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PubMed1981

Cyclic AMP-dependent protein kinase I: cyclic nucleotide binding, structural changes, and release of the catalytic subunits.

Type I cyclic AMP (cAMP)-dependent protein kinase is composed of a dimeric regulatory subunit (R(2)) and two catalytic subunits (C subunits). The R(2) dimer binds four cAMP molecules to release the two C subunits. To characterize the cAMP binding sites and elucidate their role in the release of the C subunits, the R(2) dimer has been studied by equilibrium methods. The cAMP titration of R(2) was monitored by endogenous tryptophan fluorescence, and the results suggest one class of binding sites. The titration plot is monotonic for saturation of four sites per R(2). A similar titration monitored by near-UV circular dichroic changes exhibited profound changes in the region of the (1)L(b) tyrosine and (1)L(a) and (1)L(b) tryptophan transitions; a plot of these data also showed a linear monotonic response. Thus, the fluorescence and circular dichroic changes show that cAMP binding to R(2) induces a conformational or structural change. The one apparent class of binding sites implies that all binding sites are characterized by similar K(d) values or by K(d) values much less than the receptor concentration. The reactivity of the cysteine sulfhydryl groups with 5,5'-dithiobis(2-nitrobenzoic acid) showed that saturation with cAMP indirectly protects one sulfhydryl group per R monomer. Analysis of cAMP activation of the holoenzyme, detected by phosphotransferase assays, showed that saturation of both cAMP binding sites per R monomer is necessary to effect the release of the C subunit. By using a fluorescent analog of cAMP, 1,N(6)-etheno-cyclic AMP (epsilon cAMP), the (epsilon cAMP)(4).R(2) complex was titrated with C subunit, causing the release of epsilon cAMP. The titration showed that the release of epsilon cAMP was a positive cooperative process; its Hill plot had a slope of 2.6 and the K(a1) and K(an) values obtained by extrapolation were 2.1 x 10(7) M(-1) and 5.0 x 10(8) M(-1), respectively. The calculated DeltaDeltaG for first and last site coupling was 1.9 kcal/mol (1 cal = 4.18 J) of holoenzyme.

Animals↗
PubMed1975

Structural changes in the membrane of vero cells infected with a paramyxovirus.

Vero cells productively infected with the Halle strain of measles virus have been studied by means of surface replication, freeze-fracturing, and surface labeling with horseradish peroxidase-measles antibody conjugate in order to examine changes in the structure of the cell membrane during viral maturation. Early in infection, the surfaces of infected cells are embossed by scattered groups of twisted strands, and diffuse patches of label for viral antigens cover regions marked by these strands. At later stages, when numerous nucleocapsids become aligned under the plasmalemmal strands, the strands increase in number and width and become more convoluted. At this stage, label for viral antigens on the surface of the cell membrane is organized into stripes lying on the crests of strands. Finally, regions of the membrane displaying twisted strands protrude to form ridges or bulges, and the freeze-fractured membrane surrounding these protrusions is characterized by an abundance of particles small than those found on the rest of the cell membrane. The fractured membranes of viral buds are continuous sheets of these small particles, and the spacing between both nucleocapsids and stripes of surface antigen in buds is less than in the surrounding cell membrane. Detached virus is covered with a continuous layer of viral antigen, has unusually large but no small particles on its membrane surfaces exposed by freeze-fracturing, and no longer has nucleocapsids aligned under its surface. Thus, surface antigens, membrane particles, and nucleocapsids attached to the cell membrane are mobile within the plane of the membrane during viral maturation. All three move simutaneously in preparation for viral budding.

Antigens, Viral↗
PubMed2002

Imaging saponin-induced structural changes in neural processes with atomic force microscopy.

Temporal changes in the structure of neuronal processes in the presence of saponin were studied by atomic force microscopy in a fluid medium. After saponin treatment, concavities were formed on the surface of some neurites and fibrous structures in other neurites were splayed. The vertical height of these splayed fibrils or fibrillar bundles ranged from 13 to 370 nm, and the horizontal width was less than 500 nm. These findings suggest that formation of concavities and separation of bundled fibrils occurred simultaneously in saponin-treated neurites.

Animals↗
PubMed2006

Osteogenic protein-1 injection into a degenerated disc induces the restoration of disc height and structural changes in the rabbit anular puncture model.

