Using averaged models from 4D ultrasound strain imaging allows to significantly differentiate local wall strains in calcified regions of abdominal aortic aneurysms

Abdominal aortic aneurysms are a degenerative disease of the aorta associated with high mortality. To date, in vivo information to characterize the individual elastic properties of the aneurysm wall in terms of rupture risk is lacking. We have used time-resolved 3D ultrasound strain imaging to calculate spatially resolved in-plane strain distributions characterized by mean and local maximum strains, as well as indices of local variations in strains. Likewise, we here present a method to generate averaged models from multiple segmentations. Strains were then calculated for single segmentations and averaged models. After registration with aneurysm geometries based on CT-A imaging, local strains were divided into two groups with and without calcifications and compared. Geometry comparison from both imaging modalities showed good agreement with a root mean squared error of 1.22 ± 0.15 mm and Hausdorff Distance of 5.45 ± 1.56 mm (mean ± sd, respectively). Using averaged models, circumferential strains in areas with calcifications were 23.2 ± 11.7% (mean ± sd) smaller and significantly distinguishable at the 5% level from areas without calcifications. For single segmentations, this was possible only in 50% of cases. The areas without calcifications showed greater heterogeneity, larger maximum strains, and smaller strain ratios when computed by use of the averaged models. Using these averaged models, reliable conclusions can be made about the local elastic properties of individual aneurysm (and long-term observations of their change), rather than just group comparisons. This is an important prerequisite for clinical application and provides qualitatively new information about the change of an abdominal aortic aneurysm in the course of disease progression compared to the diameter criterion.

     

Escherichia coli LPS-induced LV dysfunction: role of toll-like receptor-4 in the adult heart

The precise molecular mechanisms responsible for sepsis-induced myocardial dysfunction remain undefined. Toll-like receptor-4 (TLR-4) engages lipopolysaccharide (LPS) and activates signaling pathways leading to the expression of proinflammatory cytokines implicated in myocardial dysfunction. We determined whether TLR-4 was necessary for LPS-induced myocardial dysfunction in vivo. The effects of LPS on left ventricular (LV) function were studied in mice with defective TLR-4 signaling (C3H/HeJ, TLR-4 deficient) and wild-type mice (C3HeB/FeJ). Mice (n = 5/group) were injected with LPS or diluent, and LV function was examined by using two-dimensional echocardiography and conductance catheters. LPS significantly decreased all indexes of LV function in wild-type mice when compared with controls; LV function was not depressed in the LPS-treated TLR-4-deficient mice relative to controls. LPS increased myocardial nitric oxide synthase-2 expression and cGMP only in wild-type mice. This study suggests that TLR-4 mediates the LV dysfunction that occurs in LPS-induced shock. Therefore, TLR-4 might be a therapeutic target for attenuating the effects of LPS on the heart.     

Evaluation of the antimicrobial activity of ebselen: role of the yeast plasma membrane H+-ATPase

Ebselen (2-phenyl-1,2-benzisoselenazol-3(2H)-one) is a selenium-containing antioxidant demonstrating anti-inflammatory and cytoprotective properties in mammalian cells and cytotoxicity in lower organisms. The mechanism underlying the antimicrobial activity of ebselen remains unclear. It has recently been proposed that, in lower organisms like yeast, the plasma membrane H+-ATPase (Pma1p) could serve as a potential target for this synthetic organoselenium compound. Using yeast and bacteria, the present study found ebselen to inhibit microbial growth in a concentration- and time-dependent manner, and yeast and Gram-positive bacteria to be more sensitive to this action (IC50 approximately 2-5 microM) than Gram-negative bacteria (IC50 < 80 microM). Washout experiments and scanning electron microscopic analysis revealed ebselen to possess fungicidal activity. In addition, ebselen was found to inhibit medium acidification by PMA1-proficient haploid yeast in a concentration-dependent manner. Additional studies comparing PMA1 (+/-) and PMA1 (+/+) diploid yeast cells revealed the mutant to be more sensitive to treatment with ebselen than the wild type. Ebselen also inhibited the ATPase activity of Pma1p from S. cerevisiae in a concentration-dependent manner. The interaction of ebselen with the sulfhydryl-containing compounds L-cysteine and reduced glutathione resulted in the complete and partial prevention, respectively, of the inhibition of Pma1p ATPase activity by ebselen. Taken together, these results suggest that the fungicidal action of ebselen is due, at least in part, to interference with both the proton-translocating function and the ATPase activity of the plasma membrane H+-ATPase.     

