Multiparametric MRI identifies subtle adaptations for demarcation of disease transition in murine aortic valve stenosis.

Aortic valve stenosis (AS) is the most frequent valve disease with relevant prognostic impact. Experimental model systems for AS are scarce and comprehensive imaging techniques to simultaneously quantify function and morphology in disease progression are lacking. Therefore, we refined an acute murine AS model to closely mimic human disease characteristics and developed a high-resolution magnetic resonance imaging (MRI) approach for simultaneous in-depth analysis of valvular, myocardial as well as aortic morphology/pathophysiology to identify early changes in tissue texture and critical transition points in the adaptive process to AS. AS was induced by wire injury of the aortic valve. Four weeks after surgery, cine loops, velocity, and relaxometry maps were acquired at 9.4 T to monitor structural/functional alterations in valve, aorta, and left ventricle (LV). In vivo MRI data were subsequently validated by histology and compared to echocardiography. AS mice exhibited impaired valve opening accompanied by significant valve thickening due to fibrotic remodelling. While control mice showed bell-shaped flow profiles, AS resulted not only in higher peak flow velocities, but also in fragmented turbulent flow patterns associated with enhanced circumferential strain and an increase in wall thickness of the aortic root. AS mice presented with a mild hypertrophy but unaffected global LV function. Cardiac MR relaxometry revealed reduced values for both T1 and T2 in AS reflecting subtle myocardial tissue remodelling with early alterations in mitochondrial function in response to the enhanced afterload. Concomitantly, incipient impairments of coronary flow reserve and myocardial tissue integrity get apparent accompanied by early troponin release. With this, we identified a premature transition point with still compensated cardiac function but beginning textural changes. This will allow interventional studies to explore early disease pathophysiology and novel therapeutic targets.

Cardiac magnetic resonance T2 mapping and feature tracking in athlete's heart and HCM.

OBJECTIVES: Distinguishing hypertrophic cardiomyopathy (HCM) from left ventricular hypertrophy (LVH) due to systematic training (athlete's heart, AH) from morphologic assessment remains challenging. The purpose of this study was to examine the role of T2 mapping and deformation imaging obtained by cardiovascular magnetic resonance (CMR) to discriminate AH from HCM with (HOCM) or without outflow tract obstruction (HNCM). METHODS: Thirty-three patients with HOCM, 9 with HNCM, 13 strength-trained athletes as well as individual age- and gender-matched controls received CMR. For T2 mapping, GRASE-derived multi-echo images were obtained and analyzed using dedicated software. Besides T2 mapping analyses, left ventricular (LV) dimensional and functional parameters were obtained including LV mass per body surface area (LVMi), interventricular septum thickness (IVS), and global longitudinal strain (GLS). RESULTS: While LVMi was not significantly different, IVS was thickened in HOCM patients compared to athlete's. Absolute values of GLS were significantly increased in patients with HOCM/HNCM compared to AH. Median T2 values were elevated compared to controls except in athlete's heart. ROC analysis revealed T2 values (AUC 0.78) and GLS (AUC 0.91) as good parameters to discriminate AH from overall HNCM/HOCM. CONCLUSION: Discrimination of pathologic from non-pathologic LVH has implications for risk assessment of competitive sports in athletes. Multiparametric CMR with parametric T2 mapping and deformation imaging may add information to distinguish AH from LVH due to HCM. KEY POINTS: • Structural analyses using T2 mapping cardiovascular magnetic resonance imaging (CMR) may help to further distinguish myocardial diseases. • To differentiate pathologic from non-pathologic left ventricular hypertrophy, CMR including T2 mapping was obtained in patients with hypertrophic obstructive/non-obstructive cardiomyopathy (HOCM/HNCM) as well as in strength-trained athletes. • Elevated median T2 values in HOCM/HNCM compared with athlete's may add information to distinguish athlete's heart from pathologic left ventricular hypertrophy.

