rat cardiomyocyte cell line  (ATCC)

Journal: Redox Biology

Article Title: Peroxiredoxin Ⅲ safeguards cardiac function against doxorubicin by regulating mitochondrial quality control via H 2 O 2 detoxification

doi: 10.1016/j.redox.2026.104176

Figure Lengend Snippet: Doxorubicin increases mitochondrial H 2 O 2 levels and causes cell death in rat cardiomyocytes. (A) H9c2 cells were treated with the indicated concentrations of DOX for 24 h. Cell death was assessed by Annexin V-FITC staining and analyzed using flow cytometer. Representative histograms of three independent experiments are shown. (B) H9c2 cells were treated with 1 μM DOX for the indicated times. Cell viability was measured using WST-1 reagent. (C-D) H9c2 cells were treated with 1 μM DOX for the indicated times and then stained with 5 μM CM-H 2 DCFDA (C) or 5 μM MitoPY-1 (D) for 15 min. Relative fluorescence intensity (RFI) was measured by flow cytometer. All data are presented as mean ± S.D. (n = 3). Statistical significance was determined by one-way ANOVA followed by Dunnett's post hoc test. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 compared to untreated control.

Article Snippet: Rat cardiomyocyte cell line, H9c2 cells (CRL-1446) were obtained from the American Type Culture Collection (Manassas, VA, USA) and cultured in DMEM high glucose (SH30022.01, Hyclone, Logan, UT, USA) supplemented with 10% FBS, 1% antibiotic-antimycotic, and 1 μg/ml puromycin.

Techniques: Staining, Flow Cytometry, Fluorescence, Control

Journal: Redox Biology

Article Title: Peroxiredoxin Ⅲ safeguards cardiac function against doxorubicin by regulating mitochondrial quality control via H 2 O 2 detoxification

doi: 10.1016/j.redox.2026.104176

Figure Lengend Snippet: PrxⅢ regulates mitochondrial H 2 O 2 accumulation and oxidative damage in DOX-treated cardiomyocytes. (A) H9c2 cells stably expressing either control (pSUPER) or PrxⅢ-targeting siRNA (pSUPER-siPrxⅢ) were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 9 h. Mitochondrial H 2 O 2 levels were assessed using the mitochondria-targeted fluorescent probe MitoPY-1 and visualized by green fluorescence. Scale bar, 75 μm. Five randomly selected microscopic fields per sample were analyzed and combined to represent one biological replicate. (B) Cells were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 18 h. FOXO3a localization was assessed by immunofluorescence using anti-FOXO3a (green) antibody and DAPI (blue). Scale bar, 25 μm Five randomly selected microscopic fields per sample were analyzed and combined to represent one biological replicate. All data are expressed as mean ± S.D. Statistical significance was determined using two-way ANOVA followed by Bonferroni's post hoc test. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001 and ns, p > 0.05.

Article Snippet: Rat cardiomyocyte cell line, H9c2 cells (CRL-1446) were obtained from the American Type Culture Collection (Manassas, VA, USA) and cultured in DMEM high glucose (SH30022.01, Hyclone, Logan, UT, USA) supplemented with 10% FBS, 1% antibiotic-antimycotic, and 1 μg/ml puromycin.

Techniques: Stable Transfection, Expressing, Control, Transduction, Fluorescence, Immunofluorescence

Journal: Redox Biology

Article Title: Peroxiredoxin Ⅲ safeguards cardiac function against doxorubicin by regulating mitochondrial quality control via H 2 O 2 detoxification

