bluuebunny/crossref_metadata_2025 huggingface dataset:
Crystal structure of nitrogen regulatory protein IIANtr from Neisseria meningitidis
Authors: Ren Jingshan , Sainsbury Sarah , Berrow Nick S , Alderton David , Nettleship Joanne E , Stammers David K , Saunders Nigel J , Owens Raymond J
Published: August 2005

Background

The NMB0736 gene of Neisseria meningitidis serogroup B strain MC58 encodes the putative nitrogen regulatory protein, IIANtr (abbreviated to NM-IIANtr). The homologous protein present in Escherichia coli is implicated in the control of nitrogen assimilation. As part of a structural proteomics approach to the study of pathogenic Neisseria spp., we have selected this protein for structure determination by X-ray crystallography.

Results

The NM-IIANtr was over-expressed in E. coli and was shown to be partially mono-phosphorylated, as assessed by mass spectrometry of the purified protein.

Crystals of un-phosphorylated protein were obtained and diffraction data collected to 2.5 Å resolution. The structure of NM-IIANtr was solved by molecular replacement using the coordinates of the E. coli nitrogen regulatory protein IIAntr [PDB: 1A6J] as the starting model. The overall fold of the Neisseria enzyme shows a high degree of similarity to the IIANtr from E. coli, and the position of the phosphoryl acceptor histidine residue (H67) is conserved. The orientation of an adjacent arginine residue (R69) suggests that it may also be involved in coordinating the phosphate group. Comparison of the structure with that of E. coli IIAmtl complexed with HPr [PDB: 1J6T] indicates that NM-IIANtr binds in a similar way to the HPr-like enzyme in Neisseria.

Conclusion

The structure of NM-IIANtr confirms its assignment as a homologue of the IIANtr proteins found in a range of other Gram-negative bacteria. We conclude that the NM- IIANtr protein functions as part of a phosphorylation cascade which, in contrast to E. coli, shares the upstream phosphotransfer protein with the sugar uptake phosphoenolpyruvate:sugar phosphotransferase system (PTS), but in common with E. coli has a distinct downstream effector mechanism.
Effect on survey response rate of hand written versus printed signature on a covering letter: randomised controlled trial [ISRCTN67566265]
Authors: McKenzie-McHarg Kirstie , Tully Lucy , Gates Simon , Ayers Sarah , Brocklehurst Peter
Published: August 2005

Background

It is important that response rates to postal surveys are as high as possible to ensure that the results are representative and to maximise statistical power. Previous research has suggested that any personalisation of approach helps to improve the response rate. This experiment tested whether personalising questionnaires by hand signing the covering letter improved the response rate compared with a non-personalised group where the investigator's signature on the covering letter was scanned into the document and printed.

Methods

Randomised controlled trial. Questionnaires about surgical techniques of caesarean section were mailed to 3,799 Members and Fellows of the Royal College of Obstetricians and Gynaecologists resident in the UK. Individuals were randomly allocated to receive a covering letter with either a computer printed signature or a hand written signature. Two reminders were sent to non-respondents. The outcome measures were the proportion of questionnaires returned and their time to return.

Results

The response rate was 79.1% (1506/1905) in the hand-signed group and 78.4% (1484/1894) in the scanned and printed signature group. There was no detectable difference between the groups in response rate or time taken to respond.

Conclusion

No advantage was detected to hand signing the covering letter accompanying a postal questionnaire to health professionals.
Menstrual function among women exposed to polybrominated biphenyls: A follow-up prevalence study
Authors: Davis Stephanie I , Blanck Heidi Michels , Hertzberg Vicki S , Tolbert Paige E , Rubin Carol , Cameron Lorraine L , Henderson Alden K , Marcus Michele
Published: August 2005

Background

Alteration in menstrual cycle function is suggested among rhesus monkeys and humans exposed to polybrominated biphenyls (PBBs) and structurally similar polychlorinated biphenyls (PCBs). The feedback system for menstrual cycle function potentially allows multiple pathways for disruption directly through the hypothalamic-pituitary-ovarian axis and indirectly through alternative neuroendocrine axes.

Methods

The Michigan Female Health Study was conducted during 1997–1998 among women in a cohort exposed to PBBs in 1973. This study included 337 women with self-reported menstrual cycles of 20–35 days (age range: 24–56 years). Current PBB levels were estimated by exponential decay modeling of serum PBB levels collected from 1976–1987 during enrollment in the Michigan PBB cohort. Linear regression models for menstrual cycle length and the logarithm of bleed length used estimated current PBB exposure or enrollment PBB exposure categorized in tertiles, and for the upper decile. All models were adjusted for serum PCB levels, age, body mass index, history of at least 10% weight loss in the past year, physical activity, smoking, education, and household income.

Results

Higher levels of physical activity were associated with shorter bleed length, and increasing age was associated with shorter cycle length. Although no overall association was found between PBB exposure and menstrual cycle characteristics, a significant interaction between PBB exposures with past year weight loss was found. Longer bleed length and shorter cycle length were associated with higher PBB exposure among women with past year weight loss.