STUDY DESIGN: In vivo study of the effect of injection of osteogenic protein-1 (OP-1) on a rabbit anular needle puncture model of intervertebral disc (IVD) degeneration. OBJECTIVE: To study radiographic, magnetic resonance imaging (MRI), biochemical, and histologic changes in the rabbit IVD after injection of OP-1 into the nucleus pulposus in a needle puncture disc degeneration model. SUMMARY OF THE BACKGROUND DATA: Growth factors, such as OP-1, have the ability to stimulate synthesis of proteoglycans and collagen in vitro. The in vivo injection of OP-1 into the normal rabbit IVD has increased disc height and proteoglycan content in the anulus fibrosus and nucleus pulposus. However, to our knowledge, no attempts have yet been made to determine the effects of these growth factors in an in vivo model of disc degeneration. METHODS: New Zealand adolescent white rabbits (n = 90, 8 for baseline evaluation, 82 at 8 times) received an anular puncture in 2 noncontiguous discs with an 18-gauge needle to induce disc degeneration. Four weeks later, either 5% lactose (10 microL) or OP-1 (100 microg in 10 microL 5% lactose) was injected into the center of the nucleus pulposus. The disc height was followed radiographically for up to 24 weeks after the injections. At the 2, 4, 8, 12, and 24-week times after the injection, rabbits were euthanized, and MRI of the harvested spinal columns was obtained to grade the degeneration. The discs injected with OP-1 or lactose and noninjected discs were subjected to biochemical and histologic analysis. The specimens at the 24-week time were limited to histologic evaluation. RESULTS: The anular puncture with a needle induced a consistent disc narrowing within 4 weeks. The injection of OP-1 induced a restoration of disc height at 6 weeks, which was sustained for the entire experimental period, up to 24 weeks after the injection. The injection of lactose alone did not change the course of disc narrowing over the same time. MRI grading score showed significant differences between the OP-1 and lactose groups at the 8, 12, and 24-week times, suggesting an increase in water content in the nucleus pulposus of the OP-1 group. The proteoglycan content of the nucleus pulposus and anulus fibrosus was significantly higher in the OP-1 group than in the control group. The degeneration grades of the punctured discs in the OP-1 group were significantly lower than those in the lactose group. CONCLUSION: The results of this study show the feasibility of restoring degenerative rabbit discs by a single injection of OP-1 into the nucleus pulposus. Importantly, the effects of the OP-1 injection on disc height were sustained for up to 24 weeks. The metabolic changes in the cells, following a single injection, might be sustained and, thus, induce long-term changes in disc structure. An efficacy study in large animals is required to show further that the intradiscal injection of OP-1, or bone morphogenetic proteins or growth factors with similar properties would be useful for the structural restoration of the IVD in humans.

Animals↗
PubMed2006

Short exposure to high levels of fluoride induces stage-dependent structural changes in ameloblasts and enamel mineralization.

We tested the hypothesis that the sensitivity of forming dental enamel to fluoride (F-) is ameloblast developmental stage-dependent and that enamel mineralization disturbances at the surface of fluorotic enamel are caused by damage to late-secretory- and transitional-stage ameloblasts. Four-day-old hamsters received a single intraperitoneal dose of 2.5-20 mg NaF/kg body weight and were examined, 24 h later, by histology and histochemistry. A single dose of >or=5 mg of NaF/kg induced the formation of a hyper- followed by a hypomineralized band in the secretory enamel, without changing the ameloblast structure. At 10 mg of NaF/kg, cystic lesions became apparent under isolated populations of distorted late-secretory- and transitional-stage ameloblasts. Staining with von Kossa stain showed that the enamel under these lesions was hypermineralized. At 20 mg of NaF/kg, cystic lesions containing necrotic cells were also found in the early stages of secretory amelogenesis and were also accompanied with hypermineralization of the enamel surface. We concluded that the sensitivity to F- is ameloblast developmental stage-dependent. Groups of transitional ameloblasts are most sensitive, followed by those at early secretory stages. These data suggest that a F-induced increase in cell death in the transitional-stage ameloblasts accompanies the formation of cystic lesions, which may explain the formation of enamel pits seen clinically in erupted teeth.

Ameloblasts↗
PubMed1999

How locomotor parameters adapt to gravity and body structure changes during gait development in children.

The principles of direct dynamics and oscillating systems have been used to study the development of gait parameters in children, with respect to their kinetic consequences on the oscillations of body center of mass (CM). In particular the equations established (a) a natural body frequency (NBF), a body parameter specific to oscillating movements which is invariant for adults and decreases with age for children, and (b) the amplitude ratio of CM to center-of-foot pressure (CP) oscillations as a parametric function of the step frequency, whose parameter is the NBF. This function was used to analyze the development of gait locomotors with respect to their kinetic effects on balance in the frontal plane. Five children were examined longitudinally during their first 5 years of independent walking (IW), and two cross-sectional groups between 5 and 7 years of IW were also considered. The results showed a shift toward the low end of step frequency bands as the NBF decreased along with invariances in the amplitudes of CM oscillation in both the frontal and sagittal planes, regardless of age and gait velocity. The biomechanical meaning of the NBF, of its decrease and of postural invariances associated with the decrease of the frequency, are discussed as well as how the programming of locomotor parameters adapts to changes in body structure during gait development.