doi: 10.1016/j.redox.2026.104176

Figure Lengend Snippet: PrxⅢ regulates oxidative damage and mitochondrial bioenergetics in DOX-treated cardiomyocytes. (A) Cells were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 12 h. Cells were labeled with 5 μM 10-NAO and relative fluorescence intensity (RFI) was analyzed by flow cytometer. Representative histograms and quantification are shown. (B) Cells were treated with 1 μM DOX for 12 h and labeled with 10 μM Rho-123 to assess mitochondrial membrane potential ( ΔΨ m ). The percentage of cells with low ΔΨ m was quantified by flow cytometry. Representative histograms and quantification are shown. (C) Cells were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 12 h. Representative bar graph showing relative ATP concentration normalized to protein contents. (D-H) Cells were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 12 h. (D) Representative tracing of the oxygen consumption rate (OCR). Arrows indicate time points when cells were treated with oligomycin, FCCP, and rotenone plus antimycin A (Rot/AA), respectively. The OCR is calculated as (E) basal respiration, (F) maximal respiration, (G) spare respiration, and (H) proton-leak. All data are expressed as mean ± S.D. (n = 3-5). Statistical significance was determined using two-way ANOVA followed by Bonferroni's post hoc test. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001.

Article Snippet: Rat cardiomyocyte cell line, H9c2 cells (CRL-1446) were obtained from the American Type Culture Collection (Manassas, VA, USA) and cultured in DMEM high glucose (SH30022.01, Hyclone, Logan, UT, USA) supplemented with 10% FBS, 1% antibiotic-antimycotic, and 1 μg/ml puromycin.

Techniques: Transduction, Labeling, Fluorescence, Flow Cytometry, Membrane, Concentration Assay

Journal: Redox Biology

Article Title: Peroxiredoxin Ⅲ safeguards cardiac function against doxorubicin by regulating mitochondrial quality control via H 2 O 2 detoxification

doi: 10.1016/j.redox.2026.104176

Figure Lengend Snippet: PrxⅢ regulates mitochondrial biogenesis in DOX-treated cardiomyocytes. (A) Cells were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 6 h. Total RNA was isolated and the mRNA levels of PGC-1α, Nrf1, and mtTFA were determined by qRT-PCR. Representative bar graphs showing relative expression normalized to GAPDH are presented. (B) Relative mRNA expression of the mitochondrial DNA-encoded cytochrome c oxidase (COX) mRNA was measured as an indicator of mtDNA copy-dependent transcription. Representative bar graphs showing relative expression normalized to GAPDH are presented. (C) Cells were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 18 h. Representative immunoblots and quantitative analyses of PGC-1α, Nrf1, and mtTFA proteins. GAPDH and α-tubulin were used as loading controls. Densitometric quantification of protein expression was performed from three independent biological replicates. All data are presented as mean ± S.D. (n = 3). Statistical significance was determined using two-way ANOVA followed by Bonferroni's post hoc test. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001.

Article Snippet: Rat cardiomyocyte cell line, H9c2 cells (CRL-1446) were obtained from the American Type Culture Collection (Manassas, VA, USA) and cultured in DMEM high glucose (SH30022.01, Hyclone, Logan, UT, USA) supplemented with 10% FBS, 1% antibiotic-antimycotic, and 1 μg/ml puromycin.

Techniques: Transduction, Isolation, Quantitative RT-PCR, Expressing, Western Blot

Journal: Redox Biology

Article Title: Peroxiredoxin Ⅲ safeguards cardiac function against doxorubicin by regulating mitochondrial quality control via H 2 O 2 detoxification

doi: 10.1016/j.redox.2026.104176

Figure Lengend Snippet: PrxⅢ protects cardiomyocytes against DOX-induced apoptotic cell death. Cells were transduced with Ad-PrxⅢ or Ad-Stuffer for 24 h and then exposed to 1 μM DOX for 18 h. (A) Cell lysates were subjected to immunoblot analyses for cleaved Caspase-3 and cleaved PARP-1 with normalization to β-actin. Densitometric quantification of protein expression was performed from three independent biological replicates. (B) Apoptotic cell death was assessed by Annexin V-FITC and 7-AAD double staining followed by flow cytometric analysis. Quantification of Annexin V and/or 7-AAD-positive cells is shown. All data are presented as mean ± S.D. Statistical significance was determined using two-way ANOVA followed by Bonferroni's post hoc test. ∗p < 0.05, ∗∗p < 0.01 and ∗∗∗p < 0.001.