Conclusion

This study suggests that PBB exposure may impact ovarian function as indicated by menstrual cycle length and bleed length. However, these associations were found among the small number of women with recent weight loss suggesting either a chance finding or that mobilization of PBBs from lipid stores may be important. These results should be replicated with larger numbers of women exposed to similar lipophilic compounds.
Reduced expression of multiple gap junction proteins is a feature of cervical dysplasia
Authors: Aasen Trond , Graham Sheila V , Edward Mike , Hodgins Malcolm B
Published: August 2005

Cervical dysplasia is a premalignant lesion associated with human papillomavirus (HPV) infection which, over time, can turn cancerous. Previous studies have indicated that loss of gap junctions may be a feature of cervical cancer and premalignant dysplasia. Loss of the gap junction protein connexin43 has been demonstrated in dysplastic cervix, but other connexins have not been investigated. In contrast we previously showed that HPV-associated cutaneous warts – and other hyperproliferative skin conditions – display a dramatic upregulation of certain connexins, in particular connexin26. By performing immunofluorescence staining after antigen retrieval of paraffin-embedded cervical tissue samples, this study reports for the first time that connexin26 and connexin30, in addition to connexin43, are expressed in differentiating cells of normal human cervical epithelia. Moreover, in dysplastic ectocervix, all connexins studied display a dramatic loss of expression compared to adjacent normal epithelia. The role of connexins in keratinocyte differentiation and carcinogenesis is discussed.
Proanthocyanidins, from Ribes nigrum leaves, reduce endothelial adhesion molecules ICAM-1 and VCAM-1
Authors: Garbacki N , Kinet M , Nusgens B , Desmecht D , Damas J
Published: August 2005

Background

The effects of proanthocyanidins (PACs), isolated from blackcurrant (Ribes nigrum L.) leaves, on neutrophil accumulation during inflammatory processes were investigated in vivo and in vitro.

Methods

In vivo studies were performed using carrageenin-induced pleurisy in rats pre-treated with PACs. Exudate volume and PMNs accumulation were measured. Leukocyte cell adhesion molecules (LFA-1, Mac-1 and VLA-4) mobilization in circulating granulocytes were analysed by flow cytometry and endothelial cell adhesion molecules (ICAM-1 and VCAM-1) were detected by immunohistochemistry on lung sections.

In vitro studies were conducted on endothelial LT2 cells, stimulated with TNF-α, to evaluate ICAM-1, IL-8 and VEGF mRNA expression upon PACs treatment.

Data sets were examined by one-way analysis of variance (ANOVA) followed by a Scheffe post-hoc test.

Results

Pretreatment of the animals with PACs (10, 30 and 60 mg/kg) inhibited dose-dependently carrageenin-induced pleurisy in rats by reducing pleural exudate formation and PMNs infliltration. Leukocyte cell adhesion molecules mobilization was not down-regulated on granulocytes by PACs. Immunohistochemistry on lung sections showed a decreased production of endothelial cell adhesion molecules.

In vitro experiments demonstrated that PACs were able to significantly inhibit ICAM-1 but not IL-8 and VEGF165 mRNA expression. Moreover, VEGF121 mRNA expression was dose-dependently enhanced.

Conclusion

This study provides evidence to support the anti-inflammatory activity of proanthocyanidins is related to an inhibition of leukocyte infiltration which can be explained at least in part by a down-regulation of endothelial adhesion molecules, ICAM-1 and VCAM-1 and that these compounds are capable of modulating TNF-α-induced VEGF transcription.
The cost-effectiveness of neonatal screening for Cystic Fibrosis: an analysis of alternative scenarios using a decision model
Authors: Simpson Neil , Anderson Rob , Sassi Franco , Pitman Alexandra , Lewis Peter , Tu Karen , Lannin Heather
Published: August 2005

Background

The use of neonatal screening for cystic fibrosis is widely debated in the United Kingdom and elsewhere, but the evidence available to inform policy is limited. This paper explores the cost-effectiveness of adding screening for cystic fibrosis to an existing routine neonatal screening programme for congenital hypothyroidism and phenylketonuria, under alternative scenarios and assumptions.

Methods

The study is based on a decision model comparing screening to no screening in terms of a number of outcome measures, including diagnosis of cystic fibrosis, life-time treatment costs, life years and QALYs gained. The setting is a hypothetical UK health region without an existing neonatal screening programme for cystic fibrosis.

Results

Under initial assumptions, neonatal screening (using an immunoreactive trypsin/DNA two stage screening protocol) costs £5,387 per infant diagnosed, or £1.83 per infant screened (1998 costs). Neonatal screening for cystic fibrosis produces an incremental cost-effectiveness of £6,864 per QALY gained, in our base case scenario (an assumed benefit of a 6 month delay in the emergence of symptoms). A difference of 11 months or more in the emergence of symptoms (and mean survival) means neonatal screening is both less costly and produces better outcomes than no screening.

Conclusion

Neonatal screening is expensive as a method of diagnosis. Neonatal screening may be a cost-effective intervention if the hypothesised delays in the onset of symptoms are confirmed. Implementing both antenatal and neonatal screening would undermine potential economic benefits, since a reduction in the birth incidence of cystic fibrosis would reduce the cost-effectiveness of neonatal screening.
Fast, visual specialization for reading in English revealed by the topography of the N170 ERP response
Authors: Maurer Urs , Brandeis Daniel , McCandliss Bruce D
Published: August 2005

Background

N170 effects associated with visual words may be related to perceptual expertise effects that have been demonstrated for faces and other extensively studied classes of visual stimuli. Although face and other object expertise effects are typically bilateral or right-lateralized, the spatial topography of reading-related N170 effects are often left-lateralized, providing potential insights into the unique aspects of reading-related perceptual expertise.

