Thrombosis in the Coronary Microvasculature Impairs Cardiac Relaxation and Induces Diastolic Dysfunction.

BACKGROUND: Heart failure with preserved ejection fraction is proposed to be caused by endothelial dysfunction in cardiac microvessels. Our goal was to identify molecular and cellular mechanisms underlying the development of cardiac microvessel disease and diastolic dysfunction in the setting of type 2 diabetes. METHODS: We used Leprdb/db (leptin receptor-deficient) female mice as a model of type 2 diabetes and heart failure with preserved ejection fraction and identified Hhipl1 (hedgehog interacting protein-like 1), which encodes for a decoy receptor for HH (hedgehog) ligands as a gene upregulated in the cardiac vascular fraction of diseased mice. RESULTS: We then used Dhh (desert HH)-deficient mice to investigate the functional consequences of impaired HH signaling in the adult heart. We found that Dhh-deficient mice displayed increased end-diastolic pressure while left ventricular ejection fraction was comparable to that of control mice. This phenotype was associated with a reduced exercise tolerance in the treadmill test, suggesting that Dhh-deficient mice do present heart failure. At molecular and cellular levels, impaired cardiac relaxation in DhhECKO mice was associated with a significantly decreased PLN (phospholamban) phosphorylation on Thr17 (threonine 17) and an alteration of sarcomeric shortening ex vivo. Besides, as expected, Dhh-deficient mice exhibited phenotypic changes in their cardiac microvessels including a prominent prothrombotic phenotype. Importantly, aspirin therapy prevented the occurrence of both diastolic dysfunction and exercise intolerance in these mice. To confirm the critical role of thrombosis in the pathophysiology of diastolic dysfunction, we verified Leprdb/db also displays increased cardiac microvessel thrombosis. Moreover, consistently, with Dhh-deficient mice, we found that aspirin treatment decreased end-diastolic pressure and improved exercise tolerance in Leprdb/db mice. CONCLUSIONS: Altogether, these results demonstrate that microvessel thrombosis may participate in the pathophysiology of heart failure with preserved ejection fraction.

Partial Mural Cell Ablation Disrupts Coronary Vasculature Integrity and Induces Systolic Dysfunction.

Background Although the critical role of pericytes in maintaining vascular integrity has been extensively demonstrated in the brain and the retina, little is known about their role in the heart. We aim to investigate structural and functional consequences of partial pericyte depletion (≈60%) in the heart of adult mice. Methods and Results To deplete pericytes in adult mice, we used platelet-derived growth factor receptor ß-Cre/ERT2; RosaDTA mice and compared their phenotype with that of control mice (RosaDTA) chosen among their littermates. Cardiac function was assessed via echocardiography and left ventricular catheterization 1 month after the first tamoxifen injection. We found mice depleted with pericytes had a reduced left ventricular ejection fraction and an increased end-diastolic pressure, demonstrating both systolic and diastolic dysfunction. Consistently, mice depleted with pericytes presented a decreased left ventricular contractility and an increased left ventricular relaxation time (dP/dtmin). At the tissue level, mice depleted of pericytes displayed increased coronary endothelium leakage and activation, which was associated with increased CD45+ cell infiltration. Consistent with systolic dysfunction, pericyte depletion was associated with an increased expression of myosin heavy chain 7 and decreased expression of ATPase sarcoplasmic/endoplasmic reticulum Ca2+ transporting 2 and connexin 43. More important, coculture assays demonstrated, for the first time, that the decreased expression of connexin 43 is likely attributable to a direct effect of pericytes on cardiomyocytes. Besides, this study reveals that cardiac pericytes may undergo strong remodeling on injury. Conclusions Cardiac pericyte depletion induces both systolic and diastolic dysfunction, suggesting that pericyte dysfunction may contribute to the occurrence of cardiac diseases.

Praliciguat Promotes Ischemic Leg Reperfusion in Leptin Receptor-Deficient Mice.

