Background Transcatheter edge-to-edge mitral valve repair using the MitraClip device is increasingly used for high surgical risk patients with severe mitral regurgitation (MR). Previous guidelines for infective endocarditis prophylaxis prior to dental procedures focused on high-risk patients, but without explicit recommendation for MitraClip recipients. We believe this could be the first reported case to identify Streptococcus oralis as the causative organism. Case presentation An 87-year-old male with severe MR treated with two MitraClip devices three months prior to index admission, presented with worsening malaise and intermittent chills on a background of multiple comorbid conditions. The patient had dental work a month prior to presentation, including a root canal procedure, without antibiotic prophylaxis. Vitals were significant for fever and hypotension. On physical examination, there was a holosystolic murmur at the apex radiating to the axilla, bilateral pitting edema in the lower extremities, and elevated jugular venous pulsation. A transthoracic echocardiogram showed severe MR with a possible echodensity on the mitral valve, prompting a transesophageal echocardiogram, which demonstrated a pedunculated, mobile mass on the posterior leaflet of the mitral valve. Five blood cultures grew gram positive cocci in pairs and chains, later identified as Streptococcus oralis , with minimum inhibitory concentration to penicillin 0.06 mg/L. Initial empiric antibiotics were switched to ceftriaxone 2 gr daily and subsequent blood cultures remained negative. However, the patient developed pulmonary edema and worsening hemodynamic instability requiring vasopressors. As surgical risk for re-operation was considered prohibitive, the decision was made to continue medical management and comfort-directed care. The patient died a week later. Conclusions Despite low incidence, infective endocarditis should be included in the differential among MitraClip recipients. The explicit inclusion of this growing patient population in the group requiring prophylaxis prior to dental procedures in the 2020 ACC/AHA valvular heart disease guidelines is an important step forward.

Angiogenesis is a multistep process that requires highly regulated endothelial cell (EC) behavior. The transcription factor Krüppel-like factor 4 (KLF4) is a critical regulator of several basic EC functions; we have recently shown that KLF4 disturbs pathological (tumor) angiogenesis by mediating the expression of members of VEGF and Notch signaling pathways. Notch signaling is central to orchestration of sprouting angiogenesis but little is known about the upstream regulation of Notch itself. To determine the role of KLF4 in normal (developmental) angiogenesis, we used a mouse retinal angiogenesis model. We found that endothelial-specific overexpression of KLF4 in transgenic mice (EC-K4 Tg) leads to increased vessel density, branching and number of tip cell filopodia as assessed on postnatal day 6 (P6). The hypertrophic vasculature seen with sustained KLF4 overexpression is not stable and undergoes prominent remodeling during P7–P12 resulting in a normal appearing retinal vasculature in adult EC-K4 Tg mice. We find that KLF4 inhibits Delta-like 4 (DLL4) expression in the angiogenic front during retinal vascular development. Furthermore, in an oxygen-induced retinopathy model, overexpression of KLF4 results in decreased vaso-obliteration and neovascular tuft formation that is similar to genetic or pharmacologic DLL4 inhibition. Mechanistically, we show that KLF4 disables the activity of the essential Notch transcriptional activator RBP-J by interfering with binding of co-activators NICD and MAML at intron 3 of the Notch ligand DLL4. In summary, our experimental results demonstrate a regulatory role of KLF4 in developmental angiogenesis through regulation of DLL4 transcription.

Loss of protein and organelle quality control secondary to reduced autophagy is a hallmark of aging. However, the physiologic and molecular regulation of autophagy in long-lived organisms remains incompletely understood. Here we show that the Kruppel-like family of transcription factors are important regulators of autophagy and healthspan in C. elegans, and also modulate mammalian vascular age-associated phenotypes. Kruppel-like family of transcription factor deficiency attenuates autophagy and lifespan extension across mechanistically distinct longevity nematode models. Conversely, Kruppel-like family of transcription factor overexpression extends nematode lifespan in an autophagy-dependent manner. Furthermore, we show the mammalian vascular factor Kruppel-like family of transcription factor 4 has a conserved role in augmenting autophagy and improving vessel function in aged mice. Kruppel-like family of transcription factor 4 expression also decreases with age in human vascular endothelium. Thus, Kruppel-like family of transcription factors constitute a transcriptional regulatory point for the modulation of autophagy and longevity in C. elegans with conserved effects in the murine vasculature and potential implications for mammalian vascular aging.KLF family transcription factors (KLFs) regulate many cellular processes, including proliferation, survival and stress responses. Here, the authors position KLFs as important regulators of autophagy and lifespan in C. elegans, a role that may extend to the modulation of age-associated vascular phenotypes in mammals.