Methods

Extending previous research in German [1], we use a high-density channel array to characterize the N170 topography for reading-related perceptual expertise in English, a language with inconsistent spelling-to-sound mapping. N170 effects related to overall reading-related expertise are defined by contrasting responses to visual words versus novel symbol strings. By contrasting each of these conditions to pseudowords, we examined how this reading-related N170 effect generalizes to well-ordered novel letter strings.

Results

A sample-by-sample permutation test computed on word versus symbol ERP topographies revealed differences during two time windows corresponding to the N170 and P300 components. Topographic centroid analysis of the word and symbol N170 demonstrated significant differences in both left-right as well as inferior-superior dimensions. Words elicited larger N170 negativities than symbols at inferior occipito-temporal channels, with the maximal effect over left inferior regions often unsampled in conventional electrode montages. Further contrasts produced inferior-superior topographic effects for the pseudoword-symbol comparison and left-lateralized topographic effects for the word-pseudoword comparison.

Conclusion

Fast specialized perception related to reading experience produces an N170 modulation detectable across different EEG systems and different languages. Characterization of such effects may be improved by sampling with greater spatial frequency recordings that sample inferior regions. Unlike in German, reading-related expertise effects in English produced only partial generalization in N170 responses to novel pseudowords. The topographic inferior-superior N170 differences may reflect general perceptual expertise for orthographic strings, as it was found for words and pseudowords across both languages. The topographic left-right N170 difference between words and pseudowords was only found in English, and may suggest that ambiguity in pronunciating novel pseudowords due to inconsistency in spelling-to-sound mapping influences early stages of letter string processing.
Preface: The Cellular Communications Maze
Authors: SIDEMAN SAMUEL
Published: August 2005

Abstract: Inter‐ and intracellular ionic and molecular communications are indispensable to the preservation of life and all organic functions. The cell constantly responds to a myriad of extracellular ionic and molecular signals, and cell behavior in single‐cell or multicellular organisms is coordinated by these signals moving into, inside, or between the cells. The signals pass through the cell's phospholipidic plasma membrane by diffusion and, mostly, via protein transporters: gates, receptors, and/or ion pumps imbedded in the membrane. Each signaling pathway is a complex cause and effect chain of events involving intricate networks of interactions. For example, the extracellular signals, which are monitored by the cell's membrane cognate receptors, form ligand‐receptor complexes. These are typically amplified by interaction with a coupling protein, mostly a G protein, and are diversified via intracellular signal transductions, either directly or via the activation of intracellular second messengers. The present volume focuses on signal‐induced interactions that trigger specific responses of the effector system. Highlighting the cellular communication framework and the major parameters encountered in these complex interactive phenomena may help to familiarize the uninitiated with the complex phenomena involved in sustaining life.
From Organ to Molecules: Steps and Consequences
Authors: BEYAR RAFAEL
Published: August 2005

Abstract: The classic cardiac research programs revolved around measurable properties such as pressures, work done, vascular blood flow, electrical propagation, and other such parameters that defined the global heart functions in health and decease. Consistently, the first Henry Goldberg Workshop, held in Haifa in 1984, focused on the interactions between cardiac mechanics, electrical activation, perfusion, and metabolism of the whole heart. Questions focused on the macroscale cardiac function and performance. These studies involved engineering science, simulation, and modeling tools that were essential for the understanding of the complex interactions within the cardiac system. Three‐dimensional imaging, ventricular structure, fiber mechanics, circulation and cardiovascular flow, electrical propagation, and blood pumping were all major foci of research at that time. However, it was soon obvious that in order to understand organ level characteristics, one must explore the complex cellular and intracellular control mechanisms; these became the foci of our subsequent workshops. Better understanding of organ level performance required integrated studies of organ and tissue structure and function with genetic, molecular, and cellular characteristics, including cellular communication and ionic and molecular signaling. Analysis of the cardiac system thus depends on continuous probing of the heart system with modern measurement techniques and on integrating data and acquired knowledge with analytical models, constantly evolving to match reality.
The Miscommunicative Cardiac Cell: When Good Proteins Go Bad
Authors: GOMES ALDRIN V. , VENKATRAMAN GAYATHRI , POTTER JAMES D.
Published: August 2005

Abstract: Troponin (Tn) is made up of three subunits, troponin T (TnT), troponin I (TnI), and troponin C (TnC). In cardiac muscle, TnI can exist as two isoforms, slow skeletal TnI (ssTnI) or cardiac TnI (cTnI), whereas TnT occurs as multiple isoforms. The predominant form of TnI in fetal cardiac muscle is ssTnI, which is derived from a different gene than cTnI. However, the predominant form of cardiac TnT (cTnT) in fetal muscle is cTnT1, which is derived from the same gene that produces the adult cTnT isoform (cTnT3). Fetal cardiac muscle is more sensitive to Ca2+ than adult muscle and this may be due in part to the fetal cTnT1 and ssTnI isoforms. cTnT1 and/or ssTnI by themselves cause a significant increase in Ca2+ sensitivity when compared to cTnT3 and/or cTnI. Mutations in the gene for cTnT can cause hypertrophic cardiomyopathy or dilated cardiomyopathy (DCM). Investigation of DCM mutations in the fetal cTnT1 isoform showed that the cTnT isoform is an important determinant of the effect of the mutation. The TnI isoform also affects the physiological function of the cardiac muscle. The presence of both the fetal TnT isoform, containing a DCM mutation, and ssTnI results in larger changes in Ca2+ sensitivity than the same DCM mutant in the adult TnT isoform and in the presence of cTnI (when compared to their respective wild‐type TnT controls). These recent results suggest that some mutations may have different severities in fetal and adult hearts.
Functional Properties of Human Embryonic Stem Cell‐Derived Cardiomyocytes
Authors: DOLNIKOV KATYA , SHILKRUT MARK , ZEEVI‐LEVIN NAAMA , DANON A.AF , GERECHT‐NIR SHARON , ITSKOVITZ‐ELDOR JOSEPH , BINAH OFER
Published: August 2005

