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PubMed2003

Expression of Clostridium difficile toxins A and B and their sigma factor TcdD is controlled by temperature.

Growth temperature was found to control the expression of toxins A and B in Clostridium difficile VPI 10463, with a maximum at 37 degrees C and low levels at 22 and 42 degrees C in both peptone yeast (PY) and defined media. The up-regulation of toxin A and B mRNA and protein levels upon temperature upshift from 22 to 37 degrees C followed the same kinetics, showing that temperature control occurred at the level of transcription. Experiments with Clostridium perfringens using gusA as a reporter gene demonstrated that both toxin gene promoters were temperature controlled and that their high activity at 37 degrees C was dependent on the alternative sigma factor TcdD. Furthermore, tcdD was found to be autoinduced at 37 degrees C. Glucose down-regulated all these responses in the C. perfringens constructs, similar to its impact on toxin production in C. difficile PY broth cultures. C. difficile proteins induced at 37 degrees C and thus coregulated with the toxins by temperature were demonstrated by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified as enzymes involved in butyric acid production and as electron carriers in oxidation-reduction reactions. The regulation of toxin production in C. difficile by temperature is a novel finding apparently reflecting an adaptation of the expression of its virulence to mammalian hosts.

Bacterial Proteins↗
PubMed1991

RNA transcription and translation in the hearts of normal and cardiomyopathic Syrian hamsters.

The cardiomyopathic Syrian hamster has an autosomal recessive defect that results in the development of an early onset cardiac myopathy leading to cardiac dysfunction and, eventually, complete heart failure. To assess the regulatory mechanisms modulating gene expression in the normal and myopathic myocardium, we investigated both RNA transcription and translation. Our results indicated that the incorporation of [3H]UMP into myocardial cell nuclear RNA decreased 10-fold from 7 to 210 days of age in the normal Syrian hamster. The incorporation of [3H]UMP was approximately 50% lower in the cardiomyopathic as compared with the normal Syrian hamster. RNA translation, as assessed by rabbit reticulocyte lysate in vitro translation, indicated that a coordinated 50% decrease in RNA translation occurred in normal Syrian hamster from 7 to 210 days of age. A further reduction of 20% in translation was found in cardiomyopathic Syrian hamster ventricular RNA translation as compared with matched random bred control groups. Two-dimensional polyacrylamide gel analysis of cell-free translated protein products demonstrated two myocardial peptides that were found to be consistently altered when the normal and cardiomyopathic Syrian hamsters were compared. These results indicate that transcription and translation decrease with age and that these processes are further downregulated, in an additive manner, with the genesis of the disease process.

Aging↗
PubMed2004

Extracellular and circulating redox- and metalloregulated eRNA and eRNP: copper ion-structured RNA cytokines (angiotropin ribokines) and bioaptamer targets imparting RNA chaperone and novel biofunctions to S100-EF-hand and disease-associated proteins.

