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PubMed1994

Characterization of the early lesion of 'degenerative' valvular aortic stenosis. Histological and immunohistochemical studies.

BACKGROUND: Nonrheumatic stenosis of trileaflet aortic valves, often termed senile or calcific valvular aortic stenosis, is considered a "degenerative" process, but little is known about the cellular or molecular factors that mediate its development. METHODS AND RESULTS: To characterize the developing aortic valvular lesion, we performed histological and immunohistochemical studies on Formalin-fixed and methanol-Carnoy's-fixed paraffin-embedded aortic valve leaflets or on frozen sections obtained at autopsy from 27 adults (age, 46 to 82 years) with normal leaflets (n = 6), mild macroscopic leaflet thickening (n = 15), or clinical aortic stenosis (n = 6). Focal areas of thickening ("early lesions") were characterized by (1) subendothelial thickening on the aortic side of the leaflet, between the basement membrane (PAS-positive) and elastic lamina (Verhoeff-van Gieson), (2) the presence of large amounts of intracellular and extracellular neutral lipids (oil red O) and fine, stippled mineralization (von Kossa), and (3) disruption of the basement membrane overlying the lesion. Regions of the fibrosa adjacent to these lesions were characterized by thickening and by protein, lipid, and calcium accumulation. Control valves showed none of these abnormalities. Immunohistochemical studies were performed using monoclonal antibodies directed against macrophages (anti-CD68 or HAM-56), and contractile proteins of smooth muscle cells or myofibroblasts (anti-alpha-actin and HHF-35) or rabbit polyclonal antiserum against T lymphocytes (anti-CD3). In normal valves, scattered macrophages were present in the fibrosa and ventricularis, and occasional muscle actin-positive cells were detected in the proximal portion of the ventricularis near the leaflet base, but no T lymphocytes were found. In contrast, early lesions were characterized by the presence of an inflammatory infiltrate composed of non-foam cell and foam cell macrophages, occasional T cells, and rare alpha-actin-positive cells. In stenotic aortic valves, a similar but more advanced lesion was seen. CONCLUSIONS: The early lesion of "degenerative" aortic stenosis is an active inflammatory process with some similarities (lipid deposition, macrophage and T-cell infiltration, and basement membrane disruption) and some dissimilarities (presence of prominent mineralization and small numbers of smooth muscle cells) to atherosclerosis.

Aged↗

PubMed1995

Sensitive detection of abnormal aortic architecture in Marfan syndrome with high-frequency ultrasonic tissue characterization.