CD73 on T Cells Orchestrates Cardiac Wound Healing After Myocardial Infarction by Purinergic Metabolic Reprogramming.

BACKGROUND: T cells are required for proper healing after myocardial infarction. The mechanism of their beneficial action, however, is unknown. The proinflammatory danger signal ATP, released from damaged cells, is degraded by the ectonucleotidases CD39 and CD73 to the anti-inflammatory mediator adenosine. Here, we investigate the contribution of CD73-derived adenosine produced by T cells to cardiac remodeling after ischemia/reperfusion and define its mechanism of action. METHODS: Myocardial ischemia (50 minutes followed by reperfusion) was induced in global CD73-/- and CD4-CD73-/- mice. Tissue injury, T-cell purinergic signaling, cytokines, and cardiac function (magnetic resonance tomography at 9.4 T over 4 weeks) were analyzed. RESULTS: Changes in functional parameters of CD4-CD73-/- mice were identical to those in global CD73 knockouts (KOs). T cells infiltrating the injured heart significantly upregulated at the gene (quantitative polymerase chain reaction) and protein (enzymatic activity) levels critical transporters and enzymes (connexin43, connexin37, pannexin-1, equilibrative nucleoside transporter 1, CD39, CD73, ecto-nucleotide pyrophosphatase/phosphodiesterases 1 and 3, CD157, CD38) for the accelerated release and hydrolysis of ATP, cAMP, AMP, and NAD to adenosine. It is surprising that a lack of CD39 on T cells (from CD39-/- mice) did not alter ATP hydrolysis and very likely involves pyrophosphatases (ecto-nucleotide pyrophosphatase/phosphodiesterases 1 and 3). Circulating T cells predominantly expressed A2a receptor (A2aR) transcripts. After myocardial infarction, A2b receptor (A2bR) transcription was induced in both T cells and myeloid cells in the heart. Thus, A2aR and A2bR signaling may contribute to myocardial responses after myocardial infarction. In the case of T cells, this was associated with an accelerated secretion of proinflammatory and profibrotic cytokines (interleukin-2, interferon-γ, and interleukin-17) when CD73 was lacking. Cytokine production by T cells from peripheral lymph nodes was inhibited by A2aR activation (CGS-21680). The A2bR agonist BAY 60-6583 showed off-target effects. The adenosine receptor agonist NECA inhibited interferon-γ and stimulated interleukin-6 production, each of which was antagonized by a specific A2bR antagonist (PSB-603). CONCLUSIONS: This work demonstrates that CD73 on T cells plays a crucial role in the cardiac wound healing process after myocardial infarction. The underlying mechanism involves a profound increase in the hydrolysis of ATP/NAD and AMP, resulting primarily from the upregulation of pyrophosphatases and CD73. We also define A2bR/A2aR-mediated autacoid feedback inhibition of proinflammatory/profibrotic cytokines by T cell-derived CD73.

Multimodal assessment of orbital immune cell infiltration and tissue remodeling during development of graves disease by 1 H19 F MRI.

PURPOSE: To evaluate key molecular and cellular features of Graves orbitopathy (GO) by simultaneous monitoring of alterations in morphology, inflammatory patterns, and tissue remodeling. METHODS: To this end, we utilized a murine model of GO induced by immunization with a human thyroid-stimulating hormone receptor A-subunit plasmid. Altogether, 52 mice were used: 27 GOs and 25 controls (Ctrl) immunized with ß-galactasidose plasmid. From these, 17 GO and 12 Ctrl mice were subjected to multimodal MRI at 9.4T, whereas 23 mice only underwent histology. Beyond anatomical hydrogen-1 (1 H) MRI, we employed transverse relaxation time (T2 ) mapping for visualization of edema, chemical exchange saturation transfer (CEST) for detection of hyaluronan, and fluorine-19 (19 F) MRI for tracking of in situ-labeled immune cells after intravenous injection of perfluorcarbons (PFCs). RESULTS: 1 H/19 F MRI demonstrated substantial infiltration of PFC-loaded immune cells in peri and retro-orbital regions of GO mice, whereas healthy Ctrls showed only minor 19 F signals. In parallel, T2 mapping indicated onset of edema in periorbital tissue and adjacent ocular glands (P = 0.038/0.017), which were associated with enhanced orbital CEST signals in GO mice (P = 0.031). Concomitantly, a moderate expansion of retrobulbar fat (P = 0.029) was apparent; however, no signs for extraocular myopathy were detectable. 19 F MRI-based visualization of orbital inflammation exhibited the highest significance level to discriminate between GO and Ctrl mice (P = 0.006) and showed the best correlation with the clinical score (P = 0.0007). CONCLUSION: The present approach permits the comprehensive characterization of orbital tissue and holds the potential for accurate GO diagnosis in the clinical setting. Magn Reson Med 80:711-718, 2018. © 2018 International Society for Magnetic Resonance in Medicine.