Abstract: Regeneration of the diseased myocardium by cardiac cell transplantation is an attractive therapeutic modality. Yet, because the transplanted cardiomyocytes should functionally integrate within the diseased myocardium, it is preferable that their properties resemble those of the host. To determine the functional adaptability of human embryonic stem cell‐derived cardiomyocytes (hESC‐CM) to the host myocardium, the authors investigated the excitation‐contraction (E‐C) coupling and the responsiveness to common physiological stimuli. The main findings are: (1) hESC‐CM readily respond to electrical pacing and generate corresponding [Ca2+]i transients (measured by fura‐2 fluorescence) and contractions (measured by video edge detector). (2) In contrast to the mature myocardium, hESC‐CM display negative force‐frequency relations. (3) The hESC‐CM contraction is dependent on [Ca2+]o and blocked by verapamil. (4) Surprisingly, ryanodine, the sarcoplasmic‐endoplasmic reticulum Ca2+‐ATPase inhibitor thapsigargin, and caffeine do not affect the [Ca2+]i transient or contraction. Collectively, these results indicate that at the developmental stage of 45 to 60 days, the contraction is largely dependent on [Ca2+]o rather than on sarcoplasmic reticulum (SR) Ca2+ stores. The results show for the first time that the E‐C coupling properties of hESC‐CM differ from the adult myocardium, probably due to immature SR function. Based on these findings, genetic manipulation of hESC‐CM toward the adult myocardium should be considered.
The Assembly of Calcium Release Units in Cardiac Muscle
Authors: FRANZINI‐ARMSTRONG CLARA , PROTASI FELICIANO , TIJSKENS PIERRE
Published: August 2005

Abstract: Calcium release units (CRUs) are constituted of specialized junctional domains of the sarcoplasmic reticulum (jSR) that bear calcium release channels, also called ryanodine receptors (RyRs). In cardiac muscle, CRUs come in three subtypes that differ in geometry, but have common molecular components. Peripheral couplings are formed by a junction of the jSR with the plasmalemma; dyads occur where the jSR is associated with transverse (T)‐tubules; corbular SR is a jSR domain that is located within the cells and bears RyRs but does not associate with either plasmalemma or T‐tubules. Using transmission electron microscopy, this study followed the formation of CRUs and their accrual of four components: the L‐type channel dihydropyridine receptors (DHPRs) of plasmalemma/T‐tubules; the RyRs of jSR; triadin (Tr) and junctin (JnC), two homologous components of the jSR membrane; and calsequestrin (CSQ), the internal calcium binding proteins. During differentiation, peripheral couplings are formed first and the others follow. RyRs and DHPRs are targeted to subdomains of the CRUs that face each other and are acquired in a concerted manner. Overexpressions of either junction (JnC or Tr) and of CSQ, singly or in conjunction, shed light on the specific role of JnC in the structural development, organization, and maintenance of jSR cisternae and on the independent synthetic pathways and targeting of JnC and CSQ. In addition, the structural cues provided by the overexpression models allow us to define sequential steps in the synthetic pathway for JnC and CSQ and their targeting to the CRUs of differentiating myocardium.
Multimodality of Ca2+ Signaling in Rat Atrial Myocytes
Authors: MORAD MARTIN , JAVAHERI ASHKAN , RISIUS TIM , BELMONTE STEVE
Published: August 2005

Abstract: It has been suggested that the multiplicity of Ca2+ signaling pathways in atrial myocytes may contribute to the variability of its function. This article reports on a novel Ca2+ signaling cascade initiated by mechanical forces induced by “puffing” of solution onto the myocytes. Cai transients were measured in fura‐2 acetoxymethyl (AM) loaded cells using alternating 340‐ and 410‐nm excitation waves at 1.2 kHz. Pressurized puffs of bathing solutions, applied by an electronically controlled micro‐barrel system, activated slowly (∼300 ms) developing Cai transients that lasted 1,693 ± 68 ms at room temperature. Subsequent second and third puffs, applied at ∼20 s intervals activated significantly smaller or no Cai transients. Puff‐triggered Cai transients could be reactivated once again following caffeine (10 mM)‐induced release of Ca2+ from sarcoplasmic reticulum (SR). Puff‐triggered Cai transients were independent of [Ca2+]o, and activation of voltage‐gated Ca2+ or cationic stretch channels or influx of Ca2+ on Na+/Ca2+exchanger, because puffing solution containing no Ca2+, 10 μM diltiazem, 1 mM Cd2+, 5 mM Ni2+, or 100 μM Gd3+ failed to suppress them. Puff‐triggered Cai transients were enhanced in paced compared to quiescent myocytes. Electrically activated Cai transients triggered during the time course of puff‐induced transients were unaltered, suggesting functionally separate Ca2+ pools. Contribution of inositol 1,4,5‐triphosphate (IP3)‐gated or mitochondrial Ca2+ pools or modulation of SR stores by nitric oxide/nitric oxide synthase (NO/NOS) signaling were evaluated using 0.5 to 500 μM 2‐aminoethoxydiphenyl borate (2‐APB) and 0.1 to 1 μM carbonylcyanide‐p‐trifluoromethoxyphenylhydrazone (FCCP), and 1 mM Nω‐Nitro‐L‐arginine methyl ester (L‐NAME) and 7‐nitroindizole, respectively. Only FCCP appeared to significantly suppress the puff‐triggered Cai transients. It was concluded that neither Ca2+ influx nor depolarization was required for activation of this signaling pathway. These studies suggest that pressurized puffs of solutions activate a mechanically sensitive receptor, which signals in turn the release of Ca2+ from a limited Ca2+ store of mitochondria. How mechanical forces are sensed and transmitted to mitochondria to induce Ca2+ release and what role such a Ca2+ signaling pathway plays in the physiology or pathophysiology of the heart remain to be worked out.
Effects of Na+‐Ca2+ Exchange Expression on Excitation‐Contraction Coupling in Genetically Modified Mice
Authors: GOLDHABER JOSHUA I. , HENDERSON SCOTT A. , REUTER HANNES , POTT CHRISTIAN , PHILIPSON KENNETH D.
Published: August 2005