BACKGROUND: Aneurysmal dilation of the aorta with subsequent rupture or dissection occurs frequently in patients with Marfan syndrome and is the primary cause of morbidity. These complications are related to the altered composition and disorganized structure of the aortic media. Our goal was to use high-frequency ultrasonic tissue characterization to identify these structural changes in abnormal aorta from patients with Marfan syndrome. We measured integrated backscatter and anisotropy of backscatter of ultrasound from specimens of aorta from patients with Marfan syndrome undergoing aortic root replacement and compared these values with those from aortic specimens of patients without clinical aortic pathology. METHODS AND RESULTS: Aortic tissue was obtained at the time of surgery from 11 patients with Marfan syndrome undergoing repair of an aortic aneurysm or dissection. Normal tissue was obtained at the time of autopsy from 8 patients without evidence of aortic disease. Acoustic microscopy at 50 MHz was performed to measure integrated backscatter from each specimen. The magnitude of ultrasonic anisotropy of backscatter for each tissue type was determined as an index of the three-dimensional (3D) organization of the vessel matrix. The collagen content of each specimen was determined with a hydroxyproline assay. Marfan aortas exhibited less backscatter than did normal aortas (-40.9 +/- 2.9 versus -32.6 +/- 2.2 dB for patients with Marfan syndrome and healthy subjects, respectively, P < .0001). No significant difference in collagen concentrations was observed between normal and Marfan aorta (262.7 +/- 52.7 versus 282.4 +/- 41.8 mg/g tissue for normal and Marfan aortas, respectively, P = .42), despite the large difference in backscatter. Histological analysis revealed striking differences in both the amount and organization of the elastin in the aortic aneurysm segments from patients with Marfan syndrome compared with normal aorta. Normal aorta was characterized by well-formed elastin fibers arranged in a lamellar pattern. The media from aneurysms in Marfan aorta exhibited a profound decrease in elastin content that was associated with loss of the highly aligned and ordered lamellar arrangement. The directional dependence of scattering, or ultrasonic anisotropy, also differed dramatically between the two tissue types. Backscatter from normal aorta decreased substantially when the media was insonified parallel compared with perpendicular to the principal axis of the elastin fibers. Marfan aorta exhibited a much smaller directional dependence of scattering. Normal aortas manifested a 14-fold greater ultrasonic anisotropy than did Marfan aortas (24.1 +/- 3.7 versus 12.4 +/- 3.3 dB for normal and Marfan aortas, P < .0001), which is indicative of the profound extent of matrix disorganization in Marfan syndrome. CONCLUSIONS: These data show that high-frequency ultrasonic tissue characterization sensitively detects changes in vessel wall composition and organization that occur in the aorta of patients with Marfan syndrome. Aortic segments from these patients manifested a significant decrease in integrated backscatter compared with normal aorta (approximately 8 dB, or greater than a 6-fold decrease in scattering). A 15-fold reduction in the ultrasonic anisotropy of Marfan tissue was observed, which suggests a marked disorganization of the 3D architecture of these aortas. These data support the hypothesis that high-frequency ultrasonic tissue characterization may be useful for identifying abnormalities of vessel wall composition, architecture, and material properties.

Adult↗

PubMed2003

Characterization of a spontaneously immortalized cell line (IOBA-NHC) from normal human conjunctiva.

PURPOSE: To characterize a new nontransfected, spontaneously immortalized epithelial cell line from normal human conjunctiva (IOBA-NHC), both morphologically and functionally, to determine whether the differentiated phenotype of conjunctival epithelial cells is preserved. METHODS: Outgrowing cells from explanted conjunctival tissue were successively passaged and preliminarily characterized at passage 3 to assess epithelial origin. The cells were further characterized at passages 15 to 20, 40, 60, and 100 by analyzing (1) proliferation and in vitro behavior (viability, plating efficiency, colony forming efficiency and colony size, and Ki-67 protein expression), (2) karyotype and G-banding, (3) epithelial marker expression (cytokeratins, desmoplakins, EGF receptor), (4) absence of contaminating cell types, (5) expression of conjunctival differentiation markers (mucin gene expression), and (6) functional capability in response to proinflammatory stimuli. IOBA-NHC cells were analyzed by light and electron (transmission and scanning) microscopy, immunohistochemistry, electrophoresis and Western blot analysis, flow cytometry, and reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: IOBA-NHC cells showed high proliferative ability in vitro and typical epithelial morphology. Cytokeratins and GalNAc, GluNAc, mannose, and sialic acid residues were immunodetected in these cells. No contaminating cell types were found. MUC1, -2, and -4, but not -5AC or -7 mucin genes were expressed in every cell passage tested. Exposure of cells to inflammatory mediators (IFNgamma and/or TNFalpha) resulted in increased expression of intercellular adhesion molecule (ICAM)-1 and HLA-DR. CONCLUSIONS: Morphologic and functional characterization of the nontransfected, spontaneously immortalized IOBA-NHC cell line shows that this new cell line may be a useful experimental tool in the field of ocular surface cell biology.

Biomarkers↗

PubMed2006

Characterizing acoustic attenuation of homogeneous media using focused impulsive acoustic radiation force.