Abnormal T2 mapping cardiovascular magnetic resonance correlates with adverse clinical outcome in patients with suspected acute myocarditis.

BACKGROUND: While most patients recover from suspected acute myocarditis (sAMC) some develop progressive disease with 5-year mortality up to 20%. Recently, parametric Cardiovascular Magnetic Resonance (CMR) approaches, quantifying native T1 and T2 relaxation time, have demonstrated the ability to increase diagnostic accuracy. However, prognostic implications of T2 values in this cohort are unknown. The purpose of the study was to investigate the prognostic relevance of elevated CMR T2 values in patients with sAMC. METHODS AND RESULTS: We carried out a prospective study in 46 patients with sAMC defined by current ESC recommendations. A combined endpoint was defined by the occurrence of at least one major adverse cardiac event (MACE) and hospitalisation for heart failure. Event rate was 24% (n = 11) for 1-year-MACE and hospitalisation. A follow-up after 11 ± 7 months was performed in 98% of the patients. Global T2 values were significantly increased at acute stage of disease compared to controls and decreased over time. During acute disease, elevated global T2 time (odds ratio 6.3, p < 0.02) as well as myocardial fraction with T2 time >80 ms (odds ratio 4.9, p < 0.04) predicted occurrence of the combined endpoint. Patients with clinical recovery revealed significantly decreased T2 relaxation times at follow-up examinations; however, T2 values were still elevated compared to healthy controls. CONCLUSION: Assessment of myocardial T2 relaxation times at initial presentation facilitates CMR-based risk stratification in patients with acute myocarditis. T2 Mapping may emerge as a new tool to monitor inflammatory myocardial injuries during the course of disease.

Multiparametric cardiac magnetic resonance imaging (CMR) for the diagnosis of Loeffler's endocarditis: a case report.

BACKGROUND: Endocarditis parietalis fibroplastica Löfflein (EPF) is a rare form of primary restrictive cardiomyopathy with poor prognosis. It is generally caused by hypereosinophilic syndrome with eosinophilic penetration of the heart. This leads to congestive heart failure in three different stages. As a frequent manifestation of neoplastic diseases, cardiac involvement means poor prognosis. CASE PRESENTATION: The present report deals with a case of EPF caused by non-specified T-cell lymphoma (T-NOS). Besides an elevated Troponin-T enzyme, the electrocardiogram and the transthoracic echocardiography did not show any characteristic results. Due to risk/benefit assessment and low thrombocyte amounts, endomyocardial biopsy and catheterization were discarded. Using cardiovascular magnetic resonance (CMR) with steady-state free precession sequences, T2-mappping, strain analysis and late gadolinium enhancement, we were able to clearly highlight cardiac involvement at different stages. These findings characterized T-NOS as a palliative situation. CONCLUSION: Multiparametric CMR can not only identify EPF but also characterize the patchy disease state. This provides an individual prognosis assessment. Aside from prognosis estimation, it can also be used for therapy monitoring.

Loss of UCP2 attenuates mitochondrial dysfunction without altering ROS production and uncoupling activity.