BACKGROUND: Lower-limb peripheral artery disease is one of the major complications of diabetes. Peripheral artery disease is associated with poor limb and cardiovascular prognoses, along with a dramatic decrease in life expectancy. Despite major medical advances in the treatment of diabetes, a substantial therapeutic gap remains in the peripheral artery disease population. Praliciguat is an orally available sGC (soluble guanylate cyclase) stimulator that has been reported both preclinically and in early stage clinical trials to have favorable effects in metabolic and hemodynamic outcomes, suggesting that it may have a potential beneficial effect in peripheral artery disease. METHODS: We evaluated the effect of praliciguat on hind limb ischemia recovery in a mouse model of type 2 diabetes. Hind limb ischemia was induced in leptin receptor-deficient (Leprdb/db) mice by ligation and excision of the left femoral artery. Praliciguat (10 mg/kg/day) was administered in the diet starting 3 days before surgery. RESULTS: Twenty-eight days after surgery, ischemic foot perfusion and function parameters were better in praliciguat-treated mice than in vehicle controls. Improved ischemic foot perfusion was not associated with either improved traditional cardiovascular risk factors (ie, weight, glycemia) or increased angiogenesis. However, treatment with praliciguat significantly increased arteriole diameter, decreased ICAM1 (intercellular adhesion molecule 1) expression, and prevented the accumulation of oxidative proangiogenic and proinflammatory muscle fibers. While investigating the mechanism underlying the beneficial effects of praliciguat therapy, we found that praliciguat significantly downregulated Myh2 and Cxcl12 mRNA expression in cultured myoblasts and that conditioned medium form praliciguat-treated myoblast decreased ICAM1 mRNA expression in endothelial cells. These results suggest that praliciguat therapy may decrease ICAM1 expression in endothelial cells by downregulating Cxcl12 in myocytes. CONCLUSIONS: Our results demonstrated that praliciguat promotes blood flow recovery in the ischemic muscle of mice with type 2 diabetes, at least in part by increasing arteriole diameter and by downregulating ICAM1 expression.

Endothelial Dysfunction in Heart Failure With Preserved Ejection Fraction: What are the Experimental Proofs?

Heart failure with preserved ejection fraction (HFpEF) has been recognized as the greatest single unmet need in cardiovascular medicine. Indeed, the morbi-mortality of HFpEF is high and as the population ages and the comorbidities increase, so considerably does the prevalence of HFpEF. However, HFpEF pathophysiology is still poorly understood and therapeutic targets are missing. An unifying, but untested, theory of the pathophysiology of HFpEF, proposed in 2013, suggests that cardiovascular risk factors lead to a systemic inflammation, which triggers endothelial cells (EC) and coronary microvascular dysfunction. This cardiac small vessel disease is proposed to be responsible for cardiac wall stiffening and diastolic dysfunction. This paradigm is based on the fact that microvascular dysfunction is highly prevalent in HFpEF patients. More specifically, HFpEF patients have been shown to have decreased cardiac microvascular density, systemic endothelial dysfunction and a lower mean coronary flow reserve. Importantly, impaired coronary microvascular function has been associated with the severity of HF. This review discusses evidence supporting the causal role of endothelial dysfunction in the pathophysiology of HFpEF in human and experimental models.

Mast Cells Are the Trigger of Small Vessel Disease and Diastolic Dysfunction in Diabetic Obese Mice.

[Figure: see text].

Full-length Dhh and N-terminal Shh act as competitive antagonists to regulate angiogenesis and vascular permeability.

AIMS: The therapeutic potential of Hedgehog (Hh) signalling agonists for vascular diseases is of growing interest. However, molecular and cellular mechanisms underlying the role of the Hh signalling in vascular biology remain poorly understood. The purpose of the present article is to clarify some conflicting literature data. METHODS AND RESULTS: With this goal, we have demonstrated that, unexpectedly, ectopically administered N-terminal Sonic Hh (N-Shh) and endogenous endothelial-derived Desert Hh (Dhh) induce opposite effects in endothelial cells (ECs). Notably, endothelial Dhh acts under its full-length soluble form (FL-Dhh) and activates Smoothened in ECs, while N-Shh inhibits it. At molecular level, N-Shh prevents FL-Dhh binding to Patched-1 (Ptch1) demonstrating that N-Shh acts as competitive antagonist to FL-Dhh. Besides, we found that even though FL-Hh ligands and N-Hh ligands all bind Ptch1, they induce distinct Ptch1 localization. Finally, we confirmed that in a pathophysiological setting, i.e. brain inflammation, astrocyte-derived N-Shh acts as a FL-Dhh antagonist. CONCLUSION: The present study highlights for the first time that FL-Dhh and N-Hh ligands have antagonistic properties especially in ECs.

Blood-brain barrier genetic disruption leads to protective barrier formation at the Glia Limitans.

