Marked Changes In Shape Research Articles

Unveiling Frequency-Dependent Electromechanical Dynamics in Ferroelectric BaTiO3 Nanofilm with a Core-Shell Structure

Diverse domain patterns significantly influence the nonlinear electromechanical behaviors of ferroelectric nanomaterials, with polarization switching under strong electric fields being inherently a frequency-dependent phenomenon. Nevertheless, research in this area remains limited. In this study, we present a phase-field investigation of frequency-dependent electromechanical dynamics of a polycrystalline BaTiO3 nanofilm with a core-shell structure, subjected to applied frequencies ranging from 1 to 80 kHz. Our findings elucidate the microstructural mechanisms underlying the electromechanical behaviors observed in these materials. The effect of the grain size and the strains effect are also taken into account. Hysteresis and butterfly loops exhibit a marked change in shape as the frequency changes. We discuss the underlying domain-switching dynamics as a basis for evaluating such frequency-dependent properties. In addition, we examine the scaling behaviors of the dynamic hysteresis and the influence of grain boundaries on the domain structure. We can also observe from hysteresis loops that the remnant polarization and coercive field significantly diminish when grain sizes decrease from 60 to 5 nm. A smaller grain size of the nanofilm yields a larger percentage of the dielectric grain boundary, which “dilutes” the overall ferroelectricity of the film. A vortex domain structure is more likely to form at low frequency and a small grain size.

Neuroplasticity in honey bee brains: An enhanced micro-computed tomography protocol for precise mushroom body volume measurement

BackgroundIn insect brains, mushroom bodies are associated with memory and learning behavior. It has been demonstrated that the volume of the mushroom bodies in the brain of a worker honey bee changes during the adult stage. Changes in mushroom body volume imply high neuroplasticity in the brains and may be related to the age polyethism of honey bees. A suitable volume measurement method is needed to understand the correlation between behavioral changes and mushroom body volume changes in honey bees. New methodWe developed a new protocol for insect micro-computed tomography by modifying a previously reported method. Permount™ mounting medium was used as the embedding medium for micro-computed tomography scanning. ResultsThis protocol can generate images with high contrast inside the brain and reduce the marked shape changes during specimen processing. From the resulting high-contrast images, we used freeware to generate a three-dimensional model and calculate the volumes of the mushroom bodies in honey bees. The measured volumes of the mushroom bodies were larger than the values reported in most previous studies. There was no significant difference between the left and right mushroom body volumes, but the volumes of honey bee mushroom bodies significantly increased with age. Comparison with existing methodsPrevious protocols for micro-computed tomography using dried samples would cause brain shrinkage; protocols using ethanol-preserved or resin-embedded samples generated images with lower contrast. ConclusionsThe embedding protocol for micro-computed tomography is suitable for calculating volume of the mushroom bodies in honey bee brains.

Genetic deletion of skeletal muscle iPLA2γ results in mitochondrial dysfunction, muscle atrophy and alterations in whole-body energy metabolism

Skeletal muscle is the major site of glucose utilization in mammals integrating serum glucose clearance with mitochondrial respiration. To mechanistically elucidate the roles of iPLA2γ in skeletal muscle mitochondria, we generated a skeletal muscle-specific calcium-independent phospholipase A2γ knockout (SKMiPLA2γKO) mouse. Genetic ablation of skeletal muscle iPLA2γ resulted in pronounced muscle weakness, muscle atrophy, and increased blood lactate resulting from defects in mitochondrial function impairing metabolic processing of pyruvate and resultant bioenergetic inefficiency. Mitochondria from SKMiPLA2γKO mice were dysmorphic displaying marked changes in size, shape, and interfibrillar juxtaposition. Mitochondrial respirometry demonstrated a marked impairment in respiratory efficiency with decreases in the mass and function of oxidative phosphorylation complexes and cytochrome c. Further, a pronounced decrease in mitochondrial membrane potential and remodeling of cardiolipin molecular species were prominent. Collectively, these alterations prevented body weight gain during high-fat feeding through enhanced glucose disposal without efficient capture of chemical energy thereby altering whole-body bioenergetics.

