PFKM inhibits doxorubicin-induced cardiotoxicity by enhancing oxidative phosphorylation and glycolysis

doi:10.21203/rs.3.rs-1664969/v1

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PFKM inhibits doxorubicin-induced cardiotoxicity by enhancing oxidative phosphorylation and glycolysis

https://doi.org/10.21203/rs.3.rs-1664969/v1

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Background: Heart failure (HF) is a global pandemic which affects about 26 million people. PFKM (Phosphofructokinase, Muscle), catalyzing the phosphorylation of fructose-6-phosphate, plays a very important role in cardiovascular diseases. However, the effect of PFKM in glycolysis and HF remains to be elucidated.

Materials and methods: H9c2 rat cardiomyocyte cells were used in this study. Cells were treated with doxorubicin (DOX) to establish injury models. Cell viability was measured using CCK-8 kits. Apoptosis was estimated by TdT-mediated dUTP Nick-End Labeling (TUNEL) staining. Levels of lactate and ATP were measured using commercial kits. Extracellular acidification rate (ECAR) and Oxygen consumption rate (OCR) were determined using an XFe24 Extracellular Flux Analyzer. Chromatin immunoprecipitation (ChIP) assays were used to study the interaction between H3K27ac or histone deacetylase 1 (HDAC1) and the PFKM promoter region. Phosphoglycerate kinase 1 (PGK1) was either silenced or overexpressed to study its role. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) and immunoblotting were used for gene expression.

Results: DOX treatment significantly inhibited PFKM expression in H9c2 cells. Overexpression of PFKM inhibited DOX-induced cell apoptosis and DOX-decreased glycolysis and oxidative phosphorylation, while silencing PFKM promoted cell apoptosis and inhibited glycolysis and oxidative phosphorylation in H9c2 cells. Moreover, PFKM regulated DOX-mediated cell viability and apoptosis through glycolysis pathway. Mechanism study showed that HDAC1 inhibited H3K27ac-induced transcription of PFKM in DOX-treated cells and regulated glycolysis.

Conclusion: PFKM could inhibit DOX-induced cardiotoxicity by enhancing oxidative phosphorylation and glycolysis, which might benefit us in developing novel therapeutics for prevention or treatment of HF.

Heart failure (HF) is a clinical syndrome caused by defects in myocardium resulting in impairment of ventricular filling or the ejection of blood [1]. HF symptoms include breathlessness, ankle swelling, and fatigue, accompanied by pulmonary rales, peripheral oedema, etc. [2]. HF is a global pandemic and its prevalence is still increasing [3]. The major HF risk factors heart disease, cardiopulmonary disease, etc. [4]. Evaluation factors for HF include physical examination, blood tests, levels of serum creatinine and glucose, liver function tests etc. [1]. Measurement of plasma concentrations of brain natriuretic peptide is a mainstay for the diagnosis of HF [5]. The burden of HF is huge, study show that estimated mean cost of HF was $11 552 in 2014 in USA [6]. More importantly, HF is still increasing in prevalence [3]. Therefore, a better understanding of the pathogenesis of HF will benefit us in the prevention and treatment of HF.

PFKM (Phosphofructokinase, Muscle) regulates glycolysis via catalyzing the phosphorylation of fructose-6-phosphate [7]. PFKM (Phosphofructokinase, Muscle) has been shown to be involved in various pathological processes. For example, Gao et al. have reported that S-nitrosylation at Cys351 of PFKM promoted cell proliferation, and increased tumor growth and metastasis of ovarian cancer [8]. A study identified PFKM as a breast cancer gene [9]. Another study indicated that celastrol directly inhibits PFKM to induce weight loss [10]. PFKM also plays a very important role in cardiovascular diseases. For example, PFKM mutation causes myopathy [7]. PFK deficiency results in a severe cardiac and hematological disorder [11]. However, the exact role of PFKM in cardiotoxicity remains elusive.

