bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantification of the Role of Glycosaminoglycans on the Tensile and Viscoelastic Properties of the Aortic Valve Leaflet
Authors: Tseng Hubert , Kim Eric J. , Patrick S. Connell , Grande-Allen K. Jane
Published: July 2012

The spongiosa is the middle of three layers of the aortic valve leaflet and contains the proteoglycan (PG) versican and glycosaminoglycan (GAG) hyaluronan (HA) in large quantities. The resulting versican-HA aggregates allow the layer to retain large quantities of water, giving the layer a gel-like consistency. This layer is the least understood layer of the leaflet as it pertains to its role in valvular function and mechanics. However, the GAGs in this layer are perceived to be important to valve function, as the loss of GAGs in bioprosthetic heart valves coincides with its failure. The functions attributed to these GAGs include: (1) lubricating shear between the outer layers during flexure and tension; and (2) filling large volumes, resisting compression, and dampening shock from valve closure. The lack of understanding of GAGs role in valvular function is due to the difficulty in its isolation from the rest of the leaflet. For our study, rather than isolate the spongiosa, we enzymatically digest GAGs from the spongiosa via hyaluronidase. Previous studies have shown that the complete enzymatic removal of GAGs from heart valve leaflets greatly increases flexural rigidity. In this study, we instead varied the amounts of GAGs in the leaflets and investigated the effects on the tensile properties of the native leaflet; native leaflets in tension have a bilinear stress-strain curve, little hysteresis and minimal relaxation. GAGs were depleted using varying concentrations (0, 1, 2, 5 U/mL) and application times (8, 24 h) to yield a gradient of 4 GAG amounts ranging between 20-40 μg GAG/mg dry weight of native tissue, although with little change in water content. GAG depletion from the spongiosa was also confirmed using Alcian blue staining, which also verified no gross changes to the outer layers. Leaflets in these GAG depletion gradient conditions are in the progress of being mechanically tested to elucidate the effects of GAGs on the tensile elastic and viscoelastic properties of the native leaflet. This work is a component of a broader effort to elucidate the important role of GAGs and PGs in the function of native and diseased valves and in the design of improved valve replacements.
Biomechanical Stimuli Effects on Valve Endothelial Cell Anti-thrombotic Mechanisms
Authors: Balaoing Liezl R. , Post Allison D. , Lin Adam Y. , Nolasco Leticia , Turner Nancy A. , Moake Joel L. , Grande-Allen K. Jane
Published: July 2012

Because of the degeneration and thrombosis in artificial heart valve implants, it is important to understand the anti-thrombotic mechanisms of cardiac valve endothelial cells (VECs). These anti-thrombotic mechanisms can be integrated into poly(ethylene glycol) diacrylate (PEGDA) tissue-engineered heart valve (TEHV) design. This work will study the effects of (1) PEGDA hydrogel stiffness and (2) specific extracellular matrix (ECM) adhesive peptides on VEC phenotype and anti-thrombotic mechanisms. PEGDA 10% (w/v) hydrogels of MWs 3.4 and 20kDa were polymerized to apply different substrate rigidities in VEC culture. Thiol-ene reactions were used to covalently bind laminin- and fibronectin- derived peptides to the acrylate groups on PEGDA hydrogel surfaces. Laminin-derived peptide motif RKRLQVQLSIRT (RKR) and fibronectin adhesive peptide RGD were modified to include an additional cysteine at the end of each sequence, introducing a free thiol to undergo the thiol-ene reaction. Thiol-PEG-fluorescein (SH-PEG-FITC) served as a negative adhesive substrate control. Porcine aortic VECs were seeded onto each of the ECM-hydrogel combinations and cultured for 2, 6, and 10 days. Cell phenotype, adhesion, and proliferation were then assessed. Analysis of specific VEC anti-thrombotic protein regulation is in progress. At each time point, samples will be analyzed for maintenance of VEC phenotype and expression of thrombotic (von Willebrand Factor [VWF]) and anti-thrombotic (VWF cleaving enzyme [ADAMTS-13], eNOS, PGI2, tPA) proteins using histochemistry and qRT-PCR. Addition of histamine has been shown to stimulate rapid release of thrombogenic ultra-large VWF (ULVWF) strings by vascular endothelial cells. This method will be used to study VEC ULVWF string production and the associated cleavage activity of ADAMTS-13. Control of the hemostatic process will be quantified via western blot and ELISAs.
The 3.4 and 20kDa MW PEGDA hydrogels had compressive moduli of 131±5 and 7.5±2kPa, respectively. Binding different concentrations of SH-PEG-FITC onto the gel surfaces showed that acrylate saturation was achieved for both MW compositions using ~5mM of peptide solution. After 2 days, the VECs on the stiffer 3.4kDa RKR gels appeared spread and elongated, whereas the 3.4kDa RGD seeded VECs had cobblestone morphology. VEC adhesion on the RKR and RGD 20kDa gels was observed, but with limited spreading. The cultured VECs may prefer the stiffer 3.4kDa gels over the softer 20kDa gels. After 10 days, VECs on the 3.4kDa RGD gels had minimal proliferation, while VECs on RKR grew confluent, were cobblestone shaped, and expressed VWF. Results suggest that both substrate rigidity and adhesive substrate greatly influence VEC survival, and likely affects anti-thrombotic regulation. Future studies include the use of basal lamina components collagen IV and perlecan peptides to evaluate changes in VEC behavior.
Biomechanical Activation of Human Valvular Interstitial Cells from Early Stage of CAVD
Authors: Poggio Paolo , Sainger Rachana , Grau Juan B. , Branchetti Emanuela , Lai Eric , Gorman Robert C. , Gorman III Joseph H. , Bavaria Joseph E. , Sacks Michael S. , Ferrari Giovanni
Published: July 2012

