the-left-pneumonectomy-combined-with-monocrotaline-or-sugen-as-model

The Left Pneumonectomy Combined with Monocrotaline or Sugen as a Model of Pulmonary Hypertension in Rats

In this protocol, we detail the correct procedural steps and necessary precautions to successfully perform a left pneumonectomy and induce PAH in rats ...

induction-characterization-pulmonary-hypertension-mice-using

Induction and Characterization of Pulmonary Hypertension in Mice using the Hypoxia/SU5416 Model

Pulmonary Hypertension (PH) is a pathophysiological condition, defined by a mean pulmonary arterial pressure exceeding 25 mm Hg at rest, as assessed by ...

Icahn School of Medicine at Mount Sinai

This review focuses on multidrug resistance protein 4 (MRP4 or ABCC4) that has recently been shown to play a role in cAMP homeostasis, a key-pathway in vascular biology and in platelet functions. In vascular system, recent data provide evidence that inhibition of MRP4 prevents human coronary artery smooth muscle cell proliferation in vitro and in vivo, as well as human pulmonary artery smooth muscle cell proliferation in vitro and pulmonary hypertension in mice in vivo. In the heart, MRP4 silencing in adult rat ventricular myocytes results in an increase in intracellular cAMP levels leading to enhanced cardiomyocyte contractility. However, a prolonged inhibition of MRP4 can promote cardiac hypertrophy. In addition, secreted cAMP, through its metabolite adenosine, prevents adrenergically induced cardiac hypertrophy and fibrosis. Finally, MRP4 inhibition in platelets induces a moderate thrombopathy. The localization of MRP4 underlines the emerging concept of cAMP compartmentalization in platelets, which is a major regulatory mechanism in other cells. cAMP storage in platelet dense granules might limit the cAMP cytosolic concentration upon adenylate cyclase activation, a necessary step to induce platelet activation. In this review, we discuss the therapeutic potential of direct pharmacological inhibition of MRP4 in atherothrombotic disease, via its vasodilating and antiplatelet effects.

MRP4 (ABCC4) as a potential pharmacologic target for cardiovascular disease.

Chronic cardiac stress induces pathologic hypertrophy and fibrosis of the myocardium. The microRNA-29 (miR-29) family has been found to prevent excess collagen expression in various organs, particularly through its function in fibroblasts. Here, we show that miR-29 promotes pathologic hypertrophy of cardiac myocytes and overall cardiac dysfunction. In a mouse model of cardiac pressure overload, global genetic deletion of miR-29 or antimiR-29 infusion prevents cardiac hypertrophy and fibrosis and improves cardiac function. Targeted deletion of miR-29 in cardiac myocytes in vivo also prevents cardiac hypertrophy and fibrosis, indicating that the function of miR-29 in cardiac myocytes dominates over that in non-myocyte cell types. Mechanistically, we found cardiac myocyte miR-29 to de-repress Wnt signaling by directly targeting four pathway factors. Our data suggests that, cell- or tissue-specific antimiR-29 delivery may have therapeutic value for pathological cardiac remodeling and fibrosis.

Cardiac myocyte miR-29 promotes pathological remodeling of the heart by activating Wnt signaling.

Myocardial fibrosis is associated with profound changes in ventricular architecture and geometry, resulting in diminished cardiac function. There is currently no information on the role of the delta-like homologue 1 (Dlk1) in the regulation of the fibrotic response. Here, we investigated whether Dlk1 is involved in cardiac fibroblast-to-myofibroblast differentiation and regulates myocardial fibrosis and explored the molecular mechanism underpinning its effects in this process.

Deletion of delta-like 1 homologue accelerates fibroblast-myofibroblast differentiation and induces myocardial fibrosis.