Morphine-Induced Preconditioning: Involvement of Protein Kinase A and Mitochondrial Permeability Transition Pore

Background

Morphine induces myocardial preconditioning (M-PC) via activation of mitochondrial large conductance Ca²⁺-sensitive potassium (mK_Ca) channels. An upstream regulator of mK_Ca channels is protein kinase A (PKA). Furthermore, mK_Ca channel activation regulates mitochondrial bioenergetics and thereby prevents opening of the mitochondrial permeability transition pore (mPTP). Here, we investigated in the rat heart in vivo whether 1) M-PC is mediated by activation of PKA, and 2) pharmacological opening of the mPTP abolishes the cardioprotective effect of M-PC and 3) M-PC is critically dependent on STAT3 activation, which is located upstream of mPTP within the signalling pathway.

Methods

Male Wistar rats were randomised to six groups (each n = 6). All animals underwent 25 minutes of regional myocardial ischemia and 120 minutes of reperfusion. Control animals (Con) were not further treated. Morphine preconditioning was initiated by intravenous administration of 0.3 mg/kg morphine (M-PC). The PKA blocker H-89 (10 μg/kg) was investigated with and without morphine (H-89+M-PC, H-89). We determined the effect of mPTP opening with atractyloside (5 mg/kg) with and without morphine (Atr+M-PC, Atr). Furthermore, the effect of morphine on PKA activity was tested in isolated adult rat cardiomyocytes. In further experiments in isolated hearts we tested the protective properties of morphine in the presence of STAT3 inhibition, and whether pharmacological prevention of the mPTP-opening by cyclosporine A (CsA) is cardioprotective in the presence of STAT3 inhibition.

Results

Morphine reduced infarct size from 64±5% to 39±9% (P<0.05 vs. Con). H-89 completely blocked preconditioning by morphine (64±9%; P<0.05 vs. M-PC), but H-89 itself had not effect on infarct size (61±10%; P>0.05 vs. Con). Also, atractyloside abolished infarct size reduction of morphine completely (65±9%; P<0.05 vs. M-PC) but had no influence on infarct size itself (64±5%; P>0.05 vs. Con). In isolated hearts STAT3 inhibitor Stattic completely abolished morphine-induced preconditioning. Administration of Stattic and mPTP inhibitor cyclosporine A reduced infarct size to 31±6% (Stat+CsA, P<0.05 vs. Con). Cyclosporine A alone reduced infarct size to 26±7% (CsA P<0.05 vs. Con). In cardiomyocytes, PKA activity was increased by morphine.

Conclusion

Our data suggest that morphine-induced cardioprotection is mediated by STAT3-activation and inhibition of mPTP, with STA3 located upstream of mPTP. There is some evidence that protein kinase A is involved within the signalling pathway.     

Induction of cyclooxygenase-2 by heat shock protein 60 in macrophages and endothelial cells