Aortic Valve (AV) leaflets are exposed to complex biomechanical stimulations, which has been linked to the progression of Calcific Aortic Valve Disease (CAVD) characterized by valvular interstitial cells (VICs) adopting an osteogenic phenotype, remodeling of the ECM and biomineralization. Initial asymptomatic phases of CAVD, named Aortic Valve Sclerosis (AVSc), include thickening of the cusps whereas advanced stages, Aortic Valve Stenosis (AVS), are associated with severe calcification. Prospective clinical studies of CAVD are hampered by the typically slow and variable progression of the disease. In addition, patients who present with AVS are already in the stage of severe calcification where damage to the AV leaflets is too severe to be reversed by drug therapy. Finally, due to its asymptomatic presentation little is known about the AVSc stage. The aim of this study is to develop tissue- and cell-based models to uncover the biomechanical forces leading to activation of VICs in early, asymptomatic AVSc patients. We investigate the impact of biological (BMP4 pathway) and mechanical (tensile stretch) forces leading to VICs osteogenic transdifferentiation, and biomineralization of the fibrosa layer.
Human, surgically resected, AV tissue from AVSc patients were characterized for cellular and extracellular markers and compared to healthy controls and AVS tissues. BMP4 and tensile stretch induce osteogenic marker expression and biomineralization of the fibrosa in non-calcified AVSc-derived tissue. VICs were isolated from patients and induced to transdifferentiate in either 2D cell culture or 3D Tissue Engineering valve model based on decellularized porcine AV scaffold repopulated with human-derived cells. Our results show that the synergistic combination of biological (BMP4) and mechanical (tensile stretch) forces is required to promote SMA, RUNX2, OPN, and ON expression of AVSc-derived cells. These results provide a novel study model of early asymptomatic AVSc that combine biological and mechanical stimulation to induce activation of AVSc-derived VICs and biomineralization of AVSc tissue. This model could be used to unveil the molecular mechanisms lading to VIC activation and to test possible future small drug to control cellular activation and tissue remodeling.
Decellularized In-Vitro Tissue-Engineered Heart Valves - First In-Vivo Results
Authors: Driessen-Mol Anita , Dijkman Petra E. , Frese Laura , Emmert Maximilian Y. , Grunenfelder Juerg , Sidler Michele , Weber Benedikt , Sanders Bart , Jenni Rolf , Baaijens Frank P.T. , Simon P. Hoerstrup Simon P. Hoerstrup
Published: July 2012

Decellularized xenogeneic and allogeneic heart valves are used as matrices for tissue regeneration. However, xenografts are associated with a risk of immunogenic reactions or disease transmission and homografts are sparse. Further, remodeling capacity of these matrices is questioned, as cell infiltration is limited. Alternatively, biodegradable synthetic materials are attractive as of their unlimited supply and freedom in valve geometry, with sufficient capacity for cell infiltration and neotissue formation and remodeling, but with cell-mediated leaflet retraction and thickening as common problem with in-vivo application. To overcome cell-mediated leaflet retraction and thickening and the limitations when using xenografts or homografts, we propose to decellularize in-vitro cultured tissue-engineered heart valves as off-the-shelf matrices for in-vivo regeneration. Tissue-engineered heart valves are grown based on ovine vascular cells seeded onto PGA/P4HB valvular shaped scaffolds and exposed to dynamic loading in bioreactors for 4 weeks. Decellularization of in-vitro cultured tissue-engineered heart valves is demonstrated feasible with efficient cell removal and preservation of the collagen architecture and tissue strength. Storage of these valves up to 18 months did not affect tissue properties. Decellularization strongly reduced leaflet retraction and, therewith, improved valvular function up to 24 hours in a valve tester. In-vivo performance of decellularized in-vitro cultured tissue-engineered heart valves after trans-apical implantation in pulmonary position in sheep was evaluated after 8 (n=1), 16 (n=1) and 24 (n=1) weeks. Complete cellular repopulation was demonstrated within 8 weeks with excellent in-vivo performance and no signs of tissue thickening. Moderate regurgitation developed after 16 weeks with leaflet prolapse after 24 weeks and a reduced leaflet area, likely due to minimal coaptation in the current valve design. Minimal coaptation makes the leaflets prone to prolapse with direct loss of coaptation when repopulating cells exert traction forces. Mechanical analyses demonstrated tissue remodeling with a trend towards the development of anisotropic tissue properties in time, demonstrating the promising nature of decellularized in-vitro cultured tissue-engineered heart valves for in-vivo regeneration. Efforts are ongoing to increase the number of samples and to optimize valve design and geometry to increase leaflet coaptation and maintain optimal valve performance.
The European Union's Seventh Framework Program is acknowledged for funding this study
Atypical expression and regulation of smooth muscle markers in human calcified valves
Authors: Latif Najma , Sarathchandra Padmini , Yacoub Magdi H. , Chester Adrian H.
Published: July 2012