Glycolysis and oxidative phosphorylation (OXPHOS) produce energy for cells [12]. OXPHOS is the process by which ATP synthesis is coupled to the movement of electrons through the mitochondrial electron transport chain and the associated consumption of oxygen [13]. Glycolysis converts glucose to lactate and provides ATP under anaerobic conditions [14]. Dysfunction of OXPHOS or glycolysis has been associated with a variety of diseases. For instance, it has been reported that defects in OXPHOS in insulin-sensitive tissues contribute to type 2 diabetes [15]. Defect of OXPHOS system has been linked to neurodegeneration including Alzheimer disease, Huntington disease, etc. [16]. An adipocyte-specific defect in OXPHOS increases systemic energy expenditure [17]. OXPHOS also regulates cardiovascular diseases. For example, coronary artery disease subjects showed suppressed function of complexes I, II and III [18]. Glycolysis also plays a role in HF. Increased glycolysis was found in failing hearts [19]. HF animals showed a remarkable increase in glycolysis [20]. Despite the advance of studies on HF, PFKM, and OXPHOS/glycolysis, the exact role of PFKM in OXPHOS/glycolysis and how it does affect HF are still unclear and remain to be elucidated.

Cell culture

The H9c2 cells, purchased from ATCC (Manassas, VA), were cultured in Dulbecco's modified eagle medium (DMEM) with 10% Fetal bovine serum (FBS) at 37°C. Cardiac injury model was induced by providing different concentrations of DOX (Chroma Bio., Chengdu, China) for indicated times with/without 2-Deoxy-D-glucose (2-DG) (5 mM) or Mocetinostat (MGCD; 0.1 µM).

Plasmids Or Sirna Transfection

Transfections were conducted with the Lipofectamine 3000 (Invitrogen) as per the supplier’s protocol. pcDNA3.1 plasmid was used to construct the PFKM overexpression vector. Small interfering RNAs targeting PFKM (siPFKM-1, GGAGGTATACAAGCTTCTA (sense), TAGAAGCTTGTATACCTCC (reverse) and siPFKM-2, GGCGAGTGTTTATCATCGA (sense), TCGATGATAAACACTCGCC (reverse)) and nonspecific siRNA were obtained from Shanghai GenePharma Co., Ltd. Empty vector and nonspecific siRNA were utilized as controls.

Cell Viability Assay

Cell growth was measured using CCK-8 kits. Briefly, 24 hours after treatment, cells were cultured in 96-well plates and incubated with CCK-8 for 1 h and OD450 was recorded.

Tdt-mediated Dutp Nick-end Labeling (Tunel) Staining

TUNEL staining was used for apoptosis [21]. Cell nucleus was stained with 4',6-diamidino-2-phenylindole (DAPI) and observed under a fluorescence microscope.

Extracellular Flux (Xf) Analysis

Twenty-four hours after treatment, glycolysis and mitochondrial respiration levels were monitored by measuring extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) using a Seahorse XF24 Extracellular Flux Analyzer [22].

Measurement Of Lactate And Atp

Twenty-four hours after treatment, cells were cultured in the 6-well plate for 24h. The lactate release and ATP content was measured with Lactic Acid assay kit) and ATP assay kit (Jiancheng Bio., Nanjing), respectively.

Reverse Transcription-polymerase Chain Reaction (Rt-pcr)

Total RNAs were extracted using TRIzol reagents and reverse transcribed to cDNA. Levels of interested genes were assessed by Quantitative RT-PCR using SYBR Green Master Mix (Roche, Shanghai) with following primers: PFKM: 5’-ATCACAGCCGAGGAGGCTAC-3’ (F), 5'-GGCGGCCCATCACTTCTAAC-3' (R); β-actin: 5’-CGGTCAGGTCATCACTATC-3’ (F), 5'-CAGGGCAGTAATCTCCTTC-3' (R). Fold change was calculated using 2^−ΔΔCt formula.