Rationale: Lymphatic vessels function to drain interstitial fluid from a variety of tissues. Although shear stress generated by fluid flow is known to trigger lymphatic expansion and remodeling, the molecular basis underlying flow-induced lymphatic growth is unknown. Objective: We aimed to gain a better understanding of the mechanism by which laminar shear stress activates lymphatic proliferation. Methods and Results: Primary endothelial cells from dermal blood and lymphatic vessels (blood vascular endothelial cells and lymphatic endothelial cells [LECs]) were exposed to low-rate steady laminar flow. Shear stress–induced molecular and cellular responses were defined and verified using various mutant mouse models. Steady laminar flow induced the classic shear stress responses commonly in blood vascular endothelial cells and LECs. Surprisingly, however, only LECs showed enhanced cell proliferation by regulating the vascular endothelial growth factor (VEGF)-A, VEGF-C, FGFR3, and p57/CDKN1C genes. As an early signal mediator, ORAI1, a pore subunit of the calcium release–activated calcium channel, was identified to induce the shear stress phenotypes and cell proliferation in LECs responding to the fluid flow. Mechanistically, ORAI1 induced upregulation of Krüppel-like factor (KLF)-2 and KLF4 in the flow-activated LECs, and the 2 KLF proteins cooperate to regulate VEGF-A, VEGF-C, FGFR3, and p57 by binding to the regulatory regions of the genes. Consistently, freshly isolated LECs from Orai1 knockout embryos displayed reduced expression of KLF2, KLF4, VEGF-A, VEGF-C, and FGFR3 and elevated expression of p57. Accordingly, mouse embryos deficient in Orai1 , Klf2 , or Klf4 showed a significantly reduced lymphatic density and impaired lymphatic development. Conclusions: Our study identified a molecular mechanism for laminar flow–activated LEC proliferation.

Historically, the relationship between exercise and the cardiovascular system was viewed as unidirectional, with a disease resulting in exercise limitation and hazard. This article reviews and explores the bidirectional nature, delineating the effects, generally positive, on the cardiovascular system and atherosclerosis. Exercise augments eNOS, affects redox potential, and favorably affects mediators of atherosclerosis including lipids, glucose homeostasis, and inflammation. There are direct effects on the vasculature as well as indirect benefits related to exercise-induced changes in body composition and skeletal muscle. Application of aerobic exercise to specific populations is described, with the hope that this knowledge will move the science forward and improve individual patient outcome.

Objective:To determine the relationships between Krüppel-like factors (KLF) 2 and 4, immune-activation, and subclinical vascular disease in HIV-infected patients on antiretroviral therapy (ART). Design:Double-blind, randomized, placebo-controlled trial. Methods:We studied 74 HIV-infected adults on ART enrolled in a randomized clinical trial of statin therapy. KLF2 and KLF4 gene expression was measured by quantitative PCR from peripheral blood mononuclear cells (PBMCs) at baseline and after 24 weeks of 10 mg daily rosuvastatin or placebo. At the same time points, T-cell and monocyte activation were assessed by flow cytometry and vascular health was assessed by cardiac computed tomography and carotid ultrasound. Results:KLF4 expression was negatively correlated with duration of ART (r = −0.351, P = 0.004) and positively correlated with measures of immune activation: proinflammatory monocytes [CD14+CD16+ (r = 0.343, P = 0.003)], patrolling monocytes [CD14dimCD16+ (r = 0.276, P = 0.017)], and activated CD8+ T-lymphocytes [CD8+DR+CD38+ (r = 0.264, P = 0.023)]. KLF2 expression was negatively correlated with subclinical atherosclerosis: mean–mean common carotid artery intima–media thickness (r = −0.231, P = 0.048), mean–max carotid artery intima-media thickness (r = −0.271, P = 0.020), and coronary artery calcium score (r = −0.254, P = 0.029). There were no statistically significant changes in KLF2/4 expression in PBMCs after 24 weeks of rosuvastatin. Conclusion:Expression of KLF4 in PBMCs positively correlates with cellular markers of immune activation, whereas KLF2 expression negatively correlates with markers of subclinical atherosclerosis in this HIV-infected population on ART. Additional studies are needed to determine if targeted interventions might alter KLF2/4 expression to reduce inflammation and vascular risk in humans.