Calcific aortic stenosis and atherosclerosis share many similar characteristics suggesting that similar mediators and pathogenic pathways are involved. In both conditions, the native cells are quiescent. However upon damage, both valve interstitial cells (VICs) and smooth muscle cells (SMCs) in the valve and the vascular wall respectively undergo remodelling and transformation. We hypothesise that the smooth muscle cell phenotype plays a role during the calcification process. 12 normal and 16 calcified valves were analysed for smooth muscle markers including smooth muscle α-actin, smooth muscle myosin, calponin, caldesmon, desmin, SM1 and SM2 by immunocytochemistry. The expression of myocardin-related transcription factors, (MRTFA/B) as key regulators of smooth muscle gene expression, was also evaluated. Normal human valves demonstrated the expression of smooth muscle markers localised to the base of the valve. Occasionally smooth muscle markers were seen in small groups of cells in the region from the base to the central region of the valve but never in the region from the central to the co-apting edge. Smooth muscle markers in all the calcified valves demonstrated an increased and aberrant expression. The expression of smooth muscle myosin, SM1, SM2, calponin and smooth muscle α-actin was found to be abundantly expressed around calcified nodules and distal to the nodules. The pattern of staining was frequently localised along the majority of the valve tissue as well as in isolated cells. Additionally, regions of the endothelium and endothelial cells lining the vessels were found to be positive for smooth muscle markers in 9/16 calcified valves. Normal valves did not demonstrate any expression of MRTFs. However, MRTF-A and MRTF-B were induced in calcified valves in a similar pattern to the expression of smooth muscle markers. They were also induced in the endothelium and the endothelial cells lining some vessels. Importantly, the expression of MRTFs was nuclear in both VICs and endothelial cells indicative of activation. TGFβ1 (10ng/ml) was able to induce and translocate the expression of MRTF-A to the nucleus in VICs within 4 hours in some isolates.
Calcified valves harbour a greatly increased number of smooth muscle marker-positive VICs and endothelial cells. This aberrant expression in endothelial cells may emulate endothelial to mesenchymal transformation (EMT) during embyonic valvulogenesis. Concomittantly, calcified valves expressed MRTFs, key regulators of smooth muscle gene expression. The presence of smooth muscle cell markers and MRTFs in calcified valves suggest a role for this cell phenotype in the development of the calcification process.
A Completely Biological, Tissue Engineered Valve Leaflet Suitable for TAVI
Authors: McAllister Todd , Dusserre Nathalie , Chronos Nicolas , L'Heureux Nicolas
Published: July 2012

Clinically available transcatheter aortic valve replacement (TAVR) technologies typically use chemically fixed bovine or equine tissues for the valve leaflets. While these fixed, xenogeneic materials have been used with success in devices placed by open surgical access, the tissue thickness (>500 microns) adds significantly to the overall crossing profile of the delivery device. Complications associated with device diameter are generally reported in at least 10-20% of clinical cases, making a reduced crossing profile one of the most critical targets for second generation TAVR devices. Another limitation associated with pericardium is fatigue induced delamination. Previously we have reported clinical results with a completely autologous tissue engineered vascular graft built using a process termed sheet-based tissue engineering. Using this approach, we were able to build small diameter blood vessels with supraphysiologic burst pressures, and demonstrated clinical durability with time points out to 3 years. Importantly, this tissue engineering approach requires no chemical fixation or exogenous biomaterials. More recently, we reported initial human use with an allogeneic version of the vessel. With time points out to 1 year, the allogeneic tissue engineered material demonstrated no evidence of immune reaction. This transition to an off-the shelf, allogeneic approach enables use the material in a variety of new clinical indications, including valve reconstruction. Valve leaflets built from a single sheet, demonstrated ultimate tensile strength in excess of that for bovine valve leaflets. Of note, the thickness of the sheet was less than 200 microns, roughly 30 percent that of bovine pericardium. The tissue can also be compressed, further reducing the thickness to approximately 75 microns. This thin, durable, single layered tissue can be assembled onto commercially available TAVR devices resulting in a reduction in crossing profile of approximately 2 Fr. The valve leaflets can be sutured easily, coapt normally, and can withstand arterial backpressure. Given the non-laminated structure of the tissue engineered leaflet, the lack of synthetic materials, and the durability demonstrated in other clinical indications, this approach may provide not only a reduced crossing profile, but also improved long term clinical results.
A Finite Element Study of Human Pulmonary Autograft Wall Stress after the Ross Procedure
Authors: Wisneski Andrew D. , Matthews Peter B. , Azadani Ali N. , Chitsaz Sam , Lui Natalie , Guccione Julius , Ge Liang , Tseng Elaine E.
Published: July 2012

Dilatation of the pulmonary autograft after the Ross procedure can lead to aortic insufficiency and/or aneurysmal pathology requiring reoperation. Autograft remodeling occurs as the autograft wall is exposed to systemic pressure and higher wall stresses, which have not been quantified in humans. The aim of the study was to develop a realistic Finite Element (FE) model of the human pulmonary autograft and to perform simulations at systemic pressure to quantify the increases in autograft wall stress immediately after the Ross procedure. Autograft geometry was generated from high-resolution micro-computed tomography images of an explanted human pulmonary root to create a mesh of hexahedral elements. Constitutive equations were used to describe the regional tissue material properties of the human pulmonary root obtained from bi-axial stretch testing. LS-DYNA (LSTC Inc., Livermore, CA) FE software was used to simulate cardiac cycles at pulmonary and systemic pressure. Autograft dilatation and wall stress distribution were determined. Correlation of LS-DYNA model material properties to actual tissue stress-strain data was performed to ensure model accuracy. Human autograft dilation from pulmonary to systemic pressure was minimal (32.1 to 33.4mm) due to the non-linearity of the material properties. Less compliance was demonstrated at greater wall stresses. Significant increases in autograft wall stresses were found at systemic pressures. Maximal wall stresses increased approximately 10-fold in diastole (12.4 to 122.3 kPa) and 5-fold in systole (48.1 to 234.2 kPa), relative to the wall stresses at pulmonary pressures. Pulmonary autograft wall stress increased by an order of magnitude at systemic pressure. Initial autograft dilation at systemic pressure was minimal as validated by clinical studies. Chronically elevated wall stress may lead to pathologic remodeling and aneurysmal formation over time. The correspondence of this model with future studies of post-dilated autografts will lead to an improved understanding of tissue remodeling, and offer necessary data for developing improvements to the Ross procedure.
Sea Fishing at Salango (Manabí Province, Ecuador) During the Middle Formative Machalilla Phase
Authors: Béarez Philippe , Gay Patrick , Lunniss Richard
Published: July 2012