Biomechanical Phenomena↗
PubMed1995

Organizational and provider structural changes and resource utilization of services for the elderly in Norway.

This paper presents recent innovations in organizing provider work and changes in organizational structure that promote: (1) reduction in fragmentation of care (2) improved continuity and quality of care and (3) improved cost-containment efforts and (4) better decision making. Data is also given on resource utilization. It is anticipated that these models of care and services are prototypes of future models of care for the elderly as this population increases in number. Data is also presented that suggests that, eventually, increases in taxes and social security contributions and/or the rationing of health care may be necessary to meet pensioner health needs.

Aged↗
PubMed1975

The sheep ovary: regulation of steroidogenic, haemodynamic and structural changes in the largest follicle and adjacent tissue before ovulation.

Important changes occur in the follicle that is destined to ovulate and also in the surrounding ovarian tissue of sheep during the 72 hr preceding ovulation. It is our purpose to describe the morphological and functional changes that take place in large follicles during the preovulatory period and to relate these to the local ovarian environment and general endocrine status prevailing at this time. A consideration of changes in the structure, blood supply and function of the ovaries in vivo, is followed by a discussion of the functional capacity of the individual ovarian components when isolated and studied in vitro. Against this background, the nature and sequence of events that occur in the preovulatory follicle and the mechanisms which regulate them will be discussed.

Adenylyl Cyclases↗
PubMed2000

Structural changes of mitochondria during free radical-induced apoptosis.

The initial proposal for apoptosis stressed nuclear change (condensation of chromatin) and the intactness of intracellular organelles, including mitochondria, based on light and electron microscopic observations. However, data have accumulated to demonstrate that the opening of megachannels of mitochondrial membranes, resulting in the swelling of the organelles, notably by Ca2+ and free radicals, is the crucial step in the apoptotic processes of the cell. Application of fluorescent dyes to mitochondria, combined with flow cytometry, has made it possible to detect subtle changes in the structure and function of the organelles related to apoptosis. The present article overviews structural aspects of mitochondria related to apoptosis, including the free radical-induced formation of megamitochondria.

Animals↗
PubMed2001

Medical physics education from the view of the possible structural changes.

Teaching subject physics at the university level represents a specific didactic transformation of the scientific field--physics. The determination of the content, extent, used methods, mutual relation to other subjects of curriculum as well as to the entrance knowledge of students are the most important parts of pedagogical activities in the educational process. Based on own experiences, successes and mistakes in teaching so-called medical physics the authors discuss didactic procedures which should support the interest and creativity of students. Some changes in the structure of physics education are recommended. The usefulness of the international collaboration in the framework of projects such as TEMPUS, ERASMUS is also remembered.

Biophysics↗
PubMed1989

[Structural changes in the jaws. Differential diagnosis].

In this work the Authors have considered the causes of changes in the structure of maxillary bones. They have also discussed the available diagnostic means useful to make a correct diagnosis and the importance of a right differential diagnosis of the causes in order to have a better treatment.

Diagnosis, Differential↗
PubMed1985

Histone phosphorylation in native chromatin induces local structural changes as probed by electric birefringence.

In order to understand how the phosphorylation of histones affects the chromatin structure, we used electron microscopy, sedimentation velocity, circular dichroism and electric birefringence to monitor the salt-induced filament reversible solenoid transition of phosphorylated and native chromatin. Phosphorylation in vitro of chicken erythrocyte chromatin by cyclic-AMP-dependent protein kinase from porcine heart led to the modification of the histones H3 and H5 only, which were modified at a level of one phosphate and about three phosphate groups per molecule, respectively. In contrast to circular dichroism and sedimentation studies, which tend to suggest that phosphorylation of H3 and H5 does not affect chromatin structure, electron microscopy reveals that phosphorylation causes a relaxation of structure at low ionic strength. Electric birefringence and relaxation time measurements clearly prove that local structural changes are induced in chromatin: we observe a decrease of the steady-state birefringence with the appearance of a negative contribution in the signal and a marked increase of the flexibility of fibres. The component with the negative birefringence presents very short relaxation times, like those exhibited by small DNA fragments or individual nucleosomes. Two possibilities are then suggested. First, the conformational change is consistent with what would be expected from the presence of DNA segments loosely associated with the core histone H3. That the length of such segments could correspond to about one to two base-pairs per nucleosome strongly suggests that phosphorylation induces changes affecting some specific H3-DNA interactions only. This result could corroborate previous observations indicating that the N-terminal region of H3, where the site of phosphorylation is located, plays a decisive role in maintaining the superstructure of chromatin. Second, phosphorylation could introduce hinge points between each nucleosome. In this case, the negative birefringence results from partial orientation of the swinging nucleosomes. A possible mode of action of phosphorylation might be to weaken structural restraints imposed by histone H3, thus facilitating further condensation of chromatin.

