Anna Folino

Apelin-13 limits infarct size and improves cardiac postischemic mechanical recovery only if given after ischemia

We studied whether apelin-13 is cardioprotective against ischemia/reperfusion injury if given as either a pre- or postconditioning mimetic and whether the improved postischemic mechanical recovery induced by apelin-13 depends only on the reduced infarct size or also on a recovery of function of the viable myocardium. We also studied whether nitric oxide (NO) is involved in apelin-induced protection and whether the reported ischemia-induced overexpression of the apelin receptor (APJ) plays a role in cardioprotection. Langendorff-perfused rat hearts underwent 30 min of global ischemia and 120 min of reperfusion. Left ventricular pressure was recorded. Infarct size and lactate dehydrogenase release were determined to evaluate the severity of myocardial injury. Apelin-13 was infused at 0.5 μM concentration for 20 min either before ischemia or in early reperfusion, without and with NO synthase inhibition by N(G)-nitro-l-arginine (l-NNA). In additional experiments, before ischemia also 1 μM apelin-13 was tested. APJ protein level was measured before and after ischemia. Whereas before ischemia apelin-13 (0.5 and 1.0 μM) was ineffective, after ischemia it reduced infarct size from 54 ± 2% to 26 ± 4% of risk area (P < 0.001) and limited the postischemic myocardial contracture (P < 0.001). l-NNA alone increased postischemic myocardial contracture. This increase was attenuated by apelin-13, which, however, was unable to reduce infarct size. Ischemia increased APJ protein level after 15-min perfusion, i.e., after most of reperfusion injury has occurred. Apelin-13 protects the heart only if given after ischemia. In this protection NO plays an important role. Apelin-13 efficiency as postconditioning mimetic cannot be explained by the increased APJ level.

Identification of IL-17F/frequent exacerbator endotype in asthma

Background Severe asthma might be associated with overexpression of Th17 cytokines, which induce neutrophil recruitment via neutrophil‐mobilizing cytokines in airways. Objective To study IL‐17–related cytokines in nasal/bronchial biopsies from controls and mild asthmatics (MAs) to severe asthmatics (SAs) in relation to exacerbation rate. Methods Inflammatory cells and IL‐17A+, IL‐17F+, IL‐21+, IL‐22+, and IL‐23+ cells were examined by immunohistochemistry in cryostat sections of bronchial/nasal biopsies obtained from 33 SAs (21 frequent exacerbators [FEs]), 31 MAs (3 FEs), and 14 controls. IL‐17F protein was also measured by ELISA in bronchial/nasal lysates and by immunohistochemistry in bronchial tissue obtained from subjects who died because of fatal asthma. Immunofluorescence/confocal microscopy was used for IL‐17F colocalization. Results Higher number (P < .05) of neutrophils, IL‐17A+, IL‐17F+, and IL‐21+ cells in bronchial biopsies and higher numbers (P < .01) of IL‐17F+ and IL‐21+ cells in nasal biopsies were observed in SAs compared with MAs. Bronchial IL‐17F+ cells correlated with bronchial neutrophils (r = 0.54), exacerbation rate (r = 0.41), and FEV1 (r = −0.46). Nasal IL‐17F+ cells correlated with bronchial IL‐17F (r = 0.35), exacerbation rate (r = 0.47), and FEV1 (r = −0.61). FEs showed increased number of bronchial neutrophils/eosinophils/CD4+/CD8+ cells and bronchial/nasal IL‐17F+ cells. Receiver operating characteristic curve analysis evidenced predictive cutoff values of bronchial neutrophils and nasal/bronchial IL‐17F for discriminating between asthmatics and controls, between MAs and SAs and between FEs and non‐FEs. IL‐17F protein increased in bronchial/nasal lysates of SAs and FEs and in bronchial tissue of fatal asthma. IL‐17F colocalized in CD4+/CD8+ cells. Conclusions IL‐17–related cytokines expression was amplified in bronchial/nasal mucosa of neutrophilic asthma prone to exacerbation, suggesting a pathogenic role of IL‐17F in FEs. Graphical abstract Figure. No Caption available.

Effect of apelin-apelin receptor system in postischaemic myocardial protection: a pharmacological postconditioning tool?