A new method to characterize a material's attenuation using acoustic radiation force is proposed. Comparison of displacement magnitudes generated in a homogeneous material by acoustic radiation force excitations can be used to estimate the material's attenuation when the excitations are applied over a range of focal depths while maintaining a constant lateral focal configuration. Acoustic attenuations are related to the inverse of the excitation focal depth that yields the greatest focal zone displacement for this protocol. Experimental studies in calibrated tissue-mimicking phantoms are presented to demonstrate the feasibility of this method. Attenuations ranging from 0.3-1.5 dB/cm/MHz were characterized over excitation focal depths ranging from 5-30 mm, with an accuracy of 0.1 +/- 0.15 dB/cm/MHz. As currently implemented, this method is limited to characterizing materials that have homogeneous material properties and acoustic attenuations. This method for characterizing acoustic attenuation can be performed using conventional diagnostic scanners without any additional hardware and could also be performed concurrently with acoustic radiation force-based imaging modalities to generate images of mechanical properties and attenuation that are spatially co-registered with B-mode images.

Acoustics↗

PubMed1999

A method for experimental characterization of the noise performance of elastographic systems.

Characterization of the noise performance of the elastographic system is necessary to evaluate the accuracy and precision of the estimated strain. The elastographic system includes the ultrasonic scanner, the computer controlled compression device and the strain estimation algorithm. In this paper, we present a method of characterizing the elastographic system experimentally using a uniformly elastic homogenous tissue-mimicking phantom. The strain response of the elastographic system is evaluated by characterizing the accuracy and precision of the strain estimates for a large range of input strains. The experimental results obtained follow the theoretical predictions obtained using the Strain Filter (SF) concept for the cross-correlation based strain estimator. In this paper, we illustrate the application of the Experimental Strain Response (ESR) to characterize strain estimation at the focus of the transducer and the axis of symmetry of the phantom.

Elasticity↗

PubMed2003

Characterization of the nodulation plasmid encoded chemoreceptor gene mcpG from Rhizobium leguminosarum.

BACKGROUND: In general, chemotaxis in Rhizobium has not been well characterized. Methyl accepting chemotaxis proteins are sensory proteins important in chemotaxis of numerous bacteria, but their involvement in Rhizobium chemotaxis is unclear and merits further investigation. RESULTS: A putative methyl accepting chemotaxis protein gene (mcpG) of Rhizobium leguminosarum VF39SM was isolated and characterized. The gene was found to reside on the nodulation plasmid, pRleVF39d. The predicted mcpG ORF displayed motifs common to known methyl-accepting chemotaxis proteins, such as two transmembrane domains and high homology to the conserved methylation and signaling domains of well-characterized MCPs. Phenotypic analysis of mcpG mutants using swarm plates did not identify ligands for this putative receptor. Additionally, gene knockouts of mcpG did not affect a mutant strain's ability to compete for nodulation with the wild type. Notably, mcpG was found to be plasmid-encoded in all strains of R. leguminosarum and R. etli examined, though it was found on the nodulation plasmid only in a minority of strains. CONCLUSIONS: Based on sequence homology R. leguminosarum mcpG gene codes for a methyl accepting chemotaxis protein. The gene is plasmid localized in numerous Rhizobium spp. Although localized to the sym plasmid of VF39SM mcpG does not appear to participate in early nodulation events. A ligand for McpG remains to be found. Apparent McpG orthologs appear in a diverse range of proteobacteria. Identification and characterization of mcpG adds to the family of mcp genes already identified in this organism.

Amino Acid Sequence↗

PubMed2006

A broadly applicable method to characterize large DNA viruses and adenoviruses based on the DNA polymerase gene.