Immunoblotting

Total protein was isolated, quantified, resolved, transferred to PVDF membranes, and probed with primary antibodies, including antiPFKM antibody (ab154804, Abcam), antiBcl-2 antibody (ab182858, Abcam), antiBax antibody (ab32503, Abcam, dilution: 1:1,0000), antiH3K27ac antibody (ab177178, Abcam), antiHDAC1 antibody (10197-1-AP, Proteintech), and anti-beta actin antibody (66009-1-Ig, Proteintech). ECL was used for visualization.

Chromatin Immunoprecipitation (Chip)

ChIP assay was performed [23] with antibody against H3K27ac or control IgG (Cell Signaling). Purified ChIP DNA was confirmed by Quantitative RT-PCR. PFKM primers sequences: 5’-CAACACCACCACTACCTT-3’ (forward), 5’-CACTGCCATCAAACAAAC-3’ (reverse).

Statistical analysis

Data were expressed as mean ± SD and analyzed by Prism8.4.2. Comparisons between 2 groups were performed with Student's t test, and multiple comparisons were performed with one-way ANOVA. P < 0.05 was defined statistically significant.

DOX treatment inhibited PFKM expression in H9c2 cells

To study the role of PFKM in cardiac injury, DOX treatment was used to induce in vitro cardiac injury. DOX treatment not only dose-dependently decreased the expression of PFKM (Fig. 1A-1C), but also time-dependently suppressed the expression of PFKM (Fig. 1D-1F), suggesting that DOX inhibited PFKM expression in H9c2 cells.

Pfkm Upregulation Abolished Dox-induced Cell Apoptosis And Dox-suppressed Oxidative Phosphorylation And Glycolysis In H9c2 Cells

To further investigate the role of PFKM, PFKM was overexpressed. PFKM overexpression significantly increased DOX-inhibited cell viability (Fig. 2A), decreased DOX-promoted apoptosis (Fig. 2B-2C), and reversed DOX-decreased PFKM, Bcl-2, and DOX-increased Bax (Fig. 2D-2E). Overexpression of PFKM also abolished DOX-inhibited OCR and ECAR (Fig. 2F-2G), and reversed DOX-decreased levels of ATP (Fig. 2H) and lactate levels (Fig. 2I). The results show that overexpressing PFKM abolished DOX-induced cell apoptosis and DOX-suppressed OXPHOS and glycolysis.

Pfkm Downregulation Promoted Cell Apoptosis And Inhibited Oxidative Phosphorylation And Glycolysis In H9c2 Cells

Next, PFKM was silenced to further study its role. Silencing PFKM significantly inhibited cell viability (Fig. 3A), promoted apoptosis (Fig. 3B-3C), and inhibited PFKM, Bcl-2, but increased Bax (Fig. 3D-3E). Silencing PFKM also suppressed OCR and ECAR (Fig. 3F-3G), and decreased levels of ATP (Fig. 3H) and lactate (Fig. 3I). The results demonstrate that PFKM downregulation promoted cell apoptosis and inhibited oxidative phosphorylation and glycolysis in H9c2 cells.

Pfkm Regulated Dox-mediated Cell Viability And Apoptosis Via Glycolysis

To find out how PFKM regulates cell viability, glycolysis inhibitor, 2-DG, was introduced. Results showed that inhibition of glycolysis not only significantly promoted DOX-suppressed cell viability, but also abolished PFKM-overexpression-increased cell viability (Fig. 4A). TUNEL staining results showed that inhibition of glycolysis not only significantly increased DOX-increased cell apoptosis, but also abolished PFKM-overexpression-decreased cell apoptosis (Fig. 4B-4C). The findings suggest that PFKM regulates DOX-mediated growth and apoptosis via the glycolysis pathway.