Bioassays for cellular differentiation and tissue morphogenesis were used to design methods for isolation of bioactive redox- and metalloregulated nucleic acids and copper ion complexes with proteins from extracellular, circulating, wound, and supernatant fluids of cultured cells. In extracellular biospheres, diversities of nucleic acids were found to be secreted by cells upon activation. They may reflect nucleic acid biolibraries with molecular imprints of cellular history. After removal of protein components, eRNA prototypes exuded by activated cells were sequenced. They are small, endogenous, highly modified and edited, redox- and metalloregulated 5'-end phosphorylated extracellular eRNA (approximately 2-200 bases) with cellular, enzymic, and bioaptamer functions. Fenton-type OH* radical redox reactions may form modified nucleotides in RNA as wobbles eRNA per se, or as copper ion-complex with protein (e.g., S100A12-EF-hand protein, angiotropin-related protein, calgranulin-C, hippocampal neurite differentiation factor) are shown to be bioactive in vivo and in vitro as cytokines (ribokines) and as nonmitogenic angiomorphogens for endothelial cell differentiation in the formation of organoid supracellular capillary structures. As bioaptamers, copper ion-structured eRNA imparts novel biofunctions to proteins that they do not have on their own. The origin of extracellular RNA and intermediate precursors (up to 500 bases) was traced to intracellular parent nucleic acids. Intermediate precursors with and without partial homology were found. This suggests that bioaptamers are not directly retranslatable gene products. Metalloregulated eRNA bioaptamer function was investigated by domains (e.g. 5'...CUG...3' hairpin loop) for folding, bioactivity, and binding of protein with copper, calcium, and alkali metal ion affinity. Vice versa, metalloregulated nucleic acid-binding domains (K3H, R3H) in proteins were identified. Interaction of protein and eRNA docking potentials were visualized by 3D-rapid prototyping of accurate molecular image models based on crystallographic or NMR data. For S100A12-homologous proteins, receptor- and metalloregulated RNA chaperone-shaped protein assemblies were investigated. They suggest insight into signaling cascades as to how eRNA transmits its cytokine (ribokine) bioinformation from the extracellular RNA biosphere into cells. Proteomics of the extracellular RNA biosphere demonstrate the presence of nucleic acid-binding domain homologies in defense-, aging-, and disease-associated neuronal and other proteins as targets for RNA orphans. By structural relationships found to transmissible processes, proteinaceous transfer ("infectivity") and feedback of bioinformation beyond the central dogma of molecular biology are considered in terms of metalloregulated RNA bioaptamer function, nucleic acid-binding domains, and protein conformation.

Amino Acid Sequence↗
PubMed2000

Interactions of regional respiratory mechanics and pulmonary ventilatory impairment in pulmonary emphysema: assessment with dynamic MRI and xenon-133 single-photon emission CT.

STUDY OBJECTIVES: Dynamic MRI and (133)Xe single-photon emission CT (SPECT) were used to directly evaluate the interaction of regional respiratory mechanics and lung ventilatory function in pulmonary emphysema. METHODS: Respiratory diaphragmatic and chest wall (D/CW) motions were analyzed by sequential MRI of fast-gradient echo pulse sequences during two to three respiratory cycles in 28 patients with pulmonary emphysema, including 9 patients undergoing lung volume reduction surgery (LVRS). The extent of air trapping in the regional lung was quantified by the (133)Xe retention index (RI) on three-dimensional (133)Xe SPECT displays. RESULTS: By contrast to healthy subjects (n = 6) with regular, synchronous D/CW motions, pulmonary emphysema patients showed reduced, irregular, or asynchronous motions in the hemithorax or location with greater (133)Xe retention, with significant decreases in the maximal amplitude of D/CW motions (MADM and MACWM; p < 0.0001 and p < 0.05, respectively). The removal of (133)Xe retention sites by LVRS effectively and regionally improved D/CW motions in nine patients, with significant increases in MADM and MACWM (p < 0.01 and p < 0.001, respectively). In a total of 40 studies of the 28 patients including post-LVRS studies, normalized MADM and MACWM correlated with percent predicted FEV(1) (r = 0.881, p < 0.0001; and r = 0.906, p < 0.0001, respectively), and also with (133)Xe RI in each hemithorax (r = -0.871, p < 0 0.0001; and r = -0.901, p < 0 0.0001, respectively.) CONCLUSIONS: This direct comparison of regional respiratory mechanics with lung ventilation demonstrated a close interaction between these impairments in pulmonary emphysema. The present techniques provide additional sensitivity for evaluating pathophysiologic compromises in pulmonary emphysema, and may also be useful for selecting resection targets for LVRS and for monitoring the effects.

Aged↗
PubMed1987

Generation of oligomeric insulin receptor forms by intramolecular sulfhydryl-disulfide exchange. Involvement of masked sulfhydryl groups.