Although the initial reparative fibrosis after myocardial infarction (MI) is crucial for preventing rupture of the ventricular wall, an exaggerated fibrotic response and reactive fibrosis outside the injured area are detrimental. Although metformin prevents adverse cardiac remodeling, as well as provides glycemic control, the underlying mechanisms remain poorly documented. This study describes the effect of mitochondrial NADPH oxidase 4 (mitoNox) and protein kinase C-alpha (PKCα) on the cardiac fibrosis and galectin 3 (Gal-3) expression. Randomly rats underwent MI, received metformin or saline solution. A model of biomechanical strain and co-culturewas used to enable cross talk between cardiomyocytes and fibroblasts. Long-term metformin treatment after MIwas associated with (1) a reduction in myocardial fibrosis and Gal-3 levels; (2) an increase in adenosine monophosphate-activated protein kinase (AMPK) α1/α2 levels; and (3) an inhibition of both mRNA expression and enzymatic activities of mitoNox and PKCα. These findings were replicated in the cellular model, where the silencing of AMPK expression blocked the ability of metformin to protect cardiomyocytes from strain. The use of specific inhibitors or small interference RNA provided evidence that PKCα is downstream of mitoNox, and that the activation of this pathway results in Gal-3 upregulation.The Gal-3 secreted by cardiomyocytes has a paracrine effect on cardiac fibroblasts, inducing their activation. In conclusion, a metformin-induced increase in AMPK improves myocardial remodeling post-MI, which is related to the inhibition of the mitoNox/PKCα/Gal-3 pathway. Manipulation of this pathway might offer new therapeutic options against adverse cardiac remodeling, in terms of preventing the activation of the present fibroblast population.

Pharmacological inhibition of the mitochondrial NADPH oxidase 4/PKCα/Gal-3 pathway reduces left ventricular fibrosis following myocardial infarction.

Pulmonary arterial hypertension (PAH) is a syndrome characterized by pulmonary vascular remodeling and vasoconstriction, leading to increased pulmonary vascular resistance, right ventricular pressure overload and, eventually, to right ventricular failure and premature death. Animal models have been an essential tool for understanding pulmonary hypertension pathophysiology and for the discovery and development of novel therapies.MCT-induced PAH in rats leads to a significant increase in RV pressure and pulmonary vascular remodeling, as well as greater RV hypertrophy. In this chapter, we describe protocols for inducing and assessing the monocrotaline (MCT) rat model, the most classical and widely used in vivo model of PAH. Using this protocol, rats reproducibly develop pulmonary hypertension with a mean pulmonary pressure of ~40 mmHg approximately 4 weeks after single MCT administration.

Pulmonary Artery Hypertension Model in Rats by Monocrotaline Administration.

Pulmonary hypertension is a rapidly progressive, life-threatening, and often fatal disease. Despite many new developments in pulmonary arterial hypertension (PAH) therapy, there is currently no cure for PAH, and new therapies are desperately needed. PAH pathobiology involves a remodeling process in pulmonary arteries that plays a critical role in elevating pulmonary arterial and right ventricle pressures. The discovery and development of new therapies requires animal models of PAH that mimic the human disease, including vascular remodeling.Here we review and describe a detailed protocol for creating an in vivo model of Sugen/Hypoxia-induced PAH in mice that is commonly used to assess the efficiency of new therapies in PAH. Severe pulmonary hypertension can be established in 1 month using this protocol. Additional protocols to evaluate the model by invasive pressure measurements and histology are provided.

The Sugen 5416/Hypoxia Mouse Model of Pulmonary Arterial Hypertension.

Despite its functional importance in various fundamental bioprocesses, studies of N-methyladenosine (m6A) in the heart are lacking. Here, we show that the FTO (fat mass and obesity-associated protein), an m6A demethylase, plays a critical role in cardiac contractile function during homeostasis, remodeling, and regeneration.

FTO-Dependent N-Methyladenosine Regulates Cardiac Function During Remodeling and Repair.

Direct measurement of left atrial and pulmonary artery pressure in rats with pulmonary hypertension.

Pulmonary arterial hypertension (PAH) results in right ventricular (RV) failure, electro-mechanical dysfunction and heightened risk of sudden cardiac death (SCD), although exact mechanisms and predisposing factors remain unclear. Because impaired chronotropic response to exercise is a strong predictor of early mortality in patients with PAH, we hypothesized that progressive elevation in heart rate can unmask ventricular tachyarrhythmias (VT) in a rodent model of monocrotaline (MCT)-induced PAH. We further hypothesized that intra-tracheal gene delivery of aerosolized AAV1.SERCA2a (AAV1.S2a), an approach which improves pulmonary vascular remodeling in PAH, can suppress VT in this model.