Formative Ecuadorian coastal societies possessed the technologies and skills necessary not only for fishing, but also for deep-sea navigation. Although marine shellfish are acknowledged as significant both for dietary and religious purposes, the importance of sea-fishing is rarely highlighted or explored. In order to help evaluate more fully the significance of the sea in the Ecuadorian Formative, this paper presents recently studied evidence, excavated at the Salango site, in the Province of Manabí, for local off-shore tuna fishing during the Middle Formative Machalilla phase (ca. 1500–900 B.C.), where Scombrids constituted 80 percent of recovered fish remains. The results are then compared with those obtained from other Machalilla sites. Finally, data from the subsequent Late Formative Engoroy phase (ca. 900–100 B.C.) indicate that while Scombrid fishing continued to predominate at Salango, species capture changed through time, with a shift from yellowfin tuna in the Machalilla phase to black skipjacks in Engoroy times. It is suggested that the decline in tuna capture reflects not so much change in fishing strategy as change in the populations of fish species reaching the local marine environment.
Non‐Covalent Self‐Assembly and Covalent Polymerization Co‐Contribute to Polydopamine Formation
Authors: Hong Seonki , Na Yun Suk , Choi Sunghwan , Song In Taek , Kim Woo Youn , Lee Haeshin
Published: July 2012

Polydopamine is the first adhesive polymer that can functionalize surfaces made of virtually all material chemistries. The material‐independent surface modification properties of polydopamine allow the functionalization of various types of medical and energy devices. However, the mechanism of dopamine polymerization has not yet been clearly demonstrated. Covalent oxidative polymerization via 5,6‐dihydroxyindole (DHI), which is similar to the mechanism for synthetic melanin synthesis, has been the clue. Here, it is reported that a physical, self‐assembled trimer of (dopamine)2/DHI exists in polydopamine, which has been known to be formed only by covalent polymerization. It is also found that the trimeric complex is tightly entrapped within polydopamine and barely escapes from the polydopamine complex. The result explains the previously reported in vitro and in vivo biocompatibility. The study reveals a different perspective of polydopamine formation, where it forms in part by the self‐assembly of dopamine and DHI, providing a new clue toward understanding the structures of catecholamines such as melanin.
Lithium‐Air Batteries: Survey on the Current Status and Perspectives Towards Automotive Applications from a Battery Industry Standpoint
Authors: Park Myounggu , Sun Heeyoung , Lee Hyungbok , Lee Junesoo , Cho Jaephil
Published: July 2012

Li‐air rechargeable batteries theoretically have advantages from both secondary batteries and fuel cells, which can be viewed as the best technological blends for automotive applications resolving the so called mileage anxiety problem due to the limited driving range of electrical vehicles based upon Li‐ion batteries; this problem is rooted in the intrinsically small energy density of Li‐ion batteries. This very scientific trait of Li‐air batteries, which is apparently suited to the requirements of batteries for future electric vehicles, has induced quite a strong surge of research recently. This occurrence has motivated the authors to undertake a thorough review in an effort to understand the current status of Li‐air battery related technologies. A comprehensive survey from a battery industry standpoint has been conducted on the fundamentals of chemistry, utilized Li‐air cell configurations (or types) vs. performance, and major components comprising Li‐air batteries using various sources of previously published peer‐reviewed journal papers, book chapters, patents, and industrial reports. The survey results are presented here.
The effect of a booster intervention to promote hearing protection behavior in operating engineers
Authors: Hong OiSaeng , Chin Dal Lae , Fiola Lauren A. , Kazanis Anamaria S.
Published: July 2012

Background

Booster interventions may be useful in promoting workers' hearing protection device (HPD) use. Previous research on the effectiveness and the optimal timeframe for boosters is limited and inconsistent.

Methods

In this randomized controlled trial, 403 workers were assigned to receive an individually tailored booster, a control booster, or no booster. The booster intervention groups were further divided by timeframe. Frequency of HPD use was measured 12 months post‐intervention.

Results

The booster intervention groups significantly accounted for the variance in HPD use in Year 2. Workers in middle‐term booster (67–94 days) group reported a significantly greater increase in HPD use in Year 2 than those in other timeframes, when potential covariates were controlled for.

