Spermidine Prevents Heart Injury in Neonatal Rats Exposed to Intrauterine Hypoxia by Inhibiting Oxidative Stress and Mitochondrial Fragmentation

Intrauterine hypoxia (IUH) is a common intrauterine dysplasia that can cause programming of the offspring cardiovascular system. In this study, we hypothesized that placental treatment with spermidine (SPD) can prevent heart injury in neonatal offspring exposed to IUH. Pregnant rats were exposed to 21% O2 or 10% O2 (hypoxia) for 7 days prior to term or were exposed to hypoxia and intraperitoneally administered SPD or SPD+difluromethylornithine (DFMO) on gestational days 15-21. Seven-day-old offspring were then sacrificed to assess several parameters. Our results demonstrated that IUH led to decreased myocardial ornithine decarboxylase (ODC) and increased spermidine/spermine N1-acetyltransferase (SSAT) expression in the offspring. IUH also resulted in decreased offspring body weight, heart weight, cardiomyocyte proliferation, and antioxidant capacity and increased cardiomyocyte apoptosis and fibrosis. Furthermore, IUH caused mitochondrial structure abnormality, dysfunction, and decreased biogenesis and led to a fission/fusion imbalance in offspring hearts. In vitro, hypoxia induced mitochondrial ROS accumulation, decreased membrane potential, and increased fragmentation. Notably, all hypoxia-induced changes analyzed in this study were prevented by SPD. Thus, in utero SPD treatment is a potential strategy for preventing IUH-induced neonatal cardiac injury.

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Fosl1 is vital to heart regeneration upon apex resection in adult Xenopus tropicalis

npj Regenerative Medicine ◽

10.1038/s41536-021-00146-y ◽

2021 ◽

Vol 6 (1) ◽

Author(s):

Hai-Yan Wu ◽

Yi-Min Zhou ◽

Zhu-Qin Liao ◽

Jia-Wen Zhong ◽

You-Bin Liu ◽

...

Keyword(s):

Early Stage ◽

Dominant Negative ◽

Luciferase Reporter ◽

Heart Regeneration ◽

Cell Cycle Regulators ◽

Cardiomyocyte Proliferation ◽

Neonatal Mice ◽

Heart Injury

AbstractCardiovascular disease is the leading cause of death in the world due to losing regenerative capacity in the adult heart. Frogs possess remarkable capacities to regenerate multiple organs, including spinal cord, tail, and limb, but the response to heart injury and the underlying molecular mechanism remains largely unclear. Here we demonstrated that cardiomyocyte proliferation greatly contributes to heart regeneration in adult X. tropicalis upon apex resection. Using RNA-seq and qPCR, we found that the expression of Fos-like antigen 1 (Fosl1) was dramatically upregulated in early stage of heart injury. To study Fosl1 function in heart regeneration, its expression was modulated in vitro and in vivo. Overexpression of X. tropicalis Fosl1 significantly promoted the proliferation of cardiomyocyte cell line H9c2. Consistently, endogenous Fosl1 knockdown suppressed the proliferation of H9c2 cells and primary cardiomyocytes isolated from neonatal mice. Taking use of a cardiomyocyte-specific dominant-negative approach, we show that blocking Fosl1 function leads to defects in cardiomyocyte proliferation during X. tropicalis heart regeneration. We further show that knockdown of Fosl1 can suppress the capacity of heart regeneration in neonatal mice, but overexpression of Fosl1 can improve the cardiac function in adult mouse upon myocardium infarction. Co-immunoprecipitation, luciferase reporter, and ChIP analysis reveal that Fosl1 interacts with JunB and promotes the expression of Cyclin-T1 (Ccnt1) during heart regeneration. In conclusion, we demonstrated that Fosl1 plays an essential role in cardiomyocyte proliferation and heart regeneration in vertebrates, at least in part, through interaction with JunB, thereby promoting expression of cell cycle regulators including Ccnt1.