PI3K-Delta Inhibitors Induce Primary Monocyte Cytotoxicity but Do Not Alter Monocyte Differentiation

It is well appreciated that an altered tumor microenvironment is a hallmark of cancer, including in chronic lymphocytic leukemia (CLL) where monocyte-derived cells known as “nurse-like cells” (NLC) in the lymph node microenvironment support CLL cell viability, survival, and resistance to drug-induced apoptosis. CLL directed therapy may include conventional chemotherapy, immunotherapy, targeted kinase inhibitors, and/or Bcl2 inhibitors. Of these options, kinase inhibitors such as PI3K-delta inhibitors are an effective therapy that have been found to disrupt NLC-CLL cell interaction in vitro and induce initial lymphocytosis thought to be related to this disrupted cellular interaction. We previously demonstrated that the PI3K-delta inhibitors idelalisib and umbralisib (TGR-1202) induce cytotoxicity, apoptosis, and inhibition of pAKT in primary CLL cells. We now investigate the effect of idelalisib and umbralisib on monocytes by evaluating cytotoxicity and cytokine-induced differentiation. We isolated monocytes from blood obtained from normal donors using negative selection (RosetteSep monocyte). For cytotoxicity experiments, primary purified monocytes were incubated ± M-CSF (10 ng/mL) ± PI3K-delta inhibitor (at 1.25 to 20 μM) for three days, and cell viability was measured with the MTS reagent after three hours. For differentiation experiments, primary purified monocytes were incubated first with M-CSF (10 ng/mL) for three days, then washed and incubated ± IL-10 (20 ng/mL) ± PI3K-delta inhibitor (10 μM) for three days. To judge monocyte differentiation, we used flow cytometry to measure expression of CD14, CD206, CD163, CD124, CD80, and CD86. We also visually assessed monocyte appearance using phase microscopy with these conditions. Both idelalisib and umbralisib induced concentration dependent cytotoxicity (N = 3), but cytotoxicity was higher with idelalisib (62% cytotoxicity at 1.25 μM) compared to umbralisib (24% cytotoxicity at 1.25 μM). The addition of M-CSF reduced PI3K-delta inhibitor-induced cytotoxicity of primary monocytes. For example, 1.25 μM idelalisib induced 62% cytotoxicity without M-CSF, compared with 5% cytotoxicity with M-CSF. Staged cytokine incubation of monocytes with M-CSF and IL-10 induced the expression of M2 differentiation markers (CD163 and CD124) and reduced expression of the M1 differentiation marker (CD86). Addition of idelalisib or umbralisib did not significantly affect monocyte surface marker expression compared to no drug (N = 4). Even though idelalisib and umbralisib were cytotoxic for monocytes, these drugs did not affect morphologic changes induced by IL-10 (marked spreading and shape change). Thus, PI3K-delta inhibitors do not appear to affect cytokine-induced monocyte differentiation, but they do induce monocyte cytotoxicity (idelalisib > umbralisib). The addition of M-CSF, which contributes to M2 differentiation, blocks these effects. Together, our results suggest that depleting monocyte or NLC number may contribute to the anti-CLL therapeutic effects of PI3K-delta inhibitors. In addition, a higher level of monocyte cytotoxicity with idelalisib compared to umbralisib could potentially contribute to the higher infectious side effect profile with idelalisib seen in clinical use, as monocyte derived cells are a key component of the innate immune system. The anti-monocyte effect of PI3K-delta inhibitors could provide rationale for using these agents in other malignancies in which monocyte-derived cells are an integral part of the tumor microenvironment. DisclosuresFriedman:TG Therapeutics: Research Funding. Maryanski:TG Therapeutics, Inc.: Employment, Equity Ownership. Miskin:TG Therapeutics, Inc.: Employment, Equity Ownership.

Sexual dimorphism of the mandibular conformational changes in aging human adults: A multislice computed tomographic study by geometric morphometrics.

