisons test. b Bar graphs represent the mRNA levels of indicated genes in wild-type control (WT, n = 8) and PGC-1α knockdown (KD, n = 8) human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 vs WT, two-tailed Student’s t-test. c Levels of ERRγ occupation on the indicated targets in WT (n = 6 for MYH6 -7 and NPPA-NPPB , n = 8 for POU5F1 , n = 5 for others), PGC-1α KD ( n = 8 for TNNI3 , TNNC1 , FABP3 , and CPT1B , n = 6 for MYBPC3 and POU5F1 , n = 7 for MYH6 -7, NPPA-NPPB , and ATP5B ), and ERRα/γ knockout (KO) hiPSC-CMs ( n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 vs WT, one-way ANOVA followed by Dunnett’s multiple comparisons test. Stem cell enhancer region on POU5F1 was used as a negative control. MYH6 -7 and NPPA-NPPB denote the distal enhancer sites upstream of each gene cluster. d Representative immunoblot images to show the interaction between FLAG-tagged ERRγ and endogenous GATA4 in FLAG-ERRγ overexpressed hiPSC-CMs. e Schematic to indicate reported naturally occurring GATA4 mutations. NLS denotes nuclear localization signal. ERRγ-Gal4 experiment was performed with PGC-1α and GATA4 natural mutants. Bar graphs indicate RLU from pG5luc construct in AD-293 cells. ** p < 0.01, *** p < 0.001, vs ERRγ, PGC-1α, and WT GATA4 transfected group, one-way ANOVA followed by Tukey multiple comparisons test. n = 6. f TNNI3 -luc reporter experiment with overexpression of ERRγ and GATA4 WT or GATA4 G296S in H9c2 myoblast. Bar graphs represent RLU of TNNI3 -luc. n = 9, **** p < 0.0001, one-way ANOVA followed by Tukey multiple comparisons test. g Venn diagram showing significant overlap (using Fisher’s exact test) between RNA-seq datasets generated in G296S GATA4 hiPSC-CMs (GSE85631) and ERRα/γ KO hiPSC-CMs (GSE165963). Bar graphs represent enrichment score for significantly enriched Gene Ontology (GO) Biological Process and GO Cellular Component with the commonly downregulated genes. All bars in a , b , c , e , and f represent the means ± SEM. n denotes independent biological replicates.

Article Snippet: The following antibodies were used: ERRα; SMYD1; VDAC; ACSL3; TNNI3 (Abcam catalog and clone #s: ab76228; EPR46Y, ab181372; EPR13574(B)-30, ab15895, ab151959, and ab47003 respectively); His tag; α-tubulin; ACSL1 (Cell Signaling Technology catalog and clone #s: 12698S; D3I1O, 3873; DM1A, and 9189; D2H5 respectively); HA tag and MYL2 (Proteintech catalog #s 51064-2-AP and 10906-1-AP); α-Actinin, FLAG M2, TNNI1, ACTB, and PGC-1α (Sigma-Aldrich catalog and clone #s: A7732-100UL; EA-53, F1804-50UG; M2, AV42117-100UL, A5316; AC-74, and ST1202; 4C1.3); GATA4 and MYBPC3 (SANTA CRUZ BIOTECHNOLOGY catalog and clone #s, sc-25310; G-4, and sc-137180; E-7).

Techniques: Construct, Control, Knockdown, Derivative Assay, Two Tailed Test, Knock-Out, Negative Control, Western Blot, Transfection, Over Expression, RNA Sequencing, Generated