BACKGROUND: Many viral pathogens are poorly characterized, are difficult to culture or reagents are lacking for confirmatory diagnoses. We have developed and tested a robust assay for detecting and characterizing large DNA viruses and adenoviruses. The assay is based on the use of degenerate PCR to target a gene common to these viruses, the DNA polymerase, and sequencing the products. RESULTS: We evaluated our method by applying it to fowl adenovirus isolates, catfish herpesvirus isolates, and largemouth bass ranavirus (iridovirus) from cell culture and lymphocystis disease virus (iridovirus) and avian poxvirus from tissue. All viruses with the exception of avian poxvirus produced the expected product. After optimization of extraction procedures, and after designing and applying an additional primer we were able to produce polymerase gene product from the avian poxvirus genome. The sequence data that we obtained demonstrated the simplicity and potential of the method for routine use in characterizing large DNA viruses. The adenovirus samples were demonstrated to represent 2 types of fowl adenovirus, fowl adenovirus 1 and an uncharacterized avian adenovirus most similar to fowl adenovirus 9. The herpesvirus isolate from blue catfish was shown to be similar to channel catfish virus (Ictalurid herpesvirus 1). The case isolate of largemouth bass ranavirus was shown to exactly match the type specimen and both were similar to tiger frog virus and frog virus 3. The lymphocystis disease virus isolate from largemouth bass was shown to be related but distinct from the two previously characterized lymphocystis disease virus isolates suggesting that it may represent a distinct lymphocystis disease virus species. CONCLUSION: The method developed is rapid and broadly applicable to cell culture isolates and infected tissues. Targeting a specific gene for in the large DNA viruses and adenoviruses provide a common reference for grouping the newly identified viruses according to relatedness to sequences of reference viruses and the submission of the sequence data to GenBank will build the database to make the BLAST analysis a valuable resource readily accessible by most diagnostic laboratories. We demonstrated the utility of this assay on viruses that infect fish and birds. These hosts are phylogenetically distant from mammals yet, sequence data suggests that the assay would work equally as well on mammalian counterparts of these groups of viruses. Furthermore, we demonstrated that obtaining genetic information on routine diagnostic samples has great potential for revealing new virus strains and suggesting the presence of new species.

Adenoviridae↗

PubMed2004

Applying a nonlinear regression model to characterize cortisol responses to corticotropin-releasing hormone challenge.

We previously applied a pharmacokinetic model to characterize stress-induced cortisol curves. In the current study, we apply this pharmacokinetic model to characterize cortisol responses to CRH challenge in children aged 6-16. Two hundred eleven sessions were completed beginning between 4 and 5 pm. Sessions included 30-40 minutes preinfusion baseline, 1 mug/kg CRH infusion, 90-180 minutes of postinfusion measures, and 9-10 plasma cortisol samples per session. A simpler version of the original oral dose model best-fit responses to the CRH challenge: Response = alpha + beta*Time + lambda*T*exp(-kappa*T). Time was standardized so that CRH infusion was at time 0. T = max (0, Time) such that the nonlinear departure from baseline only operated in the postinfusion phase. Alpha represents baseline cortisol at time of infusion, beta is the slope of the linear approximation of the diurnal rhythm, lambda controls magnitude of peak change from baseline, and kappa is reactivity rate. The model converged and appeared to provide a good fit to "real-world" data in healthy children. Our proposed model provides a detailed characterization of cortisol response to CRH infusion and should allow for increased flexibility in characterizing the influence of individual and situational variables on specific aspects of cortisol response curves.

Adolescent↗

PubMed2007

Image analysis by pulse coupled neural networks (PCNN)--a novel approach in granule size characterization.