This invited review summarizes work presented in the Russell Ross lecture delivered at the 2012 proceedings of the American Heart Association. We begin with a brief overview of the structural, cellular, and molecular biology of Krüppel-like factors. We then focus on discoveries during the past decade, implicating Krüppel-like factors as key determinants of vascular cell function in atherosclerotic vascular disease.

Background: The transcription factor Krüppel-like factor 4 (KLF4) is a critical regulator of endothelial cell biology. Results: Sustained expression of endothelial KLF4 limits tumor growth by creating ineffective angiogenesis. Conclusion: KLF4 is an upstream regulator of angiogenesis in part by mediating Notch expression and activity. Significance: KLF4 regulates sprouting angiogenesis and may be a therapeutic target in regulation of tumor angiogenesis. Regulation of endothelial cell biology by the Notch signaling pathway (Notch) is essential to vascular development, homeostasis, and sprouting angiogenesis. Although Notch determines cell fate and differentiation in a wide variety of cells, the molecular basis of upstream regulation of Notch remains poorly understood. Our group and others have implicated the Krüppel-like factor family of transcription factors as critical regulators of endothelial function. Here, we show that Krüppel-like factor 4 (KLF4) is a central regulator of sprouting angiogenesis via regulating Notch. Using a murine model in which KLF4 is overexpressed exclusively in the endothelium, we found that sustained expression of KLF4 promotes ineffective angiogenesis leading to diminished tumor growth independent of endothelial cell proliferation or cell cycling effects. These tumors feature increased vessel density yet are hypoperfused, leading to tumor hypoxia. Mechanistically, we show that KLF4 differentially regulates expression of Notch receptors, ligands, and target genes. We also demonstrate that KLF4 limits cleavage-mediated activation of Notch1. Finally, we rescue Notch target gene expression and the KLF4 sprouting angiogenesis phenotype by supplementation of DLL4 recombinant protein. Identification of this hitherto undiscovered role of KLF4 implicates this transcription factor as a critical regulator of Notch, tumor angiogenesis, and sprouting angiogenesis.

This invited review summarizes work presented in the Russell Ross lecture delivered at the 2012 proceedings of the American Heart Association. We begin with a brief overview of the structural, cellular, and molecular biology of Krüppel-like factors. We then focus on discoveries during the past decade, implicating Krüppel-like factors as key determinants of vascular cell function in atherosclerotic vascular disease.

Krüppel-like factor 4 (KLF4) is a transcription factor expressed in the vascular endothelium, where it promotes anti-inflammatory and anticoagulant states, and increases endothelial nitric oxide synthase expression. We examined the role of endothelial KLF4 in pulmonary arterial (PA) hypertension (PAH). Mice with endothelial KLF4 knockdown were exposed to hypoxia for 3 weeks, followed by measurement of right ventricular and PA pressures, pulmonary vascular muscularization, and right ventricular hypertrophy. The effect of KLF4 on target gene expression was assessed in lungs from these mice, verified in vitro by small interfering RNA (siRNA) knockdown of KLF4, and further studied at the promoter level with cotransfection experiments. KLF4 expression was measured in lung tissue from patients with PAH and normal control subjects. We found that, after hypoxia, right ventricular and PA pressures were significantly higher in KLF4 knockdown animals than controls. Knockdown animals also had more severe pulmonary vascular muscularization and right ventricular hypertrophy. KLF4 knockdown resulted in increased pulmonary expression of endothelin-1 and decreased expression of endothelial nitric oxide synthase, endothelin receptor subtype B, and prostacyclin synthase. Concordant findings were observed in vitro, both with siRNA knockdown of KLF4 and promoter activity assays. Finally, KLF4 expression was reduced in lungs from patients with PAH. In conclusion, endothelial KLF4 regulates the transcription of genes involved in key pathways implicated in PAH, and its loss exacerbates pulmonary hypertension in response to chronic hypoxia in mice. These results introduce a novel transcriptional modulator of PAH, with the potential of becoming a new therapeutic target.

