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Dramatic Differences (dramatic + difference)
Selected AbstractsUse tests: ROAT (repeated open application test)/PUT (provocative use test): an overviewCONTACT DERMATITIS, Issue 1 2000Tokio Nakada As one step in defining the clinical relevance of exposure to an allergen identified with patch testing, use tests (provocative use test (PUT), and repeated open application test (ROAT)) have been used. In 1/2 of the cases of seemingly reliable patch tests, use tests are negative, suggesting that the patient's biologic threshold of response had not been reached with open application dosing. Dramatic differences exist in regional skin reactivity and percutaneous penetration. Negative results of use tests on normal skin may become positive on diseased skin. To refine this assay further, more controlled observations and analysis of reaction differences between normal and damaged skin, and among regional anatomic sites might be performed. In addition, we require a standardized measurement for the results. Use testing has significant potential in refinement of the evidence-based diagnosis of clinical relevance. However, for general validation, we should fill the deficiencies described above. [source] Ictal Perfusion Changes During Occipital Lobe Seizures in Infancy: Report of Two Serial Ictal ObservationsEPILEPSIA, Issue 2 2001Andras Hollo Summary: Serial-ictal single-photon-emission computed tomography (SPECT) examinations are presented in two infants (ages 1 and 2 years), with early ictal and ictal in one, and ictal and late ictal images in the other. Both had pharmacoresistant occipital epilepsy, due to focal cortical dysplasia. In the first case, size of ictal hyperperfusion increased in the course of the seizure from early ictal to ictal state. A concomitant ictal hypoperfusion was observed around the hyperperfused area. In the second patient, there was a dramatic difference between ictal and late ictal images. In the late ictal state, the previous occipital ictal hyperperfusion and extraoccipital ictal hypoperfusion disappeared, together with homolateral posterotemporal and contralateral occipital hyperperfusion, corresponding to seizure propagation. Ictal extratemporal blood-flow changes are therefore highly dynamic, particularly in very young children. [source] Ethene Polymerization Behavior of MAO-Activated Dichloridotitanium Complexes Bearing Bi- and Tetradentate Salicylaldimine DerivativesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 2 2010Antti Pärssinen Abstract New chiral bridged tetradentate (N2O2)TiIVCl2 -type complexes bearing dimethylbiphenyl (1-Ti,3-Ti) and previously published binaphthyl-bridged (4-Ti) complex were synthesized with high yields. This was achieved by treating the corresponding Schiff-base ligand (H2L) precursors with Ti(NMe2)4, followed by conversion of these diamido complexes to LTiCl2 derivatives by the addition of excess of Me3SiCl. A series of unbridged titanium complexes 5-Ti,8-Ti with similar substituents at the phenoxy group were studied and their polymerization properties, after methylaluminoxane (MAO) activation, compared with the above bridged complexes. It was found that the catalysts bearing chiral tetradentate biaryl-bridged salicylaldimine ligands produce multimodal polyethylene (PE) with low activity [below 10 kgPE/(molTi,h,bar)] while their unbridged analogues provide activities that are 10,1000 times greater under similar reaction conditions. The reasons for this dramatic difference in polymerization activities are discussed based on the stabilities of the different cationic species configurations. [source] Synthetic peptide vaccine development: measurement of polyclonal antibody affinity and cross-reactivity using a new peptide capture and release system for surface plasmon resonance spectroscopyJOURNAL OF MOLECULAR RECOGNITION, Issue 6 2004Paul J. Cachia Abstract A method has been developed for measurement of antibody affinity and cross-reactivity by surface plasmon resonance spectroscopy using the EK-coil heterodimeric coiled-coil peptide capture system. This system allows for reversible capture of synthetic peptide ligands on a biosensor chip surface, with the advantage that multiple antibody-antigen interactions can be analyzed using a single biosensor chip. This method has proven useful in the development of a synthetic peptide anti- Pseudomonas aeruginosa (PA) vaccine. Synthetic peptide ligands corresponding to the receptor binding domains of pilin from four strains of PA were conjugated to the E-coil strand of the heterodimeric coiled-coil domain and individually captured on the biosensor chip through dimerization with the immobilized K-coil strand. Polyclonal rabbit IgG raised against pilin epitopes was injected over the sensor chip surface for kinetic analysis of the antigen-antibody interaction. The kinetic rate constants, k(on) and k(off), and equilibrium association and dissociation constants, KA