Although mitochondrial dysfunction is often accompanied by excessive reactive oxygen species (ROS) production, we previously showed that an increase in random somatic mtDNA mutations does not result in increased oxidative stress. Normal levels of ROS and oxidative stress could also be a result of an active compensatory mechanism such as a mild increase in proton leak. Uncoupling protein 2 (UCP2) was proposed to play such a role in many physiological situations. However, we show that upregulation of UCP2 in mtDNA mutator mice is not associated with altered proton leak kinetics or ROS production, challenging the current view on the role of UCP2 in energy metabolism. Instead, our results argue that high UCP2 levels allow better utilization of fatty acid oxidation resulting in a beneficial effect on mitochondrial function in heart, postponing systemic lactic acidosis and resulting in longer lifespan in these mice. This study proposes a novel mechanism for an adaptive response to mitochondrial cardiomyopathy that links changes in metabolism to amelioration of respiratory chain deficiency and longer lifespan.

Noninvasive Imaging of Early Venous Thrombosis by 19F Magnetic Resonance Imaging With Targeted Perfluorocarbon Nanoemulsions.

BACKGROUND: Noninvasive detection of deep venous thrombi and subsequent pulmonary thromboembolism is a serious medical challenge, since both incidences are difficult to identify by conventional ultrasound techniques. METHODS AND RESULTS: Here, we report a novel technique for the sensitive and specific identification of developing thrombi using background-free 19F magnetic resonance imaging, together with α2-antiplasmin peptide (α2AP)-targeted perfluorocarbon nanoemulsions (PFCs) as contrast agent, which is cross-linked to fibrin by active factor XIII. Ligand functionality was ensured by mild coupling conditions using the sterol-based postinsertion technique. Developing thrombi with a diameter<0.8 mm could be visualized unequivocally in the murine inferior vena cava as hot spots in vivo by simultaneous acquisition of anatomic matching 1H and 19F magnetic resonance images at 9.4 T with both excellent signal-to-noise and contrast-to-noise ratios (71±22 and 17±5, respectively). Furthermore, α2AP-PFCs could be successfully applied for the diagnosis of experimentally induced pulmonary thromboembolism. In line with the reported half-life of factor XIIIa, application of α2AP-PFCs>60 minutes after thrombus induction no longer resulted in detectable 19F magnetic resonance imaging signals. Corresponding results were obtained in ex vivo generated human clots. Thus, α2AP-PFCs can visualize freshly developed thrombi that might still be susceptible to pharmacological intervention. CONCLUSIONS: Our results demonstrate that 1H/19F magnetic resonance imaging, together with α2AP-PFCs, is a sensitive, noninvasive technique for the diagnosis of acute deep venous thrombi and pulmonary thromboemboli. Furthermore, ligand coupling by the sterol-based postinsertion technique represents a unique platform for the specific targeting of PFCs for in vivo 19F magnetic resonance imaging.

Ecto-5'-nucleotidase on immune cells protects from adverse cardiac remodeling.

RATIONALE: Ecto-5'-nucleotidase (CD73) on immune cells is emerging as a critical pathway and therapeutic target in cardiovascular and autoimmune disorders. OBJECTIVE: Here, we investigated the role of CD73 in postinfarction inflammation, cardiac repair, and remodeling in mice after reperfused myocardial infarction (50-minute ischemia). METHODS AND RESULTS: We found that compared with control mice (1) cardiac function in CD73(-/-) mice more severely declined after infarction (systolic failure with enhanced myocardial edema formation) as determined by MRI and was associated with the persistence of cardiac immune cell subsets, (2) cardiac adenosine release was augmented 7 days after ischemia/reperfusion in control mice but reduced by 90% in CD73 mutants, (3) impaired healing involves M1-driven immune response with increased tumor necrosis factor-α and interleukin-17, as well as decreased transforming growth factor-ß and interleukin-10, and (4) CD73(-/-) mice displayed infarct expansion accompanied by an immature replacement scar and diffuse ventricular fibrosis. Studies on mice after bone marrow transplantation revealed that CD73 present on immune cells is a major determinant promoting cardiac healing. CONCLUSIONS: These results, together with the upregulation of CD73 on immune cells after ischemia/reperfusion, demonstrate the crucial role of purinergic signaling during cardiac healing and provide groundwork for novel anti-inflammatory strategies in treating adverse cardiac remodeling.