The aging process has an impact on mandibular bone morphology and can therefore affect shape sexual dimorphism. Understanding the effect of senescence on mandibular shape changes is particularly important to correctly estimate the sex of an individual and predict age-related conformational modifications. The purpose of this study was to assess age-related changes in mandibular shape and sexual dimorphism. The study sample comprised 160 Multi Slice Computed Tomography examinations of individuals aged 40 to 79 years. Geometric morphometric analysis of fourteen osteometric landmarks was used to examine sexual dimorphism and patterns of mandibular shape variation with age. Results showed that mandibular sexual dimorphism of shape remained significant with aging. Conformational changes occurred between 50 and 70 years and were different for male and female individuals. Females presented earlier and more marked age-related shape changes than males. These observations suggest that mandibular senescence is a sexually dimorphic process since its onset, rate, and the areas subjected to conformational changes differ from male to female individuals. Senescence-related changes present substantial variability, and further investigation is required to determine precisely the age that marks their onset.

Antibacterial activity and mechanism of action of some Iranian honeys compared to manuka honey against multidrug-resistant respiratory and urinary infections

Increasing antibiotic resistance among bacterial pathogens requires the identification of alternative therapeutic agents, especially the natural ones. The importance of honey in health care was document from the oldest medical literature. Currently, the antimicrobial properties of honey, especially for the prevention of resistant strains, has been considered. However, from mechanistic insights, this activity has received less attention. In this study, the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of 6 Iranian honey samples in comparison with Manuka honey against reference strains of Staphylococcus aureus ATCC 29737, Pseudomonas aeruginosa ATCC 27853, Klebsiella pneumonia ATCC 10031, Escherichia coli ATCC 10536 and 22 multidrug-resistant clinical strains of respiratory and urinary infections were determined. The survival of bacteria and their ability to produce biofilm in the presence of a bactericidal concentration of honey were monitored. The effect of the most powerful honey and Manuka on the structure of reference strains of the Staphylococcus aureus and Pseudomonas aeruginosa was investigated using Field Emission Scanning Electron Microscopy (FESEM) and flow cytometry analysis. Exposure of bacteria to the studied honey generally resulted in high antimicrobial and anti-biofilm activity of Iranian honey compared to Manuka honey. Using SEM, division inhibition accompanied by loss of structure and marked changes in cell shape and surface were observed in the presence of honey. Analysis of flow cytometric charts revealed synchronous increasing and decreasing in the size of honey-treated strains. Present results indicated that Iranian honey samples possess the potential to be an effective inhibitor of multi-drug resistant strains.

Right Ventricular Shape Distortion in Tricuspid Regurgitation.

Tricuspid regurgitation (TR) is a failure in right-sided AV valve function which, if left untreated, leads to marked cardiac shape changes and heart failure. However, the specific right ventricular shape changes resulting from TR are unknown. The goal of this study is to characterize the RV shape changes of patients with severe TR. RVs were segmented from CINE MRI images. Using particle-based shape modeling (PSM), a dense set of homologous landmarks were placed with geometric consistency on the endocardial surface of each RV, via an entropy-based optimization of the information content of the shape model. Principal component analysis (PCA) identified the significant modes of shape variation across the population. These modes were used to create a patient-prediction model. 32 patients and 6 healthy controls were studied. The mean RV shape of TR patients demonstrated increased sphericity relative to controls, with the three most dominant modes of variation showing significant widening of the short axis of the heart, narrowing of the base at the RV outflow tract (RVOT), and blunting of the RV apex. By PCA, shape changes based on the first three modes of variation correctly identified patient vs. control hearts 86.5% of the time. The shape variation may further illuminate the mechanics of TR-induced RV failure and recovery, providing potential targets for therapies including novel devices and surgical interventions.

Antibacterial properties of selected Malaysian Tualang honey against Pseudomonas aeruginosa and Streptococcus pyogenes.

