Role of fms-like tyrosine kinase 3 in cardiac health and disease

The fms-like tyrosine kinase 3 receptor (flt3) and ligand (flt3L) are important regulators of early haematopoiesis (1). Uncontrolled activity of flt3 is associated with acute myeloid leukaemia, making flt3 a major target of receptor tyrosine kinase inhibitors (TKIs) (2). TKIs are currently used in a variety of malignancies and patients have experienced cardiotoxicity under some of them. We recently demonstrated that exogenous activation of flt3 through intramyocardial injection of flt3L is cardioprotective and improves post-myocardial infarction remodeling and function in mice (3). Based on this background, we wanted to assess how endogenous disruption of flt3-signaling affects the healthy and the injured heart. We also hypothesized that lack of flt3L has an impact on the adult cardiac progenitor cell pool, and, hence, on their contribution to cardiac homeostasis. We isolated cardiac progenitor cells (CPCs) according to the side population (SP) phenotype (SP-CPCs) from 12 week-old wt and flt3L-/- mice (4) and we further expanded the SP-CPC fraction positive for Sca1 and negative for CD31 in culture (5). Our in vivo findings showed a reduced number of SP-CPCs in flt3L-/- hearts, suggesting an untimely activation of a proportion of this physiologically quiescent population. In line with this finding, transcriptional comparison revealed a reduced CD31- progenitor cell signature in flt3L-/- SP-CPCs, with downregulation of transcripts involved in cell cycle arrest and stemness. Expanded SP-CPCs do express functional flt3 receptor and secrete flt3L allowing for an autocrine and/or paracrine activation of the receptor. SP-CPCs maintained the capacity to differentiate into different cardiac lineages when expanded under progenitor potential-preserving culturing conditions. However, in this context, flt3L-/- SP-CPCs showed enhanced proliferation capacity associated with reduced differentiation potential towards all three major cardiac lineages, i.e., cardiomyocytes, smooth muscle and endothelial cells. Echocardiography showed that disruption of flt3-signaling leads to significant systolic dysfunction and subtle structural alterations of the heart. Specifically, flt3L-/- hearts are smaller and exhibit reduced microvascularization within the ventricular wall. Together, these findings support that flt3-signaling is important for the maintenance of a functional CPC pool and cardiac homeostasis of the healthy heart. We further investigated the implications of the lack of a functional flt3 system in a model of myocardial infarction. Given the importance of flt3 in cardiomyocyte survival (3) and myocardial microvascularization, we hypothesized that flt3-disruption may aggravate post-infarct myocardial damage and dysfunction. Contrary to this hypothesis, however, flt3L ablation mitigated the functional decline during the acute and post-acute phase after myocardial infarction. This protective effect may involve potential immunomodulatory properties related to flt3-signaling as supported by our most recent RNA sequencing data. In summary, this thesis work shows that in a context of a potential prospective use of CPCs for cardiac cell therapy, careful adjustments of culturing conditions are pivotal to preserve the CPC potential, and thus the capability for multi-lineage commitment and differentiation, during ex vivo expansion. Furthermore, this work uncovers a so-far unrecognized role of flt3 signaling in the regulation of adult CPC homeostasis and function. In particular we showed that flt3 contributes to the maintenance of a pool of quiescent, functional cardiac progenitor cells in vivo and their balanced proliferation and differentiation in vitro. Finally, our most recent data suggest that the systemic inhibition of flt3 may be protective during the early phase post-myocardial infarction. In view of the increasing use of TK targeting cancer therapies, improved understanding of the roles of such TK in cardiac health and disease is important in order to anticipate potential cardiac side effects. This work uncovers so far unknown functions of flt3 signaling in the healthy and injured heart that raise both awareness of potential cardiac toxicity, but also of a possible therapeutic effect of the systemic inhibition of flt3 in the post-acute phase after myocardial infarction

Regenerative therapy for cardiovascular disease

Recent insights into myocardial biology uncovered a hereto unknown regenerative capacity of the adult heart. The discovery of dividing cardiomyocytes and the identification and characterization of cardiac stem and progenitor cells with myogenic and angiogenic potential have generated new hopes that cardiac regeneration and repair might become a therapeutic option. During the past decade, multiple candidate cells have been proposed for cardiac regeneration, and their mechanisms of action in the myocardium have been explored. Initial clinical trials have focused on the use of bone marrow-derived cells to promote myocardial regeneration in ischemic heart disease and have yielded very mixed results, with no clear signs of clinically meaningful functional improvement. Although the efficiency of bona fide cardiomyocyte generation is generally low, stem cells delivered into the myocardium act mainly via paracrine mechanisms. More recent studies taking advantage of cardiac committed cells (eg, resident cardiac progenitor cells or primed cardiogenic mesenchymal stem cells) showed promising results in first clinical pilot trials. Also, transplantation of cardiomyogenic cells generated by induced pluripotent stem cells and genetic reprogramming of dividing nonmyocytes into cardiomyocytes may constitute attractive new regenerative approaches in cardiovascular medicine in the future. We discuss advantages and limitations of specific cell types proposed for cell-based therapy in cardiology and give an overview of the first clinical trials using this novel therapeutic approach in patients with cardiovascular disease

