MLP (muscle LIM protein)-deficient mice count among the first mouse models for dilated cardiomyopathy (DCM), yet the exact role of MLP in cardiac signalling processes is still enigmatic. Elevated PKCα signalling activity is known to be an important contributor to heart failure. Here we show that MLP directly inhibits the activity of PKCα. In end-stage DCM, PKCα is concentrated at the intercalated disc of cardiomyocytes, where it is sequestered by the adaptor protein CARP in a multiprotein complex together with PLCβ1. In mice deficient for both MLP and CARP the chronic PKCα signalling chain at the intercalated disc is broken and they remain healthy. Our results suggest that the main role of MLP in heart lies in the direct inhibition of PKCα and that chronic uninhibited PKCα activity at the intercalated disc in the absence of functional MLP leads to heart failure.

Patients with obstructive sleep apnea, who experience episodic hypoxia and hypercapnia during sleep, often demonstrate increased inflammation, oxidative stress, and dyslipidemia. We hypothesized that sleep apnea patients would be predisposed to the development of atherosclerosis. To dissect the mechanisms involved, we developed an animal model in mice whereby we expose mice to intermittent hypoxia/hypercapnia (IHH) in normobaric environments. Two- to three-month-old low-density lipoprotein receptor deficient (Ldlr(-/-)) mice were fed a high-fat diet for 8 or 16 wk while being exposed to IHH for either 10 h/day or 24 h/day. Plasma lipid levels, pulmonary artery and aortic atherosclerotic lesions, and cardiac function were then assayed. Surprisingly, atherosclerosis in the aorta of IHH mice was similar compared with controls. However, in IHH mice, atherosclerosis was markedly increased in the trunk and proximal branches of the pulmonary artery of exposed mice; even though plasma cholesterol and triglycerides were lower than in controls. Hemodynamic analysis revealed that right ventricular maximum pressure and isovolumic relaxation constant were significantly increased in IHH exposed mice and left ventricular % fractional shortening was reduced. In conclusion, 1) Intermittent hypoxia/hypercapnia remarkably accelerated atherosclerotic lesions in the pulmonary artery of Ldlr(-/-) mice and 2) increased lesion formation in the pulmonary artery was associated with right and left ventricular dysfunction. These findings raise the possibility that patients with obstructive sleep apnea may be susceptible to atherosclerotic disease in the pulmonary vasculature, an observation that has not been previously recognized.

Statins, which reduce LDL-cholesterol by inhibition of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, are among the most widely prescribed drugs. Skeletal myopathy is a known statin-induced adverse effect associated with mitochondrial changes. We hypothesized that similar effects would occur in cardiac myocytes in a lipophilicity-dependent manner between 2 common statins: atorvastatin (lipophilic) and pravastatin (hydrophilic). Neonatal cardiac ventricular myocytes were treated with atorvastatin and pravastatin for 48 h. Both statins induced endoplasmic reticular (ER) stress, but only atorvastatin inhibited ERK1/2, Akt, and mammalian target of rapamycin signaling; reduced protein abundance of caveolin-1, dystrophin, epidermal growth factor receptor, and insulin receptor-β; decreased Ras homolog gene family, member A activation; and induced apoptosis. In cardiomyocyte-equivalent HL-1 cells, atorvastatin, but not pravastatin, reduced mitochondrial oxygen consumption. When male mice underwent atorvastatin and pravastatin administration per os for up to 7 mo, only long-term atorvastatin, but not pravastatin, induced elevated serum creatine kinase; swollen, misaligned, size-variable, and disconnected cardiac mitochondria; alteration of ER structure; repression of mitochondria- and endoplasmic reticulum-related genes; and a 21% increase in mortality in cardiac-specific vinculin-knockout mice during the first 2 months of administration. To our knowledge, we are the first to demonstrate in vivo that long-term atorvastatin administration alters cardiac ultrastructure, a finding with important clinical implications.-Godoy, J. C., Niesman, I. R., Busija, A. R., Kassan, A., Schilling, J. M., Schwarz, A., Alvarez, E. A., Dalton, N. D., Drummond, J. C., Roth, D. M., Kararigas, G., Patel, H. H., Zemljic-Harpf, A. E. Atorvastatin, but not pravastatin, inhibits cardiac Akt/mTOR signaling and disturbs mitochondrial ultrastructure in cardiac myocytes.

Objetives: Assess the role of the catalase antioxidant enzyme in the vascular calcification process associated with chronic renal failure (CRF) and its effect on bone mass. Material and methods: Wild type C57/BL6J mice (WT) and transgenic mice (TG) were used, that overexpress the catalase enzyme, to which CRF was induced. Control WT and TG mice were used in simulated intervention. After 16 weeks, the mice were sacrificed, with serum samples taken for biochemical markers as well as residual pieces of kidney, aorta and tibias. An in vitro model of primary culture of smooth vascular muscle cells (SVMC) taken from the WT and TG aorta which underwent eight days of 3 mM phosphorus and 2 mM calcium calcifying medium. Results: A significant increase in Runx2 gene expression, calcium renal deposit and bone structure deterioration at trabecular level was only detected in WT mice with CRF. This was not observed in TG mice with CRF. Only in the case of WT mice SVMC, did added calcification medium raise calcium levels, proteic Runx2 expression and the reactive oxygen species of mitochondria with low catalase enzyme. Conclusions: Calcifying catalase over-expression was observed in both in vivo and in vitro, with in vivo showing that this reduction was accompanied by an improvement in bone parameters under study.

The small size of the mouse heart frequently imparts technical challenges when applying conventional in vivo imaging methods for assessing heart function. Here, we describe the use of high-frequency ultrasound imaging in conjunction with a size-tuned blood pool contrast agent for quantitatively assessing myocardial perfusion in living mice. A perflurocarbon microbubble formulation exhibiting a narrow size distribution was developed, and echogenicity was assessed at 18 MHz in vitro. Adult mice were subjected to permanent ligation of the left anterior descending artery. Ultrasound imaging was performed on day 7, and a cohort of intact mice was used as a control. Parasternal long-axis cine clips were acquired at 18 MHz before and after contrast administration. Reduced ejection fraction and increased end-systolic volume were observed in infarcted compared with control mice. In control animals, washin of the contrast agent was visible in all myocardial segments. Reduced contrast enhancement was observed in apical-posterolateral regions of all infarcted mice. A novel method for reslicing of the imaging data through the time domain provided a two-dimensional presentation of regional contrast agent washin, enabling convenient identification of locations exhibiting altered perfusion. Myocardial segments exhibiting diminished contractility were observed to have correspondingly low relative myocardial perfusion. The contrast agent formulation and methods demonstrated here provide the basis for simplifying routine in vivo estimation of infarct size in mice and may be particularly useful in longitudinal evaluation of revascularization interventions and assessment of peri-infarct ischemia. NEW & NOTEWORTHY Murine myocardial contrast echocardiography frequently suffers from poor sensitivity to contrast. Here, we formulated a novel size-tuned microbubble contrast agent and validated it for use with ultra-high-frequency ultrasound. A novel data method for evaluating myocardial perfusion based on reslicing the imaging data through the time domain is presented.