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Abstract 416: B-Raf Loss Suppresses Extracellular-Regulated Kinase Activation and Cardiomyocyte Proliferation

Circulation Research ◽

10.1161/res.117.suppl_1.416 ◽

2015 ◽

Vol 117 (suppl_1) ◽

Author(s):

Natasha N Chattergoon ◽

Sara McCrohan ◽

Kent L Thornburg ◽

Philip Stork

Keyword(s):

Cell Cycle ◽

Heart Weight ◽

Fetal Life ◽

Cardiomyocyte Proliferation ◽

P53 Expression ◽

Erk Activation ◽

The Face ◽

Kinase Cascade ◽

Extracellular Regulated Kinase

Objectives: The postnatal heart does not retain the proliferative capacity it had during fetal life. Unlike large mammals, murine cardiomyocytes (CM) continue to divide into the first week of life before terminal differentiation and binucleation. We hypothesized that B-Raf regulates ERK activation in newborn CM and that loss of B-Raf suppresses cyclin levels and reduced proliferation. To test this, we determined whether loss of B-Raf disrupts the ERK (extracellular-regulated kinase) cascade and impairs CM growth. Methods: CM specific knockout (KO) of B-Raf was generated using CRE/lox (floxed B-Raf x αMHC CRE) resulting in a truncated, unstable B-Raf and a null phenotype. KO mice (α-MHC-CRE / B-Raf lox/lox ) were compared to CRE negative / B-Raf lox/lox mice (wild type; WT). Hearts from 3d and 8d old pups were harvested for molecular analysis of B-Raf signaling and cell cycle markers. Hearts from 3d old pups were harvested and CMs isolated for culture using a trypsin/DNAse digestion. The cells were treated with Isoproterenol (Iso;10uM), forskolin (20uM), and IGF-1 (1ng/ml) for 15 min to determine if the loss of B-Raf results in reduced activation of ERK. Results: Heart weight to body weight (HW/BW) ratio was less in 3d KO versus 3d WT (n=50, p<0.05). HW/BW ratio became greater in 8d KO; there was no difference in 3d and 8d HW/BW in WT animals. Baseline B-Raf and phosphorylated ERK levels were reduced in KO hearts (*p<0.05). Cell cycle inhibitors p21 and p53 were increased in 3d KO hearts with decreased levels of all cyclins (p<0.05). In 8d KO hearts, increased p21, p27, and p53 expression was accompanied with increased cyclin levels (p<0.05). In vitro ERK activation was blunted in KO CMs by forskolin and Iso compared to IGF-1. Conclusions: ERK activation was suppressed in KO hearts resulting in smaller newborn hearts but which exceeded normal HW/BW by 8d. This is may represent premature hypertrophy as the proliferative period of CM development had ended. Cell cycle analysis supports reduced CM mitotis among 8d CM. Such early disturbances in normal CM growth may increase susceptibility for reduced cardiac function in the face of increased postnatal load stress.

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Abstract 310: Role of Cardiac Fibroblast in Cardioprotective Effects of Prior Transient ACE Inhibition

Circulation Research ◽

10.1161/res.113.suppl_1.a310 ◽

2013 ◽

Vol 113 (suppl_1) ◽

Author(s):

Taben M Hale ◽

Lauren A Biwer ◽

Karen M D’Souza

Keyword(s):

