Background-Postnatal growth of the heart chiefly involves nonproliferative cardiomyocyte enlargement. Cardiac hypertrophy exists in a "physiological" form that is an adaptive response to long-term exercise training and as a "pathological" form that often is a maladaptive response to provocative stimuli such as hypertension and aortic valvular stenosis. A signaling cascade that includes the protein kinase Akt regulates the growth and survival of many cell types, but the precise role of Akt1 in either form of cardiac hypertrophy is unknown. Methods and Results-To evaluate the role of Akt1 in physiological cardiac growth, akt1 Ϫ/Ϫ adult murine cardiac myocytes (AMCMs) were treated with IGF-1, and akt1 Ϫ/Ϫ mice were subjected to exercise training. akt1 Ϫ/Ϫ AMCMs were resistant to insulin-like growth factor-1-stimulated protein synthesis. The akt1 Ϫ/Ϫ mice were found to be resistant to swimming training-induced cardiac hypertrophy. To evaluate the role of Akt in pathological cardiac growth, akt1

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Bone homeostasis requires stringent regulation of osteoclasts, which secrete proteolytic enzymes to degrade the bone matrix. Despite recent progress in understanding how bone resorption occurs, the mechanisms regulating osteoclast secretion, and in particular the trafficking route of cathepsin K vesicles, remain elusive. Using a genetic approach, we describe the requirement for PKCδ in regulating bone resorption by affecting cathepsin K exocytosis. Importantly, PKCδ deficiency does not perturb formation of the ruffled border or trafficking of lysosomal vesicles containing the v-ATPase. Mechanistically, we find that cathepsin K exocytosis is controlled by PKCδ through modulation of the actin bundling protein MARCKS. The relevance of our finding is emphasized in vivo as PKCδ−/− mice exhibit increased bone mass and are protected from pathological bone loss in a model of experimental post-menopausal osteoporosis. Collectively, our data provide novel mechanistic insights into the pathways that selectively promote secretion of cathepsin K lysosomes independently of ruffled border formation, providing evidence for the presence of multiple mechanisms that regulate lysosomal exocytosis in osteoclasts.

Fibroblast growth factor (FGF) signaling is cardioprotective in various models of myocardial infarction. FGF receptors (FGFRs) are expressed in multiple cell types in the adult heart, but the cell type-specific FGFR signaling that mediates different cardioprotective endpoints is not known. To determine the requirement for FGFR signaling in endothelium in cardiac ischemia-reperfusion injury, we conditionally inactivated the Fgfr1 and Fgfr2 genes in endothelial cells with Tie2-Cre (Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice). Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice had normal baseline cardiac morphometry, function, and vessel density. When subjected to closed-chest, regional cardiac ischemia-reperfusion injury, Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice showed a significantly increased hypokinetic area at 7 days, but not 1 day, after reperfusion. Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice also showed significantly worsened cardiac function compared with controls at 7 days but not 1 day after reperfusion. Pathophysiological analysis showed significantly decreased vessel density, increased endothelial cell apoptosis, and worsened tissue hypoxia in the peri-infarct area at 7 days following reperfusion. Notably, Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice showed no impairment in the cardiac hypertrophic response. These data demonstrate an essential role for FGFR1 and FGFR2 in endothelial cells for cardiac functional recovery and vascular remodeling following in vivo cardiac ischemia-reperfusion injury, without affecting the cardiac hypertrophic response. This study suggests the potential for therapeutic benefit from activation of endothelial FGFR pathways following ischemic injury to the heart.

14-3-3 family members are intracellular dimeric phosphoserine-binding proteins that regulate signal transduction, cell cycle, apoptotic, and metabolic cascades. Previous work with global 14-3-3 protein inhibitors suggested that these proteins play a critical role in antagonizing apoptotic cell death in response to provocative stimuli. To determine the specific role of one family member in apoptosis, mice were generated with targeted disruption of the 14-3-3τ gene. 14-3-3τ−/− mice did not survive embryonic development, but haploinsufficient mice appeared normal at birth and were fertile. Cultured adult cardiomyocytes derived from 14-3-3τ+/− mice were sensitized to apoptosis in response to hydrogen peroxide or UV irradiation. 14-3-3τ+/− mice were intolerant of experimental myocardial infarction and developed pathological ventricular remodeling with increased cardiomyocyte apoptosis. ASK1, c-jun NH2-terminal kinase, and p38 mitogen-activated protein kinase (MAPK) activation was increased, but extracellular signal-regulated kinase MAPK activation was reduced, in 14-3-3τ+/− cardiac tissue. Inhibition of p38 MAPK increased survival in 14-3-3τ+/− mice subjected to myocardial infarction. These results demonstrate that 14-3-3τ plays a critical antiapoptotic function in cardiomyocytes and that therapeutic agents that increase 14-3-3τ activity may be beneficial to patients with myocardial infarction.