The endothelium regulates vascular homeostasis, and endothelial dysfunction is a proximate event in the pathogenesis of atherothrombosis. Stimulation of the endothelium with proinflammatory cytokines or exposure to hemodynamic-induced disturbed flow leads to a proadhesive and prothrombotic phenotype that promotes atherothrombosis. In contrast, exposure to arterial laminar flow induces a gene program that confers a largely antiadhesive, antithrombotic effect. The molecular basis for this differential effect on endothelial function remains poorly understood. While recent insights implicate Kruppel-like factors (KLFs) as important regulators of vascular homeostasis, the in vivo role of these factors in endothelial biology remains unproven. Here, we show that endothelial KLF4 is an essential determinant of atherogenesis and thrombosis. Using in vivo EC-specific KLF4 overexpression and knockdown murine models, we found that KLF4 induced an antiadhesive, antithrombotic state. Mechanistically, we demonstrated that KLF4 differentially regulated pertinent endothelial targets via competition for the coactivator p300. These observations provide cogent evidence implicating endothelial KLFs as essential in vivo regulators of vascular function in the adult animal.

A healthy organism demands much from its endothelium; constance in homeostatic control of blood fluidity, vascular tone, interaction with circulating blood cells, maintenance of barrier functions, and, when appropriate, creation of new blood vessels.Breakdown in any of these functions, anywhere in the 60,000 miles of vessels in the human body, leads to or exacerbates a vast array of diseases including those plaguing modern man: atherosclerosis, diabetes, hypertension, cancer, and clotting disorders.In recent years, 2 endothelial transcription factors, Kruppel-like factor 2 (KLF2) and Kruppel-like factor 4 (KLF4), have garnered attention as guardians of endothelial health.2][3][4] Thus, increased knowledge of the regulation of these regulators is of great interest.In this issue of Arteriosclerosis, Thrombosis, and Vascular Biology, Fang and Davies demonstrate that a single microRNA, miR92, regulates both KLF2 and KLF4 -a finding that has exciting scientific and potentially therapeutic implications.

Current paradigms suggest that two macrophage subsets, termed M1 and M2, are involved in inflammation and host defense. While the distinct functions of M1 and M2 macrophages have been intensively studied - the former are considered proinflammatory and the latter antiinflammatory - the determinants of their speciation are incompletely understood. Here we report our studies that identify Krüppel-like factor 4 (KLF4) as a critical regulator of macrophage polarization. Macrophage KLF4 expression was robustly induced in M2 macrophages and strongly reduced in M1 macrophages, observations that were recapitulated in human inflammatory paradigms in vivo. Mechanistically, KLF4 was found to cooperate with Stat6 to induce an M2 genetic program and inhibit M1 targets via sequestration of coactivators required for NF-κB activation. KLF4-deficient macrophages demonstrated increased proinflammatory gene expression, enhanced bactericidal activity, and altered metabolism. Furthermore, mice bearing myeloid-specific deletion of KLF4 exhibited delayed wound healing and were predisposed to developing diet-induced obesity, glucose intolerance, and insulin resistance. Collectively, these data identify KLF4 as what we believe to be a novel regulator of macrophage polarization.