Background and Objectives:Tualang honey (TH) is a Malaysian multifloral jungle honey. In recent years, there has been a marked increase in the number of studies published in medical databases regarding its potential health benefits. The study aimed to investigate the effect of TH against Pseudomonas aeruginosa and Streptococcus pyogenes.Materials and Methods:The effect of TH on both bacteria was investigated using MIC, MBC, growth curve, time-kill curve, scanning electron microscopy (SEM) and RT-qPCR.Results:The MIC of TH against P. aeruginosa and S. pyogenes was 18.5% (w/v) and 13% (w/v) respectively and MBC was 25% (w/v) for both bacteria. Spectrophotometric readings of at least 90% inhibition yielded MIC90 values of TH, 18.5% (w/v) and 15% (w/v) for P. aeruginosa and S. pyogenes respectively. A time–kill curve demonstrated a bactericidal with a 4-log reduction estimated within 8 hours. Using SEM, loss of structural integrity and marked changes in cell shape were observed. RT-qPCR analysis showed that TH reduced the pattern of gene expression in both bacteria, with a trend toward reduced expression of the virulence genes of interest.Conclusion:This study suggests that TH could potentially be used as an alternative therapeutic agent for microbial infection particularly against these two organisms.

3D Muscle Fiber Architecture of Avian Jaw Muscles and their Significance for Avian cranial functional morphology evolution

Birds evolved a diversity of head shapes, feeding behaviors and jaw muscles which drive the feeding apparatus. However, capturing the anatomical and functional diversity of these muscles remains challenging and we know little about how jaw muscles develop, function and evolve among lineages of the clade and this hampers our ability to understand skeletal tissue biology, joint function, and patterns of evolution in bird skulls. Here we use contrast imaging, 3D muscle fiber tracking and new data visualization methods to explore the morphology, architecture and functional significance of the jaw muscles of a sample of birds to highlight the potential of these approaches. First, we show differences in functional morphology of jaw muscles between a hatchling and adult Mallard duck (Anas platyrhynchos). Ducks show a marked change in adductor chamber shape during ontogeny including an elongated postorbital process, quadrate orbital process and retroarticular process along with a broad temporal fossa which likely track with changes in muscle function and cranial kinesis. Indeed, we found m. depressor mandibulae in baby and adult ducks shifts from a rostrocadual orientation to a more dorsoventral orientation that offers additional mechanical advantage to jaw opening and propalinal mandible movement as well as concomitant changes in the 3D resultant and pennation of other jaw muscle bellies. Second, we show differences between a long‐ and short‐faced, dabbling birds (mallard, green‐wing teal), long faced orthally‐biting bird (belted kingfisher) and a short‐faced, hard‐biting bird, the grey parrot (Psittacus erithacus). We expected position, architecture and biomechanics of homologous muscles to differ substantially between these avian species given their different behaviors and overall cranial morphology. Although some muscles showed marked differences in resultant and pennation, others, including protractor pterygoideus, the primary actuator of kinesis, were remarkably similar in morphology suggesting a mosaic of changes happening during avian evolution. Together, these new, 3D high fidelity anatomical data on jaw muscle functional morphology in birds will not only better illuminate ecomorphological evolution but also illustrate how other clades of birds and vertebrates adapt their muscle architecture to meet functional demands.

Sensing of red blood cells with decreased membrane deformability by the human spleen

AbstractThe current paradigm in the pathogenesis of several hemolytic red blood cell disorders is that reduced cellular deformability is a key determinant of splenic sequestration of affected red cells. Three distinct features regulate cellular deformability: membrane deformability, surface area-to-volume ratio (cell sphericity), and cytoplasmic viscosity. By perfusing normal human spleens ex vivo, we had previously showed that red cells with increased sphericity are rapidly sequestered by the spleen. Here, we assessed the retention kinetics of red cells with decreased membrane deformability but without marked shape changes. A controlled decrease in membrane deformability (increased membrane rigidity) was induced by treating normal red cells with increasing concentrations of diamide. Following perfusion, diamide-treated red blood cells (RBCs) were rapidly retained in the spleen with a mean clearance half-time of 5.9 minutes (range, 4.0-13.0). Splenic clearance correlated positively with increased membrane rigidity (r = 0.93; P < .0001). To determine to what extent this increased retention was related to mechanical blockade in the spleen, diamide-treated red cells were filtered through microsphere layers that mimic the mechanical sensing of red cells by the spleen. Diamide-treated red cells were retained in the microsphilters (median, 7.5%; range, 0%-38.6%), although to a lesser extent compared with the spleen (median, 44.1%; range, 7.3%-64.0%; P < .0001). Taken together, these results have implications for understanding the sensitivity of the human spleen to sequester red cells with altered cellular deformability due to various cellular alterations and for explaining clinical heterogeneity of RBC membrane disorders.