Article Snippet: Rat cardiomyocyte cell line, H9c2 cells (CRL-1446) were obtained from the American Type Culture Collection (Manassas, VA, USA) and cultured in DMEM high glucose (SH30022.01, Hyclone, Logan, UT, USA) supplemented with 10% FBS, 1% antibiotic-antimycotic, and 1 μg/ml puromycin.

Techniques: Transduction, Western Blot, Expressing, Double Staining

Journal: Pharmaceuticals

Article Title: Comparison of Quercetin and Isoquercitrin’s Anti-Heart Failure Activity via MAPK Inflammatory Pathway and Caspase Apoptosis Pathway

doi: 10.3390/ph18101447

Figure Lengend Snippet: IQ and Que comparably protected H9c2 cells against Ang II induced damage. ( A ) The cytotoxicity of IQ was evaluated in H9c2 cells using CCK-8 assay. ( B ) The cytotoxicity of Que was evaluated in H9c2 cells using CCK-8 assay. ( C ) Ang II concentration optimization in H9c2 cells. ( D ) Protective effects of IQ and Que against Ang II induced cytotoxicity in H9c2 cardiomyocytes. ( E ) IQ and Que attenuate Ang II induced ROS production in H9c2 cells. (n = 3 data were presented as mean ± SEM, ++ p < 0.01, +++ p < 0.001, ++++ p < 0.0001 vs. Con, #### p < 0.0001 vs. Ang II, ** p < 0.01, *** p < 0.001, **** p < 0.0001, IQ vs. Que).

Article Snippet: The H9c2 rat cardiomyocyte cell line was purchased from Procell Life Science and Technology Company Limited Wuhan China.

Techniques: CCK-8 Assay, Concentration Assay

Journal: Pharmaceuticals

Article Title: Comparison of Quercetin and Isoquercitrin’s Anti-Heart Failure Activity via MAPK Inflammatory Pathway and Caspase Apoptosis Pathway

doi: 10.3390/ph18101447

Figure Lengend Snippet: IQ and Que treatments exerted a dose-dependent inhibitory effect on in vitro apoptosis. ( A ) Hoechst 33342 staining of apoptotic cells in IQ and Que treated groups (The scale bar represents 200 μm). ( B ) PI staining for Ang II induced apoptosis in H9c2 cells treated with IQ/Que (The scale bar represents 400 μm). ( C , D ) Quantification of Hoechst 33342 and PI fluorescence intensity. Data are presented as mean ± SD. (n = 3 data were presented as mean ± SEM, # p < 0.05, ## p < 0.01, vs. Ang II, ns, p > 0.05, IQ vs. Que).

Article Snippet: The H9c2 rat cardiomyocyte cell line was purchased from Procell Life Science and Technology Company Limited Wuhan China.

Techniques: In Vitro, Staining, Fluorescence

Journal: Pharmaceuticals

Article Title: Comparison of Quercetin and Isoquercitrin’s Anti-Heart Failure Activity via MAPK Inflammatory Pathway and Caspase Apoptosis Pathway

doi: 10.3390/ph18101447

Figure Lengend Snippet: WB analysis of inflammatory and apoptotic proteins in cardiomyocytes treated with IQ and Que in vitro. ( A ) IQ and Que regulate Ang II induced expression of apoptosis related proteins (Caspase-3/Bax/CytoC/Bcl-2) in H9c2 cells. Grayscale quantification of apoptotic markers (Bax/Bcl-2/Caspase-3/CytoC). ( B ) IQ and Que modulate Ang II induced expression of JNK/ERK/P38 and their phosphorylated forms in H9c2 cells. Quantification of inflammatory protein expression by densitometric analysis. Data are presented as mean ± SD. (n = 3 data were presented as mean ± SEM, # p < 0.05, ## p < 0.01, ### p < 0.001, #### p < 0.0001, vs. Ang II, ** p < 0.01, IQ vs. Que, ns, p > 0.05, IQ vs. Que).

Article Snippet: The H9c2 rat cardiomyocyte cell line was purchased from Procell Life Science and Technology Company Limited Wuhan China.