Conclusions

Booster intervention letters mailed between 67 and 94 days post‐initial intervention may promote HPD use among participants. Future research should explore additional factors associated with optimal booster design, including more innovative mobile and internet‐based approaches. Am. J. Ind. Med. 56:258–266, 2013. © 2011 Wiley Periodicals, Inc.
Estimating age from adult occlusal wear: A modification of the miles method
Authors: Gilmore Cassandra C. , Grote Mark N.
Published: July 2012

The Miles method of age estimation relies on molar wear to estimate age and is widely used in bioarcheological contexts. However, because the method requires physical seriation and a sample of subadults to estimate wear rates it cannot be applied to many samples. Here, we modify the Miles method by scoring occlusal wear and estimating molar wear rates from adult wear gradients in 311 hunter‐gatherers and provide formulae to estimate the error associated with each age estimate. A check of the modified method in a subsample (n = 22) shows that interval estimates overlap in all but one case with age categories estimated from traditional methods; this suggests that the modifications have not hampered the ability of the Miles method to estimate age even in heterogeneous samples. As expected, the error increases with age and in populations with smaller sample sizes. These modifications allow the Miles method to be applied to skeletal samples of adult crania that were previously only amenable to cranial suture age estimation, and importantly, provide a measure of uncertainty for each age estimate. Am J Phys Anthropol 149:181–192, 2012. © Wiley Periodicals, Inc.
When Unconscious Wishes Become Laws: Policing Memory and Identity in Israel and Palestine
Authors: Cohen Shuki J.
Published: July 2012

This paper proposes a psychoanalytically‐informed model for the policing of memory and identity in Israel and Palestine. Borrowing from both empirically‐ and clinically‐validated insights into psychopathology, it purports to account for the increasingly frequent attempts of the Israeli government to suppress alternatives to the extant Zionist narrative using legislative and administrative means. The model explains why, counter‐intuitively, these attempts to impose an idealized Zionist narrative have markedly increased in the past several years, at a time when Israel's military power, geographical expansion and economic prosperity are arguably at an all‐time high. Supported by examples from both world history and Israeli documents, the proposed model suggests a dynamic link between trauma, annihilation anxiety, hyper‐vigilance and defensive behavior on a nation‐wide level, which runs as a leitmotiv in both Israeli government actions and in sentiments expressed by a considerable portion of Israeli society, from the foundation of the state of Israel in 1948. The model further uses psychodynamic insights to account for the cognitive‐emotional rigidity underpinning the discrepancy in the perceptions of reality between Israel's narrative and its sundry worldwide alternatives. The model posits that this growing discrepancy between worldwide public opinion and Israeli internal reasoning may underlie the growing disapproval and isolation of Israel in both diplomatic circles and world media. Finally, following psychodynamically‐oriented therapeutic practices, the explanatory power of the model is harnessed to suggest potentially effective remedial attitudes and interventions, whose mutative powers may be of use to psychodynamically‐trained mental health professionals who may be involved in future reconciliation efforts between Israel and Palestine. Copyright © 2012 John Wiley & Sons, Ltd.
Applied Psychoanalysis in Biblical Interpretation
Authors: Peri Tuvia
Published: July 2012

Applied psychoanalysis is the term used to denote the practice of applying psychoanalytic theoretical concepts to a variety of social and cultural domains. This work focuses on the application of psychoanalytic concepts to the understanding of great works of art or literature. Despite its inherent limitations stemming from the lack of an actual meeting with the patient, character analyses of heroes of dramatic plays or art works is part of the history of psychoanalysis. This work argues for the possible advantages of the use of psychoanalysis to the understanding of enigmatic behaviors of biblical central figures. Ancient rabbinic midrash is suggested as an auxiliary tool that help to uncover unconscious layers of the biblical text. This work uses the enigmatic story of the binding of Isaac and the exegetical midrash about it as an example for such a work. A clinical vignette demonstrates the potential utilization of this analytic interpretation for clinical work and for the teaching of complicated psychoanalytic concepts. Copyright © 2012 John Wiley & Sons, Ltd.
Lack of Tir Ubiquitylation Contributes to Enteropathogenic E. coli Remaining Extracellular During Nonphagocytic Cell Infections
Authors: Lin Ann E. , Guttman Julian A.
Published: July 2012

Enteropathogenic Escherichia coli (EPEC) is an extracellular pathogen that alters many host subcellular components during its infectious processes. We have previously shown that EPEC hijacks a large assortment of host cell endocytic components and uses these proteins to form protruding structures called “pedestals” rather than triggering internalization of the bacteria. Other invasive pathogens that also recruit similar endocytic components have been shown to enter their host cells on the ubiquitylation of their host cell receptors. Therefore, we hypothesize that EPEC remains extracellular by maintaining its receptor, translocated intimin receptor (Tir), in an unubiquitylated state. Using immunoprecipitation‐Western blots, we demonstrate no association of ubiquitin with Tir. To further elucidate the effect Tir ubiquitylation would have on EPEC during their infections, we engineered Tir‐ubiquitin fusion constructs, expressed them in host epithelial cells, and infected them with Δtir EPEC. We found these cells induced a significant increase in EPEC invasion as compared with cells that expressed the Tir construct that lacked ubiquitin conjugation. Our results indicate that the lack of EPEC receptor ubiquitylation is a contributing factor that these microbes use to prevent their internalization into epithelial cells. Anat Rec, 2012. © 2012 Wiley Periodicals, Inc.
A Distributed State Estimation Approach to Condition Monitoring of Nonlinear Electric Power Systems
Authors: Rigatos G. , Siano P. , Zervos N.
Published: July 2012