Polo-Like Kinase 2 is Dynamically Regulated to Coordinate Proliferation and Early Lineage Specification Downstream of Yes-Associated Protein 1 in Cardiac Progenitor Cells

Background-Recent studies suggest that adult cardiac progenitor cells (CPCs) can produce new cardiac cells. Such cell formation requires an intricate coordination of progenitor cell proliferation and commitment, but the molecular cues responsible for this regulation in CPCs are ill defined. Methods and Results-Extracellular matrix components are important instructors of cell fate. Using laminin and fibronectin, we induced two slightly distinct CPC phenotypes differing in proliferation rate and commitment status and analyzed the early transcriptomic response to CPC adhesion (< 2 hours). Ninety-four genes were differentially regulated on laminin versus fibronectin, consisting of mostly downregulated genes that were enriched for Yes-associated protein (YAP) conserved signature and TEA domain family member 1 (TEAD1)-related genes. This early gene regulation was preceded by the rapid cytosolic sequestration and degradation of YAP on laminin. Among the most strongly regulated genes was polo-like kinase 2 (Plk2). Plk2 expression depended on YAP stability and was enhanced in CPCs transfected with a nuclear-targeted mutant YAP. Phenotypically, the early downregulation of Plk2 on laminin was succeeded by lower cell proliferation, enhanced lineage gene expression (24 hours), and facilitated differentiation (3 weeks) compared with fibronectin. Finally, overexpression of Plk2 enhanced CPC proliferation and knockdown of Plk2 induced the expression of lineage genes. Conclusions-Plk2 acts as coordinator of cell proliferation and early lineage commitment in CPCs. The rapid downregulation of Plk2 on YAP inactivation marks a switch towards enhanced commitment and facilitated differentiation. These findings link early gene regulation to cell fate and provide novel insights into how CPC proliferation and differentiation are orchestrated

NOX1 mediates metabolic heart disease in mice and is upregulated in monocytes of humans with diastolic dysfunction

AIMS: Microvascular inflammation plays an important role in the pathogenesis of diastolic dysfunction (DD) and metabolic heart disease. NOX1 is expressed in vascular and immune cells and has been implicated in the vascular pathology of metabolic disease. However, its contribution to metabolic heart disease is less understood. METHODS AND RESULTS: NOX1-deficient mice (KO) and male wild-type (WT) littermates were fed a high-fat high-sucrose diet (HFHS) and injected streptozotocin (75 mg/kg i.p.) or control diet (CTD) and sodium citrate. Despite similar weight gain and increase in fasting blood glucose and insulin, only WT-HFHS but not KO-HFHS mice developed concentric cardiac hypertrophy and elevated left ventricular filling pressure. This was associated with increased endothelial adhesion molecule expression, accumulation of Mac-2-, IL-1β-, and NLRP3-positive cells and nitrosative stress in WT-HFHS but not KO-HFHS hearts. Nox1 mRNA was solidly expressed in CD45(+) immune cells isolated from healthy mouse hearts but was negligible in cardiac CD31(+) endothelial cells. However, in vitro, Nox1 expression increased in response to lipopolysaccharide (LPS) in endothelial cells and contributed to LPS-induced upregulation of Icam-1. Nox1 was also upregulated in mouse bone marrow-derived macrophages in response to LPS. In peripheral monocytes from age- and sex-matched symptomatic patients with and without DD, NOX1 was significantly higher in patients with DD compared to those without DD. CONCLUSIONS: NOX1 mediates endothelial activation and contributes to myocardial inflammation and remodelling in metabolic disease in mice. Given its high expression in monocytes of humans with DD, NOX1 may represent a potential target to mitigate heart disease associated with DD

NOX1 mediates metabolic heart disease in mice and is upregulated in monocytes of humans with diastolic dysfunction