Gene Expression ◽

Cellular Proliferation ◽

Cardiac Fibroblasts ◽

Cardiac Injury ◽

Ace Inhibition ◽

Cardiac Fibroblast ◽

Heart Weight ◽

Nitric Oxide Synthase Inhibitor

Prior treatment with the ACE inhibitor enalapril followed by washout protects against nitric oxide synthase inhibitor (L-NAME) induced fibrosis, cellular proliferation, and cardiac dysfunction. The present study investigated i) whether in vivo L-NAME administration induces a change in cardiac fibroblast phenotype that persists in vitro, ii) whether prior ACE inhibition protects against L-NAME induced changes in cardiac fibroblasts. SHR were divided into 3 groups: Control, L-NAME (C+L: 7d), enalapril+L-NAME (E+L: 14d enalapril + 14d washout + 7d L-NAME). MAP was measured by radiotelemetry (n=5-9), injury assessed by histology (n=6-10), and heart weight to body weight (HW/BW) was determined after 0 or 7 days of L-NAME in C+L and E+L (n=6-10). In separate rats cardiac fibroblasts were isolated after 7 days of L-NAME (C+L, E+L) or placebo (Con) and cultured to passage 1 (n=10-12). Gene expression was measured by quantitative real-time PCR. L-NAME increased MAP in C+L (22±4.1%) and E+L (21±3.6%) rats. Prior enalapril induced a persistent 13% reduction in HW/BW. L-NAME increased heart mass in E+L (7%) but not C+L; however, HW/BW remained 8% lower than C+L at sacrifice. L-NAME induced infarct in 70% of C+L and 40% of E+L hearts. Cardiac fibroblasts demonstrated a significant increase in proliferation rate in C+L, but not E+L, relative to control (C+L: 1.75-fold vs. con; E+L 1.09-fold vs. con). Fibroblasts from C+L hearts tended to have increased Collagen I and III gene expression. Despite hypertension, cardiac injury, and increased HW/BW; fibroblasts isolated from E+L proliferated at the same rate as those from control. In contrast, those isolated from C+L were hyperproliferative with a tendency toward increased capacity for collagen production. It may be that the fibroblast phenotype from E+L hearts would protect against infarct expansion and account, in part, for the previously reported cardioprotection in these rats.

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Rosuvastatin protects against coronary microembolization-induced cardiac injury via inhibiting NLRP3 inflammasome activation

Cell Death and Disease ◽

10.1038/s41419-021-03389-1 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Ao Chen ◽

Zhangwei Chen ◽

You Zhou ◽

Yuan Wu ◽

Yan Xia ◽

...

Keyword(s):

Nlrp3 Inflammasome ◽

Molecular Mechanisms ◽

Cardiac Injury ◽

Serum Lactate Dehydrogenase ◽

Inflammasome Activation ◽

H9c2 Cells ◽

Iodide Uptake ◽

Coronary Microembolization ◽

Mitochondrial Ros

AbstractCoronary microembolization (CME), a common reason for periprocedural myocardial infarction (PMI), bears very important prognostic implications. However, the molecular mechanisms related to CME remain largely elusive. Statins have been shown to prevent PMI, but the underlying mechanism has not been identified. Here, we examine whether the NLRP3 inflammasome contributes to CME-induced cardiac injury and investigate the effects of statin therapy on CME. In vivo study, mice with CME were treated with 40 mg/kg/d rosuvastatin (RVS) orally or a selective NLRP3 inflammasome inhibitor MCC950 intraperitoneally (20 mg/kg/d). Mice treated with MCC950 and RVS showed improved cardiac contractile function and morphological changes, diminished fibrosis and microinfarct size, and reduced serum lactate dehydrogenase (LDH) level. Mechanistically, RVS decreased the expression of NLRP3, caspase-1, interleukin-1β, and Gasdermin D N-terminal domains. Proteomics analysis revealed that RVS restored the energy metabolism and oxidative phosphorylation in CME. Furthermore, reduced reactive oxygen species (ROS) level and alleviated mitochondrial damage were observed in RVS-treated mice. In vitro study, RVS inhibited the activation of NLRP3 inflammasome induced by tumor necrosis factor α plus hypoxia in H9c2 cells. Meanwhile, the pyroptosis was also suppressed by RVS, indicated by the increased cell viability, decreased LDH and propidium iodide uptake in H9c2 cells. RVS also reduced the level of mitochondrial ROS generation in vitro. Our results indicate the NLRP3 inflammasome-dependent cardiac pyroptosis plays an important role in CME-induced cardiac injury and its inhibitor exerts cardioprotective effect following CME. We also uncover the anti-pyroptosis role of RVS in CME, which is associated with regulating mitochondrial ROS.

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Interleukin-35 pretreatment attenuates lipopolysaccharide-induced heart injury by inhibition of inflammation, apoptosis and fibrosis

10.1101/2020.01.23.916890 ◽

2020 ◽

Author(s):

Yang Fu ◽

Huan Hu ◽

Meng Li ◽

Huasong Xia ◽

Yue Liu ◽

...