Hdac1 Inhibited H3k27ac-induced Transcription Of Pfkm In Dox-induced H9c2 Cells

To figure out the mechanism by which PFKM is regulated, levels of H3K27ac and HDAC1 in DOX-treated H9c2 cells were measured. Results showed that DOX suppressed H3K27ac in a time-dependent manner, but increased the expression of HDAC1 (Fig. 5A-5B). ChIP assay revealed that DOX treatment significantly suppressed the interaction between H3K27ac and the PFKM promoter (Fig. 5C). HDAC inhibitor, mocetinostat (MGCD), significantly increased the interaction between H3K27ac and the PFKM promoter (Fig. 5D). Inhibition of HDAC1 also significantly increased PKFM and H3K27ac expression (Fig. 5E-5G). Together, these results suggest that HDAC1 inhibited H3K27ac-induced transcription of PFKM in DOX-induced H9c2 cells.

Hdac1-induced Pfkm Transcriptional Repression Regulated Dox-mediated Oxidative Phosphorylation And Glycolysis

To further study the role of HDAC1, MGCD and DOX were used to treat H9c2 cells transfected with PFKM small interfering RNA (siRNA) or nonspecific siRNA (siNC). Results showed that MGCD treatment abolished DOX-suppressed cell viability (Fig. 6A), and ameliorated DOX-inhibited OCR and ECAR (Fig. 6B-6C). MGCD treatment also reversed DOX-decreased levels of ATP (Fig. 6D) and lactate (Fig. 6E), suggesting that HDAC1-induced PFKM transcriptional repression regulated DOX-mediated oxidative phosphorylation and glycolysis in H9c2 cells.

We revealed that DOX treatment significantly inhibited PFKM expression in H9c2 cells. Overexpressing of PFKM inhibited DOX-induced cell apoptosis and DOX-decreased glycolysis, while silencing PFKM promoted cell apoptosis and inhibited oxidative phosphorylation and glycolysis in H9c2 cells. Moreover, PFKM regulated DOX-mediated cell growth and apoptosis via glycolysis pathway. Data also supports that the expression of PFKM was suppressed by HDAC1 through regulating H3K27 acetylation. For the first time, we show that HDAC1-mediated PFKM down-regulation promoted cell apoptosis and inhibited oxidative phosphorylation in H9c2 cells through regulating glycolysis, which may provide novel directions for new drug development.

Glycolysis regulated energy metabolism [24]. Glucose is converted into pyruvate, NADH, and ATP by glycolysis [25]. Glycolysis involves in many biological and pathological processes. For example, glycolysis promotes tumor growth [26]. Preclinical studies demonstrate that some small molecules such as 3-bromopyruvate suppresses cancer via targeting glycolysis [27]. Another study showed that Smad4 depletion in podocytes protects mice from glomerulosclerosis [28]. Inhibition of aerobic glycolysis causes depression of cardiac excitability and can lead to Ca²⁺ alternant in cardiac tissue [29]. Increased glycolysis is the earliest energy metabolic change during heart failure with preserved ejection fraction [30]. Other studies have demonstrated that glycolysis affects sarcoplasmic reticulum (SR) function and SR Ca²⁺ release not only through generation of ATP but also through direct interactions of glycolytic intermediates and products with the Ca²⁺ release channel itself [29]. In this study, we demonstrated that suppressing glycolysis remarkably ameliorated the effect of PFKM on DOX-mediated cell growth and apoptosis. These results reveal a very important role of glycolysis in regulating DOX-mediated cell growth and apoptosis and improve our knowledge of the role of glycolysis in cardiotoxicity and HF.

PFK catalyzes the rate-limiting phosphorylation of fructose-6-phosphate and sustains a high rate of glycolysis [7]. It has 3 isoforms: platelet (PFKP), muscle (PFKM), and liver (PFKL) [31]. PFKM gene has 24 exons [32]. PFK deficiency belongs to glycogen storage disease characterized by weakness with spasms and cramping on exercise [32]. Ristow et al. have reported that deficiency of PFKM results in insulin resistance, contributing to diabetes [33]. Studies also indicate that PFKM plays a very important role in cardiovascular diseases. For instance, Garcia et al. indicated that PFK deficiency causes a cardiac and hematological disorder [11]. Two-month-old PFKM knockout mice developed cardiac hypertrophy and evident cardiomegaly with age [11]. Our findings indicate a very important role of PFKM in regulating the proliferation of cardiomyoblasts and cardiotoxicity and improve our knowledge of FPKM in the pathogenesis of HF.