This paper analyzes distributed state estimation methods for condition monitoring of electric power transmission and distribution systems. When a fault occurs in such large‐scale systems, it is usually difficult to detect it and to determine its exact position. Moreover, due to the cost of installation and maintenance of measurement devices and due to the excessive size of the electric power grid, the complete monitoring of the associated infrastructure is impractical. Therefore, to monitor the condition of the power grid, some form of estimation is required. As suitable approaches for distributed state estimation this paper proposes the extended information filter (EIF) and the unscented information filter (UIF). The Extended Information Filter is actually an implementation of distributed extended Kalman filtering while the unscented information filter is an implementation of distributed unscented Kalman filtering. With the use of the aforementioned filtering algorithms on processing units located at different parts of the power grid, one can produce local estimates of the system's state vector which in turn can be fused into an aggregate state estimation. The produced global state estimate enables continuous monitoring of the condition of the electric power system and early fault diagnosis if used by a suitable fault detection and isolation algorithm.
Rational Approximation and Analog Realization of Fractional Order Transfer Function with Multiple Fractional Powered Terms
Authors: Khanra M. , Pal J. , Biswas K.
Published: July 2012

The concept of model order reduction of high integer order systems using approximate moment matching has been generalized and extended to rational approximation of fractional order transfer function (FOTF). This approximation method converts an FOTF into characteristic numeric values and thus can be applied to rational approximation of FOTF, irrespective of its form of representation. This is very effective when FOTF has multiple fractional powered terms. Two circuit configurations for analog realization of FOTF through rational approximation are discussed and have been shown to be useful in hardware implementation. As a case study, a half order differentiator has been implemented in hardware and can be extended to any FOTF.
An amperometric enzyme biosensor for real‐time measurements of cellobiohydrolase activity on insoluble cellulose
Authors: Cruys‐Bagger Nicolaj , Ren Guilin , Tatsumi Hirosuke , Baumann Martin J. , Spodsberg Nikolaj , Andersen Heidi Delcomyn , Gorton Lo , Borch Kim , Westh Peter
Published: July 2012

An amperometric enzyme biosensor for continuous detection of cellobiose has been implemented as an enzyme assay for cellulases. We show that the initial kinetics for cellobiohydrolase I, Cel7A from Trichoderma reesei, acting on different types of cellulose substrates, semi‐crystalline and amorphous, can be monitored directly and in real‐time by an enzyme‐modified electrode based on cellobiose dehydrogenase (CDH) from Phanerochaete chrysosporium (Pc). PcCDH was cross‐linked and immobilized on the surface of a carbon paste electrode which contained a mediator, benzoquinone. An oxidation current of the reduced mediator, hydroquinone, produced by the CDH‐catalyzed reaction with cellobiose, was recorded under constant‐potential amperometry at +0.5 V (vs. Ag/AgCl). The CDH‐biosensors showed high sensitivity (87.7 µA mM−1 cm−2), low detection limit (25 nM), and fast response time (t95% ∼ 3 s) and this provided experimental access to the transient kinetics of cellobiohydrolases acting on insoluble cellulose. The response from the CDH‐biosensor during enzymatic hydrolysis was corrected for the specificity of PcCDH for the β‐anomer of cello‐oligosaccharides and the approach were validated against HPLC. It is suggested that quantitative, real‐time data on pure insoluble cellulose substrates will be useful in attempts to probe the molecular mechanism underlying enzymatic hydrolysis of cellulose. Biotechnol. Bioeng. 2012; 109: 3199–3204. © 2012 Wiley Periodicals, Inc.
Automated dynamic fed‐batch process and media optimization for high productivity cell culture process development
Authors: Lu Franklin , Toh Poh Choo , Burnett Iain , Li Feng , Hudson Terry , Amanullah Ashraf , Li Jincai
Published: July 2012

Current industry practices for large‐scale mammalian cell cultures typically employ a standard platform fed‐batch process with fixed volume bolus feeding. Although widely used, these processes are unable to respond to actual nutrient consumption demands from the culture, which can result in accumulation of by‐products and depletion of certain nutrients. This work demonstrates the application of a fully automated cell culture control, monitoring, and data processing system to achieve significant productivity improvement via dynamic feeding and media optimization. Two distinct feeding algorithms were used to dynamically alter feed rates. The first method is based upon on‐line capacitance measurements where cultures were fed based on growth and nutrient consumption rates estimated from integrated capacitance. The second method is based upon automated glucose measurements obtained from the Nova Bioprofile FLEX® autosampler where cultures were fed to maintain a target glucose level which in turn maintained other nutrients based on a stoichiometric ratio. All of the calculations were done automatically through in‐house integration with a Delta V process control system. Through both media and feed strategy optimization, a titer increase from the original platform titer of 5 to 6.3 g/L was achieved for cell line A, and a substantial titer increase of 4 to over 9 g/L was achieved for cell line B with comparable product quality. Glucose was found to be the best feed indicator, but not all cell lines benefited from dynamic feeding and optimized feed media was critical to process improvement. Our work demonstrated that dynamic feeding has the ability to automatically adjust feed rates according to culture behavior, and that the advantage can be best realized during early and rapid process development stages where different cell lines or large changes in culture conditions might lead to dramatically different nutrient demands. Biotechnol. Bioeng. 2013; 110: 191–205. © 2012 Wiley Periodicals, Inc.
Outdoor cultivation of temperature‐tolerant Chlorella sorokiniana in a column photobioreactor under low power‐input
Authors: Béchet Quentin , Muñoz Raul , Shilton Andy , Guieysse Benoit
Published: July 2012