Myocardial T2 mapping reveals age- and sex-related differences in volunteers.

BACKGROUND: T2 mapping indicates to be a sensitive method for detection of tissue oedema hidden beyond the detection limits of T2-weighted Cardiovascular Magnetic Resonance (CMR). However, due to variability of baseline T2 values in volunteers, reference values need to be defined. Therefore, the aim of the study was to investigate the effects of age and sex on quantitative T2 mapping with a turbo gradient-spin-echo (GRASE) sequence at 1.5 T. For that reason, we studied sensitivity issues as well as technical and biological effects on GRASE-derived myocardial T2 maps. Furthermore, intra- and interobserver variability were calculated using data from a large volunteer group. METHODS: GRASE-derived multiecho images were analysed using dedicated software. After sequence optimization, validation and sensitivity measurements were performed in muscle phantoms ex vivo and in vivo. The optimized parameters were used to analyse CMR images of 74 volunteers of mixed sex and a wide range of age with typical prevalence of hypertension and diabetes. Myocardial T2 values were analysed globally and according to the 17 segment model. Strain-encoded (SENC) imaging was additionally performed to investigate possible effects of myocardial strain on global or segmental T2 values. RESULTS: Ex vivo studies in muscle phantoms showed, that GRASE-derived T2 values were comparable to those acquired by a standard multiecho spinecho sequence but faster by a factor of 6. Besides that, T2 values reflected tissue water content. The in vivo measurements in volunteers revealed intra- and interobserver correlations with R2=0.91 and R2=0.94 as well as a coefficients of variation of 2.4% and 2.2%, respectively. While global T2 time significantly decreased towards the heart basis, female volunteers had significant higher T2 time irrespective of myocardial region. We found no correlation of segmental T2 values with maximal systolic, diastolic strain or heart rate. Interestingly, volunteers´ age was significantly correlated to T2 time while that was not the case for other coincident cardiovascular risk factors. CONCLUSION: GRASE-derived T2 maps are highly reproducible. However, female sex and aging with typical prevalence of hypertension and diabetes were accompanied by increased myocardial T2 values. Thus, sex and age must be considered as influence factors when using GRASE in a diagnostic manner.

Multifunctional MR monitoring of the healing process after myocardial infarction.

Healing of the myocardium after infarction comprises a variety of local adaptive processes which contribute to the functional outcome after the insult. Therefore, we aimed to establish a setting for concomitant assessment of regional alterations in contractile function, morphology, and immunological state to gain prognostic information on cardiac recovery after infarction. For this, mice were subjected to myocardial ischemia/reperfusion (I/R) and monitored for 28 days by cine MRI, T2 mapping, late gadolinium enhancement (LGE), and (19)F MRI. T2 values were calculated from gated multi-echo sequences. (19)F-loaded nanoparticles were injected intravenously for labelling circulating monocytes and making them detectable by (19)F MRI. In-house developed software was used for regional analysis of cine loops, T2 maps, LGE, and (19)F images to correlate local wall movement, tissue damage as well as monocyte recruitment over up to 200 sectors covering the left ventricle. This enabled us to evaluate simultaneously zonal cardiac necrosis, oedema, and inflammation patterns together with sectional fractional shortening (FS) and global myocardial function. Oedema, indicated by a rise in T2, showed a slightly better correlation with FS than LGE. Regional T2 values increased from 19 ms to above 30 ms after I/R. In the course of the healing process oedema resolved within 28 days, while myocardial function recovered. Infiltrating monocytes could be quantitatively tracked by (19)F MRI, as validated by flow cytometry. Furthermore, (19)F MRI proved to yield valuable insight on the outcome of myocardial infarction in a transgenic mouse model. In conclusion, our approach permits a comprehensive surveillance of key processes involved in myocardial healing providing independent and complementary information for individual prognosis.