Intra-tracheal gene delivery of aerosolized SERCA2a to the lung suppresses ventricular arrhythmias in a model of pulmonary arterial hypertension.

Cyclic adenosine monophosphate (cAMP) is the predominant intracellular second messenger that transduces signals from Gs-coupled receptors. Intriguingly, there is evidence from various cell types that an extracellular cAMP pathway is active in the extracellular space. Herein, we investigated the role of extracellular cAMP in the lung and examined whether it may act on pulmonary vascular cell proliferation and pulmonary vasculature remodeling in the pathogenesis of pulmonary hypertension (PH).

A novel secreted-cAMP pathway inhibits pulmonary hypertension via a feed-forward mechanism.

Coronary microembolization is one of the main causes of the "no-reflow" phenomenon, which commonly occurs after reperfusion of an occluded coronary artery. Given its high incidence and the fact that it has been proven to be an independent predictor of cardiac morbidity and mortality, there is an imperative need to study its underlying mechanisms and pathophysiology. Large animal models are essential to perform translational studies. Currently there is no animal model that recapitulates a clinical scenario of thrombogenic microembolism with preceding myocardial ischemia. Therefore, the goal of this study was to develop and characterize a novel pig model of coronary microembolization using autologous thrombus injection (CMET). Twenty-three pigs underwent myocardial infarction through percutaneous balloon occlusion of the left anterior descending artery (LAD). Each animal was enrolled in one of two groups: (1) the CMET group, in which the LAD occlusion was followed by delivery of autologous clotted blood in the LAD (distal to the balloon occlusion) and reperfusion; (2) the ischemic reperfusion (I/R) group, in which the LAD ischemia was followed by reperfusion. Surviving animals underwent functional and morphological characterization at 1-week post-procedure. Three sham operated animals were used as a control. CMET resulted in impaired left ventricular function compared to I/R pigs at 1 week. Three-dimensional echocardiography demonstrated reduced ejection fraction in the CMET group (CMET vs. I/R: 35.6 ± 4.2% vs. 47.6 ± 2.4%,  = 0.028). Invasive hemodynamic measurements by Swan-Ganz and left ventricular pressure-volume catheters revealed that CMET impaired left ventricular contractility and diastolic function. This was confirmed by both load-dependent indices including cardiac output (CMET vs. I/R: 2.7 ± 0.2 l/min, vs. 4.0 ± 0.1 l/min,  = 0.002) and load independent indices including preload-recruitable stroke work (CMET vs. I/R: 25.8 ± 4.0 vs. 47.5 ± 6.5 mmHg,  = 0.05) and end-diastolic pressure-volume relationship (slope, 0.68 ± 0.07 vs. 0.40 ± 0.11 mmHg/ml,  = 0.01). Our unique closed-chest model of coronary microembolization using autologous thrombus injection resembles the clinical condition of thrombogenic coronary microembolization in I/R injury. This model offers opportunities to conduct translational studies for understanding and treating coronary microembolization in myocardial infarction.

A Novel Large Animal Model of Thrombogenic Coronary Microembolization.

Inhibition of pulmonary fibrosis (PF) by restoring sarco/endoplasmic reticulum calcium ATPase 2a isoform (SERCA2a) expression using targeted gene therapy may be a potentially powerful new treatment approach for PF. Here, we found that SERCA2a expression was significantly decreased in lung samples from patients with PF and in the bleomycin (BLM) mouse model of PF. In the BLM-induced PF model, intratracheal aerosolized adeno-associated virus serotype 1 (AAV1) encoding for human SERCA2a (AAV1.hSERCA2a) reduces lung fibrosis and associated vascular remodeling. SERCA2a gene therapy also decreases right ventricular pressure and hypertrophy in both prevention and curative protocols. In vitro, we observed that SERCA2a overexpression inhibits fibroblast proliferation, migration, and fibroblast-to-myofibroblast transition induced by transforming growth factor β (TGF-β1). Thus, pro-fibrotic gene expression is prevented by blocking nuclear factor κB (NF-κB)/interleukin-6 (IL-6)-induced signal transducer and activator of transcription 3 (STAT3) activation. This effect is signaled toward an inhibitory mechanism of small mother against decapentaplegic (SMAD)/TGF-β signaling through the repression of OTU deubiquitinase, ubiquitin aldehyde binding 1 (OTUB1) and Forkhead box M1 (FOXM1). Interestingly, this cross-inhibition leads to an increase of SKI and SnoN expression, an auto-inhibitory feedback loop of TGF-β signaling. Collectively, our results demonstrate that SERCA2a gene transfer attenuates bleomycin (BLM)-induced PF by blocking the STAT3/FOXM1 pathway and promoting the SNON/SKI Axis. Thus, SERCA2a gene therapy may be a potential therapeutic target for PF.