Temperature‐tolerant Chlorella sorokiniana was cultivated in a 51‐L column photobioreactor with a 1.1 m2 illuminated area. The reactor was operated outdoors under tropical meteorological conditions (Singapore) without controlling temperature and the culture was mixed at a power input of 7.5 W/m3 by sparging CO2‐enriched air at 1.2 L/min (gas hold‐up of 0.02). Biomass productivity averaged 10 ± 2.2 g/${\rm m}_{{\rm illuminated}\,{\rm area}}^{{\rm 2}} {\rm \hbox{-} day}$ over six batch studies, yielding an average photosynthetic efficiency (PE) of 4.8 ± 0.5% of the total solar radiation (P = 0.05, N = 6). This demonstrates that temperature‐tolerant microalgae can be cultivated at high PE under a mixing input sevenfold to ninefold lower than current operational guidelines (50–70 W/m3) and without the need for temperature control (the culture broth temperature reached 41°C during operation). In this study, the PE value was determined based on the amount of solar radiation actually reaching the algae and this amount was estimated using a mathematical model fed with onsite solar irradiance data. This determination was found to be particularly sensitive to the value of the atmospheric diffusion coefficient, which generated a significant uncertainty in the PE calculation. The use of the mathematical model, however, confirmed that the vertical reactor geometry supported efficient photosynthesis by reducing the duration and intensity of photoinhibition events. The model also revealed that all three components of direct, diffuse, and reflected solar radiation were quantitatively important for the vertical column photobioreactor, accounting for 14%, 65%, and 21% of the total solar radiation reaching the culture, respectively. The accurate prediction of the discrete components of solar radiation reaching the algae as a function of climatic, geographic, and design parameters is therefore crucial to optimize the individual reactor geometry and the layout/spacing between the individual reactors in a reactor farm. Biotechnol. Bioeng. 2013; 110: 118–126. © 2012 Wiley Periodicals, Inc.
Simultaneous quantification of metformin and glipizide in human plasma by high‐performance liquid chromatography–tandem mass spectrometry and its application to a pharmacokinetic study
Authors: Li Ning , Deng Ying , Qin Feng , Yu Jia , Li Famei
Published: July 2012

A selective, sensitive and rapid high‐performance liquid chromatography–tandem mass spectrometry (HPLC‐MS/MS) method was developed and validated to determine metformin and glipizide simultaneously in human plasma using phenacetin as internal standard (IS). After one‐step protein precipitation of 200 μL plasma with methanol, metformin, glipizide and IS were separated on a Kromasil Phenyl column (4.6 × 150 mm, 5 µm) at 40°C with an isocratic mobile phase consisting of methanol–10 mmol/L ammonium acetate (75:25, v/v) at a flow rate of 0.35 mL/min. Electrospray ionization source was applied and operated in the positive mode. Multiple reaction monitoring using the precursor → product ion combinations of m/z 130 → m/z 71, m/z 446 → m/z 321 and m/z 180 → m/z 110 were used to quantify metformin, glipizide and IS, respectively. The linear calibration curves were obtained over the concentration ranges 4.10–656 ng/mL for metformin and 2.55–408 ng/mL for glipizide. The relative standard deviation of intra‐day and inter‐day precision was below 10% and the relative error of accuracy was between −7.0 and 4.6%. The presented HPLC‐MS/MS method was proved to be suitable for the pharmacokinetic study of metformin hydrochloride and glipizide tablets in healthy volunteers after oral administration. Copyright © 2012 John Wiley & Sons, Ltd.
Carnosine prevents necrotic and apoptotic death of rat thymocytes via ouabain‐sensitive Na/K‐ATPase
Authors: Smolyaninova Larisa V. , Dergalev Alexander A. , Kulebyakin Konstantin Y. , Carpenter David O. , Boldyrev Alexander A.
Published: July 2012

It is known that ouabain, a selective inhibitor of Na/K‐ATPase, not only can cause the activation of signal cascades, which regulate the cell viability, but also can cause the accumulation of free radicals, which can evoke the oxidative stress. We have shown that the nanomolar concentrations of ouabain result in the temporary increase in the level of intracellular free radicals, but the millimolar concentration of ouabain induces a stable intracellular accumulation of free radicals in rat thymocytes. The increasing level of free radicals resulting from both low and high concentrations of ouabain can be attenuated by the antioxidant, carnosine. Moreover, the long‐term incubation with ouabain leads to the cell death by necrosis and apoptosis. Ouabain‐mediated apoptosis and necrosis were also abolished by carnosine. Copyright © 2012 John Wiley & Sons, Ltd.
A Modified Approach for Generating Column Grand Composite Curves
Authors: Wei Z. , Wu S. , Zhang B. , Chen Q. , Hui C.‐W.
Published: July 2012

In generating a column grand composite curve (CGCC) the enthalpy values calculated by top‐down and bottom‐up approaches are different at the feed stage. An insight analysis of material and energy balances indicates that these differences are due to the feed stage treated as a stripping stage in the top‐down approach and as a rectifying stage in the bottom‐up approach. To consider the practical feed process and overcome the shortcomings in the existing CGCC generation approaches, a modified approach is proposed, dividing the column into three sections: the rectifying section (from the 1st to the f‐2th stage, the condenser being the 1st stage), the feeding section (the f‐1th and the fth stage) and the stripping section (from the f+1th to the nth stage, the reboiler being the nth stage). The modification is mainly focused on the feeding section stages. An indicator, the reflux energy‐saving potential rate, is defined and used to evaluate these approaches. A benzene‐toluene column is investigated to demonstrate the performance of the proposed approach, and a comparison is made with Aspen Plus Column Targeting. The results show that the modified approach is able to eliminate the ambiguity at the feed stage. Compared to Aspen Plus Column Targeting and the existing approaches, the modified approach could give better results.
A Nucleobase‐Discriminating Pyrrolo‐dC Click Adduct Designed for DNA Fluorescence Mismatch Sensing
Authors: Ming Xin , Seela Frank
Published: July 2012