Techniques: In Vitro, Expressing

Journal: Frontiers in Pharmacology

Article Title: Mechanisms of sorafenib-induced cardiotoxicity: ER stress induces upregulation of ATF3, leading to downregulation of NDUFS1 expression and mitochondrial dysfunction

doi: 10.3389/fphar.2025.1593290

Figure Lengend Snippet: Sorafenib reduces viability in H9C2 cells and induces apoptosis. (A,B) Calcein AM/PI double staining was employed to evaluate sorafenib’s effect on H9C2 cell viability. Green: live cells (Calcein AM); red: dead cells (PI). n = 3 per group. Scale bar = 100 μm. (C) CCK-8 assays further assessed sorafenib’s impact on H9C2 cell viability (n = 6 per group). (D,E) Cleaved caspase-9 expression was analyzed using Western blot (n = 3 per group). (F) The apoptosis rate in rat cardiac H9c2 cells was assessed via fluorescence microscopy, with a scale bar of 100 μm. Statistical significance was denoted as ns for non-significance, * P < 0.05, ** P < 0.01, and *** P < 0.001 relative to the control group.

Article Snippet: The H9C2 rat cardiomyocyte cell line (CRL-1446) was obtained from the American Type Culture Collection (ATCC, United States) and cultured in a CO 2 incubator with 5% CO 2 using Dulbecco’s Modified Eagle’s Medium (DMEM, GIBCO, United States) supplemented with 10% fetal bovine serum (FBS, GIBCO, United States).

Techniques: Double Staining, CCK-8 Assay, Expressing, Western Blot, Fluorescence, Microscopy, Control

Journal: Frontiers in Pharmacology

Article Title: Mechanisms of sorafenib-induced cardiotoxicity: ER stress induces upregulation of ATF3, leading to downregulation of NDUFS1 expression and mitochondrial dysfunction

doi: 10.3389/fphar.2025.1593290

Figure Lengend Snippet: Endoplasmic reticulum stress inhibitor mitigates sorafenib-induced cardiomyocyte cell death. (A) Western blot analysis and quantification of P-PERK, PERK, P-eIF2α, eIF2α, GRP78, ATF4, and GAPDH. H9C2 cardiomyocytes were treated with 5 μM sorafenib (or vehicle) for 48 h. For rescue experiments, 1 μM GSK2606414 was co-administered. (B) Immunofluorescent staining was performed on cultured cells to analysis and quantification of P-eIF2α. (C) CCK-8 assays was performed to study how endoplasmic reticulum stress affects cell viability. (D) Calcein AM/PI staining assessed ER stress–induced changes in cell viability by distinguishing live and dead cells. Green: live cells (Calcein AM); red: dead cells (PI). Scale bar, 100 μm. Statistical significance was denoted as ns for non-significance, * P < 0.05, ** P < 0.01, and *** P < 0.001 relative to the control group.

Article Snippet: The H9C2 rat cardiomyocyte cell line (CRL-1446) was obtained from the American Type Culture Collection (ATCC, United States) and cultured in a CO 2 incubator with 5% CO 2 using Dulbecco’s Modified Eagle’s Medium (DMEM, GIBCO, United States) supplemented with 10% fetal bovine serum (FBS, GIBCO, United States).

Techniques: Western Blot, Staining, Cell Culture, CCK-8 Assay, Control

Journal: Frontiers in Pharmacology

Article Title: Mechanisms of sorafenib-induced cardiotoxicity: ER stress induces upregulation of ATF3, leading to downregulation of NDUFS1 expression and mitochondrial dysfunction

doi: 10.3389/fphar.2025.1593290

Figure Lengend Snippet: Inhibitors of endoplasmic reticulum stress can reduce cardiomyocyte apoptosis caused by sorafenib. (A) Cell apoptosis rate in rat cardiac H9c2 cells determined by flow cytometry. (B) The apoptosis rate in rat cardiac H9c2 cells was assessed via fluorescence microscopy, with a scale bar of 100 μm. (C) Western blot analysis was used to evaluate the expression of cleaved caspase-9 and cleaved caspase-3. Statistical significance was denoted as ns for non-significance, * P < 0.05, ** P < 0.01, and *** P < 0.001 relative to the control group.