Insulin receptors from rat liver membranes were labelled with a 125I-labelled photoreactive insulin analogue or by iodination using lactoperoxidase and analysed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Under nonreducing conditions different receptor forms with Mr 400,000 (alpha 2 beta 2), 360,000 (alpha 2 beta beta'), 330,000 (alpha 2 beta' beta'), 320,000 (alpha 2 beta), 280,000 (alpha 2 beta'), 240,000 (alpha 2), 210,000 (alpha beta), 165,000 (alpha beta') and 115,000 (alpha) were detected. The subunit composition of these receptor forms was determined by two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis in the absence and presence of dithioerythritol. During denaturation in sodium dodecyl sulfate in the absence of reductants, the Mr 400,000 receptor form (alpha 2 beta 2) was converted into the Mr 320,000 (alpha 2 beta) and Mr 240,000 (alpha 2) receptor form. This conversion was prevented either by N-ethylmaleimide, oxidants, or low pH. In contrast, alkylation of the receptor with N-ethylmaleimide under non-denaturing conditions did not prevent the appearance of intermediate-sized receptor forms. Furthermore, the inhibition of receptor cleavage by N-ethylmaleimide during denaturation was also observed when the amount of free sulfhydryl groups was reconstituted by the addition of an unlabelled and non-alkylated receptor sample to the alkylated and photoaffinity-labelled receptor. These results suggest, that the generation of different oligomeric receptor forms detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis is due at least in part to the cleavage of one or both beta-subunits from the insulin receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Affinity Labels↗
PubMed2000

[A tridimensional study of lateral grasping and pointing in spatial neglect].

The aim of this study was to analyze the ipsilateral upper limb and head kinematics in patients presenting with spatial neglect, using pointing and grasping to object. Four patients were included and compared to an equivalent number of normal subjects. Head, shoulder, and wrist movements were recorded using a three-dimensional VICON system. Objects to be pointed or grasped were localized in the right or left space facing the subject. Patients presented with a reduction in the mean wrist velocity, which was more obvious when directed to the left side, reduction in the peak velocity, and a severe increase of the time between the peak velocity and the end of the movement. The amplitude and mean velocity of left head movements were comparable in both groups. The analysis of the intersegmental co-ordination showed that the sequence of activation of corporeal segments was similar in patients and controls: head movement, shoulders rotation, upper limb extension. Co-ordination between head and wrist movements was assessed using the cross-correlation method. An increase in time lag and a reduction in crossed correlations between the head and wrist movements was observed in leftward movements of neglect patients, but the co-ordination between the shoulder and wrist movements was preserved. Thus, we demonstrated a global disorder of intentional movements in patients, which was relatively similar in pointing and grasping to object, and predominated on the approach phase, which is associated to important visuo-motor adjustments. This study also demonstrated desynchronization of head and wrist movements, which can be an important problem in neglect patients.

Adult↗
PubMed2001

Implicit learning of sequences of tasks.

Task sets can be configured in advance of performing a new task. However, the degree to which advance information is actually used for advance configuration depends on the nature of the available information. The role of implicit learning was explored in 2 experiments by means of a modified serial reaction time task with repeated sequences of 4 dimensionally organized tasks. Although there was clear evidence for implicit learning of the sequence (of length 8), the learning was not associated with a reduction of shift costs, either with a short (200 ms) or with a long (1,200 ms) response-stimulus interval. In contrast, a reduction of shift costs was observed when external precues were introduced in a 3rd experiment. According to these results, the sequences of stimulus features that serve as cues for the tasks to perform on the stimuli are learned, but the representation of the features is void of their task-associated meanings.

Adult↗
PubMed1990

[Effects of magnesium pidolate on cardiovascular hemodynamics].

Aim of this study is to define the hemodynamic effects of magnesium (Mg) in patients with ischemic heart disease. Thus, the effects of 1 g of Mg pidolate injected over 3 min iv were evaluated in 10 patients by analysis of pressure data from a micromanometer catheter in the left ventricle and of volume data derived from Doppler and 2-dimensional echocardiography. Recordings were made before and immediately after the injection, at 3, 5, and 8 min. At 5 min, Mg induced a significant reduction in left ventricular peak systolic pressure (from 132.4 +/- 19.4 to 122.7 +/- 17.5 mmHg, p = 0.015, with early filling notably augmented (E wave of Doppler curve; from 39.4 +/- 8.5 to 43.9 +/- 9.7 cm/s, p = 0.013) and a slight increase in cardiac output (from 6.23 +/- 1.4 to 6.9 +/- 1.7 l/min, p = 0.092). Heart rate increased (from 75.9 +/- 9.6 to 78.13 +/- 8.4 b/min, p = 0.062) and the constant of isovolumetric pressure decay decreased (from 48.9 +/- 13.2 to 45.7 +/- 14.9 ms, p = 0.065), probably as a final effect of aortic pressure reduction. The diastolic pressure-volume relationship was insignificantly displaced downward (p = 0.11) and to the right (p = 0.62). In conclusion, Mg unloads the ischemic left ventricle through a reduction of systemic aortic pressure, with a secondary increase in early filling; this increase may derive from a rise in the atrioventricular diastolic gradient, subsequent to an improvement in ventricular emptying. Active relaxation is improved, and passive diastolic properties are not affected, in spite of the reported calcium-antagonist action of the drug.