and KD, were calculated. Antibody affinities ranged from 1.14,×,10,9 to 1.60,×,10,5,M. The results suggest that the carrier protein and adjuvant used during immunization make a dramatic difference in antibody affinity and cross-reactivity. Antibodies raised against the PA strain K pilin epitope conjugated to keyhole limpet haemocyanin using Freund's adjuvant system were more broadly cross-reactive than antibodies raised against the same epitope conjugated to tetanus toxoid using Adjuvax adjuvant. The method described here is useful for detailed characterization of the interaction of polyclonal antibodies with a panel of synthetic peptide ligands with the objective of obtaining high affinity and cross-reactive antibodies in vaccine development. Copyright © 2004 John Wiley & Sons, Ltd. [source] Tellurium-Based Polymeric Surfactants as a Novel Seleno-Enzyme Model with High ActivityMACROMOLECULAR RAPID COMMUNICATIONS, Issue 24 2006Xin Huang Abstract Summary: A tellurium-based polymeric sufactant as a seleno-enzyme model has been constructed by employing 11-acryloyloxyundecyltriethylammonium bromide (AUTEAB, 4) and a tellurium-containing compound (1). It demonstrates strong substrate binding ability for thiols and high glutathione peroxidase (GPx) activity about 6 orders of magnitude more efficient than the well-known GPx mimic PhSeSePh in an ArSH assay system. More importantly, a series of tellurium-based polymeric micelle catalysts with the catalytic tellurium center located at various positions in the micelle have been constructed, and the dramatic difference in activity indicates that the exact match of the catalytic center and binding site plays a key role in enzyme catalytic efficiency. Schematic representation of the proposed mode of the telluro-micelle catalysts. [source] Supramolecular selectivity of poly(ethylene oxide) in semi-crystalline polymer nanocompositesPOLYMER INTERNATIONAL, Issue 12 2007Li Zhou Abstract Semi-crystalline polymer nanocomposites were prepared using successive meltings and recrystallizations techniques by intercalation of small guest molecules such as 4-chlorotoluene (PCT), 4-bromotoluene (PBT) and 1,4-dibromobenzene (PDBB) into poly(ethylene oxide) (PEO) crystals. Differential scanning calorimetry, Fourier transform infrared spectroscopy and wide-angle X-ray diffraction experimental results show that supramolecular selectivity exists for the PEO,PDBB/PBT ternary system, while there is no supramolecular selectivity for PEO,PCT/PBT ternary nanocomposites. The interactions between PEO chains and small guest molecules have an important influence on the polymer conformation, which results in the dramatic difference in intercalation behavior. Copyright © 2007 Society of Chemical Industry [source] Temperature-induced reversible conformational change in the first 100 residues of ,-synucleinPROTEIN SCIENCE, Issue 3 2006Brian C. McNulty Abstract Natively disordered proteins are a growing class of anomalies to the structure,function paradigm. The natively disordered protein ,-synuclein is the primary component of Lewy bodies, the cellular hallmark of Parkinson's disease. We noticed a dramatic difference in dilute solution 1H- 15N Heteronuclear Single Quantum Coherence (HSQC) spectra of wild-type ,-synuclein and two disease-related mutants (A30P and A53T), with spectra collected at 35°C showing fewer cross-peaks than spectra acquired at 10°C. Here, we show the change to be the result of a reversible conformational exchange linked to an increase in hydrodynamic radius and secondary structure as the temperature is raised. Combined with analytical ultracentrifugation data showing ,-synuclein to be monomeric at both temperatures, we conclude that the poor quality of the 1H- 15N HSQC spectra obtained at 35°C is due to conformational fluctuations that occur on the proton chemical shift time scale. Using a truncated variant of ,-synuclein, we show the conformational exchange occurs in the first 100 amino acids of the protein. Our data illustrate a key difference between globular and natively disordered proteins. The properties of globular proteins change little with solution conditions until they denature cooperatively, but the properties of natively disordered proteins can vary dramatically with solution conditions. [source] Detection of arthritis-susceptibility loci, including Ncf1, and variable effects of the major histocompatibility complex region depending on genetic background in ratsARTHRITIS & RHEUMATISM, Issue 2 2009Carola Rintisch Objective To characterize the arthritis-modulating effects of 3 non,major histocompatibility complex (MHC) quantitative trait loci (QTLs) in rat experimental arthritis in the disease-resistant E3 strain, and to investigate the disease-modulating effects of the MHC region (RT1) in various genetic backgrounds. Methods A congenic fragment containing Ncf1 along with congenic fragments containing the strongest remaining loci, Pia5/Cia3 and Pia7/Cia13 on