AAV1.SERCA2a Gene Therapy Reverses Pulmonary Fibrosis by Blocking the STAT3/FOXM1 Pathway and Promoting the SNON/SKI Axis.

The adult mammalian heart has limited regenerative capacity, mostly attributable to postnatal cardiomyocyte cell cycle arrest. In the last 2 decades, numerous studies have explored cardiomyocyte cell cycle regulatory mechanisms to enhance myocardial regeneration after myocardial infarction. Pkm2 (Pyruvate kinase muscle isoenzyme 2) is an isoenzyme of the glycolytic enzyme pyruvate kinase. The role of Pkm2 in cardiomyocyte proliferation, heart development, and cardiac regeneration is unknown.

ArticleTotal : 2	Year
Publication title Cited by 27	2019
Publication title Cited by 7	2020

Article	Year
MRP4 (ABCC4) as a potential pharmacologic target for cardiovascular disease. Pharmacological research\| PubMed ID: 27063943	2016
Cardiac myocyte miR-29 promotes pathological remodeling of the heart by activating Wnt signaling. Nature communications\| PubMed ID: 29158499	2017
Deletion of delta-like 1 homologue accelerates fibroblast-myofibroblast differentiation and induces myocardial fibrosis. European heart journal\| PubMed ID: 29668883	2019
Pharmacological inhibition of the mitochondrial NADPH oxidase 4/PKCα/Gal-3 pathway reduces left ventricular fibrosis following myocardial infarction. Translational research : the journal of laboratory and clinical medicine\| PubMed ID: 29753686	2018
Pulmonary Artery Hypertension Model in Rats by Monocrotaline Administration. Methods in molecular biology (Clifton, N.J.)\| PubMed ID: 29987824	2018
The Sugen 5416/Hypoxia Mouse Model of Pulmonary Arterial Hypertension. Methods in molecular biology (Clifton, N.J.)\| PubMed ID: 29987825	2018
FTO-Dependent N-Methyladenosine Regulates Cardiac Function During Remodeling and Repair. Circulation\| PubMed ID: 29997116	2019
Direct measurement of left atrial and pulmonary artery pressure in rats with pulmonary hypertension. The Journal of thoracic and cardiovascular surgery\| PubMed ID: 30119283	2018
Intra-tracheal gene delivery of aerosolized SERCA2a to the lung suppresses ventricular arrhythmias in a model of pulmonary arterial hypertension. Journal of molecular and cellular cardiology\| PubMed ID: 30502350	2019
A novel secreted-cAMP pathway inhibits pulmonary hypertension via a feed-forward mechanism. Cardiovascular research\| PubMed ID: 31529026	2019
A Novel Large Animal Model of Thrombogenic Coronary Microembolization. Frontiers in cardiovascular medicine\| PubMed ID: 31750316	2019
AAV1.SERCA2a Gene Therapy Reverses Pulmonary Fibrosis by Blocking the STAT3/FOXM1 Pathway and Promoting the SNON/SKI Axis. Molecular therapy : the journal of the American Society of Gene Therapy\| PubMed ID: 31879190	2020
Pkm2 Regulates Cardiomyocyte Cell Cycle and Promotes Cardiac Regeneration. Circulation\| PubMed ID: 32078387	2020

Yassine Sassi

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