Adult↗
PubMed1982

Effect of interferons on protein synthesis in human lymphocytes: enhanced synthesis of eight specific peptides in T cells and activation-dependent inhibition of overall protein synthesis.

Lymphoblastoid and fibroblast IFN inhibited PHA stimulation of overall protein synthesis in human lymphocytes by ca. 30%. Inhibition occurred within the first 6 hr of PHA treatment and was not progressive. DNA synthesis at 48 hr was inhibited to the same extent. Overall protein synthesis in resting lymphocytes was not detectably inhibited by IFN concentrations up to 1000 U/ml. Thus, inhibition of protein synthesis and subsequent reduction of cell proliferation by IFN require certain early events in mitogen activation. Resting lymphocytes were not unresponsive to IFN treatment, however. Two-dimensional electrophoretic analysis of newly synthesized proteins after IFN treatment showed enhanced synthesis of a specific set of eight peptides (I-peptides), which were shown to be synthesized in T lymphocytes. This enhancement was produced by both IFN-alpha and IFN-beta after 4 to 6 hr of exposure and was identical for all lymphocyte donors studied. After growth stimulation, IFN treatment produced no enhancements of additional peptides, although the original eight I-peptides were enhanced as usual. It is concluded that the biochemical activities of the I-peptides, which remain to be determined, cannot inhibit protein synthesis in resting lymphocytes, but may do so after mitogen activation, when the major physiologic restriction of lymphocyte protein synthesis is released. Alternatively, the I-peptides may be unrelated to regulation of protein synthesis but may be involved in viral protection or enhancement of NK activity.

Blood Proteins↗
PubMed1995

[Structure of an acidic polysaccharide found in a bacteriolytic complex of lysoamidase].

A structural study of an acidic polysaccharide, a component of the lysoamidase bacteriolytic complex has been carried out. Analysis of the monosaccharide composition of the original polysaccharide, of the product of carboxyl groups reduction and of the 13C-NMR and 1H-NMR spectra of the original and the O-deacetylated polysaccharides using two-dimensional NMR spectroscopy has made it possible to establish the structure of the repeating trisaccharide unit of the polysaccharide as follows: [formula: see text] where ManNAcA and GalNAcA are 2-acetamido-2-deoxymannuronic acid and 2-acetamido-2-deoxygalacturonic acid, respectively. Also small amount of a neutral polysaccharide containing of rhamnose is present in the lysoamidase preparation.

Bacteria↗
PubMedDate not supplied

A three-dimensional finite element study on two versus four implants in an edentulous mandible.

The difference in stress distribution in the bone around dental implants was determined under conditions where either two or four implants were placed in the interforaminal region of the mandible. The purpose was also to determine the influence of the height of the mandible on the stress distribution. The stress distribution around the implants was investigated using three-dimensional finite element models of the interforaminal region of two mandibles with two different stages of resorption. It was concluded that there is no reduction of the extreme principal stresses when the load is distributed over an increasing number of implants. There is a tendency for a gradual stress increase as mandibular height decreases.

Alveolar Bone Loss↗
PubMed1993

[Intraarticular fractures of the calcaneus: pathological description. Contribution of x-ray computed tomography].