chromosome 4, were transferred from the arthritis-susceptible DA strain into the background of the completely resistant E3 strain. The arthritis-regulatory potential of the transferred alleles was evaluated by comparing the susceptibility to experimental arthritis in congenic rats with that in E3 rats. The RT1u haplotype from the E3 strain was transferred into the susceptible DA strain (RT1av1), and various F1 and F2 hybrids were generated to assess the effects of RT1 on arthritis susceptibility. Results The DA allele of Ncf1 did not break the arthritis resistance of the E3 rats, although it led to enhanced autoimmune B cell responses, as indicated by significantly elevated levels of anticollagen antibodies in congenic rats. Introgressing Pia5 and Pia7 loci on chromosome 4 broke the resistance to arthritis, and the MHC locus on chromosome 20 in DA rats enhanced arthritis when RT1 interacted with E3 genes. Conclusion The findings in these congenic lines confirm the existence of 3 major QTLs that regulate the severity of arthritis and are sufficient to induce the transformation of a completely arthritis-resistant rat strain into an arthritis-susceptible strain. This study also reveals a dramatic difference in the arthritis-regulatory potential of the rat MHC depending on genetic background, suggesting that strong epistatic interactions occur between MHC and non-MHC genes. [source] 4151: Epidemiology of uveitis in the Middle East and North AfricaACTA OPHTHALMOLOGICA, Issue 2010M KHAIRALLAH Purpose Numerous studies have examined the pattern of uveitis around the world. Most of them are from western countries, including the USA and countries in Europe, and Eastern Asia. The aim of this presentation is to review the epidemiological characteristics of uveitis in the the Middle East and North Africa. Methods The epidemiologic data on uveitis available from the Middle East and North Africa were reviewed. Results Several recent studies addressed the pattern of uveitis in different countries, including Iran, Saudi Arabia, Turkey, and Tunisia. Uveitis was most often seen in adults with a peak age at presentation in the third and fourth decades. There was no dramatic difference in gender distribution. Anterior uveitis was the most common anatomic form of uveitis, but a high rate of posterior uveitis and panuveitis was reported. A definitive or presumed specific diagnosis could be established for 57-87% of patients. The most common infectious entities were herpetic anterior uveitis, toxoplasmosis, and tuberculosis (Saudi Arabia). The most common identifiable non-infectious entities included Behçet's disease and Vogt-Koyanagi-Harada disease. Conclusion Herpetic infection, toxoplasmosis, and tuberculosis are the most common infectious causes of uveitis in the Middle East and North Africa. Behçet's diease and Vogt-Koyanagi-Harada disease are the most common non-infectious uveitic entities.HLA-B27 acute anterior uveitis, ocular sarcoidosis, and juvenile idiopathic arthritis associated uveitis are less common than in western countries. [source] An in vivo comparison of photoactivatable fluorescent proteins in an avian embryo modelDEVELOPMENTAL DYNAMICS, Issue 6 2007Danny A. Stark Abstract Tracing the lineage or neighbor relationships of cells in a migratory population or deep within an embryo is difficult with current methods. The recent explosion of photoactivatable fluorescent proteins (PAFPs) offers a unique cell labeling tool kit, yet their in vivo performance in intact embryos and applicability have not been thoroughly explored. We report a comparison study of PAGFP, PSCFP2, KikGR, and Kaede analyzed in the avian embryo using confocal and 2-photon microscopy. PAFPs were introduced into the chick neural tube by electroporation and each photoconverted in the neural crest or cells in the neural tube with exposure to 405 nm light, but showed dramatic differences in photoefficiency and photostability when compared at the same 2% laser power. KikGR and Kaede photoconverted with ratios only slightly lower than in vitro results, but cells rapidly photobleached after reaching maximal photoefficiency. PSCFP2 had the lowest photoefficiency and photoconverted nearly 70 times slower than the other dual-color PAFPs tested, but was effective at single-cell marking, especially with 2-photon excitation at 760 nm. The dual-color PAFPs were more effective to monitor cell migratory behaviors, since non-photoconverted neighboring cells were fluorescently marked with a separate color. However, photoconverted cells were limited in all cases to be visually distinguishable for long periods, with PSCFP2 visible from background the longest (48 hr). Thus, photoactivation in embryos has the potential to selectively mark less accessible cells with laser accuracy and may provide an effective means to study cell,cell interactions and short-term cell lineage in developmental and stem cell biology. Developmental Dynamics 236:1583,1594, 2007. © 2007 