Abstract: We have created genetically altered mice to investigate how expression of the Na+‐Ca2+ exchange protein alters excitation‐contraction (E‐C) coupling. Whereas low levels of exchanger overexpression have minimal effects on E‐C coupling properties, high levels of overexpression in homozygous animals results in susceptibility to hypertrophy and heart failure, along with a significant reduction in E‐C coupling gain. While global knockout of the exchanger in mice is embryonic‐lethal, conditional knockout mice live to adulthood. Cardiac function is surprisingly normal in seven‐week‐old mice, but E‐C coupling gain is apparently increased. Thus, genetic modification of exchanger expression has a major influence on E‐C coupling.
Mitochondrial Calcium Signaling and Energy Metabolism
Authors: NGUYEN MY‐HANH T. , JAFRI M. SALEET
Published: August 2005

Abstract: A computational model of energy metabolism in the mammalian ventricular myocyte is developed to study the effect of cytosolic calcium (Ca2+) transients on adenosine triphosphate (ATP) production. The model couples the Jafri‐Dudycha model for tricarboxylic acid cycle regulation to a modified version of the Magnus‐Keizer model for the mitochondria. The fluxes associated with Ca2+ uptake and efflux (i.e., the Ca2+ uniporter and Na+‐Ca2+ exchanger) and the F1F0‐ATPase were modified to better model heart mitochondria. Simulations were performed at steady state and with Ca2+ transients at various pacing frequencies generated by the Rice‐Jafri‐Winslow model for the guinea pig ventricular myocyte. The effects of the Ca2+ transients for mitochondria both adjacent to the dyadic space and in the bulk myoplasm were studied. The model shows that Ca2+ activation of both the tricarboxylic acid cycle and the F1F0‐ATPase are necessary to produce increases in ATP production. The model also shows that in mitochondria located near the subspace, the large Ca2+ transients can depolarize the mitochondrial membrane potential sufficiently to cause a transient decline in ATP production. However, this transient is of short duration, minimizing its impact on overall ATP production.
Caveolae and Lipid Rafts: G Protein‐Coupled Receptor Signaling Microdomains in Cardiac Myocytes
Authors: INSEL PAUL A. , HEAD BRIAN P. , OSTROM RENNOLDS S. , PATEL HEMAL H. , SWANEY JAMES S. , TANG CHIH‐MIN , ROTH DAVID M.
Published: August 2005

Abstract: A growing body of data indicates that multiple signal transduction events in the heart occur via plasma membrane receptors located in signaling microdomains. Lipid rafts, enriched in cholesterol and sphingolipids, form one such microdomain along with a subset of lipid rafts, caveolae, enriched in the protein caveolin. In the heart, a key caveolin is caveolin‐3, whose scaffolding domain is thought to serve as an anchor for other proteins. In spite of the original morphologic definition of caveolae (“little caves”), most work related to their role in compartmenting signal transduction molecules has involved subcellular fractionation or immunoprecipitation with anti‐caveolin antibodies. Use of such approaches has documented that several G protein‐coupled receptors (GPCR), and their cognate heterotrimeric G proteins and effectors, localize to lipid rafts/caveolae in neonatal cardiac myocytes. Our recent findings support the view that adult cardiac myocytes appear to have different patterns of localization of such components compared to neonatal myocytes and cardiac fibroblasts. Such results imply the existence of multiple subcellular microdomains for GPCR‐mediated signal transduction in cardiac myocytes, in particular adult myocytes, and raise a major unanswered question: what are the precise mechanism(s) that determine co‐localization of GPCR and post‐receptor components with lipid rafts/caveolae in cardiac myocytes and other cell types?
Cardiac Neurobiology of Nitric Oxide Synthases
Authors: DANSON EDWARD J. , PATERSON DAVID J.
Published: August 2005