Age-specific environmental differences on the otolith shape of the bastard grunt (Pomadasys incisus) in the north-western Mediterranean

The effects of sex, age, and environment on the shape of the bastard grunt (Pomadasys incisus) otoliths from the north-western Mediterranean were investigated. Specimens of this species were collected from two separate sampling areas in the north-western Mediterranean with different thermal regimes. Sex, growth rates, and age of P. incisus were determined by using gonad histology techniques, biometric analyses, and otolith microstructure analyses, respectively. The shape was described using normalized Elliptic Fourier descriptors (EFDs), and studied by means of multivariate statistics as predictive variables with age-specific discriminant analyses. There were no consistent differences found between sexes, but otolith shapes varied significantly between environments within different age classes. Total classification success varied between 87.3% and 89.2% between environments for the different age classes and provided a phenotypic basis for P. incisus population separation within an environmental gradient in determining its otolith shape. In addition, significant differences were observed between sampling areas in von Bertalanffy’s growth parameters, as well as in the fish length-weight (LWR) and fish length-otolith radius (TL-Ro) relationships. Data was discussed considering that the physical habitat variability could underlie a marked change in otolith shape during the animals’ growth. In this matter, we discussed the relative importance of both ontogenetic and environmental conditions (such as water temperature) on otolith shape.

Action Potential Modulation in CA1 Pyramidal Neuron Axons Facilitates OLM Interneuron Activation in Recurrent Inhibitory Microcircuits of Rat Hippocampus

Oriens-lacunosum moleculare (O-LM) interneurons in the CA1 region of the hippocampus play a key role in feedback inhibition and in the control of network activity. However, how these cells are efficiently activated in the network remains unclear. To address this question, I performed recordings from CA1 pyramidal neuron axons, the presynaptic fibers that provide feedback innervation of these interneurons. Two forms of axonal action potential (AP) modulation were identified. First, repetitive stimulation resulted in activity-dependent AP broadening. Broadening showed fast onset, with marked changes in AP shape following a single AP. Second, tonic depolarization in CA1 pyramidal neuron somata induced AP broadening in the axon, and depolarization-induced broadening summated with activity-dependent broadening. Outside-out patch recordings from CA1 pyramidal neuron axons revealed a high density of α-dendrotoxin (α-DTX)-sensitive, inactivating K+ channels, suggesting that K+ channel inactivation mechanistically contributes to AP broadening. To examine the functional consequences of axonal AP modulation for synaptic transmission, I performed paired recordings between synaptically connected CA1 pyramidal neurons and O-LM interneurons. CA1 pyramidal neuron–O-LM interneuron excitatory postsynaptic currents (EPSCs) showed facilitation during both repetitive stimulation and tonic depolarization of the presynaptic neuron. Both effects were mimicked and occluded by α-DTX, suggesting that they were mediated by K+ channel inactivation. Therefore, axonal AP modulation can greatly facilitate the activation of O-LM interneurons. In conclusion, modulation of AP shape in CA1 pyramidal neuron axons substantially enhances the efficacy of principal neuron–interneuron synapses, promoting the activation of O-LM interneurons in recurrent inhibitory microcircuits.

‘Is it normal?’ A simple question that often lacks an easy answer

Anyone experiencing a change in bodily function must ask ‘is this normal or is it a manifestation of a serious disease’? Often the answer will be obvious, but not always; individuals may delay seeking medical advice until they believe their condition is ‘doctorable’, worthy of utilising a physician’s time. 1 The problems are greatest at the extremes of age. First, any problems will arise in bodies that are anyway undergoing a series of often quite marked changes that are entirely normal. Young children experience marked changes in body size and shape and in the ability to do things, such as walk and control their bodily functions. Very old people may also experience changes, but in the opposite direction as they lose body mass from muscles and bones; they become unable to do things that once were easy, again including walking and controlling their bodily functions. Second, those at the extremes of age are also often dependent on others to recognise the changes to their bodies, to interpret them, and to determine which may require responses by health professionals and which are ‘normal’.

