Tissue kallikrein (TK), the major kinin-forming enzyme, is synthesized in several organs, including the kidney and arteries. A loss-of-function polymorphism of the human TK gene (R53H) induces a substantial decrease in enzyme activity. As inactivation of the TK gene in the mouse induces endothelial dysfunction, we investigated the vascular, hormonal, and renal phenotypes of carriers of the 53H allele. In a crossover study, 30 R53R-homozygous and 10 R53H-heterozygous young normotensive white males were randomly assigned to receive both a low sodium–high potassium diet to stimulate TK synthesis and a high sodium–low potassium diet to suppress TK synthesis, each for 1 week. Urinary kallikrein activity was 50–60% lower in R53H subjects than in R53R subjects. Acute flow-dependent vasodilatation and endothelium-independent vasodilatation of the brachial artery were both unaffected in R53H subjects. In contrast, R53H subjects consistently exhibited an increase in wall shear stress and a paradoxical reduction in artery diameter and lumen compared with R53R subjects. Renal and hormonal adaptation to diets was unaffected in R53H subjects. The partial genetic deficiency in TK activity is associated with an inward remodeling of the brachial artery, which is not adapted to a chronic increase in wall shear stress, indicating a new form of arterial dysfunction affecting 5–7% of white people.
Michel Azizi, Pierre Boutouyrie, Alvine Bissery, Mohsen Agharazii, Francis Verbeke, Nora Stern, Alessandra Bura-Rivière, Stéphane Laurent, François Alhenc-Gelas, Xavier Jeunemaitre
We have identified a subpopulation of stem cells within adult human BM, isolated at the single-cell level, that self-renew without loss of multipotency for more than 140 population doublings and exhibit the capacity for differentiation into cells of all 3 germ layers. Based on surface marker expression, these clonally expanded human BM-derived multipotent stem cells (hBMSCs) do not appear to belong to any previously described BM-derived stem cell population. Intramyocardial transplantation of hBMSCs after myocardial infarction resulted in robust engraftment of transplanted cells, which exhibited colocalization with markers of cardiomyocyte (CMC), EC, and smooth muscle cell (SMC) identity, consistent with differentiation of hBMSCs into multiple lineages in vivo. Furthermore, upregulation of paracrine factors including angiogenic cytokines and antiapoptotic factors, and proliferation of host ECs and CMCs, were observed in the hBMSC-transplanted hearts. Coculture of hBMSCs with CMCs, ECs, or SMCs revealed that phenotypic changes of hBMSCs result from both differentiation and fusion. Collectively, the favorable effect of hBMSC transplantation after myocardial infarction appears to be due to augmentation of proliferation and preservation of host myocardial tissues as well as differentiation of hBMSCs for tissue regeneration and repair. To our knowledge, this is the first demonstration that a specific population of multipotent human BM-derived stem cells can induce both therapeutic neovascularization and endogenous and exogenous cardiomyogenesis.
Young-sup Yoon, Andrea Wecker, Lindsay Heyd, Jong-Seon Park, Tengiz Tkebuchava, Kengo Kusano, Allison Hanley, Heather Scadova, Gangjian Qin, Dong-Hyun Cha, Kirby L. Johnson, Ryuichi Aikawa, Takayuki Asahara, Douglas W. Losordo
Advanced congestive heart failure is associated with activation of the renin-angiotensin system and skeletal muscle wasting. We previously showed that angiotensin II infusion in rats produces cachexia secondarily to increased muscle proteolysis and also decreases levels of circulating and skeletal muscle IGF-1. Here we show that angiotensin II markedly downregulates phospho-Akt and activates caspase-3 in skeletal muscle, leading to actin cleavage, an important component of muscle proteolysis, and to increased apoptosis. These changes are blocked by muscle-specific expression of IGF-1, likely via the Akt/mTOR/p70S6K signaling pathway. We also demonstrate that mRNA levels of the ubiquitin ligases atrogin-1 and muscle ring finger–1 are upregulated in angiotensin II–infused WT, but not in IGF-1–transgenic, mice. These findings strongly suggest that angiotensin II downregulation of IGF-1 in skeletal muscle is causally related to angiotensin II–induced wasting. Because the renin-angiotensin system is activated in many catabolic conditions, our findings have broad implications for understanding mechanisms of skeletal muscle wasting and provide a rationale for new therapeutic approaches.
Yao-Hua Song, Yangxin Li, Jie Du, William E. Mitch, Nadia Rosenthal, Patrick Delafontaine
The angiogenic mechanism and therapeutic potential of PDGF-CC, a recently discovered member of the VEGF/PDGF superfamily, remain incompletely characterized. Here we report that PDGF-CC mobilized endothelial progenitor cells in ischemic conditions; induced differentiation of bone marrow cells into ECs; and stimulated migration of ECs. Furthermore, PDGF-CC induced the differentiation of bone marrow cells into smooth muscle cells and stimulated their growth during vessel sprouting. Moreover, delivery of PDGF-CC enhanced postischemic revascularization of the heart and limb. Modulating the activity of PDGF-CC may provide novel opportunities for treating ischemic diseases.