Abstract: Nitric oxide (NO) is a potent modulator of cardiac and vascular regulation. Its role in cardiac‐autonomic neural signaling has received much attention over the last decade because of the ability of NO to alter cardiac sympathovagal balance to favor more anti‐arrhythmic states. Complexity and controversy have arisen, however, because of the numerous sources of NO in the brain, peripheral nerves, and cardiomyocytes, all of which are potential regulators of cardiac excitability and calcium signaling. This review addresses the integrative role of NO as a relatively ubiquitous signaling molecule with respect to cardiac neurobiology. The present idea, that divergent NO‐signaling pathways from multiple sources within the heart and nervous system converge to modulate cardiac excitability and impact on morbidity and mortality in health and disease, is discussed.
Regulation of Cardiac Energetics: Role of Redox State and Cellular Compartmentation during Ischemia
Authors: CABRERA MARCO E. , ZHOU LUFANG , STANLEY WILLIAM C. , SAIDEL GERALD M.
Published: August 2005

Abstract: The heart is capable of altering its metabolic rate during exercise or ischemia. Under most state transitions, the heart maintains the concentration of adenosine triphosphate (ATP) at relatively constant values, in spite of large fluctuations in metabolic rate or in the delivery of fuels and oxygen. However, the mechanisms responsible for the regulation of cardiac energetics under conditions of increased demand or reduced supply are still under debate. To improve quantitative understanding of the regulation of glycolysis and oxidative phosphorylation under physiological and pathological conditions, it is essential to assess the dynamics of cytosolic and mitochondrial nicotinamide adenine dinucleotide (NAD+) and its reduced form (NADH) during stress (e.g., ischemia, exercise). However, at present there are no reliable methods to measure the dynamics of redox state in vivo in these subcellular compartments. In the present study, computer simulations with a mathematical model of myocardial energy metabolism are used to investigate the role of cytosolic and mitochondrial redox states in regulating cardiac energetics during reduced myocardial blood flow.
Measuring and Mapping Cardiac Fiber and Laminar Architecture Using Diffusion Tensor MR Imaging
Authors: HELM PATRICK , BEG MIRZA FAISAL , MILLER MICHAEL I. , WINSLOW RAIMOND L.
Published: August 2005

Abstract: The ventricular myocardium is known to exhibit a complex spatial organization, with fiber orientation varying as a function of transmural location. It is now well established that diffusion tensor magnetic resonance imaging (DTMRI) may be used to measure this fiber orientation at high spatial resolution. Cardiac fibers are also known to be organized in sheets with surface orientation varying throughout the ventricles. This article reviews results on use of DTMRI for measuring ventricular fiber orientation, as well as presents new results providing strong evidence that the tertiary eigenvector of the diffusion tensor is aligned locally with the cardiac sheet surface normal. Considered together, these data indicate that DTMRI may be used to reconstruct both ventricular fiber and sheet organization. This article also presents the large deformation diffeomorphic metric mapping (LDDMM) algorithm and shows that this algorithm may be used to bring ensembles of imaged and reconstructed hearts into correspondence (e.g., registration) so that variability of ventricular geometry, fiber, and sheet orientation may be quantified. Ventricular geometry and fiber structure is known to be remodeled in a range of disease processes; however, descriptions of this remodeling have remained subjective and qualitative. We anticipate that use of DTMRI for reconstruction of ventricular anatomy coupled with application of the LDDMM method for image volume registration will enable the detection and quantification of changes in cardiac anatomy that are characteristic of specific disease processes in the heart. Finally, we show that epicardial electrical mapping and DTMRI imaging may be performed in the same hearts. The anatomic data may then be used to simulate electrical conduction in a computational model of the very same heart that was mapped electrically. This facilitates direct comparison and testing of model versus experimental results and opens the door to quantitative measurement, modeling, and analysis of the ways in which remodeling of ventricular microanatomy influences electrical conduction in the heart.
Species‐ and Preparation‐Dependence of Stretch Effects on Sino‐Atrial Node Pacemaking
Authors: COOPER PATRICIA J. , KOHL PETER
Published: August 2005

Abstract: Acute dilation of the right atrium (e.g., via increased venous return) raises spontaneous beating rate (BR) of the heart in many species. Neural mechanisms contribute to this behavior in vivo, but a positive chronotropic response to stretch can also be observed in isolated right atrial tissue preparations and even at the level of single sino‐atrial node (SAN) cells. The underlying mechanism has previously been reported to be compatible with stretch‐activation of cation nonselective ion channels (SAC). This review reports species peculiarities in the chronotropic response of isolated SAN tissue strips to stretch: in contrast to guinea pig, murine SAN preparations respond to distension with a reduction in spontaneous BR. This differential response need not necessarily involve disparate (sub‐)cellular mechanisms, as SAC activation would occur against the background of very different SAN electrophysiology in the two species. On the basis of single SAN cell action potential recordings, this review illustrates how this may give rise to potentially opposing effects on spontaneous BR. Interestingly, streptomycin (a useful SAC blocker in isolated cells) has no effect on stretch‐induced chronotropy in situ, and this is interpreted as an indication of protection of SAC, in native tissue, from interaction with the drug.
Dependence of Electrical Coupling on Mechanical Coupling in Cardiac Myocytes: Insights Gained from Cardiomyopathies Caused by Defects in Cell‐Cell Connections
Authors: SAFFITZ JEFFREY E.
Published: August 2005