Wiley-Liss, Inc. [source] Diet dynamics of the juvenile piscivorous fish community in Spirit Lake, Iowa, USA, 1997,1998ECOLOGY OF FRESHWATER FISH, Issue 4 2001M. E. Pelham Abstract , We assessed temporal dynamics and variation among species and age-classes in the diets of age 0 and age 1 piscivorous fish species in Spirit Lake, Iowa, USA during 1997 and 1998. Species included walleye Stizostedion vitreum, yellow perch Perca flavescens, smallmouth bass Micropterus dolomieui, largemouth bass Micropterus salmoides, black crappie Pomoxis nigromaculatus and white bass Morone chrysops. Thirty taxa were identified in diets, including 12 species of fish. We found dramatic differences in diets among species, among age-classes within species and over time. Walleye, largemouth bass, smallmouth bass and white bass were piscivorous at age 0. Black crappie began piscivory at age 1. Yellow perch also began piscivory at age 1, but fish were a very small fraction of age-1 diets. The primary temporal pattern, seen in several species and age-classes, was an increase in piscivory from spring to fall. This pattern was due to the lack of small, age-0 prey fish in spring. Although some patterns were evident, the taxonomic composition of the diets of all species was highly variable over time, making generalizations difficult. A surprising result was the absence of yellow perch in the diet of age-0 walleye, despite their abundance in Spirit Lake and prominence in diets of age-1 walleye and other age 1-piscivores. Age-0 yellow perch were consistently too large to be eaten by age-0 piscivores, which preyed primarily on invertebrates and smaller fish such as johnny darters Etheostoma nigrum and age 0 bluegill Lepomis macrochirus. This finding suggests that predator-prey interactions and resulting population dynamics may be quite different in Spirit Lake than in other systems dominated by walleye and yellow perch., [source] Innate and Learned Shoaling Preferences Based on Body Coloration in Juvenile Mollies, Poecilia latipinnaETHOLOGY, Issue 11 2008Jessica M. Ledesma Shoaling offers fish enhanced protection from predators through a phenomenon known as the confusion effect. This phenomenon depends on a high degree of phenotypic homogeneity within a shoal, which may confuse predators that have difficulty in targeting a single individual as prey. Accordingly, fish typically choose shoalmates with similar phenotypic characteristics to themselves. In the molly (Poecilia latipinna), dramatic differences in body coloration have been shown to affect shoalmate choice in adults. Here, we show that juvenile mollies (50 d old) were capable of shoaling and that early experience impacted shoalmate choice. When raised in isolation, mollies chose shoalmates with similar body coloration to their own. When raised with other juvenile mollies, test fish chose to associate with individuals of the same coloration as the fish with whom they had been reared. These results show that P. latipinna are capable of the behavioral plasticity that has been demonstrated in other fish species, and that early experience affects the impact of body coloration on shoaling decisions in this species. [source] Plant fructans stabilize phosphatidylcholine liposomes during freeze-dryingFEBS JOURNAL, Issue 2 2000Dirk K. Hincha Fructans have been implicated as protective agents in the drought and freezing tolerance of many plant species. A direct proof of their ability to stabilize biological structures under stress conditions, however, is still lacking. Here we show that inulins (linear fructose polymers) isolated from chicory roots and dahlia tubers stabilize egg phosphatidylcholine large unilamellar vesicles during freeze-drying, while another polysaccharide, hydroxyethyl starch, was completely ineffective. Liposome stability was assessed after rehydration by measuring retention of the soluble fluorescent dye carboxyfluorescein and bilayer fusion. Inulin was an especially effective stabilizer in combination with glucose. Analysis by HPLC showed that the commercial inulin preparations used in our study contained no low molecular mass sugars that could be responsible for the observed stabilizing effect of the fructans. Fourier transform infrared spectroscopy showed a reduction of the gel to liquid-crystalline phase transition temperature of dry egg PtdCho by more than 20 °C in the presence of inulin. A direct interaction of inulin with the phospholipid in the dry state was also indicated by dramatic differences in the phosphate asymmetric stretch region of the infrared spectrum between samples with and without the polysaccharide. [source] Adipose tissue gene expression in obese dogs after weight lossJOURNAL OF ANIMAL PHYSIOLOGY AND NUTRITION, Issue 3 2008V. Leray Summary Body weight (BW) mainly depends on a balance between fat storage (lipogenesis) and fat mobilization (lipolysis) in adipocytes. BW changes play a role in insulin resistance (IR), the inability of insulin target