Viability and Regeneration of Chondrocytes after Laser Cartilage Reshaping Using 1,460 nm Diode Laser

ObjectivesCartilage reshaping by laser irradiation is used to correct septal and auricular cartilage deformities. Chondrocyte viability following laser irradiation and reshaping has been well established. However, the regeneration process of chondrocyte after laser irradiation has not been revealed yet. The aims of this study were to determine the mechanism of cartilaginous thermal injury and the regenerative process of damaged cartilage following laser irradiation.MethodsLaser irradiation was performed on human septal cartilage and rabbit auricular cartilage using a 1,460-nm diode laser. We observed change in the shape of cartilage and evaluated the extent of cartilage injury using live/dead cell assay via confocal microscopy. Hoechst and propidium iodide (PI) staining was used to evaluate the mechanism of chondrocyte injury after laser irradiation. To evaluate the regeneration of cartilage, laser irradiated cartilages were reimplanted into a subperichondrial pocket and were harvested at 1, 2, and 4 weeks after reimplantation for viability assessment and histologic examination.ResultsLaser irradiation using a 1,460-nm diode laser produced a marked shape change in both human septal and rabbit auricular cartilages. Thermal damage on cartilage was correlated with the exposure time and the laser power. Hoechst and PI staining showed that chondrocyte death by laser irradiation was due to mainly necrosis, rather than apoptosis. In lower power treatment group (0.3 W and 0.5 W), all the chondrocytes regenerated within 4 weeks, however, in 1 W treatment group, chondrocytes could not regenerate until 4 weeks.ConclusionReshaping of cartilage using 1,460 nm diode laser was attained concurrently with the thermal injury to the chondrocytes. The extent of thermal damage on chondrocytes was dependent on the exposure time and the laser power and the damaged chondrocytes irradiated with lower level of laser power could be regenerated after reimplantation into subperichondrial pocket.

T-PA–specific modulation of a human blood-brain barrier model involves plasmin-mediated activation of the Rho kinase pathway in astrocytes

AbstractTissue-type plasminogen activator (t-PA) can modulate permeability of the neurovascular unit and exacerbate injury in ischemic stroke. We examined the effects of t-PA using in vitro models of the blood-brain barrier. t-PA caused a concentration-dependent increase in permeability. This effect was dependent on plasmin formation and potentiated in the presence of plasminogen. An inactive t-PA variant inhibited the t-PA–mediated increase in permeability, whereas blockade of low-density lipoprotein receptors or exposed lysine residues resulted in similar inhibition, implying a role for both a t-PA receptor, most likely a low-density lipoprotein receptor, and a plasminogen receptor. This effect was selective to t-PA and its close derivative tenecteplase. The truncated t-PA variant reteplase had a minor effect on permeability, whereas urokinase and desmoteplase were ineffective. t-PA also induced marked shape changes in both brain endothelial cells and astrocytes. Changes in astrocyte morphology coincided with increased F-actin staining intensity, larger focal adhesion size, and elevated levels of phosphorylated myosin. Inhibition of Rho kinase blocked these changes and reduced t-PA/plasminogen–mediated increase in permeability. Hence plasmin, generated on the cell surface selectively by t-PA, modulates the astrocytic cytoskeleton, leading to an increase in blood-brain barrier permeability. Blockade of the Rho/Rho kinase pathway may have beneficial consequences during thrombolytic therapy.

Implications of Acute Left Ventricular Remodeling during Squatting Stress Echocardiography