Abstract: Cardiac myocytes are electrically coupled by large gap junctions to ensure safe conduction. Membrane regions containing gap junction channels are rigid and susceptible to fragmentation in response to shear stress. Thus, gap junctions in cardiac myocytes are located in close proximity to points of cell‐cell adhesion within the intercalated disk. These adhesion junctions mechanically stabilize the sarcolemmas of adjacent cells to allow formation and maintenance of large arrays of intercellular channels. It has been proposed that the extent to which cardiac myocytes are coupled mechanically at cell‐cell adhesion junctions is an important determinant of the extent to which cells can become electrically coupled at gap junctions. This hypothesis has been tested by analyzing gap junctions in human cardiomyopathies caused by mutations in plakoglobin and desmoplakin, intracellular proteins that link adhesion molecules at cell‐cell junctions to the cardiac myocyte cytoskeleton. Marked remodeling of cardiac gap junctions, despite the presence of normal intracellular levels of connexin43, was observed. This suggests that defects in cell‐cell adhesion, or the presence of discontinuities between adhesion junctions and the cytoskeleton, destabilize gap junctions and diminish electrical coupling. This could contribute to the high incidence of ventricular arrhythmias and sudden death known to occur in these cardiomyopathies.
Spatial Nonuniformity of Contraction Causes Arrhythmogenic Ca2+ Waves in Rat Cardiac Muscle
Authors: TER KEURS HENK E.D.J. , WAKAYAMA YUJI , MIURA MASAHITO , STUYVERS BRUNO D. , BOYDEN PENELOPE A. , LANDESBERG AMIR
Published: August 2005

Abstract: Landesberg and Sideman's four state model of the cardiac cross‐bridge (XB) hypothesizes a feedback of force development to Ca2+ binding by troponin C (TnC). We have further modeled this behavior and observed that the force (F)‐Ca2+ relationship as well as the F‐sarcomere length (SL) relationship and the time course of F and Ca2+ transients in cardiac muscle can be reproduced faithfully by a single effect of F on deformation of the TnC‐Ca complex and, thereby, on the dissociation rate of Ca2+. Furthermore, this feedback predicts that rapid decline of F in the activated sarcomere causes release of Ca2+ from TnC‐Ca2+, which is sufficient to initiate arrhythmogenic Ca2+ release from the sarcoplasmic reticulum (SR). This work investigated the initiation of Ca2+ waves underlying triggered propagated contractions (TPCs) in rat cardiac trabeculae under conditions that simulate functional nonuniformity caused by mechanical or ischemic local damage of the myocardium. A mechanical discontinuity along the trabeculae was created by exposing the preparation to a small constant flow jet of solution that reduces excitation‐contraction coupling in myocytes within that segment. Force was measured, and SL as well as [Ca2+]i were measured regionally. When the jet contained caffeine, 2,3‐butanedione monoxime or low‐[Ca2+], muscle‐twitch F decreased and the sarcomeres in the exposed segment were stretched by shortening the normal regions outside the jet. During relaxation, the sarcomeres in the exposed segment shortened rapidly. Short trains of stimulation at 2.5 Hz reproducibly caused Ca2+ waves to rise from the borders exposed to the jet. Ca2+ waves started during F relaxation of the last stimulated twitch and propagated into segments both inside and outside of the jet. Arrhythmias, in the form of nondriven rhythmic activity, were triggered when the amplitude of the Ca2+ wave increased by raising [Ca2+]o. The arrhythmias disappeared when the muscle uniformity was restored by turning the jet off. These results show that nonuniform contraction can cause Ca2+ waves underlying TPCs, and suggest that Ca2+ dissociated from myofilaments plays an important role in the initiation of arrhythmogenic Ca2+ waves.
Stem Cells for Cardiomyocyte Regeneration: State of the Art
Authors: PESCHLE CESARE , CONDORELLI GIANLUIGI
Published: August 2005

Abstract: A wide range of stem/progenitor cell types have been used for the regeneration of the infarcted heart. This review details the current status of progress concerning different strategies that have been used to manipulate cardiomyocyte cell growth in vitro, for their use in heart failure. The current status of this field involves different types of regenerating cells. Embryonic stem (ES) cells, hematopoietic stem and progenitor cells, mesenchymal stem and progenitor cells, and resident cardiac “stem” cells are discussed here.
Multiscale Modeling of Cardiac Cellular Energetics
Authors: BASSINGTHWAIGHTE JAMES B. , CHIZECK HOWARD J. , ATLAS LES E. , QIAN HONG
Published: August 2005

Abstract: Multiscale modeling is essential to integrating knowledge of human physiology starting from genomics, molecular biology, and the environment through the levels of cells, tissues, and organs all the way to integrated systems behavior. The lowest levels concern biophysical and biochemical events. The higher levels of organization in tissues, organs, and organism are complex, representing the dynamically varying behavior of billions of cells interacting together. Models integrating cellular events into tissue and organ behavior are forced to resort to simplifications to minimize computational complexity, thus reducing the model's ability to respond correctly to dynamic changes in external conditions. Adjustments at protein and gene regulatory levels shortchange the simplified higher‐level representations. Our cell primitive is composed of a set of subcellular modules, each defining an intracellular function (action potential, tricarboxylic acid cycle, oxidative phosphorylation, glycolysis, calcium cycling, contraction, etc.), composing what we call the “eternal cell,” which assumes that there is neither proteolysis nor protein synthesis. Within the modules are elements describing each particular component (i.e., enzymatic reactions of assorted types, transporters, ionic channels, binding sites, etc.). Cell subregions are stirred tanks, linked by diffusional or transporter‐mediated exchange. The modeling uses ordinary differential equations rather than stochastic or partial differential equations. This basic model is regarded as a primitive upon which to build models encompassing gene regulation, signaling, and long‐term adaptations in structure and function. During simulation, simpler forms of the model are used, when possible, to reduce computation. However, when this results in error, the more complex and detailed modules and elements need to be employed to improve model realism. The processes of error recognition and of mapping between different levels of model form complexity are challenging but are essential for successful modeling of large‐scale systems in reasonable time. Currently there is to this end no established methodology from computational sciences.
PCTAIRE protein kinases interact directly with the COPII complex and modulate secretory cargo transport
Authors: Palmer Krysten J. , Konkel Joanne E. , Stephens David J.
Published: August 2005