The 60-kDa heat shock protein (HSP60), an endogenous ligand for the toll-like 4 receptor, is generated in response to inflammation, tissue injury, and/or stress and stimulates macrophages to produce cytotoxic and proinflammatory mediators including nitric oxide, tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-12. In the present studies we report that HSP60 is an effective inducer of cyclooxygenase-2 (COX-2) in macrophages, as well as endothelial cells. In both cell types, the synthesis of COX-2 was coordinate with induction of nitric oxide synthase (NOS)-2 and with nitric oxide production. With the use of promoter constructs in transient transfection assays, optimal expression of COX-2 in macrophages was found to require nuclear factor (NF)-kappaB, the cAMP-response element (CRE), and NF-IL-6, but not the E-box. Mobility shift assays revealed that HSP60 induced NF-kappaB and CRE binding activity, while CCAAT/enhancer binding protein (C/EBP), which binds to NF-IL-6, was constitutively active in the cells. Both c-Jun and CRE binding protein (CREB) bound to the CRE, while C/EBP-beta bound to NF-IL-6. These data indicate that NF-kappaB, C/EBP-beta, c-Jun, and CREB are important in HSP60-induced expression of COX-2. The c-Jun-NH(2)-terminal kinase (JNK), p44/42 mitogen-activated protein (MAP) kinase [extracellular signal-regulated kinase 1/2 (ERK1/2)], and p38 MAP kinase were rapidly activated by HSP60 in the macrophages. PD-98059, an inhibitor of phosphorylation of ERK1/2, caused a marked inhibition of HSP60-induced COX-2 and NOS-2 expression. Unexpectedly, SB-203580, a p38 kinase antagonist, was found to block HSP60-induced expression of COX-2, but not NOS-2. These data indicate that both ERK1/2 kinase and p38 kinase play a role in regulating HSP60-induced expression of COX-2.     

In Vitro evaluation of the cytotoxic and anti-proliferative properties of resveratrol and several of its analogs

Resveratrol (RES), a component of red wine, possesses anti-inflammatory properties. The studies described in the present work were aimed at evaluating the potential for RES and related stilbene analogs (piceatannol, PIC; pterostilbene, TPS; trans-stilbene, TS; and trans-stilbene oxide, TSO) to exhibit toxicity towards RAW 264.7 mouse macrophages. The effect of TS, TSO, RES and TPS on RAW 264.7 macrophage viability was determined by two standard methods: (a) the MTT assay and (b) the trypan blue dye exclusion test. Whereas macrophages were more sensitive to PIC (LC_{50 trypan} ∼ 1.3 μM) and to TPS (LC_{50 trypan} ∼ 4.0 μM and LC_{50 MTT} ∼ 8.3 μM) than to RES (LC_{50 trypan} ∼ 8.9 μM and LC_{50 MTT} ∼ 29.0 μM), they were relatively resistant to TSO (LC_{50 trypan} ∼ 61.0 μM and LC_{50 MTT} > 100 μM) and to TS (LC_{50 trypan} ≥ 5.0 μM and LC_{50 MTT} ≥ 5.0 μM). The ability of selected stilbenes (RES, TPS and PIC) to exhibit growth inhibitory effects was also examined. Although RES and TPS were observed to inhibit cell proliferation in macrophages (IC₅₀ ≤ 25 μM), these cells were resistant to growth inhibition by PIC (IC₅₀ ≥ 50 μM). The data obtained in the present analysis demonstrate that substituted stilbene compounds such as RES have the capacity to exhibit cytotoxic and anti-proliferative activities in macrophages.     

Cyclic three-dimensional wall motion of the human ascending and abdominal aorta characterized by time-resolved three-dimensional ultrasound speckle tracking

Biomechanics and Modeling in Mechanobiology - The aim of this study was to measure, characterize, and compare the time-resolved three-dimensional wall kinematics of the ascending and the abdominal...     

Ebselen Reduces the Toxicity of Mechlorethamine in A‐431 Cells via Inhibition of Apoptosis

A series of test compounds were evaluated for an ability to reduce the toxicity of the nitrogen mustard mechlorethamine (HN2) in vitro. The test compounds included resveratrol, pterostilbene, vitamin C, ebselen, ebselen diselenide, and ebselen‐sulfur. Among them, ebselen demonstrated the highest degree of protection against HN2 toxicity. To this end, pretreatment of the cells with ebselen offered protection against the toxicant whereas no protection was observed when cells were first incubated with HN2 and then treated with ebselen. Significant increases in caspase 3 and caspase 9 activities were observed in response to HN2, and ebselen was found to reduce these effects. Taken together, the data presented here indicate that ebselen is an effective countermeasure to nitrogen mustard in vitro, which is worthy of future investigation in vivo. © 2013 Wiley Periodicals, Inc. J BiochemMol Toxicol 27:313‐322, 2013; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.21490