Protein acetylation is the process that the acetyl group is transferred to a polypeptide chain [34]. Acetylation alters protein function [34]. Protein acetylation plays a very important role in diverse physiological processes [35, 36]. H3K27ac involves in the higher activation of transcription [37]. It is elevated in mammary cancer and administration of H3K27ac inhibitor repressed tumor formation [38]. Felice et al. demonstrated that hypoacetylation of H3K27 involves in intestinal inflammation [39]. H3K27ac also involves in cardiovascular diseases. For example, a study indicated that H3K27ac acetylation status regulates phenotypic response in HF [40]. Papait et al. reported that H3K27ac was decreased in mice with transverse aortic constriction [41]. Our findings suggested that HDAC1 significantly decreased the level of H3K27ac to suppress the transcription of PFKM and regulate oxidative phosphorylation and glycolysis. These findings indicated the significance of HDAC1/H3K27ac/PFKM axis in cardiotoxicity and HF, which may benefit the study of HF and cardiovascular diseases. Keep in mind that only in vitro cell experiments were used in this study. Future studies with animals or even clinical samples will definitely supply more meaningful data. Nevertheless, our study revealed new roles of PFKM and glycolysis in HF.

In conclusion, PFKM could inhibit doxorubicin-induced cardiotoxicity by enhancing oxidative phosphorylation and glycolysis. The findings demonstrate significance of PFKM and glycolysis which might benefit us in developing novel therapeutics for prevention or treatment of HF.

Author contributions

Min Zhou wrote the main manuscript text, Xiao Sun and Chunli Wang prepared figures 1-3 and participated in statistical analyses, Fengdan Wang and Chuibi Fang prepared figures 4-6 and participated in molecular biology experiments, Zhenlei Hu participated in the conceptualization of this study. All authors reviewed this manuscript.

Competing interests

The authors declare no competing interests.

Data availability

We confirm that all data generated or analyzed during this study are available from the corresponding author on reasonable request.