New pyrrolo‐dC click adducts (4 and 5) tethered with a 1,2,3‐triazole skeleton were synthesized and oligonucleotides were prepared. The triazole system was either directly linked to the pyrrolo moiety (5) or connected via an n‐butyl linker (4). The quantum yield of nucleoside 5 (Φ=0.32), which is 10 times higher than those of 8‐methylpyrrolo‐dC (1 b, Φ=0.026) or the long linker derivative 4 (Φ=0.03), is maintained in oligonucleotides. Compound 5 was used as a nucleobase‐discriminating fluorescence sensor in duplex DNA. Excellent mismatch discrimination was observed when 5 was positioned opposite the four canonical nucleosides. Compound 5 has the potential to be used for SNP detection in long DNA targets when conventional techniques such as high resolution melt analysis fail.
High Relaxivity Supramolecular Adducts Between Human‐Liver Fatty‐Acid‐Binding Protein and Amphiphilic GdIII Complexes: Structural Basis for the Design of Intracellular Targeting MRI Probes
Authors: D'Onofrio Mariapina , Gianolio Eliana , Ceccon Alberto , Arena Francesca , Zanzoni Serena , Fushman David , Aime Silvio , Molinari Henriette , Assfalg Michael
Published: July 2012

Gadolinium complexes linked to an apolar fragment are known to be efficiently internalized into various cell types, including hepatocytes. Two lipid‐functionalized gadolinium chelates have been investigated for the targeting of the human liver fatty acid binding protein (hL‐FABP) as a means of increasing the sensitivity and specificity of intracellular‐directed MRI probes. hL‐FABP, the most abundant cytosolic lipid binding protein in hepatocytes, displays the ability to interact with multiple ligands involved in lipid signaling and is believed to be an obligate carrier to escort lipidic drugs across the cell. The interaction modes of a fatty acid and a bile acid based gadolinium complex with hL‐FABP have been characterized by relaxometric and NMR experiments in solution with close‐to‐physiological protein concentrations. We have introduced the analysis of paramagnetic‐induced protein NMR signal intensity changes as a quantitative tool for the determination of binding stoichiometry and of precise metal‐ion‐center positioning in protein–ligand supramolecular adducts. A few additional NMR‐derived restraints were then sufficient to locate the ligand molecules in the protein binding sites by using a rapid data‐driven docking method. Relaxometric and 13C NMR competition experiments with oleate and the gadolinium complexes revealed the formation of heterotypic adducts, which indicates that the amphiphilic compounds may co‐exist in the protein cavity with physiological ligands. The differences in adduct formation between fatty acid and bile acid based complexes provide the basis for an improved molecular design of intracellular targeted probes.
Strong Aphicidal Activity of GlcNAc(β1→4)Glc Disaccharides: Synthesis, Physiological Effects, and Chitinase Inhibition
Authors: Dussouy Christophe , Bultel Laurent , Saguez Julien , Cherqui Anas , Khelifa Mounia , Grand Eric , Giordanengo Philippe , Kovensky José
Published: July 2012

The synthesis of four GlcNAc(β1→4)Glc disaccharides containing 2‐O‐acetyl and/or 6‐sulfate groups was performed in high yields with total 1,2‐trans stereoselectivity. These disaccharides were evaluated as candidates for insect chitinase inhibition and aphicidal activity. All the compounds prepared displayed physiological effects on M. persicae aphids; however, the inhibition of chitinases of different sources (bacteria, fungus, and aphid) followed different patterns according to subtle structural characteristics.
ChemInform Abstract: Hierarchical Porous NiCo2O4 Nanowires for High‐Rate Supercapacitors.
Authors: Jiang Hao , Ma Jan , Li Chunzhong
Published: July 2012

The title nanowires are synthesized from mixtures of Ni(NO3)2, Co(NO3)2, H2C2O4, P123, EtOH, and polyethylene glycol at room temperature (10 min) followed by annealing the obtained precipitates under air at 250 °C for 3 h.
ChemInform Abstract: Rapid Microwave Hydrothermal Synthesis of ZnGa2O4 with High Photocatalytic Activity Toward Aromatic Compounds in Air and Dyes in Liquid Water.
Authors: Sun Meng , Li Danzhen , Zhang Wenjuan , Chen Zhixin , Huang Hanjie , Li Wenjuan , He Yunhui , Fu Xianzhi
Published: July 2012

The title compound is synthesized by a microwave hydrothermal method from an aqueous solution of Ga(NO3)3, ZnCl2, and ammonia (microwave system, 200 W, 2.45 GHz, autoclave, 160 °C, 20 min).
ChemInform Abstract: Dumbbells of Five‐Connected Ge Atoms and Superconductivity in CaGe3.
Authors: Schnelle Walter , Ormeci Alim , Wosylus Aron , Meier Katrin , Grin Yuri , Schwarz Ulrich
Published: July 2012

CaGe3 is synthesized by annealing mixtures of CaGe2 (obtained by reacting the elements in an arc furnace) and Ge at 10 GPa and 1200 K for 1 h.
ChemInform Abstract: A Strategy for the Synthesis of Mesostructured Metal Oxides with Lower Oxidation States.
Authors: Tueysuez Harun , Weidenthaler Claudia , Schueth Ferdi
Published: July 2012

Ordered mesoporous CoO and Fe3O4 are synthesized by reduction of Co3O4 and ferrihydrite (Fe2O3·nH2O), respectively, using glycerol/water vapor as a reducing agent (320 °C, 2 h).