Keyword(s):

Cardiac Injury ◽

Cardiac Inflammation ◽

Barr Virus ◽

Promising Therapeutic Target ◽

Tnf Α ◽

Immunosuppressive Cytokine ◽

Heart Injury ◽

Epstein Barr

ABSTRACTPrevious studies have demonstrated that targeting inflammation is a promising strategy for treating lipopolysaccharide (LPS)-induced sepsis and related heart injury. Interleukin-35 (IL-35), which consists of two subunits, Epstein–Barr virus-induced gene 3 (EBI3) and p35, is an immunosuppressive cytokine of the IL-12 family and exhibits strong anti-inflammatory activity. However, the role of IL-35 in LPS-induced heart injury remains obscure. In this study, we explored the role of IL-35 in heart injury induced by LPS and its potential mechanisms. Mice were treated with a plasmid encoding IL-35 (pIL-35) and then injected intraperitoneally (ip) with LPS (10 mg/kg). Cardiac function was assessed by echocardiography 12 h later. LPS apparently decreased the expression of EBI3 and p35 and caused cardiac dysfunction and pathological changes, which were significantly improved by pIL-35 pretreatment. Moreover, pIL-35 pretreatment significantly decreased the levels of cardiac proinflammatory cytokines including TNF-α, IL-6, and IL-1β, and the NLRP3 inflammasome. Furthermore, increased BCL-2 levels and decreased BAX levels inhibited apoptosis, and LPS-induced upregulation of the expression of pro-fibrotic genes (MMP2 and MMP9) was inhibited. Further investigation indicated that pIL-35 pretreatment suppressed the activation of the cardiac NF-κBp65 and TGF-β1/Smad2/3 signaling pathways in LPS-treated mice. Similar cardioprotective effects of IL-35 pretreatment were observed in mouse myocardial fibroblasts challenged with LPS in vitro. In summary, IL-35 pretreatment can attenuate cardiac inflammation, apoptosis, and fibrosis induced by LPS, implicating IL-35 as a promising therapeutic target in sepsis-related cardiac injury.

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Protective Effects of Oroxylin A against Doxorubicin-Induced Cardiotoxicity via the Activation of Sirt1 in Mice

Oxidative Medicine and Cellular Longevity ◽

10.1155/2021/6610543 ◽

2021 ◽

Vol 2021 ◽

pp. 1-11

Author(s):

Wen-Bin Zhang ◽

Yong-Fa Zheng ◽

Yao-Gui Wu

Keyword(s):

Inflammatory Diseases ◽

Cardiac Injury ◽

Life Quality ◽

Sirtuin 1 ◽

Heart Weight ◽

Protective Effects ◽

Oroxylin A ◽

Beneficial Effects

Doxorubicin- (DOX-) related cardiac injury impairs the life quality of patients with cancer. This largely limited the clinical use of DOX. It is of great significance to find a novel strategy to reduce DOX-related cardiac injury. Oroxylin A (OA) has been identified to exert beneficial effects against inflammatory diseases and cancers. Here, we investigated whether OA could attenuate DOX-induced acute cardiotoxicity in mice. A single dose of DOX was used to induce acute cardiac injury in mice. To explore the protective effects, OA was administered to mice for ten days beginning from five days before DOX injection. The data in our study indicated that OA inhibited DOX-induced heart weight loss, reduction in cardiac function, and the elevation in myocardial injury markers. DOX injection resulted in increased oxidative damage, inflammation accumulation, and myocardial apoptosis in vivo and in vitro, and these pathological alterations were alleviated by treatment of OA. OA activated the sirtuin 1 (Sirt1) signaling pathway via the cAMP/protein kinase A, and its protective effects were blocked by Sirt1 deficiency. OA treatment did not affect the tumor-killing action of DOX in tumor-bearing mice. In conclusion, OA protected against DOX-related acute cardiac injury via the regulation of Sirt1.