Antiphospholipid syndrome is characterized by thrombosis and/or recurrent pregnancy loss in the presence of antiphospholipid antibodies (APLAs). The majority of APLAs are directed against phospholipid-binding proteins, particularly β₂-glycoprotein I (β₂GPI). Anti-β₂GPI antibodies activate endothelial cells in a β₂GPI-dependent manner through a pathway that involves NF-κB. Krüppel-like factors (KLFs) play a critical role in regulating the endothelial response to inflammatory stimuli. We hypothesized that activation of endothelial cells by APLA/anti-β₂GPI antibodies might be associated with decreased expression of KLFs, which in turn might facilitate cellular activation mediated through NF-κB. Our experimental results confirmed this hypothesis, demonstrating markedly decreased expression of KLF2 and KLF4 after incubation of cells with APLA/anti-β₂GPI antibodies. Restoration of KLF2 or KLF4 levels inhibited NF-κB transcriptional activity and blocked APLA/anti-β₂GPI-mediated endothelial activation despite NF-κB p65 phosphorylation. Chromatin immunoprecipitation analysis demonstrated that inhibition of NF-κB transcriptional activity by KLFs reflects sequestration of the cotranscriptional activator CBP/p300, making this cofactor unavailable to NF-κB. These findings suggest that the endothelial response to APLA/anti-β₂GPI antibodies reflects competition between KLFs and NF-κB for their common cofactor, CBP/p300. Taken together, these observations are the first to implicate the KLFs as novel participants in the endothelial proinflammatory response to APLA/anti-β₂GPI antibodies.

Objective—Grb2-associated binder 1 (Gab1), a scaffolding adaptor protein, plays an important role in transmitting key signals that control cell growth, differentiation, and function from multiple tyrosine kinase receptors. The study was designed to investigate the role of endothelial Gab1 in angiogenesis and its underlying molecular mechanisms. Methods and Results—Using Cre-Lox recombination technology, we generated endothelial-specific Gab1 knockout (Gab1-ecKO) mice. Gab1-ecKO mice are viable and showed no obvious developmental defects in the vascular system. To analyze the role of Gab1 in postnatal angiogenesis, we used hindlimb ischemia and Matrigel plug models. We found that loss of endothelial Gab1 in mice dramatically impaired postnatal angiogenesis. Gab1-ecKO mice had impaired ischemia-initiated blood flow recovery, exhibited reduced angiogenesis, and were associated with marked limb necrosis. We further observed significant endothelial cell (EC) death in the ischemic hindlimb of Gab1-ecKO mice. Matrigel plug assay showed that hepatocyte growth factor (HGF)–mediated angiogenesis was inhibited in Gab1-ecKO mice. In vitro studies showed that Gab1 was required for HGF-induced EC migration, tube formation, and microvessel sprouting. Mechanistically, HGF stimulated Gab1 tyrosine phosphorylation in ECs, leading to activation of extracellular regulated MAP kinase 1/2 and Akt, which are angiogenic and survival signaling. Conclusion—Gab1 is essential for postnatal angiogenesis through mediating angiogenic and survival signaling.

A complex and diverse vascular system is requisite for the survival of higher organisms. The process of vascular development is highly regulated, involving the de novo formation of vessels (vasculogenesis), followed by expansion and remodeling of the primitive vasculature (angiogenesis), culminating in differentiation of endothelial phenotypes, as found in the mature vascular system. Over the last decade, significant advances have been made in understanding the molecular regulation of endothelial cell development and differentiation. Endothelial development, in particular the mechanisms in play during vasculogenesis and angiogenesis, is discussed in a sister review to this article. This review highlights the key pathways governing in endothelial differentiation, with a focus on the major molecular mechanisms of endothelial specification and heterogeneity.

A complex and diverse vascular system is requisite for the survival of higher organisms. The process of vascular development is highly regulated, involving the de novo formation of vessels (vasculogenesis), followed by expansion and remodeling of the primitive vasculature (angiogenesis), culminating in differentiation of endothelial phenotypes, as found in the mature vascular system. Over the last decade, significant advances have been made in understanding the molecular regulation of endothelial cell development and differentiation. Endothelial development, in particular the mechanisms in play during vasculogenesis and angiogenesis, is discussed in a sister review to this article. This review highlights the key pathways governing in endothelial differentiation, with a focus on the major molecular mechanisms of endothelial specification and heterogeneity.

In this issue of Blood , Zhang et al identify the Src-PLD1-PKCγ axis as critically involved in the process that causes ROP, highlighting new potential targets for therapy.[1][1] Vascular endothelial growth factor (VEGF) is acknowledged as the predominant regulator of angiogenesis; blockade of VEGF