Inamdar, A.A. and A.C. Inamdar, Heart Failure: Diagnosis, Management and Utilization. J Clin Med, 2016. 5(7).
McDonagh, T.A., et al., 2021 ESC Guidelines for the diagnosis and treatment of acute and chronic heart failure. Eur Heart J, 2021. 42(36): p. 3599–3726.
Savarese, G. and L.H. Lund, Global Public Health Burden of Heart Failure. Card Fail Rev, 2017. 3(1): p. 7–11.
Khatibzadeh, S., et al., Worldwide risk factors for heart failure: a systematic review and pooled analysis. Int J Cardiol, 2013. 168(2): p. 1186–94.
Metra, M. and J.R. Teerlink, Heart failure. Lancet, 2017. 390(10106): p. 1981–1995.
Jackson, S.L., et al., National Burden of Heart Failure Events in the United States, 2006 to 2014. Circ Heart Fail, 2018. 11(12): p. e004873.
Vasconcelos, O., et al., Nonsense mutation in the phosphofructokinase muscle subunit gene associated with retention of intron 10 in one of the isolated transcripts in Ashkenazi Jewish patients with Tarui disease. Proc Natl Acad Sci U S A, 1995. 92(22): p. 10322–6.
Gao, W., et al., The role of S-nitrosylation of PFKM in regulation of glycolysis in ovarian cancer cells. Cell Death Dis, 2021. 12(4): p. 408.
Ahsan, H., et al., A genome-wide association study of early-onset breast cancer identifies PFKM as a novel breast cancer gene and supports a common genetic spectrum for breast cancer at any age. Cancer Epidemiol Biomarkers Prev, 2014. 23(4): p. 658–69.
Pfuhlmann, K., et al., Celastrol-Induced Weight Loss Is Driven by Hypophagia and Independent From UCP1. Diabetes, 2018. 67(11): p. 2456–2465.
Garcia, M., et al., Phosphofructo-1-kinase deficiency leads to a severe cardiac and hematological disorder in addition to skeletal muscle glycogenosis. PLoS Genet, 2009. 5(8): p. e1000615.
Wang, Z., et al., Glycolysis and Oxidative Phosphorylation Play Critical Roles in Natural Killer Cell Receptor-Mediated Natural Killer Cell Functions. Front Immunol, 2020. 11: p. 202.
Wilson, D.F., Oxidative phosphorylation: regulation and role in cellular and tissue metabolism. J Physiol, 2017. 595(23): p. 7023–7038.
Kim, Y.H., T. Nakayama, and J. Nayak, Glycolysis and the Hexosamine Biosynthetic Pathway as Novel Targets for Upper and Lower Airway Inflammation. Allergy Asthma Immunol Res, 2018. 10(1): p. 6–11.
Wang, M., et al., Impaired mitochondrial oxidative phosphorylation in multiple insulin-sensitive tissues of humans with type 2 diabetes mellitus. J Int Med Res, 2010. 38(3): p. 769–81.
Koopman, W.J., et al., OXPHOS mutations and neurodegeneration. EMBO J, 2013. 32(1): p. 9–29.
Choi, M.J., et al., An adipocyte-specific defect in oxidative phosphorylation increases systemic energy expenditure and protects against diet-induced obesity in mouse models. Diabetologia, 2020. 63(4): p. 837–852.
Ait-Aissa, K., et al., Mitochondrial Oxidative Phosphorylation defect in the Heart of Subjects with Coronary Artery Disease. Sci Rep, 2019. 9(1): p. 7623.
Karwi, Q.G., et al., Loss of Metabolic Flexibility in the Failing Heart. Front Cardiovasc Med, 2018. 5: p. 68.
Riehle, C., et al., PGC-1beta deficiency accelerates the transition to heart failure in pressure overload hypertrophy. Circ Res, 2011. 109(7): p. 783–93.
Gao, L., et al., Novel role for caspase-activated DNase in the regulation of pathological cardiac hypertrophy. Hypertension, 2015. 65(4): p. 871–81.
Guo, Y., et al., Resibufogenin suppresses tumor growth and Warburg effect through regulating miR-143-3p/HK2 axis in breast cancer. Mol Cell Biochem, 2020. 466(1–2): p. 103–115.
Zhu, W., et al., FKBP3 Promotes Proliferation of Non-Small Cell Lung Cancer Cells through Regulating Sp1/HDAC2/p27. Theranostics, 2017. 7(12): p. 3078–3089.
Brooks, G.A., Cell-cell and intracellular lactate shuttles. J Physiol, 2009. 587(Pt 23): p. 5591–600.
Tran, D.H. and Z.V. Wang, Glucose Metabolism in Cardiac Hypertrophy and Heart Failure. J Am Heart Assoc, 2019. 8(12): p. e012673.
Bonnet, S., et al., A mitochondria-K + channel axis is suppressed in cancer and its normalization promotes apoptosis and inhibits cancer growth. Cancer Cell, 2007. 11(1): p. 37–51.
Akins, N.S., T.C. Nielson, and H.V. Le, Inhibition of Glycolysis and Glutaminolysis: An Emerging Drug Discovery Approach to Combat Cancer. Curr Top Med Chem, 2018. 18(6): p. 494–504.
Li, J., et al., Smad4 promotes diabetic nephropathy by modulating glycolysis and OXPHOS. EMBO Rep, 2020. 21(2): p. e48781.
Kockskamper, J., A.V. Zima, and L.A. Blatter, Modulation of sarcoplasmic reticulum Ca2 + release by glycolysis in cat atrial myocytes. J Physiol, 2005. 564(Pt 3): p. 697–714.
Fillmore, N., et al., Uncoupling of glycolysis from glucose oxidation accompanies the development of heart failure with preserved ejection fraction. Mol Med, 2018. 24(1): p. 3.
Lee, J.H., et al., Stabilization of phosphofructokinase 1 platelet isoform by AKT promotes tumorigenesis. Nat Commun, 2017. 8(1): p. 949.
Vives-Corrons, J.L., et al., First description of phosphofructokinase deficiency in spain: identification of a novel homozygous missense mutation in the PFKM gene. Front Physiol, 2013. 4: p. 393.
Ristow, M., et al., Deficiency of phosphofructo-1-kinase/muscle subtype in humans impairs insulin secretion and causes insulin resistance. J Clin Invest, 1997. 100(11): p. 2833–41.
Drazic, A., et al., The world of protein acetylation. Biochim Biophys Acta, 2016. 1864(10): p. 1372–401.
Son, S.M., et al., Autophagy regulation by acetylation-implications for neurodegenerative diseases. Exp Mol Med, 2021. 53(1): p. 30–41.
Christensen, D.G., et al., Mechanisms, Detection, and Relevance of Protein Acetylation in Prokaryotes. mBio, 2019. 10(2).
Herrera-Uribe, J., et al., Changes in H3K27ac at Gene Regulatory Regions in Porcine Alveolar Macrophages Following LPS or PolyIC Exposure. Front Genet, 2020. 11: p. 817.
Li, Q.L., et al., The hyper-activation of transcriptional enhancers in breast cancer. Clin Epigenetics, 2019. 11(1): p. 48.
Felice, C., et al., Intestinal Inflammation is Linked to Hypoacetylation of Histone 3 Lysine 27 and can be Reversed by Valproic Acid Treatment in Inflammatory Bowel Disease Patients. Cell Mol Gastroenterol Hepatol, 2021. 11(3): p. 889–891 e6.
Pei, J., et al., H3K27ac acetylome signatures reveal the epigenomic reorganization in remodeled non-failing human hearts. Clin Epigenetics, 2020. 12(1): p. 106.
Liu, C.F. and W.H.W. Tang, Epigenetics in Cardiac Hypertrophy and Heart Failure. JACC Basic Transl Sci, 2019. 4(8): p. 976–993.