In the heart, a great part of ischaemia and reperfusion injuries occurs mainly during the first minutes of reperfusion. The opening of the mitochondrial permeability transition pores is the end point of the cascade to myocardial damage. Also, oxidative stress contributes to cell death. Postconditioning is a protective maneuver that can be selectively timed at the beginning of reperfusion. It is hypothesized that it acts via the reperfusion injury salvage kinase pathway, which includes nitric oxide-dependent and nitric oxide-independent cascades. Apelin is an endogenous peptide that can protect the heart from reperfusion injury if given at the beginning of reperfusion but not before ischaemia. It is hypothesized that it may trigger the reperfusion injury salvage kinase pathway via a specific apelin receptor. Apelin can also limit the oxidative stress by the activation of superoxide dismutase. Apelin and apelin receptor expression increase early after ischaemia and at the beginning of an ischaemic heart failure. These observations suggest that the endogenous release of the peptide can limit the severity of an infarction and ameliorate myocardial contractility compromised by the appearance of the failure. Due to its protective activities, apelin could be a therapeutic tool if administered with the same catheter used for angioplasty or after the maneuvers aimed at bypassing a coronary occlusion.

Omega 3 has a beneficial effect on ischemia/reperfusion injury, but cannot reverse the effect of stressful forced exercise

BACKGROUND AND AIM The beneficial effects of exercise in reducing the incidence of cardiovascular diseases are well known. Several studies have demonstrated that forced exercise (FE) could activate a stress response similar to a restrain stress. Previous studies suggest that heart protection to ischemic events would be improved by an omega 3 free fatty acid (omega3-FFA)-enriched diet. Here, we investigate the impact of stressful FE and an omega 3-FFA-enriched diet on cardiac tolerance to ischemic events over one month. METHODS AND RESULTS Twenty-four Wistar rats were randomly assigned to one of the following protocols: 1) Sedentary (SED) animals who were regularly fed; 2) sedentary animals who were given 1ml/day of fish oil for one month; 3) FE+omega3-FFA rats who were given 1ml/day of fish oil and forced to run on a motorized wheel for 30min every day, both for one month; and 4) FE animals were forced to exercise as group 3 and fed with a regular diet. At the end of the treatments an isolated heart preparation was performed. After a 30min global ischemic event and 2h reperfusion, hearts of sedentary-omega3 animals recovered about 37% of left ventricular developed pressure, whereas FE, omega3+FE and CTRL-SED animals recovered only about 15%, 5% and 8% respectively. Similarly, heart infarct size was significantly lower in sedentary-omega3 animals compared to animals in the three other groups. CONCLUSIONS Results indicate that one month of treatment with an omega3-FFA-enriched diet improves cardioprotection upon ischemic events, whereas FE leads to a reduced heart tolerance to ischemic events, which cannot be reversed by an omega3-FFA diet.

The effect of bioartificial constructs that mimic myocardial structure and biomechanical properties on stem cell commitment towards cardiac lineage

Despite the enormous progress in the treatment of coronary artery diseases, they remain the most common cause of heart failure in the Western countries. New translational therapeutic approaches explore cardiomyogenic differentiation of various types of stem cells in combination with tissue-engineered scaffolds. In this study we fabricated PHBHV/gelatin constructs mimicking myocardial structural properties. Chemical structure and molecular interaction between material components induced specific properties to the substrate in terms of hydrophilicity degree, porosity and mechanical characteristics. Viability and proliferation assays demonstrated that these constructs allow adhesion and growth of mesenchymal stem cells (MSCs) and cardiac resident non myocytic cells (NMCs). Immunofluorescence analysis demonstrated that stem cells cultured on these constructs adopt a distribution mimicking the three-dimensional cell alignment of myocardium. qPCR and immunofluorescence analyses showed the ability of this construct to direct initial MSC and NMC lineage specification towards cardiomyogenesis: both MSCs and NMCs showed the expression of the cardiac transcription factor GATA-4, fundamental for early cardiac commitment. Moreover NMCs also acquired the expression of the cardiac transcription factors Nkx2.5 and TBX5 and produced sarcomeric proteins. This work may represent a new approach to induce both resident and non-resident stem cells to cardiac commitment in a 3-D structure, without using additional stimuli.

Balance of nitric oxide and reactive oxygen species in myocardial reperfusion injury and protection.

Abstract: Depending on their concentrations, both nitric oxide (NO) and reactive oxygen species (ROS) take part either in myocardial ischemia reperfusion injury or in protection by ischemic and pharmacological preconditioning (Ipre) and postconditioning (Ipost). At the beginning of reperfusion, a transient release of NO is promptly scavenged by ROS to form the highly toxic peroxynitrite, which is responsible for a further increase of ROS through endothelial nitric oxide synthase uncoupling. The protective role of NO has suggested the use of NO donors to mimic Ipre and Ipost. However, NO donors have not always given the expected protection, possibly because they are responsible for the production of different amounts of ROS that depend on the amount of released NO. This review is focused on the role of the balance of NO and ROS in myocardial injury and its prevention by Ipre and Ipost and after the use of NO donors given with or without antioxidant compounds to mimic Ipre and Ipost.