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Macrophages stimulate epicardial VEGFaa to trigger cardiomyocyte proliferation in larval zebrafish heart regeneration

10.1101/2021.06.15.448575 ◽

2021 ◽

Author(s):

Finnius A Bruton ◽

Aryan Kaveh ◽

Katherine Ross-Stewart ◽

Gianfranco Matrone ◽

Magdalena EM Oremek ◽

...

Keyword(s):

Cardiac Injury ◽

Cardiac Regeneration ◽

Cellular Heterogeneity ◽

Cardiomyocyte Proliferation ◽

Larval Zebrafish ◽

Laser Injury ◽

Structural Recovery ◽

Injury Model ◽

Heart Injury ◽

Zebrafish Heart

Cardiac injury induces a sustained innate immune response in both zebrafish and mammals. Macrophages, highly plastic immune cells, perform a range of both beneficial and detrimental functions during mammalian cardiac repair yet their precise roles in zebrafish cardiac regeneration are not fully understood. Here we characterise cardiac regeneration in the rapidly regenerating larval zebrafish laser injury model and use macrophage ablation and macrophage-less irf8 mutants to define the requirement of macrophages for key stages of regeneration. We found macrophages to display cellular heterogeneity and plasticity in larval heart injury as in mammals. Live heartbeat-synchronised imaging and RNAseq revealed an early proinflammatory macrophage phase which then resolves to an anti-inflammatory, profibrotic phase. Macrophages are required for cardiomyocyte proliferation but not for functional or structural recovery following injury. Macrophages are specifically recruited to the epicardial-myocardial niche, triggering the expansion of the epicardium which upregulates mitogen VEGFaa. Experimental perturbation of VEGF signalling confirmed VEGFaa to be an important inducer of cardiomyocyte proliferation revealing a previously unrecognised mechanism by which macrophages aid cardiac regeneration.

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Prrx1b restricts fibrosis and promotes Nrg1-dependent cardiomyocyte proliferation during zebrafish heart regeneration

Development ◽

10.1242/dev.198937 ◽

2021 ◽

Author(s):

Dennis E.M. de Bakker ◽

Mara Bouwman ◽

Esther Dronkers ◽

Filipa C. Simões ◽

Paul R. Riley ◽

...

Keyword(s):

Transcription Factor ◽

Cardiac Injury ◽

Lineage Tracing ◽

Heart Regeneration ◽

Cardiomyocyte Proliferation ◽

Fibrotic Scar ◽

Ligand Expression ◽

Zebrafish Heart

Fibroblasts are activated to repair the heart following injury. Fibroblast activation in the mammalian heart leads to a permanent fibrotic scar that impairs cardiac function. In other organisms, like zebrafish, cardiac injury is followed by transient fibrosis and scar-free regeneration. The mechanisms that drive scarring versus scar-free regeneration are not well understood. Here we show that the homeo-box containing transcription factor Prrx1b is required for scar-free regeneration of the zebrafish heart as the loss of Prrx1b results in excessive fibrosis and impaired cardiomyocyte proliferation. Through lineage tracing and single-cell RNA-sequencing we find that Prrx1b is activated in epicardial-derived cells (EPDCs) where it restricts TGF-β ligand expression and collagen production. Furthermore, through combined in vitro experiments in human fetal EPDCs and in vivo rescue experiments in zebrafish, we conclude that Prrx1 stimulates Nrg1 expression and promotes cardiomyocyte proliferation. Collectively, these results indicate that Prrx1 is a key transcription factor that balances fibrosis and regeneration in the injured zebrafish heart.