The export of secretory cargo from the endoplasmic reticulum is mediated by the COPII complex. In common with other aspects of intracellular transport, this step is regulated by protein kinase signalling. Recruitment of the COPII complex to the membrane is known to require ATP and to be blocked by the protein kinase inhibitor H-89. The identity of the specific protein kinase or kinases involved remains equivocal. Here we show that the Sec23p subunit of COPII interacts with PCTAIRE protein kinases. This interaction is shown using two-hybrid screening, direct binding and immunoprecipitation. Inhibition of PCTAIRE kinase activity by expression of a kinase-dead mutant, or specific depletion of PCTAIRE using RNAi, leads to defects in early secretory pathway function including cargo transport, as well as vesicular-tubular transport carrier (VTC) and Golgi localization. These data show a role for PCTAIRE protein kinase function in membrane traffic through the early secretory pathway.
Neurogenic potential of human mesenchymal stem cells revisited: analysis by immunostaining, time-lapse video and microarray
Authors: Bertani Nicoletta , Malatesta Paolo , Volpi Giorgia , Sonego Paolo , Perris Roberto
Published: August 2005

The possibility of generating neural cells from human bone-marrow-derived mesenchymal stem cells (hMSCs) by simple in vitro treatments is appealing both conceptually and practically. However, whether phenotypic modulations observed after chemical manipulation of such stem cells truly represent a genuine trans-lineage differentiation remains to be established. We have re-evaluated the effects of a frequently reported biochemical approach, based on treatment with butylated hydroxyanisole and dimethylsulphoxide, to bring about such phenotypic conversion by monitoring the morphological changes induced by the treatment in real time, by analysing the expression of phenotype-specific protein markers and by assessing the modulation of transcriptome. Video time-lapse microscopy showed that conversion of mesenchymal stem cells to a neuron-like morphology could be reproduced in normal primary fibroblasts as well as mimicked by addition of drugs eliciting cytoskeletal collapse and disruption of focal adhesion contacts. Analysis of markers revealed that mesenchymal stem cells constitutively expressed multi-lineage traits, including several pertaining to the neural one. However, the applied `neural induction' protocol neither significantly modulated the expression of such markers, nor induced de novo translation of other neural-specific proteins. Similarly, global expression profiling of over 21,000 genes demonstrated that gene transcription was poorly affected. Most strikingly, we found that the set of genes whose expression was altered by the inductive treatment did not match those sets of genes differentially expressed when comparing untreated mesenchymal stem cells and immature neural tissues. Conversely, by comparing these gene expression profiles with that obtained from comparisons between the same cells and an unrelated non-neural organ, such as liver, we found that the adopted neural induction protocol was no more effective in redirecting human mesenchymal stem cells toward a neural phenotype than toward an endodermal hepatic pathway.
Limitations Arising from Optical Saturation in Fluorescence and Thermal Lens Spectrometries Using Pulsed Laser Excitation: Application to the Determination of the Fluorescence Quantum Yield of Rhodamine 6G
Authors: Georges J. , Arnaud N. , Parise L.
Published: August 2005

The high incident irradiances available with pulsed lasers can lead to a significant depletion of the ground-state population of the chromophore and to optical saturation effects. As a result, the optical absorption coefficient decreases as a function of the excitation energy and, because the amount of energy released by radiative and nonradiative relaxation processes depends on the amount of energy absorbed, nonlinear energy-dependent signals are obtained. Therefore, large errors can be introduced when fluorescence and photothermal data are used to determine fluorescence quantum yields. This work provides experimental results describing the effects of optical saturation on fluorescence and thermal lens measurements for rhodamine 6G in various media and over a wide energy range. It is shown that, when optical saturation is avoided, the photothermal method gives accurate absolute values of Φf ranging from 0.93 to 0.95, depending on the solvent. On the contrary, fluorescence measurements seem to be sensitive to additional experimental artifacts that are more difficult to characterize and to eliminate. Index Headings: Optical saturation; Thermal lens spectrometry; Fluorescence; Rhodamine 6G; Fluorescence quantum yield.
New Standardless Method of Pb Analysis by Electrothermal Atomic Absorption Spectroscopy in Air Aerosols: Comparison with the Official Method
Authors: Locatelli Clinio , Reschiglian Pierluigi , Torsi Giancarlo , Fagioli Francesco , Rossi Nicola , Melucci Dora
Published: August 2005

A standardless analysis method with electrothermal atomization/atomic absorption spectroscopy (ETA-AAS) measurements is proposed and applied to the determination of Pb in an urban environment (Bologna, Italy) and compared with results obtained in the same place and time, with the method dictated by Italian law, which is derived by European Community Directives. It was found that the difference previously observed between the two methods is due to the low efficiency of the 0.45-μm-pore-size paper filters, dictated by the official method. The uncovering of the error source validates the proposed method and highlights its accuracy. Moreover, the two methods can be conveniently combined to give two series of data relevant to pollutants associated with low- and high-dimension particles. This capability gives a much more precise picture of the hazard of a pollutant.