Xuri Li, Marc Tjwa, Lieve Moons, Pierre Fons, Agnes Noel, Annelii Ny, Jian Min Zhou, Johan Lennartsson, Hong Li, Aernout Luttun, Annica Pontén, Laetitia Devy, Ann Bouché, Hideyasu Oh, Ann Manderveld, Silvia Blacher, David Communi, Pierre Savi, Françoise Bono, Mieke Dewerchin, Jean-Michel Foidart, Monica Autiero, Jean-Marc Herbert, Désiré Collen, Carl-Henrik Heldin, Ulf Eriksson, Peter Carmeliet
Cardiac-restricted overexpression of the Ca2+-binding protein S100A1 has been shown to lead to increased myocardial contractile performance in vitro and in vivo. Since decreased cardiac expression of S100A1 is a characteristic of heart failure, we tested the hypothesis that S100A1 gene transfer could restore contractile function of failing myocardium. Adenoviral S100A1 gene delivery normalized S100A1 protein expression in a postinfarction rat heart failure model and reversed contractile dysfunction of failing myocardium in vivo and in vitro. S100A1 gene transfer to failing cardiomyocytes restored diminished intracellular Ca2+ transients and sarcoplasmic reticulum (SR) Ca2+ load mechanistically due to increased SR Ca2+ uptake and reduced SR Ca2+ leak. Moreover, S100A1 gene transfer decreased elevated intracellular Na+ concentrations to levels detected in nonfailing cardiomyocytes, reversed reactivated fetal gene expression, and restored energy supply in failing cardiomyocytes. Intracoronary adenovirus-mediated S100A1 gene delivery in vivo to the postinfarcted failing rat heart normalized myocardial contractile function and Ca2+ handling, which provided support in a physiological context for results found in myocytes. Thus, the present study demonstrates that restoration of S100A1 protein levels in failing myocardium by gene transfer may be a novel therapeutic strategy for the treatment of heart failure.
Patrick Most, Sven T. Pleger, Mirko Völkers, Beatrix Heidt, Melanie Boerries, Dieter Weichenhan, Eva Löffler, Paul M.L. Janssen, Andrea D. Eckhart, Jeffrey Martini, Matthew L. Williams, Hugo A. Katus, Andrew Remppis, Walter J. Koch
PPARα, β/δ, and γ regulate genes involved in the control of lipid metabolism and inflammation and are expressed in all major cell types of atherosclerotic lesions. In vitro studies have suggested that PPARs exert antiatherogenic effects by inhibiting the expression of proinflammatory genes and enhancing cholesterol efflux via activation of the liver X receptor–ABCA1 (LXR-ABCA1) pathway. To investigate the potential importance of these activities in vivo, we performed a systematic analysis of the effects of PPARα, β, and γ agonists on foam-cell formation and atherosclerosis in male LDL receptor–deficient (LDLR–/–) mice. Like the PPARγ agonist, a PPARα-specific agonist strongly inhibited atherosclerosis, whereas a PPARβ-specific agonist failed to inhibit lesion formation. In concert with their effects on atherosclerosis, PPARα and PPARγ agonists, but not the PPARβ agonist, inhibited the formation of macrophage foam cells in the peritoneal cavity. Unexpectedly, PPARα and PPARγ agonists inhibited foam-cell formation in vivo through distinct ABCA1-independent pathways. While inhibition of foam-cell formation by PPARα required LXRs, activation of PPARγ reduced cholesterol esterification, induced expression of ABCG1, and stimulated HDL-dependent cholesterol efflux in an LXR-independent manner. In concert, these findings reveal receptor-specific mechanisms by which PPARs influence macrophage cholesterol homeostasis. In the future, these mechanisms may be exploited pharmacologically to inhibit the development of atherosclerosis.
Andrew C. Li, Christoph J. Binder, Alejandra Gutierrez, Kathleen K. Brown, Christine R. Plotkin, Jennifer W. Pattison, Annabel F. Valledor, Roger A. Davis, Timothy M. Willson, Joseph L. Witztum, Wulf Palinski, Christopher K. Glass
Mitral valve prolapse (MVP) is a common human phenotype, yet little is known about the pathogenesis of this condition. MVP can occur in the context of genetic syndromes, including Marfan syndrome (MFS), an autosomal-dominant connective tissue disorder caused by mutations in fibrillin-1. Fibrillin-1 contributes to the regulated activation of the cytokine TGF-β, and enhanced signaling is a consequence of fibrillin-1 deficiency. We thus hypothesized that increased TGF-β signaling may contribute to the multisystem pathogenesis of MFS, including the development of myxomatous changes of the atrioventricular valves. Mitral valves from fibrillin-1–deficient mice exhibited postnatally acquired alterations in architecture that correlated both temporally and spatially with increased cell proliferation, decreased apoptosis, and excess TGF-β activation and signaling. In addition, TGF-β antagonism in vivo rescued the valve phenotype, suggesting a cause and effect relationship. Expression analyses identified increased expression of numerous TGF-β–related genes that regulate cell proliferation and survival and plausibly contribute to myxomatous valve disease. These studies validate a novel, genetically engineered murine model of myxomatous changes of the mitral valve and provide critical insight into the pathogenetic mechanism of such changes in MFS and perhaps more common nonsyndromic variants of mitral valve disease.