Article Snippet: The H9C2 rat cardiomyocyte cell line (CRL-1446) was obtained from the American Type Culture Collection (ATCC, United States) and cultured in a CO 2 incubator with 5% CO 2 using Dulbecco’s Modified Eagle’s Medium (DMEM, GIBCO, United States) supplemented with 10% fetal bovine serum (FBS, GIBCO, United States).

Techniques: Flow Cytometry, Fluorescence, Microscopy, Western Blot, Expressing, Control

Journal: Frontiers in Pharmacology

Article Title: Mechanisms of sorafenib-induced cardiotoxicity: ER stress induces upregulation of ATF3, leading to downregulation of NDUFS1 expression and mitochondrial dysfunction

doi: 10.3389/fphar.2025.1593290

Figure Lengend Snippet: Proteomic analysis reveals ATF3 binds to the Ndufs1 promoter and negatively regulates its transcription. (A,B) Differentially expressed proteins in E45 and E52 were analyzed, with Venn diagrams showing their overlap. (C) Scatter plots from the GEPIA website, using GTEx database data, depict the correlation between Nudfs1 and ATF3 protein levels in H9C2 cells. (D) The regulatory sensitivity of ATF3 is evaluated. (E,F) The sequence logo of the NDUFS1 promoter region highlights ATF3 binding sites.

Article Snippet: The H9C2 rat cardiomyocyte cell line (CRL-1446) was obtained from the American Type Culture Collection (ATCC, United States) and cultured in a CO 2 incubator with 5% CO 2 using Dulbecco’s Modified Eagle’s Medium (DMEM, GIBCO, United States) supplemented with 10% fetal bovine serum (FBS, GIBCO, United States).

Techniques: Sequencing, Binding Assay

Journal: Frontiers in Pharmacology

Article Title: Mechanisms of sorafenib-induced cardiotoxicity: ER stress induces upregulation of ATF3, leading to downregulation of NDUFS1 expression and mitochondrial dysfunction

doi: 10.3389/fphar.2025.1593290

Figure Lengend Snippet: ATF3 binds to the Ndufs1 promoter suppressing its transcription and inducing apoptosis and mitochondrial dysfunction. (A) Western blot was used to assess the effect of ATF3 knockout on Ndufs1 and cleaved caspase-9 expressio (n = 3 per group). (B) RealTime-qPCR was used to assess the effect of ATF3 knockout on ATF3 and Ndufs1 (n = 3 per group). (C) The apoptosis rate in rat cardiac H9c2 cells was assessed via fluorescence microscopy, with a scale bar of 100 μm. (D) ChIP assays with an ATF3-specific antibody were conducted on H9C2 cells, and the precipitated DNA was quantified using real-time PCR (n = 3 per group). (E) Mitochondrial complex I activity was quantitatively analyzed in H9C2 cells (n = 4 per group). (F) ATP levels in H9C2 cells were quantified using an ATP Colorimetric Assay kit (n = 4 per group). (G,H) Mitochondrial membrane potential was evaluated via JC-1 staining. Statistical significance was denoted as ns for non-significance, * P < 0.05, ** P < 0.01, and *** P < 0.001 relative to the control group.

Article Snippet: The H9C2 rat cardiomyocyte cell line (CRL-1446) was obtained from the American Type Culture Collection (ATCC, United States) and cultured in a CO 2 incubator with 5% CO 2 using Dulbecco’s Modified Eagle’s Medium (DMEM, GIBCO, United States) supplemented with 10% fetal bovine serum (FBS, GIBCO, United States).

Techniques: Western Blot, Knock-Out, Fluorescence, Microscopy, Real-time Polymerase Chain Reaction, Activity Assay, Colorimetric Assay, Membrane, Staining, Control