A biologically inspired spiking neural network model, the pulse coupled neural network (PCNN), has been applied for the first time in bulk particle characterization, and specifically in the characterization of pharmaceutical granule size distributions. The PCNN was trained on surface images of pharmaceutical granule beds, and the adjustable parameters (radius neuron interconnection, r0, linking weight coefficient, beta, local threshold potential, VTheta, and number of iterations) were successfully optimized using design of experiments. As demonstrated with size fractions of granules, it was found that the PCNN produced granule size-dependent signals. In general, a first highest and relatively narrow peak located in the region of two to twelve iterations corresponded to smaller particle size, while larger particles resulted in wider peaks and in highest (not first) peak at a range between 13 and 25 iterations. Better predictions, i.e. lower RMSEP (root mean squared error of prediction) values, were obtained using high beta value, low r0 and VTheta values, while the number of iterations had to exceed 110 and the optimized model (RMSEP lower than 5) corresponded to PCNN variables: r0=1, beta=0.4, VTheta=2, and number of iterations=150. The coefficient of determination (R2) of the model was 0.94 and the predicted variation (Q2) was 0.91, while the Pearson correlation coefficient between the predicted and the measured mean particle size by sieving for eight test batches was 0.98. These findings could be characterized as promising and encouraging for the further use of image analysis by PCNNs in pharmaceutical bulk particle size and shape characterization.

Image Processing, Computer-Assisted↗

PubMed1999

Three-dimensional surface characterization for orthopaedic joint prostheses.

This study attempts to investigate a range of 'better' methods for the characterization of the three-dimensional (3D) surface topography of orthopaedic joint prostheses. In this paper, a new characterization tool for the comprehensive identification and evaluation of functional features of these surface topographies is presented. For identification, the surface topography is investigated in a space-scale space, by employing wavelet analysis. The roughness, waviness and form involved in surface topography are consequently separated and recovered respectively. The multiscalar topographical features are identified and captured. The errors caused as a consequence of three-dimensional measurement methods can be reduced. After identification, the three-dimensional surface assessment techniques previously reported by Stout and co-workers are used for the quantitative evaluation of various surface roughness features of the orthopaedic joint prostheses. Moreover, the functional properties, such as bearing area, material volume and void volume which are significantly effected by large peaks, pits and scratches are studied and the location of isolated peaks, pits and scratches in the different scales is also clearly characterized. In this work, measurement of the femoral heads and acetabular cups is carried out to demonstrate the applicability of the characterization technique for the three-dimensional surface topography of orthopaedic joint prostheses.

Hip Prosthesis↗

PubMed1993

Complex mixtures of air pollutants: characterizing the cancer risk of polycyclic organic matter.

Complex mixtures of polycyclic organic matter (POM) are used to illustrate the scientific problems and issues associated with characterizing the comparative risk of related complex mixtures. The complexity of mixtures in which the active components are not well characterized present special challenges, which include identifying the critical components of mixtures, their sources, and the appropriate biomarker(s) of exposure and dose; developing the appropriate experimental models for dose-response assessment; species extrapolation; and developing a scientific basis for predicting from one mixture to another. Strategies for addressing these issues include bioassay-directed chemical characterization of bioactive components of complex mixtures, apportionment methods to determine the source of biological activity and risk, DNA adduct methods to determine tissue exposure and target dose of mixtures, and comparative approaches to determining the relative similarity, potency, and risk of complex mixtures. Epidemiological data are available for humans exposed to POM from coke ovens, coal roofing tar, coal smoke, aluminum smelters, and cigarette smoke. These emissions are characterized and compared to POM from automotive emissions (diesel and gasoline), woodstove emissions, residential oil furnace emissions, and ambient air particles. The tumor potency and estimated cancer risks for these POM mixtures ranges over nearly three orders of magnitude.

Air Pollutants↗

PubMed1995

Approaches for exposure characterization and data needs for hazardous waste site assessment.

This article provides an understanding of the approaches for determining exposure and dose to populations in the vicinity of hazardous waste sites. A review of the federal legislation and jurisdiction for assessments is provided, and the approaches of the U.S. Environmental Protection Agency and the Agency for Toxic Substances and Disease Registry are compared. These methods strive to aid in the evaluation of public health impacts of contaminants that were, are, or may be released to the community, and they are concerned with various aspects of the contaminant fate, human contact, and toxic response for chemicals of concern. Such approaches have been designed for generic contamination scenarios, but they aim to be applicable to a wide range of chemicals and sites in the real world. Along with any modeling framework for exposure and dose characterization, detailed information or real data are requisite for the completion of any site-specific assessment. What kinds of data are needed and where they may be found are also discussed. A comprehensive framework for exposure characterization, recently proposed by Georgopoulos and Lioy, is outlined. The framework is one employing the following elements: chemodynamic analyses of sources and receptors; characterization of the target population; toxicokinetic/toxicodynamic analyses; uncertainty/error analyses; and evaluation of the characterization performance.