Probing different perfluorocarbons for in vivo inflammation imaging by 19F MRI: image reconstruction, biological half-lives and sensitivity.

Inflammatory processes can reliably be assessed by (19)F MRI using perfluorocarbons (PFCs), which is primarily based on the efficient uptake of emulsified PFCs by circulating cells of the monocyte-macrophage system and subsequent infiltration of the (19)F-labeled cells into affected tissue. An ideal candidate for the sensitive detection of fluorine-loaded cells is the biochemically inert perfluoro-15-crown-5 ether (PFCE), as it contains 20 magnetically equivalent (19)F atoms. However, the biological half-life of PFCE in the liver and spleen is extremely long, and so this substance is not suitable for future clinical applications. In the present study, we investigated alternative, nontoxic PFCs with predicted short biological half-lives and high fluorine content: perfluorooctyl bromide (PFOB), perfluorodecalin (PFD) and trans-bis-perfluorobutyl ethylene (F-44E). Despite the complex spectra of these compounds, we obtained artifact-free images using sine-squared acquisition-weighted three-dimensional chemical shift imaging and dedicated reconstruction accomplished with in-house-developed software. The signal-to-noise ratio of the images was maximized using a Nutall window with only moderate localization error. Using this approach, the retention times of the different PFCs in murine liver and spleen were determined at 9.4 T. The biological half-lives were estimated to be 9 days (PFD), 12 days (PFOB) and 28 days (F-44E), compared with more than 250 days for PFCE. In vivo sensitivity for inflammation imaging was assessed using an ear clip injury model. The alternative PFCs PFOB and F-44E provided 37% and 43%, respectively, of the PFCE intensities, whereas PFD did not show any signal in the ear model. Thus, for in vivo monitoring of inflammatory processes, PFOB emerges as the most promising candidate for possible future translation of (19)F MR inflammation imaging to human applications.

Growth and metastasis of B16-F10 melanoma cells is not critically dependent on host CD73 expression in mice.

BACKGROUND: Recent studies have suggested that adenosine generated by ecto-5'-nucleotidase (CD73) in the tumor microenvironment plays a major role in promoting tumor growth by suppressing the immune response and stimulating angiogenesis via A2A and A2B receptors. However, adenosine has also been reported to inhibit tumor growth acting via A1 and A3 receptors. Therefore the aim of this study was to clarify the role of host CD73, which catalyzes the extracellular hydrolysis of AMP to adenosine, on tumor growth and metastasis of B16-F10 melanoma cells. METHODS: CD73 and alkaline phosphatase (AP) activity of B16-F10 melanoma cells were measured by HPLC. Tumor cells were injected either subcutaneously or intradermally in WT and CD73-/- mice and tumor growth was monitored by MRI at 9.4 T. Immune cell subpopulations within tumors were assessed by FACS after enzymatic digestion. An endothelium specific CD73-/- was created using Tie2-Cre+ mice and CD73flox/flox (loxP) mice. Chimeric mice lacking CD73-/- on hematopoietic cells was generated by bone marrow transplantation. Lung metastatic spread was measured after intravenous B16-F10 application. RESULTS: B16-F10 cells showed very little CD73 and negligible AP activity. Neither complete loss of host CD73 nor specific knockout of CD73 on endothelial cells or hematopoietic cells affected tumor growth after subcutaneous or intradermal tumor cell application. Only peritumoral edema formation was significantly attenuated in global CD73-/- mice in the intradermal model. Immune cell composition revealed no differences in the different transgenic mice models. Also lung metastasis after intravenous B16-F10 injection was not altered in CD73-/- mice. CONCLUSIONS: CD73 expression on host cells, particularly on endothelial and hematopoietic cells, does not modulate tumor growth and metastatic spread of B16-F10 melanoma cells most likely because of insufficient adenosine formation by the tumor itself.