tissue to respond to physiological levels of insulin. This results in inhibition of lipogenesis and stimulation of lipolysis. Weight gain leads to IR whereas, weight loss improves insulin sensitivity (IS). The aim of this study was to evaluate the effect of weight loss and recovery of IS on the expression of genes involved in lipogenesis and lipolysis in weight losing dogs. Gene expression was studied in both subcutaneous and visceral adipose tissue. Obese dogs received a hypoenergetic low fat high protein diet (0.6 × NRC recommendation). Before and after weight loss, IS was assessed using the euglycaemic hyperinsulinaemic clamp. Gene expression of IRS-2, SREBP, intracellular insulin effectors, ACC, FAS, FABP, ADRP, PEPCK, lipogenesis key proteins, perilipin and HSL, lipolysis key proteins were quantified using real-time RT-PCR in subcutaneous and visceral fat. BW decreased from 15.2 ± 0.5 to 11.4 ± 0.4 kg (p < 0.05) over 78 ± 8 days. When obese, dogs were insulin resistant. After weight loss, IS was improved. In the subcutaneous adipose tissue, the expression of only the IRS-2 was increased. In the visceral adipose tissue, the expression of the genes involved in the lipogenesis was decreased whereas one of the genes implied in the lipolysis did not change. The expression profile of genes involved in lipid metabolism, as measured after weight loss, is indicative for a lower lipogenesis after weight loss than in obese dogs. Our results also confirm dramatic differences in the lipid metabolism of visceral and subcutaneous fat. They should be completed by comparing gene expression during weight losing and normal weight steady state. [source] Genome size and recombination in angiosperms: a second lookJOURNAL OF EVOLUTIONARY BIOLOGY, Issue 2 2007J. ROSS-IBARRA Abstract Despite dramatic differences in genome size , and thus space for recombination to occur , previous workers found no correlation between recombination rate and genome size in flowering plants. Here I re-investigate these claims using phylogenetic comparative methods to test a large data set of recombination data in angiosperms. I show that genome size is significantly correlated with recombination rate across a wide sampling of species and that change in genome size explains a meaningful proportion (,20%) of variation in recombination rate. I show that the strength of this correlation is comparable with that of several characters previously linked to evolutionary change in recombination rate, but argue that consideration of processes of genome size change likely make the observed correlation a conservative estimate. And finally, although I find that recombination rate increases less than proportionally to change in genome size, several mechanistic and theoretical arguments suggest that this result is not unexpected. [source] Influence of aldehyde fixation on the morphology of endosomes and lysosomes: quantitative analysis and electron tomographyJOURNAL OF MICROSCOPY, Issue 1 2003J. L. A. N. Murk Summary Cryoimmobilization is regarded as the most reliable method to preserve cellular ultrastructure for electron microscopic analysis, because it is both fast (milliseconds) and avoids the use of harmful chemicals on living cells. For immunolabelling studies samples have to be dehydrated by freeze-substitution and embedded in a resin. Strangely, although most of the lipids are maintained, intracellular membranes such as endoplasmic reticulum, Golgi and mitochondrial membranes are often poorly contrasted and hardly visible. By contrast, Tokuyasu cryosectioning, based on chemical fixation with aldehydes is the best established and generally most efficient method for localization of proteins by immunogold labelling. Despite the invasive character of the aldehyde fixation, the Tokuyasu method yields a reasonably good ultrastructural preservation in combination with excellent membrane contrast. In some cases, however, dramatic differences in cellular ultrastructure, especially of membranous structures, could be revealed by comparison of the chemical with the cryofixation method. To make use of the advantages of the two different approaches a more general and quantitative knowledge of the influence of aldehyde fixation on ultrastructure is needed. Therefore, we have measured the size and shape of endosomes and lysosomes in high-pressure frozen and aldehyde-fixed cells and found that aldehyde fixation causes a significant deformation and reduction of endosomal volume without affecting the membrane length. There was no considerable influence on the lysosomes. Ultrastructural changes caused by aldehyde fixation are most dramatic for endosomes with tubular extensions, as could be visualized with electron tomography. The implications for the interpretation of immunogold localization studies on chemically fixed cells are discussed. [source] Functional integrin subunits regulating cell,matrix interactions in the intervertebral discJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 