Connie M. Ng, Alan Cheng, Loretha A. Myers, Francisco Martinez-Murillo, Chunfa Jie, Djahida Bedja, Kathleen L. Gabrielson, Jennifer M.W. Hausladen, Robert P. Mecham, Daniel P. Judge, Harry C. Dietz
Homozygosity for mutations in ABC transporter A1 (ABCA1) causes Tangier disease, a rare HDL-deficiency syndrome. Whether heterozygosity for genetic variation in ABCA1 also contributes to HDL cholesterol (HDL-C) levels in the general population is presently unclear. We determined whether mutations or single-nucleotide polymorphisms (SNPs) in ABCA1 were overrepresented in individuals with the lowest 1% (n = 95) or highest 1% (n = 95) HDL-C levels in the general population by screening the core promoter and coding region of ABCA1. For all nonsynonymous SNPs identified, we determined the effect of genotype on lipid traits in 9,259 individuals from the general population. Heterozygosity for ABCA1 mutations was identified in 10% of individuals with low HDL-C only. Three of 6 nonsynonymous SNPs (V771M, V825I, and R1587K) were associated with increases or decreases in HDL-C in women in the general population and some with consistent trends in men, determined as isolated single-site effects varying only at the relevant SNP. Finally, these results were consistent over time. In conclusion, we show that at least 10% of individuals with low HDL-C in the general population are heterozygous for mutations in ABCA1 and that both mutations and SNPs in ABCA1 contribute to HDL-C levels in the general population.
Ruth Frikke-Schmidt, Børge G. Nordestgaard, Gorm B. Jensen, Anne Tybjærg-Hansen
Calcineurin, which binds to the Z-disc in cardiomyocytes via α-actinin, promotes cardiac hypertrophy in response to numerous pathologic stimuli. However, the endogenous mechanisms regulating calcineurin activity in cardiac muscle are not well understood. We demonstrate that a muscle-specific F-box protein called atrogin-1, or muscle atrophy F-box, directly interacts with calcineurin A and α-actinin-2 at the Z-disc of cardiomyocytes. Atrogin-1 associates with Skp1, Cul1, and Roc1 to assemble an SCFatrogin-1 complex with ubiquitin ligase activity. Expression of atrogin-1 decreases levels of calcineurin A and promotes its ubiquitination. Moreover, atrogin-1 attenuates agonist-induced calcineurin activity and represses calcineurin-dependent transactivation and NFATc4 translocation. Conversely, downregulation of atrogin-1 using adenoviral small interfering RNA (siRNA) expression enhances agonist-induced calcineurin activity and cardiomyocyte hypertrophy. Consistent with these cellular observations, overexpression of atrogin-1 in hearts of transgenic mice reduces calcineurin protein levels and blunts cardiac hypertrophy after banding of the thoracic aorta. These studies indicate that the SCFatrogin-1 ubiquitin ligase complex interacts with and represses calcineurin by targeting calcineurin for ubiquitin-mediated proteolysis, leading to inhibition of cardiac hypertrophy in response to pathologic stimuli.
Hui-Hua Li, Vishram Kedar, Chunlian Zhang, Holly McDonough, Ranjana Arya, Da-Zhi Wang, Cam Patterson
The Raf/MEK/extracellular signal–regulated kinase (ERK) signaling pathway regulates diverse cellular processes such as proliferation, differentiation, and apoptosis and is implicated as an important contributor to the pathogenesis of cardiac hypertrophy and heart failure. To examine the in vivo role of Raf-1 in the heart, we generated cardiac muscle–specific Raf-1–knockout (Raf CKO) mice with Cre-loxP–mediated recombination. The mice demonstrated left ventricular systolic dysfunction and heart dilatation without cardiac hypertrophy or lethality. The Raf CKO mice showed a significant increase in the number of apoptotic cardiomyocytes. The expression level and activation of MEK1/2 or ERK showed no difference, but the kinase activity of apoptosis signal–regulating kinase 1 (ASK1), JNK, or p38 increased significantly compared with that in controls. The ablation of ASK1 rescued heart dysfunction and dilatation as well as cardiac fibrosis. These results indicate that Raf-1 promotes cardiomyocyte survival through a MEK/ERK–independent mechanism.
Osamu Yamaguchi, Tetsuya Watanabe, Kazuhiko Nishida, Kazunori Kashiwase, Yoshiharu Higuchi, Toshihiro Takeda, Shungo Hikoso, Shinichi Hirotani, Michio Asahi, Masayuki Taniike, Atsuko Nakai, Ikuko Tsujimoto, Yasushi Matsumura, Jun-ichi Miyazaki, Kenneth R. Chien, Atsushi Matsuzawa, Chiharu Sadamitsu, Hidenori Ichijo, Manuela Baccarini, Masatsugu Hori, Kinya Otsu