Visualization of immune cell infiltration in experimental viral myocarditis by (19)F MRI in vivo.

OBJECTIVE: This paper introduces a new approach permitting for the first time a specific, non-invasive diagnosis of myocarditis by visualizing the infiltration of immune cells into the myocardium. MATERIALS AND METHODS: The feasibility of this approach is shown in a murine model of viral myocarditis. Our study uses biochemically inert perfluorocarbons (PFCs) known to be taken up by circulating monocytes/macrophages after intravenous injection. RESULTS: In vivo (19)F MRI at 9.4 T demonstrated that PFC-loaded immune cells infiltrate into inflamed myocardial areas. Because of the lack of any fluorine background in the body, detected (19)F signals of PFCs are highly specific as confirmed ex vivo by flow cytometry and histology. CONCLUSION: Since PFCs are a family of compounds previously used clinically as blood substitutes, the technique described in our paper holds the potential as a new imaging modality for the diagnosis of myocarditis in man.

Data on common carotid artery occlusion inducing focalized stroke lesions after Pertussis toxin injection.

This article contains raw and processed data related to research published by Vega et al. (2022). This complementary dataset provides further insight into the experimental validation of a single common carotid artery occlusion (CCAO) model upon pretreatment with pertussis toxin (PTX). We present data showing the extent of different PTX concentrations on neurological severity measured by Bederson score following CCAO. In addition, data indicate a protective effect of isoflurane on cerebral infarction and neurological deficits, as well as the consequences of PTX pretreatment on reperfusion after occlusion using time-of-flight magnetic resonance angiography. With these data, we aim to provide detailed experimental settings of this newly described model.

IGF-IR signaling attenuates the age-related decline of diastolic cardiac function.

Insulin-like growth factor (IGF-I) signaling has been implicated to play an important role in regulation of cardiac growth, hypertrophy, and contractile function and has been linked to the development of age-related congestive heart failure. Here, we address the question to what extent cardiomyocyte-specific IGF-I signaling is essential for maintenance of the structural and functional integrity of the adult murine heart. To investigate the effects of IGF-I signaling in the adult heart without confounding effects due to IGF-I overexpression or adaptation during embryonic and early postnatal development, we inactivated the IGF-I receptor (IGF-IR) by a 4-hydroxytamoxifen-inducible Cre recombinase in adult cardiac myocytes. Efficient inactivation of the IGF-IR (iCMIGF-IRKO) as assessed by Western analysis and real-time PCR went along with reduced IGF-I-dependent Akt and GSK3ß phosphorylation. Functional analysis by conductance manometry and MRI revealed no functional alterations in young adult iCMIGF-IRKO mice (age 3 mo). However, when induced in aging mice (11 mo) diastolic cardiac function was depressed. To address the question whether insulin signaling might compensate for the defective IGF-IR signaling, we inactivated ß-cells by streptozotocin. However, the diabetes-associated functional depression was similar in control and iCMIGF-IRKO mice. Similarly, analysis of the cardiac gene expression profile on 44K microarrays did not reveal activation of overt adaptive processes. Endogenous IGF-IR signaling is required for conservation of cardiac function of the aging heart, but not for the integrity of cardiac structure and function of young hearts.

Development of a growing rat model for the in vivo assessment of engineered aortic conduits.