Environmental Exposure↗

PubMed2003

Characterization of C4-2 prostate cancer bone metastases and their response to castration.

UNLABELLED: New well-characterized preclinical models of prostate cancer (CaP) bone metastases are needed to improve our understanding of the development of CaP-related bone disease in patients. Here we describe characterization of a model consisting of direct injection of C4-2 cells into tibias. INTRODUCTION: Prostate cancer (CaP) has a high proclivity to metastasize to bone. Development and characterization of preclinical models of CaP bone metastases are of high interest. The objective of this study was to characterize C4-2 bone metastases and their response to castration. MATERIALS AND METHODS: Cell suspensions of C4-2, a subline of LNCaP, were injected directly into the tibias of intact male mice. In groups A (n = 7) and B (n = 5), animals were killed 3 and 8 weeks after injection of C4-2 cells, respectively. In group C (n = 7), animals were castrated 3 weeks after injection and killed 5 weeks after castration. Serum prostate-specific antigen (PSA) levels and bone mineral density (BMD) were measured, and bone histomorphometric analysis was performed. RESULTS: C4-2 cells decreased BMD of the injected tibias by 36.1% and bone volume by 74.1% versus normal tibias. Castration caused a 32.3% drop in serum PSA (p = 0.0438), with a nadir at day 14, after which it began to rise again. Bone destruction in the tumorous tibias of castrated animals was decreased by 15.9% versus tumorous tibias of intact animals (p = 0.0392). However, BMD in the tumorous tibias of castrated mice was still lower than in normal tibias of intact animals. Castration also decreased BMD and bone volume in nontumorous tibias (p = 0.0406 and 0.0232, respectively). CONCLUSIONS: The C4-2 model of bone metastasis recapitulates the response to androgen deprivation observed in CaP patients with bone metastases and is suitable for study of interactions between tumor and bone cells and evaluation of new therapeutic modalities.

Animals↗

PubMed2004

Immunohistochemical characterization of cloned lamb nephropathy.

Kidneys from lambs derived by nuclear transfer are frequently abnormal and are characterized by an enlarged pelvis and narrow medulla, consistent with lower urinary tract obstruction and development of variable hydronephrosis. The precise pathogenesis of this entity is unknown. Immunohistochemical staining for intermediate filaments was used to further characterize the lesions seen in this condition and was compared with age-matched control tissue. Major findings were upregulation of cytokeratin on damaged tubules, desmin and vimentin in undifferentiated mesenchyme, and smooth muscle actin in mesenchyme and on smooth muscle "collars" around dilated tubules. In addition, some cases showed reexpression of vimentin and desmin on proximal tubular epithelial cells. Taken together, these findings provide a valuable database for tracking the expression of intermediate filaments throughout renal development in sheep and have further characterized the nature of the response to injury by the developing kidney, a response that is characterized by proliferation of mesenchyme and both reexpression and upregulation of intermediate filaments within renal cells. In addition, the study has confirmed that the changes in cloned lamb nephropathy are established by day 85 of development.

Actins↗

PubMed2003

Detection and characterization of cellular immune responses using peptide-MHC microarrays.