Inflammation of the central nervous system (CNS) induces endothelial blood-brain barrier (BBB) opening as well as the formation of a tight junction barrier between reactive astrocytes at the Glia Limitans. We hypothesized that the CNS parenchyma may acquire protection from the reactive astrocytic Glia Limitans not only during neuroinflammation but also when BBB integrity is compromised in the resting state. Previous studies found that astrocyte-derived Sonic hedgehog (SHH) stabilizes the BBB during CNS inflammatory disease, while endothelial-derived desert hedgehog (DHH) is expressed at the BBB under resting conditions. Here, we investigated the effects of endothelial Dhh on the integrity of the BBB and Glia Limitans. We first characterized DHH expression within endothelial cells at the BBB, then demonstrated that DHH is down-regulated during experimental autoimmune encephalomyelitis (EAE). Using a mouse model in which endothelial Dhh is inducibly deleted, we found that endothelial Dhh both opens the BBB via the modulation of forkhead box O1 (FoxO1) transcriptional activity and induces a tight junctional barrier at the Glia Limitans. We confirmed the relevance of this glial barrier system in human multiple sclerosis active lesions. These results provide evidence for the novel concept of "chronic neuroinflammatory tolerance" in which BBB opening in the resting state is sufficient to stimulate a protective barrier at the Glia Limitans that limits the severity of subsequent neuroinflammatory disease. In summary, genetic disruption of the BBB generates endothelial signals that drive the formation under resting conditions of a secondary barrier at the Glia Limitans with protective effects against subsequent CNS inflammation. The concept of a reciprocally regulated CNS double barrier system has implications for treatment strategies in both the acute and chronic phases of multiple sclerosis pathophysiology.

Desert Hedgehog-Driven Endothelium Integrity Is Enhanced by Gas1 (Growth Arrest-Specific 1) but Negatively Regulated by Cdon (Cell Adhesion Molecule-Related/Downregulated by Oncogenes).

OBJECTIVE: Evidences accumulated within the past decades identified hedgehog signaling as a new regulator of endothelium integrity. More specifically, we recently identified Dhh (desert hedgehog) as a downstream effector of Klf2 (Kruppel-like factor 2) in endothelial cells (ECs). The purpose of this study is to investigate whether hedgehog coreceptors Gas1 (growth arrest-specific 1) and Cdon (cell adhesion molecule-related/downregulated by oncogenes) may be used as therapeutic targets to modulate Dhh signaling in ECs. Approach and Results: We demonstrated that both Gas1 and Cdon are expressed in adult ECs and relied on either siRNAs- or EC-specific conditional knockout mice to investigate their role. We found that Gas1 deficiency mainly phenocopies Dhh deficiency especially by inducing VCAM-1 (vascular cell adhesion molecule 1) and ICAM-1 (intercellular adhesion molecule 1) overexpression while Cdon deficiency has opposite effects by promoting endothelial junction integrity. At a molecular level, Cdon prevents Dhh binding to Ptch1 (patched-1) and thus acts as a decoy receptor for Dhh, while Gas1 promotes Dhh binding to Smo (smoothened) and as a result potentiates Dhh effects. Since Cdon is upregulated in ECs treated by inflammatory cytokines, including TNF (tumor necrosis factor)-α and Il (interleukin)-1ß, we then tested whether Cdon inhibition would promote endothelium integrity in acute inflammatory conditions and found that both fibrinogen and IgG extravasation were decreased in association with an increased Cdh5 (cadherin-5) expression in the brain cortex of EC-specific Cdon knockout mice administered locally with Il-1ß. CONCLUSIONS: Altogether, these results demonstrate that Gas1 is a positive regulator of Dhh in ECs while Cdon is a negative regulator. Interestingly, Cdon blocking molecules may then be used to promote endothelium integrity, at least in inflammatory conditions.

Identification of variables influencing pharmaceutical interventions to improve medication review efficiency.

Background Clinical pharmacists' involvement has improved patients' care, by suggesting therapeutic optimizations. However, budget restrictions require a prioritization of these activities to focus resources on patients more at risk of medication errors. Objective The aim of our study was to identify variables influencing the formulation of pharmaceutical to improve medication review efficiency. Setting This study was conducted in medical wards of a 643-acute beds hospital in Paris, France. Methods All hospital medical prescriptions of all patients admitted within four medical wards (cardiology, rheumatology, neurology, vascular medicine) were analyzed. The study was conducted in each ward for 2 weeks, during 4 weeks. For each patient, variables prospectively collected were: age, gender, weight, emergency admission, number of high-alert medications and of total drugs prescribed, care unit, serum creatinine. Number of pharmaceutical interventions (PIs) and their type were reported. Main outcome measures Variables influencing the number of pharmaceutical interventions during medication review were identified using simple and multiple linear regressions. Results A total of 2328 drug prescriptions (303 patients, mean age 70.6 years-old) were analyzed. Mean number of hospital drug prescriptions was 7.9. A total of 318 PIs were formulated. Most frequent PIs were drug omission (n = 88, 27.7%), overdosing (n = 69, 21.7%), and underdosing (n = 51, 16.0%). Among variables studied, age, serum creatinine level, number of high-alert medications prescribed and total number of drugs prescribed were significantly associated with the formulation of pharmaceutical interventions (adjusted R2 = 0.34). Conclusions This study identified variables (age, serum creatinine level, number of high-alert medication, number of prescribed drugs) that may help institutions/pharmacists target their reviews towards patients most likely to require pharmacist interventions.