We previously demonstrated that squatting induces left ventricular (LV) wall motion abnormalities (WMA) in areas subtended by stenotic coronary arteries. In addition, it was observed that some subjects developed acute changes in LV shape (acute left ventricular remodeling [ALVRM]) during squatting. This study tested the hypothesis that patients with ALVRM during squatting echocardiography have higher incidences of severe coronary artery disease (CAD). Echocardiography was performed in all standard views during standing and squatting. End-systolic frames in the apical four-chamber view were analyzed. The subjects were divided into three groups. Group 1 consisted of 12 subjects who developed squatting-induced ALVRM with apical and distal posterior septal akinesis, dilation of the apex and marked LV shape change at end-systole. Group 2 consisted of 20 subjects with distal posterior septal and apical akinesis without ALVRM, during squatting. Group 3 consisted of 64 subjects who developed WMA in areas other than the apex (n = 49), or normal wall motion (n = 15) during squatting. Coronary angiography in group 1 revealed that 6 subjects had left main coronary artery stenosis (LMCAS ≥ 50%), two had severe three vessel disease (≥ 90% stenosis), and one had 100% left anterior descending coronary artery occlusion. Severe CAD was defined for purpose of this study as the presence of LMCAS, or severe three vessel disease (≥ 90% stenosis). Six subjects in group 2 had LMCAS and none had severe three vessel disease (P < 0.05 vs. group 1 for LMCAS and/or three vessel disease). In group 3, eight had LMCAS and none had severe three vessel disease (P < 0.0001 vs. group 1). Patients with ALVRM have severe CAD. Therefore, patients who develop ALVRM during squatting require urgent evaluation for revascularization therapy.

Abstract LB-190: Novel cell cycle regulation through the WW-domain of the co-chaperone BAG3

Abstract The BAG family of anti-apoptotic co-chaperones regulates protein folding and aggregation during cellular stress. The C-terminal BAG domain directly binds the ATPase domain of Hsp70/Hsc70 and prevents the release of client proteins that are otherwise directed to the proteasome for degradation. BAG3 is a unique member of the BAG family that also contains an evolutionarily conserved N-terminal WW-domain with undefined function. We removed the BAG3 WW-domain and observed a marked change in cell shape and cell cycle progression. This led us to hypothesize the BAG3 WW-domain is critical in regulating cellular division through a cytoskeletal association in a cell cycle-dependent fashion. Immunoblot analysis of synchronized HeLa (p53-null) and U2OS (human osteosarcoma, wild-type p53) cell lysates revealed that BAG3 expression was significantly increased during S phase and precipitated in the insoluble cell fraction at the G2/M transition. Indirect immunofluorescence demonstrated that BAG3 colocalized with the Golgi apparatus in the perinuclear region throughout interphase, and with leading lamellipodia exclusively during G2 phase. Co-immunoprecipitation of BAG3 with F-actin confirmed association with microfilaments and was increased during G2 phase relative to asynchronous lysates. A GST-hBAG3-WW-domain recombinant protein pulldown confirmed a specific F-actin association, suggesting a unique BAG3-WW-domain interaction with the cytoskeleton. We also generated a series of N-terminal, V5 tag-labeled domain-deletion clones of BAG3 with the WW (ΔWW) domain removed. ΔWW-BAG3 cells contained degenerate F-actin organization, abnormal morphology, and loss of BAG3 in lamellipodia. A 3.2-fold increase in DNA content of ΔWW-BAG3 cells was shown by flow cytometry, relative to empty vector (pCI Neo) and BAG3 overexpressing (FL-BAG3) transfected controls (p&amp;lt;0.001, n=3). Furthermore, centrosome amplification was increased in asynchronous cells: 2.43 ± 0.12/ΔWW-BAG3 cell, 1.86 ± 0.06/pCI Neo cell, and 1.79 ± 0.04/FL-BAG3 (p&amp;lt;0.05, n=4). Cell division in real time was followed using GFP-histone 2B-tagged live cell imaging. We confirmed polyploidy development and absence of cytokinesis in ΔWW-BAG3 cells, and also observed mitotic delay relative to pCI Neo (1260 min. vs. 110 min.) and FL-BAG3 cells (vs. 90 min., n=2). These data indicate a novel requirement for BAG3 for normal mitotic progression via BAG3 WW-domain association with the actin cytoskeleton. To our knowledge, this is the first reported activity of differential co-chaperone protein behavior based on cell cycle phase, independent of p53 status. These data identify a novel regulatory point of G2/M progression and suggest the WW-domain of BAG3 is a potential therapeutic target to deter cancer proliferation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-190. doi:10.1158/1538-7445.AM2011-LB-190