Computed tomography (CT) is of cardinal importance in the study of displaced intra-articular fractures of the calcaneus, as with this imaging technique the primary fracture line of Palmer can be followed and its various locations visualised. If the three-stage classification system of these fractures as vertical, horizontal or mixed fractures, based on their radiological appearance, is adopted, a correlation can be established between the location of Palmer's line and these three anatomical types. The fundamental fracture line is medial in the vertical types, lateral in the horizontal types and is located in the centre of the posterior facet in the mixed types. It can be seen that the fundamental fracture line separates a laterally detached fragment which is always tilted, giving a vertical slope to the surface of the posterior facet which it supports and thus giving a vertical image. This fragment is itself separated from a medially detached fragment which is tilted horizontally and produces a horizontal image on the lateral view. The CT sections must be examined for the presence of a secondary sagittal line completely isolating the horizontally tilted fragment. The importance of the location of the fundamental fracture line, which is on the borderline between tilting and horizontalisation, suggests that pronation-supination of the foot is a factor in the causal mechanism of these fractures. A clear understanding of the three-dimensional position of the fragments and of their displacement, essentially defined by the location of the fracture line, is a prequisite before attempting surgical reduction of calcaneal fractures.

Calcaneus↗
PubMed2005

Does three-dimensional computer simulation improve results of scaphoid nonunion surgery?

The purpose of this study was to clarify the effectiveness of the three-dimensional computer simulations in scaphoid nonunion surgery. Seven consecutive clinical patients with scaphoid nonunion at the middle third comprised the study group. Surface models of the scaphoid were constructed on the computer using computed tomography data of the bilateral wrists in neutral position. The distal and proximal fragments of the nonunion model were matched to the mirror image of the contralateral scaphoid model. The rotation of the distal fragment relative to the proximal fragment was calculated, and reduction of the displaced fragment of the scaphoid nonunion was simulated. Similarly, the estimated bone defect and the appropriate site and direction of the screw insertion were simulated. Full-sized hard models of the bone, including a model with simulated reduction and screw insertion, then were made using stereolithography based on the computer data. In the actual surgery, reduction, bone grafting, and screw insertion were achieved using the hard models as guides. All the patients obtained solid bone fusion and substantial clinical improvement with normalized scapholunate and radiolunate angles after surgery. Three-dimensional computer simulations were found as useful for accurate correction of scaphoid nonunions and proper screw placement, which consequently leads to good clinical results.

Adolescent↗
PubMed2005

Analysis of Lupinus albus storage proteins by two-dimensional electrophoresis and mass spectrometry.

A laboratory-prepared total protein extract (TPE) and a lupin protein isolate (LPI-E) produced in a pilot plant were submitted to a detailed two-dimensional (2DE) proteomic investigation. Recent findings have indicated that in an established rodent model of hyperlipidemia, moderate daily intakes of LPI-Es lead to a reduction of total and low-density lipoprotein cholesterol levels, and the knowledge of the actual composition of the protein sample used in that study is at the basis of further structure/action investigations. The experimental results indicate that the semi-industrial procedure used for the production of LPI-E damages only marginally the proteins. It does, however, cleave some disulfide bridges and induce mild proteolysis, as confirmed by the higher number of resolved protein spots in the low Mr and acidic pI region of the 2DE map. Out of 72 spots submitted to mass spectrometry and compared with available protein databases, 42 correspond to fragments of beta-conglutin, the 7S globulin of lupin, spanning between positions 37 and 495 of the protein sequence. Using the bioinformatic tool BlastP, these peptides were compared to the alpha'-subunit of beta-conglycinin, the 7S globulin of soybean, this being the most active hypocholesterolemic component of soybean protein, as shown by in vitro and in vivo experiments. At least 18 peptides derived from beta-conglutin, having a percentage identity higher than 50% and a similarity percentage higher than 70% vs the alpha'-subunit of beta-conglycinin, are likely candidates to be the biologically active components of lupin protein.

Amino Acid Sequence↗
PubMed2006

The effects of statins on high-density lipoproteins.