AIMS: Microvascular inflammation plays an important role in the pathogenesis of diastolic dysfunction (DD) and metabolic heart disease. NOX1 is expressed in vascular and immune cells and has been implicated in the vascular pathology of metabolic disease. However, its contribution to metabolic heart disease is less understood. METHODS AND RESULTS: NOX1-deficient mice (KO) and male wild-type (WT) littermates were fed a high-fat high-sucrose diet (HFHS) and injected streptozotocin (75 mg/kg i.p.) or control diet (CTD) and sodium citrate. Despite similar weight gain and increase in fasting blood glucose and insulin, only WT-HFHS but not KO-HFHS mice developed concentric cardiac hypertrophy and elevated left ventricular filling pressure. This was associated with increased endothelial adhesion molecule expression, accumulation of Mac-2-, IL-1β- and NLRP3-positive cells and nitrosative stress in WT-HFHS but not KO-HFHS hearts. Nox1 mRNA was solidly expressed in CD45+ immune cells isolated from healthy mouse hearts, but was negligible in cardiac CD31+ endothelial cells. However, in vitro, Nox1 expression increased in response to LPS in endothelial cells and contributed to LPS-induced upregulation of Icam-1. Nox1 was also upregulated in mouse bone marrow-derived macrophages in response to LPS. In peripheral monocytes from age- and sex-matched symptomatic patients with and without DD, NOX1 was significantly higher in patients with DD compared to those without DD. CONCLUSIONS: NOX1 mediates endothelial activation and contributes to myocardial inflammation and remodeling in metabolic disease in mice. Given its high expression in monocytes of humans with DD, NOX1 may represent a potential target to mitigate heart disease associated with DD. TRANSLATIONAL PERSPECTIVE: In their multifactorial pathogenesis, diastolic dysfunction (DD) and heart failure with preserved ejection fraction (HFpEF) still remain poorly understood. They frequently occur in patients with obesity and metabolic syndrome. Microvascular inflammation and dysfunction have recently been recognized as major driving forces. We show that genetic deletion of Nox1 prevents cardiac inflammation, remodeling and dysfunction in metabolic disease in mice and find NOX1 upregulated in peripheral monocytes of patients with DD. These findings add to our understanding how obesity, inflammation and heart disease are linked, which is a prerequisite to find therapeutic strategies beyond the control of co-morbidities in HFpEF

Dis-eguaglianza e mercato

Il rapporto tra diseguaglianza e mercato pu\uf2 essere affrontato da molteplici punti di vista. Sul piano economico, coinvolge alcune tra le pi\uf9 controverse problematiche relative al conflitto tra efficienza ed equit\ue0 nell\u2019ambito dell\u2019economia di mercato: problematiche che, paradossalmente, hanno finito per assumere toni sempre pi\uf9 polemici con il progressivo arricchirsi delle societ\ue0. Sul piano politico, comprende l\u2019ampio ventaglio delle azioni pubbliche volte a ridurre le diseguaglianze sociali prodotte dalle dinamiche di mercato: azioni che sono il frutto di scelte del legislatore e delle amministrazioni circa i tempi e i modi dell\u2019intervento pubblico nell\u2019economia (a monte, all\u2019interno o a valle del mercato; in forma di regolazione, gestione o perequazione). Sul piano giuridico, riguarda tutto l\u2019insieme delle norme e delle istituzioni che in un determinato ordinamento disciplinano il funzionamento del mercato, cercando di contemperare adeguatamente libert\ue0 individuale ed equit\ue0 sociale. La prospettiva adottata in questa sede \ue8 quella del diritto costituzionale. Dopo avere sinteticamente ricostruito il modello economico delineato dalla Costituzione italiana, nei suoi presupposti teorici e nei suoi sviluppi dottrinali e giurisprudenziali, si cerca di comprendere se esso sia stato effettivamente attuato o sia stato in tutto o in parte disatteso. Partendo dai fondamenti del pensiero economico e di quello politico per arrivare alle loro applicazioni pratiche, si valuta quindi la compatibilit\ue0 con il sistema costituzionale \u2013 e, segnatamente, con il principio di eguaglianza formale e sostanziale di cui all\u2019art. 3 Cost. \u2013 dell\u2019odierno \uabcapitalismo di mercato\ubb

Distinct HR expression patterns significantly affect the clinical behavior of metastatic HER2+ breast cancer and degree of benefit from novel anti-HER2 agents in the real world setting

We analyzed data from 738 HER2-positive metastatic breast cancer (mbc) patients treated with pertuzumab-based regimens and/or T-DM1 at 45 Italian centers. Outcomes were explored in relation to tumor subtype assessed by immunohistochemistry (IHC). The median progression-free survival at first-line (mPFS1) was 12 months. Pertuzumab as first-line conferred longer mPFS1 compared to other first-line treatments (16 vs. 9 months, p = 0.0001), regardless of IHC subtype. Median PFS in second-line (mPFS2) was 7 months, with no difference by IHC subtype, but it was more favorable with T-DM1 compared to other agents (7 vs. 6 months, p = 0.03). There was no PFS2 gain in patients with tumors expressing both hormonal receptors (HRs; p = 0.17), while a trend emerged for tumors with one HR (p = 0.05). Conversely, PFS2 gain was significant in HRs-negative tumors (p = 0.04). Median overall survival (mOS) was 74 months, with no significant differences by IHC subtypes. Survival rates at 2 and 3 years in patients treated with T-DM1 in second-line after pertuzumab were significantly lower compared to pertuzumab-naive patients (p = 0.01). When analyzed by IHC subtype, the outcome was confirmed if both HRs or no HRs were expressed (p = 0.02 and p = 0.006, respectively). Our results confirm that HRs expression impacts the clinical behavior and novel treatment-related outcomes of HER2-positive tumors when treatment sequences are considered. Moreover, multivariate analysis showed that HRs expression had no effect on PFS and OS. Further studies are warranted to confirm our findings and clarify the interplay between HER2 and estrogen receptor pathways in HER2-positive (mbc) patients