Injured cardiomyocytes promote dental pulp mesenchymal stem cell homing

BACKGROUND The heart is unable to regenerate its tissues after severe injuries. Stem cell therapy appears to be one of the most promising approaches, though preclinical results are hitherto contradictory and clinical trials scanty and/or limited to phase-I. The limited knowledge about stem cell early homing in infarcted cardiac tissues can concur to this scenario. METHODS The stem cell migration was assessed in in-vitro and ex-vivo models of heart ischemia, employing a rat dental pulp stem cell line (MUR-1) that shares the same ontogenic progenitors with portions of the heart, expresses markers typical of cardiac/vascular-like progenitors and is able to differentiate into cardiomyocytes in-vitro. RESULTS Here, we demonstrated that the MUR-1 can reach the injured cells/tissue and make contacts with the damaged cardiomyocytes, likely through Connexin 43, N-cadherin and von Willebrand Factor mediated cell-cell interactions, both in in-vitro and ex-vivo models. Furthermore, we found that SDF-1, FGF-2 and HGF, but not VEGF are involved as chemotactic factors in MUR-1 migration, notifying a similarity with neural crest cell behavior during the organogenesis of both the splanchnocranium and the heart. CONCLUSIONS Herein we found a similarity between what happens during the heart organogenesis and the early migration and homing of MUR-1 cells in ischemic models. GENERAL SIGNIFICANCE The comprehension of molecular aspects underlying the early phases of stem cell migration and interaction with damaged organ contributes to the future achievement of the coveted stem cell-mediated organ regeneration and function preservation in-vivo.

Self-Renewal and Multipotency Coexist in a Long-Term Cultured Adult Rat Dental Pulp Stem Cell Line: An Exception to the Rule?

The stemness state is characterized by self-renewal and differentiation properties. However, stem cells are not able to preserve these characteristics in long-term culture because of the intrinsic fragility of their phenotype easily undergoing senescence or neoplastic transformation. Furthermore, although isolated from the same original tissue using similar protocols, adult stem cells can display dissimilar phenotypes and important cell clone/species contamination. Finally, the lack of a clear standardization contributes to complicate the comprehension about the stemness condition. In this context, cell lines displaying a particularly stable phenotype must be identified to define one or multiple benchmarks against which other stem cell lines could be reliably assessed. The present paper demonstrates that it is possible to isolate from the rat dental pulp a stem cell line (MUR-1) that does not display neoplastic transformation in long-term culture. MUR-1 cells stably express a broad range of stemness markers and are able to differentiate into adipogenic, osteogenic, chondrogenic, neurogenic, and cardiomyogenic lineages independently of the culture passages. Moreover, serial in vitro passages have not changed their immunophenotype, proliferation capacity, or differentiation potential. The uniqueness of these characteristics candidates MUR-1 as a model to reliably improve the understanding of the mechanisms governing the stem cell fate in the same as well as in other stem cell populations.

Apelin-induced cardioprotection against ischaemia/reperfusion injury: roles of epidermal growth factor and Src

Apelin, the ligand of the G‐protein‐coupled receptor (GPCR) APJ, exerts a post‐conditioning‐like protection against ischaemia/reperfusion injury through activation of PI3K‐Akt‐NO signalling. The pathway connecting APJ to PI3K is still unknown. As other GPCR ligands act through transactivation of epidermal growth factor receptor (EGFR) via a matrix metalloproteinase (MMP) or Src kinase, we investigated whether EGFR transactivation is involved in the following three features of apelin‐induced cardioprotection: limitation of infarct size, suppression of contracture and improvement of post‐ischaemic contractile recovery.

Bradykinin in asthma: modulation of airway inflammation and remodelling

Abstract Bradykinin, a pro‐inflammatory molecule, and its related peptides have been studied for their effects on acute reactions in upper and lower airways, where they can be synthesised and metabolized after exposure to different stimuli including allergens and viral infection. Bradykinin B1 and B2 receptors are constitutively expressed in the airways on several residential and/or immune cells. Their expression can also be induced by inflammatory mediators, usually associated with eosinophil and neutrophil recruitment, such as IL‐4, IL‐13, TNF‐&agr;, IL‐6 and IL‐8, via intracellular MAPK and NF‐&kgr;B signalling. In turn, the latters up‐regulate both bradykinin receptors. Bradykinin activates epithelial/endothelial and immune cells, neurons and mesenchymal cells (such as fibroblasts, myofibroblasts and smooth muscle cells), which are implicated in the development of airway chronic inflammation, responsiveness and remodelling (a major feature of severe asthma). This review highlights the role of bradykinin and its receptors in respect to chronic inflammatory response involving eosinophils/neutrophils and to vascular/matrix‐related airway remodelling in asthmatic airways. This scenario is especially important for understanding the mechanisms involved in the pathogenesis of eosinophilic and/or neutrophilic asthma and hence their therapeutic approach.