Ontogenetic and environmental effects on otolith shape variability in three Mediterranean European eel ( Anguilla anguilla, L.) local stocks

Otolith morphology is an efficient tool for the discrimination of fish stocks, populations and species when comparative genetic data are not available. Currently, the relationship between environmental factors and otolith shape is poorly characterized for the European eel ( Anguilla anguilla), a highly migratory catadromous species constituting a single, randomly mating stock. The present study analyses the differences in otolith morphology between three Mediterranean eel local stocks from different environmental contexts (i.e. two brackish lagoons and one river). The relationship between otolith shape and otolith size was studied by means of Elliptic Fourier analysis and multivariate statistics. Otolith profile was digitally acquired and Cartesian coordinates were extracted. Partial Least Square (PLS) analysis pointed to continuous allometric growth in size and shape in otoliths from all three sites. In the three environments, shape variations occurred during growth as indicated by the presence of a significant and positive relationship between otolith size and the first PLS latent vector (i.e. which bears most of the information regarding otolith outline). Differences between smaller and larger sized otoliths were investigated using PLS Discriminant Analysis (PLSDA) and cluster analysis. Results indicate that otolith shape is highly uniform at smaller than at larger sizes. These shape differences apparently overlap the initial differentiation of the small otolith outlines acquired by eels during the growing phase as elvers in the marine environment. Data were discussed considering that the physical and chemical habitat variability in brackish lagoons and river could underlie a marked change in otolith shape during the animals' growth.

The effect of manuka honey on the structure of Pseudomonas aeruginosa

The purpose of this study was to investigate the effects of manuka honey on the structural integrity of Pseudomonas aeruginosa ATCC 27853. The minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) of manuka honey for P. aeruginosa were determined by a microtitre plate method, and the survival of bacteria exposed to a bactericidal concentration of manuka honey was monitored. The effect of manuka honey on the structure of the bacteria was investigated using scanning and transmission electron microscopy (SEM and TEM, respectively). The MIC and MBC values of manuka honey against P. aeruginosa were 9.5% (w/v) and 12% (w/v) respectively; a time-kill curve demonstrated a bactericidal rather than a bacteriostatic effect, with a 5 log reduction estimated within 257 min. Using SEM, loss of structural integrity and marked changes in cell shape and surface were observed in honey-treated cultures. With TEM, these changes were confirmed, and evidence of extensive cell disruption and lysis was found. Manuka honey does not induce the same structural changes in P. aeruginosa as those observed in staphylococci. Our results indicate that manuka honey has the potential to be an effective inhibitor of P. aeruginosa.

Control of Relative Direction and Amplitude in Extension/Contraction Motions of Molecular Strands Induced by Ion Binding

The shape of ligand strands composed of six-membered aza-heterocycles (het) connected at the alpha and alpha' positions by hydrazone (hyz) units is determined in a predictable fashion by the nature of the heterocyclic groups (pyridine, pyrimidine, pyrazine etc.), and covers the range from extended linear to compact helical structures. The binding of metal ions to the coordination subunits, defined by the het-hyz sequences, leads to marked shape changes by inter-converting bent and linear conformations of the subunits, thus inducing relative motions of strand domains either in the same (con-sense, "twirling") or in opposite (dis-sense, "flapping") directions. The amplitude of the motion induced by metal-ion binding and release and the relative directions of the formal motions can be controlled by the nature of the heterocyclic units. Thus, motions around a central 4,6-disubstituted pyrimidine are dis-sense motions, whereas there are con-sense motions around a central 2,5-disubstituted pyrazine unit, as illustrated by model ligands 1 and 2, respectively. The more extended helical 3 and undulating (zigzag shape) 4 ligands undergo larger-amplitude motions combining the relative displacements displayed by 1 and 2. Ligands 3 and 4 form linear tetranuclear Pb(II) and Zn(II) complexes, thus producing an extension motion. The same holds for [Ru(4)(terpy)(4)](PF(6))(8) (terpy=terpyridine). Reversible acid-base-triggered molecular motions have been generated with [Zn(4)(4)(OTf)(8)] (TfOH=triflic acid).