6 2007Christopher L. Gilchrist Abstract Cellular interactions with the extracellular matrix are key factors regulating cell survival, differentiation, and response to environmental stimuli in cartilagenous tissues. Much is known about the extracellular matrix proteins in the intervertebral disc (IVD) and their variations with region, age, or degenerative state of the tissue. In contrast, little is known of the integrin cell surface receptors that directly bind to and interact with these matrix proteins in the IVD. In almost all tissues, these integrin-mediated cell,matrix interactions are important for transducing environmental cues arising from mechanical stimuli, matrix degradation fragments, and cytokines into intracellular signals. In this study, cells from the nucleus pulposus and anulus fibrosus regions of porcine IVDs were analyzed via flow cytometry to quantify integrin expression levels upon isolation and after monolayer culture. Assays of cell attachment to collagens, fibronectin, and laminin were performed after functional blocking of select integrin subunits to evaluate the role of specific integrins in cell attachment. In situ distribution and co-localization of integrins and laminin were also characterized. Results identify integrin receptors critical for IVD cell interactions with collagens (,1,1) and fibronectin (,5,1). Additionally, dramatic differences in cell,laminin interactions were observed between cells of the nucleus and anulus regions, including differences in ,6 integrin expression, cell adhesion to laminin, and in situ pericellular environments. These findings suggest laminin,cell interactions may be important and unique to the nucleus pulposus region of the IVD. The results of this study provide new information on functional cell,matrix interactions in tissues of the IVD. © 2006 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 25: 829,840, 2007 [source] Sibling death clustering in India: state dependence versus unobserved heterogeneityJOURNAL OF THE ROYAL STATISTICAL SOCIETY: SERIES A (STATISTICS IN SOCIETY), Issue 4 2006Wiji Arulampalam Summary., Data from a range of environments indicate that the incidence of death is not randomly distributed across families but, rather, that there is a clustering of death among siblings. A natural explanation of this would be that there are (observed or unobserved) differences across families, e.g. in genetic frailty, education or living standards. Another hypothesis that is of considerable interest for both theory and policy is that there is a causal process whereby the death of a child influences the risk of death of the succeeding child in the family. Drawing language from the literature on the economics of unemployment, the causal effect is referred to here as state dependence (or scarring). The paper investigates the extent of state dependence in India, distinguishing this from family level risk factors that are common to siblings. It offers some methodological innovations on previous research. Estimates are obtained for each of three Indian states, which exhibit dramatic differences in socio-economic and demographic variables. The results suggest a significant degree of state dependence in each of the three regions. Eliminating scarring, it is estimated, would reduce the incidence of infant mortality (among children who are born after the first child) by 9.8% in the state of Uttar Pradesh, 6.0% in West Bengal and 5.9% in Kerala. [source] Fungal infections associated with HIV infectionORAL DISEASES, Issue 2002LP Samaranayake Oral candidiasis is perhaps the commonest infection seen in HIV disease. The aim of this workshop was to provide a sketch of the multifarious aspects of the disease from a global perspective. To this end the panellists addressed issues such as the virulence of Candida, emergence of antifungal resistance, management of candidiasis and other exotic, oral mycotic diseases. An all-pervasive theme was the dramatic differences in the management of fungal infections consequential to the availability (or the lack) of anti-HIV drugs in the developed and the developing world. Further, the social stigmata associated with the HIV disease in many developing regions in Africa and Asia appears to modify the therapeutic strategies. Additionally, the lesser-known regional variations in the disease manifestations and therapeutic approaches were stark. Further work is direly needed to address these issues. [source] Characterisation of organellar proteomes: A guide to subcellular proteomic fractionation and analysisPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 21 2006Edwin Ho Abstract Subcellular fractionation is being widely used to increase our understanding of the proteome. Fractionation is often coupled with 2-DE, thus allowing the visualisation of proteins and their subsequent identification and characterisation by MS. Whilst this strategy should be effective, to date, there has been little