The detection and characterization of antigen-specific T cell populations is critical for understanding the development and physiology of the immune system and its responses in health and disease. We have developed and tested a method that uses arrays of peptide-MHC complexes for the rapid identification, isolation, activation, and characterization of multiple antigen-specific populations of T cells. CD4(+) or CD8(+) lymphocytes can be captured in accordance with their ligand specificity using an array of peptide-MHC complexes printed on a film-coated glass surface. We have characterized the specificity and sensitivity of a peptide-MHC array using labeled lymphocytes from T cell receptor transgenic mice. In addition, we were able to use the array to detect a rare population of antigen-specific T cells following vaccination of a normal mouse. This approach should be useful for epitope discovery, as well as for characterization and analysis of multiple epitope-specific T cell populations during immune responses associated with viral and bacterial infection, cancer, autoimmunity, and vaccination.

Animals↗

PubMed2006

Dynamic phenotypes: time series analysis techniques for characterizing neuronal and behavioral dynamics.

We consider quantitative measures of behavioral and neuronal dynamics as a means of characterizing phenotypes. Such measures are important from a scientific perspective; because understanding brain function is contingent on understanding the link between the dynamics of the nervous system and behavioral dynamics. They are also important from a biomedical perspective because they provide a contrast to purely psychological characterizations of phenotype or characterizations via static brain images or maps, and are a potential means for differential diagnoses of neuropsychiatric illnesses. After a brief presentation of background work and some current advances, we suggest that more attention needs to be paid to dynamic characterizations of phenotypes. We will discuss some of the relevant time series analysis tools.

Action Potentials↗

PubMed2006

Exact analysis of heterotropic interactions in proteins: Characterization of cooperative ligand binding by isothermal titration calorimetry.

Intramolecular interaction networks in proteins are responsible for heterotropic ligand binding cooperativity, a biologically important, widespread phenomenon in nature (e.g., signaling transduction cascades, enzymatic cofactors, enzymatic allosteric activators or inhibitors, gene transcription, or repression). The cooperative binding of two (or more) different ligands to a macromolecule is the underlying principle. To date, heterotropic effects have been studied mainly kinetically in enzymatic systems. Until now, approximate approaches have been employed for studying equilibrium heterotropic ligand binding effects, except in two special cases in which an exact analysis was developed: independent binding (no cooperativity) and competitive binding (maximal negative cooperativity). The exact analysis and methodology for characterizing ligand binding cooperativity interactions in the general case (any degree of cooperativity) using isothermal titration calorimetry are presented in this work. Intramolecular interaction pathways within the allosteric macromolecule can be identified and characterized using this methodology. As an example, the thermodynamic characterization of the binding interaction between ferredoxin-NADP+ reductase and its three substrates, NADP+, ferredoxin, and flavodoxin, as well as the characterization of their binding cooperativity interaction, is presented.

Algorithms↗

PubMed2005

Genome-wide characterization of tetrahymena thermophila chromosome breakage sites. I. Cloning and identification of functional sites.

The chromosomes of the macronuclear (expressed) genome of Tetrahymena thermophila are generated by developmental fragmentation of the five micronuclear (germline) chromosomes. This fragmentation is site specific and directed by a conserved 15-bp chromosome breakage sequence (Cbs element). This article reports the construction of a library enriched for chromosome breakage junctions and the development of a successful scheme for the genome-wide isolation and characterization of functional Cbs junctions. Twenty-three new Cbs junctions were characterized and each was assigned to a specific micronuclear chromosome or chromosome arm. Two distinct previously unreported variant chromosome breakage sequences were found, each in two or more functional Cbs elements. Analysis of natural Cbs junctions confirmed that microheterogeneity in the macronuclear telomere addition site is associated with chromosome fragmentation. The physical and genetic characterization of these functional chromosome breakage junctions is reported in the accompanying article in this issue. The whole-genome shotgun sequencing and auto-assembly phase of the Tetrahymena Genome Initiative has recently been completed at The Institute for Genome Research (TIGR). By providing unique sequence from the natural ends of macronuclear chromosomes, Cbs junctions characterized in the work reported here will serve as useful sequence tags for relating macro- and micronuclear genetic, physical, and sequence maps.

Animals↗