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Norepinephrine sensitivity of mesenteric veins in pregnant rats

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1990.259.4.r753 ◽

1990 ◽

Vol 259 (4) ◽

pp. R753-R759 ◽

Cited By ~ 3

Author(s):

M. Hohmann ◽

T. M. Keve ◽

G. Osol ◽

M. K. McLaughlin

Keyword(s):

Nerve Stimulation ◽

Sympathetic Nerves ◽

Transmural Pressure ◽

Neuronal Uptake ◽

Sprague Dawley ◽

Adrenergic Stimulation ◽

Pregnant Rats ◽

Mesenteric Veins ◽

Induced Changes

This study was designed to test the hypothesis that during the course of pregnancy there is a decrease in the venous response to adrenergic stimulation that is characterized by a decrease in venoconstriction to both exogenous norepinephrine (NE) and to transmural electric stimulation of endogenous sympathetic nerves. Capacitance-size mesenteric veins were removed from nonpregnant and early- (7/8 day) and late-pregnant (18/19 and 20/21 day) Sprague-Dawley rats and studied in vitro under pressurized conditions. Lumen diameter was measured continuously by a video-electronic method. There was a marked increase in the sensitivity of the veins to exogenous NE stimulation at the end of pregnancy, which was most dramatic at a transmural pressure of 6 compared with 2 mmHg. The increase in exogenous NE sensitivity was associated with a progressive decline in the response to transmural nerve stimulation during pregnancy. Cocaine, an inhibitor of neuronal uptake, resulted in a greater potentiation of the response to transmural nerve stimulation in the pregnant rats compared to controls, suggesting an increased reuptake mechanism during pregnancy. Additional studies in the pseudopregnant, lactating, and nonlactating rat suggested that the conceptus was necessary for the alterations in neural response but not for the increase in exogenous NE sensitivity. In conclusion there is a dramatic change in venous function during pregnancy in the rat that is characterized by a difference between endogenous and exogenous NE sensitivity. The fact that transmural pressure can profoundly affect exogenous NE sensitivity suggests that pregnancy-induced changes in venous volume could contribute to changes in venous reactivity.

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P53 Contributes to Cisplatin Induced Renal Oxidative Damage via Regulating P66shc and MnSOD

Cellular Physiology and Biochemistry ◽

10.1159/000430247 ◽

2015 ◽

Vol 37 (4) ◽

pp. 1240-1256 ◽

Cited By ~ 25

Author(s):

Yanggang Yuan ◽

Hui Wang ◽

Yingyi Wu ◽

Bo Zhang ◽

Ningning Wang ◽

...

Keyword(s):

Cell Injury ◽

Kidney Injury ◽

Therapeutic Window ◽

Protein Levels ◽

Cisplatin Nephrotoxicity ◽

Mitochondrial Ros ◽

Potential Strategy ◽

Hk2 Cells

Background/Aims: Cisplatin is widely used to treat malignancies. However, its major limitation is the development of dose-dependent nephrotoxicity. The precise mechanisms of cisplatin-induced kidney damage remain unclear. Previous study demonstrated the central role of mitochondrial ROS (mtROS) in the pathogenesis of cisplatin nephrotoxicity. The purpose of this study was to explore the mechanism of mtROS regulation in cisplatin nephrotoxicity. Methods: p53, MnSOD and p66shc were detected at mRNA and protein levels by qPCR and western blot in HK2 cells. mtROS levels were determined by DCFDA and MitoSOX staining. Cell viability and cell apoptosis were accessed by CCK-8 assay, TUNEL assay and flow cytometry, respectivesly. siRNAs were used to knock down p53 and p66shc expression and subsequent changes were observed. In vivo assays using a mouse model of cisplatin-induced acute kidney injury were used to validate the in vitro results. Results: In HK2 cells, cisplatin exposure decreased the MnSOD and increased the expression of p53 and p66shc. MnTBAP, a MnSOD mimic, blocked cisplatin-induced the generation of mtROS and cell injury. P66shc and p53 siRNAs rendered renal cells resistant to cisplatin-induced mtROS production and cell death. Furthermore, knockdown of p53 restored MnSOD and inhibiting p66shc. Consistent with these results, we revealed that p53 inhibitor reduced cisplatin-induced oxidative stress and apoptosis by regulating MnSOD and p66shc in the kidney of cisplatin-treated mice. Conclusion: Our study identifies activation of p53 signalling as a potential strategy for reducing the nephrotoxicity associated with cisplatin treatments and, as a result, broadens the therapeutic window of this chemotherapeutic agent.

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