Background-Angioplasty and stent delivery are performed to treat atherosclerotic vascular disease but often cause deleterious neointimal lesion formation. Previously, growth factor receptor-bound protein 2 (Grb2), an intracellular linker protein, was shown to be essential for neointima formation and for p38 mitogen-activated protein kinase (MAPK) activation in vascular smooth muscle cells (SMCs). In this study, the role of vascular SMC p38␣ MAPK in neointimal development was examined. Methods and Results-Compound transgenic mice were generated with doxycycline-inducible SMC-specific expression of dominant-negative p38␣ MAPK (DN-p38␣). Doxycycline treatment resulted in the expression of DN-p38␣ mRNA and protein in transgenic arteries. Doxycycline-treated compound transgenic mice were resistant to neointima formation 21 days after carotid injury and showed reduced arterial p38 MAPK activation. To explore the mechanism by which p38␣ MAPK promotes neointima formation, an in vitro SMC culture system was used. Inhibition of p38␣ MAPK in cultured SMCs by treatment with SB202190 or small interfering RNA blocked platelet-derived growth factor-induced SMC proliferation, DNA replication, phosphorylation of the retinoblastoma protein, and induction of minichromosome maintenance protein 6. Conclusions-SMC p38␣ MAPK activation is required for neointima formation, perhaps because of its ability to promote retinoblastoma protein phosphorylation and minichromosome maintenance protein 6 expression. (Circulation. 2008; 118:658-666.)

Calcineurin inhibitors (CNI) are powerful immunomodulatory agents that produce marked renal dysfunction due in part to endothelin-1-mediated reductions in renal blood flow. Ligand-stimulated Gq protein signaling promotes the contraction of smooth muscle cells via phospholipase Cβ-mediated stimulation of cytosolic calcium release. RGS4 is a GTPase activating protein that promotes the deactivation of Gq and Gi family members. To investigate the role of G protein-mediated signaling in the pathogenesis of CNI-mediated renal injury, we used mice deficient for RGS4 (rgs4−/−). Compared to congenic wild type control animals, rgs4−/− mice were intolerant of the CNI, cyclosporine (CyA), rapidly developing fatal renal failure. Rgs4−/− mice exhibited markedly reduced renal blood flow after CyA treatment when compared to congenic wild type control mice as measured by magnetic resonance imaging (MRI). Hypoperfusion was reversed by coadministration of CyA with the endothelin antagonist, bosentan. The MAPK/ERK pathway was activated by cyclosporine administration and was inhibited by cotreatment with bosentan. These results show that endothelin-1-mediated Gq protein signaling plays a key role in the pathogenesis of vasoconstrictive renal injury and that RGS4 antagonizes the deleterious effects of excess endothelin receptor activation in the kidney.

Objective— Grb2 is a ubiquitously expressed linker protein that couples growth factor receptor activation to downstream mitogen-activated protein kinase (MAPK) cascades. Macrophage proliferation and uptake of modified lipoproteins are critical components of atherogenesis which require MAPK activation. However, the precise role of upstream signaling factors and the interrelationship of various MAPK cascades in the pathogenesis of atherosclerosis remains uncertain. Complete deletion of Grb2 in mice results in early embryonic lethality. However, Grb2 heterozygous mice appear normal at birth. To test the role of the Grb2 adapter protein in atherosclerotic lesion formation, we generated Grb2 +/− mice in the apoE −/− genetic background. Methods and Results— Grb2 +/− apoE −/− and apoE −/− mice exhibited similar body weight and serum lipid profiles. However, Grb2 +/− apoE −/− mice on a Western diet had reduced lesion formation compared with apoE −/− mice by aortic sinus and en face assays. Transplantation of apoE −/− mice with Grb2 +/− apoE −/− or apoE −/− bone marrow indicated that Grb2 haploinsufficiency in blood-borne cells confers resistance to Western diet–induced atherosclerosis. Cell culture experiments with bone marrow–derived macrophages showed that Grb2 is required for oxidized low density lipoprotein (oxLDL)-induced MAPK activation and foam cell formation. Conclusions— Grb2 is required for atherosclerotic lesion formation and uptake of oxidized LDL by macrophages.