or no consideration given to differences in the mass, pI, hydropathy or abundance of proteins in the organelles and how analytical strategies can be tailored to match the idiosyncrasies of proteins in each particular compartment. To address this, we analysed 3962 Saccharomyces cerevisiae proteins, previously localised to one or more of 22 subcellular compartments. Different compartments showed significantly different distributions of protein pI and hydropathy. Mitochondrial and ER proteins showed the most dramatic differences to other organelles, in their protein pIs and hydropathy, respectively. We show that organelles can be clustered by similarities in these physicochemical protein characteristics. Interestingly, the distribution of protein abundance was also significantly different between many organelles. Our results show that to fully explore subcellular fractions of the proteome, specific analytical strategies should be employed. We outline strategies for all 22 subcellular compartments. [source] Computerized analysis of cytochemical reactions for dehydrogenases and oxygraphic studies as methods to evaluate the function of the mitochondrial sheath in rat spermatozoaANDROLOGIA, Issue 1 2001M. Piasecka Cytochemical reactions for mitochondrial NADH-dependent dehydrogenases (diaphorase/NADH which is related to flavoprotein), NAD-dependent dehydrogenases (isocitrate, malate) and succinate dehydrogenase were carried out in rat spermatozoa. In addition to a morphological evaluation, the intensity of the reactions was assessed using a computer image analysing system (Quantimet 600 S). The intensity of the reactions was examined in sperm midpieces by measuring integrated optical density (IOD) and mean optical density (MOD). The activity of mitochondrial respiratory chain complexes was also analysed using the polarographic method. In the population of spermatozoa studied, all whole spermatozoa midpieces were completely filled with formazans, the product of the cytochemical reaction. These morphological findings corresponded to the values obtained for IOD and MOD for the given enzymes. In the oxygraphic studies, the spermatozoa demonstrated consumption of oxygen in the presence of substrates for I, II and IV complexes and their mitochondria revealed normal integrity and sensitivity to the substrates and inhibitors. However, the oxygraphic studies revealed differences between the sperm and somatic cells. These differences concerned the stimulation of pyruvate oxidation by malate, the lack of an effect of malonic acid on phenazine methosulphate (an acceptor of electrons) oxidation and the lack of an effect of cytochrome c on ascorbate oxidation. The cytochemical method, together with densitometric measurements, enables: (1) the reaction intensity to be determined objectively; (2) subtle and dramatic differences in reaction intensity to be revealed between spermatozoa that do not differ under morphological evaluation of the intensity; (3) possible defects within the mitochondrial sheath to be located and assessed in a large number of spermatozoa. This method can be used as a screening method alongside the routine morphological examination of spermatozoa. On the other hand, the oxygraphic method in the inner membrane of mitochondria can reveal functional changes which are related to the action of respiratory chain complexes and display characteristic features of mitochondria energy metabolism. The methods used are complementary and allow the complex evaluation of mitochondria in spermatozoa. Both methods can be used in experimental and clinical studies. [source] Mitochondrial Membrane Potential Selects Hybridomas Yielding High Viability in Fed-Batch CulturesBIOTECHNOLOGY PROGRESS, Issue 1 2002Brian D. Follstad Prior research (Follstad, B. D.; Wang, D. I. C.; Stephanopoulos, G. Mitochondrial membrane potential differentiates cells resistant to apoptosis in hybridoma cultures. Eur. J. Biochem. 2000, 267, 6534,6540.) identified mitochondrial membrane potential (MMP) as a marker of hybridoma subpopulations resistant to apoptosis caused by a variety of apoptosis inducers. In this study, we investigated the viability of hybridoma cell cultures inoculated with cells of varying MMP in regular fed-batch operation. A hybridoma cell population was separated using FACS into subpopulations based on their mean mitochondrial membrane potential (MMP) as measured using the common mitochondrial stain, Rhodamine 123 (Rh123). These subpopulations showed dramatic differences in their apoptotic death kinetics. Fed-batches inoculated with a high MMP subpopulation reached higher viable cell concentrations and viabilities that were maintained for prolonged periods of time relative to fed-batches inoculated with low MMP subpopulations. These results underline the heterogeneous nature of hybridoma cell cultures and suggest that mitochondrial physiology is a critical parameter determining culture performance. [source] | |||||||||||||||||||