Statins inhibit cholesterol synthesis and are effective in lowering total cholesterol levels in plasma or serum due to reductions in low-density lipoprotein and very low-density lipoproteins, as well as reducing progression of coronary atherosclerosis, coronary heart disease, and stroke morbidity and mortality. These agents also modestly raise levels of high-density lipoprotein (HDL) cholesterol and its major protein, apolipoprotein (apo) A-I. The more effective statins can also raise the levels of large alpha-1 HDL particles as assessed by two-dimensional gel electrophoresis. High levels of these particles promote reverse cholesterol transport and protect against coronary heart disease and progression of coronary atherosclerosis. The mechanism whereby statins alter HDL and its subspecies appears to be due to reduction of triglyceride-rich lipoproteins, with a secondary decrease in cholesteryl ester transfer protein activity, and less transfer of HDL cholesterol to triglyceride-rich lipoprotein acceptor particles. Increasingly, statins will be combined with other agents such as ezetimibe, fibrates, niacin, and cholesteryl ester transfer protein inhibitors to optimize the entire lipoprotein profile to alter not only low-density lipoprotein, but also HDL, triglycerides, lipoprotein(a), and C-reactive protein, and also to reduce cardiovascular morbidity and mortality.

Cardiovascular Diseases↗
PubMed2003

Three-dimensional edge-preserving image enhancement for computed tomography.

Computed tomography (CT) images exhibit a variable amount of noise and blur, depending on the physical characteristics of the apparatus and the selected reconstruction method. Standard algorithms tend to favor reconstruction speed over resolution, thereby jeopardizing applications where accuracy is critical. In this paper, we propose to enhance CT images by applying half-quadratic edge-preserving image restoration (or deconvolution) to them. This approach may be used with virtually any CT scanner, provided the overall point-spread function can be roughly estimated. In image restoration, Markov random fields (MRFs) have proven to be very flexible a priori models and to yield impressive results with edge-preserving penalization, but their implementation in clinical routine is limited because they are often viewed as complex and time consuming. For these practical reasons, we focused on numerical efficiency and developed a fast implementation based on a simple three-dimensional MRF model with convex edge-preserving potentials. The resulting restoration method provides good recovery of sharp discontinuities while using convex duality principles yields fairly simple implementation of the optimization. Further reduction of the computational load can be achieved if the point-spread function is assumed to be separable. Synthetic and real data experiments indicate that the method provides significant improvements over standard reconstruction techniques and compares well with convex-potential Markov-based reconstruction, while being more flexible and numerically efficient.

Algorithms↗
PubMed2004

Dose and volume reduction for normal lung using intensity-modulated radiotherapy for advanced-stage non-small-cell lung cancer.

PURPOSE: To investigate dosimetric improvements with respect to tumor-dose conformity and normal tissue sparing using intensity-modulated radiotherapy (IMRT) compared with three-dimensional conformal radiotherapy (3D-CRT) for advanced-stage non-small-cell lung cancer (NSCLC). METHODS AND MATERIALS: Forty-one patients with Stage III-IV and recurrent NSCLC who previously underwent 3D-CRT were included. IMRT plans were designed to deliver 63 Gy to 95% of the planning target volume using nine equidistant coplanar 6-MV beams. Inverse planning was performed to minimize the volumes of normal lung, heart, esophagus, and spinal cord irradiated above their tolerance doses. Dose distributions and dosimetric indexes for the tumors and critical structures in both plans were computed and compared. RESULTS: Using IMRT, the median absolute reduction in the percentage of lung volume irradiated to >10 and >20 Gy was 7% and 10%, respectively. This corresponded to a decrease of >2 Gy in the total lung mean dose and of 10% in the risk of radiation pneumonitis. The volumes of the heart and esophagus irradiated to >40-50 Gy and normal thoracic tissue volume irradiated to >10-40 Gy were reduced using the IMRT plans. A marginal increase occurred in the spinal cord maximal dose and lung volume >5 Gy in the IMRT plans, which could be have resulted from the significant increase in monitor units and thus leakage dose in IMRT. CONCLUSION: IMRT planning significantly improved target coverage and reduced the volume of normal lung irradiated above low doses. The spread of low doses to normal tissues can be controlled in IMRT with appropriately selected planning parameters. The dosimetric benefits of IMRT for advanced-stage non-small-cell lung cancer must be evaluated further in clinical trials.

Adult↗