No competing interests reported.

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DOX treatment inhibited PFKM expression in H9c2 cells. (A-C) PFKM expression in DOX-treated H9c2 cells. (D-F) PFKM expression in DOX-treated H9c2 cells for different time points. * P < 0.05, *** P < 0.001 vs 0 µM or 0 h.

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You are reading this latest preprint version

PFKM inhibits doxorubicin-induced cardiotoxicity by enhancing oxidative phosphorylation and glycolysis

Status:

Version 1

Abstract

Figures

Introduction

Materials And Methods

Cell culture

Plasmids Or Sirna Transfection

Cell Viability Assay

Tdt-mediated Dutp Nick-end Labeling (Tunel) Staining

Extracellular Flux (Xf) Analysis

Measurement Of Lactate And Atp

Reverse Transcription-polymerase Chain Reaction (Rt-pcr)

Immunoblotting

Chromatin Immunoprecipitation (Chip)

Statistical analysis

Results

DOX treatment inhibited PFKM expression in H9c2 cells

Pfkm Upregulation Abolished Dox-induced Cell Apoptosis And Dox-suppressed Oxidative Phosphorylation And Glycolysis In H9c2 Cells

Pfkm Downregulation Promoted Cell Apoptosis And Inhibited Oxidative Phosphorylation And Glycolysis In H9c2 Cells

Pfkm Regulated Dox-mediated Cell Viability And Apoptosis Via Glycolysis

Hdac1 Inhibited H3k27ac-induced Transcription Of Pfkm In Dox-induced H9c2 Cells

Hdac1-induced Pfkm Transcriptional Repression Regulated Dox-mediated Oxidative Phosphorylation And Glycolysis

Discussion

Declarations

References

Competing Interests

Supplementary Files

Status:

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