BACKGROUND: Numerous limitations of aortic valve grafts currently used in pediatric patients cause the need for alternative prostheses. For the purpose of in vivo evaluation of novel engineered aortic conduit grafts, we aimed at downsizing a previously described model to create a growing rodent model. MATERIALS AND METHODS: U-shaped aortic conduits were sutured to the infrarenal aorta of young Wistar rats (70-80 g, n = 10) in an end-to-side manner. Functional assessment was performed by Doppler sonography and high resolution rodent MRI. Histology and immunohistochemistry followed after 8 wk. RESULTS: Postoperative recovery rate was 80%. Conforming to clinical observations, postoperative MRI (d 5) and Doppler sonography (wk 8) revealed unimpaired conduit perfusion. Explanted implants were luminally completely covered by an endothelial cell layer with local hyperplasia and accumulation of α-smooth muscle actin (+) cells. Moreover microcalcification of the decellularized scaffolds was observed. CONCLUSIONS: Our downsized model of aortic conduit transplantation enables overall characterization with detailed analysis of maturation of engineered aortic grafts in a growing organism.

Methods employed for induction and analysis of experimental myocardial infarction in mice.

Myocardial ischemia und subsequent reperfusion is followed by a complex sequence of pathophysiological responses involving inflammatory cell infiltration and cytokine release as well as postinfarction wound healing and myocardial tissue remodeling. With the development of gene targeted mice the contribution of individual gene products to the pathophysiology of myocardial ischemia and reperfusion can be defined leading to an increasing interest in the widely-used mouse model of myocardial infarction. This methological paper describes in detail the required equipment, surgical instruments, drugs and additional material, the methods of anesthesia and analgesia, the procedures involved in preparation of the animal, tracheotomy, intubation, thoracotomy, occlusion of the left descending artery, removal of the heart, determination of infarct size, analysis of cardiac functional parameters with echocardiography and magnetic resonance imaging (MRI) as well as determination of the morphological consequences utilizing gelatin zymography, histology and immunohistochemistry.

Left ventricular dilation in toll-like receptor 2 deficient mice after myocardial ischemia/reperfusion through defective scar formation.

Restoration of myocardial blood flow after ischemia triggers an inflammatory response involving toll-like receptors (TLRs). TLR2(-/-)-mice show short-term advantages upon reperfusion injury as compared with WT controls. Accordingly, it has been shown that transient TLR2-blockade prior to reperfusion is associated with improved left-ventricular performance after myocardial scar formation. We present here adverse myocardial remodeling due to a chronic lack of TLR2 expression. Myocardial ischemia/reperfusion (MI/R) was surgically induced in C3HeN-mice by ligation of the left anterior descending coronary artery for 20 min, followed by 24 h or 28 days of reperfusion. TLR2(-/-)-mice and TLR2-Ab treated (T2.5) WT-mice displayed a reduction of infarct size, plasma troponin T concentrations, and leukocyte infiltration as compared with untreated controls after 24 h of reperfusion. After 28 days, however, magnetic resonance imaging revealed a marked left ventricular dilation in TLR2(-/-)-animals, which was associated with pronounced matrix remodeling characterized by reduced collagen and decorin density in the infarct scar. Our data show adverse effects on myocardial remodeling in TLR2(-/-)-mice. Although interception with TLR2 signaling is a promising concept for the prevention of reperfusion injury after myocardial ischemia, these data give cause for serious concern with respect to the time-point and duration of the potential treatment.

Multi-targeted 1H/19F MRI unmasks specific danger patterns for emerging cardiovascular disorders.

Prediction of the transition from stable to acute coronary syndromes driven by vascular inflammation, thrombosis with subsequent microembolization, and vessel occlusion leading to irreversible myocardial damage is still an unsolved problem. Here, we introduce a multi-targeted and multi-color nanotracer platform technology that simultaneously visualizes evolving danger patterns in the development of progressive coronary inflammation and atherothrombosis prior to spontaneous myocardial infarction in mice. Individual ligand-equipped perfluorocarbon nanoemulsions are used as targeting agents and are differentiated by their specific spectral signatures via implementation of multi chemical shift selective 19F MRI. Thereby, we are able to identify areas at high risk of and predictive for consecutive development of myocardial infarction, at a time when no conventional parameter indicates any imminent danger. The principle of this multi-targeted approach can easily be adapted to monitor also a variety of other disease entities and constitutes a technology with disease-predictive potential.