Animals↗
PubMed1986

Does feeding of hexachlorobenzene promote structural changes in rat-liver porphyrinogen carboxy-lyase?

This study aimed to determine whether drug treatment produced structural changes in porphyrinogen carboxy-lyase enzymatic protein, leading to altered properties. Rat-liver enzyme was obtained from normal animals and from those with hexachlorobenzene (HCB)-induced porphyria, and several of its properties were comparatively studied. The enzymes from both sources were purified 110-fold. They were similar in subcellular distribution, ammonium sulfate fractionation, calcium phosphate gel adsorption, storage stability, requirements of incubation media, effects of salts and photo-oxidizing agents. The enzymes differed with respect to effect of incubation temperature, pH, aerobiosis, chelating agents, dithiothreitol, methylene blue, heat stability and chromatographic behaviour on DEAE-cellulose and Sephadex G-100. These differences would appear to indicate structural differences between the two types of enzymatic protein. Since the porphyrinogenic action of HCB can be mediated through a metabolite, structural differences could arise through the binding of an inhibitory metabolite to the whole enzyme or through modifications to the protein during its synthesis.

Animals↗
PubMed2001

pH-dependent structural changes at the Heme-Copper binuclear center of cytochrome c oxidase.

The resonance Raman spectra of the aa3 cytochrome c oxidase from Rhodobacter sphaeroides reveal pH-dependent structural changes in the binuclear site at room temperature. The binuclear site, which is the catalytic center of the enzyme, possesses two conformations at neutral pH, assessed from their distinctly different Fe-CO stretching modes in the resonance Raman spectra of the CO complex of the fully reduced enzyme. The two conformations (alpha and beta) interconvert reversibly in the pH 6-9 range with a pKa of 7.4, consistent with Fourier transform infrared spectroscopy measurements done at cryogenic temperatures (D.M. Mitchell, J.P. Sapleigh, A.M.Archer, J.O. Alben, and R.B.Gennis, 1996, Biochemistry 35:9446-9450). It is postulated that the different structures result from a change in the position of the Cu(B) atom with respect to the CO due to the presence of one or more ionizable groups in the vicinity of the binuclear center. The conserved tyrosine residue (Tyr-288 in R. sphaeroides, Tyr-244 in the bovine enzyme) that is adjacent to the oxygen-binding pocket or one of the histidines that coordinate Cu(B) are possible candidates. The existence of an equilibrium between the two conformers at physiological pH and room temperature suggests that the conformers may be functionally involved in enzymatic activity.

Animals↗
PubMed2005

Patency and structural changes in cryopreserved arterial grafts used as vessel substitutes in the rat.

OBJECTIVE: To evaluate the patency and structural changes that occur in the short- and mid-term when cryopreserved syngenic arterial grafts are implanted in an experimental animal model. MATERIAL AND METHODS: Segments of iliac artery from the Spraque-Dawley rat were cryopreserved in a biological freezer according a controlled, computerized freezing protocol whereby the specimens are cooled at a rate of 1 degrees C/min. After storage at -145 degrees C in liquid N2 vapor for 30 days, the cryografts were slowly thawed. These vessels were grafted to the common iliac artery in syngenic animals. The following study groups were established: group I (GI), non-implanted cryografts; group II (GII), autografts; and group III (GIII), cryoisografts. The control group (CG) was comprised of fresh iliac arteries. The animals were sacrificed 14, 30, or 90 days post-surgery. At each of these follow-up times, graft specimens were morphologically evaluated by light and scanning and transmission electron microscopy and immunolabeling of endothelial cells (vWf). Cell damage attributed to the cryopreservation or grafting process was also determined. RESULTS: At the time of sacrifice, graft patency was 100% for the autografts, while 26.6% of the cryoisografts showed fully occlusive thrombosis. Among other complications, two pseudoaneurysms were detected. After cryopreservation, the grafts (GI) showed patches of endothelial denudation and good cellularity of the medial layer. The intimal hyperplasia observed in autografts implanted for 14 days (GII) was significantly delayed until day 30 when the graft was cryopreserved (GIII). Cryoisografts showed general thinning of the arterial wall and degeneration accompanied by medial layer cell loss. These grafts showed most cell damage at 90 days post-implant. Expression of the vWf in all specimens showing intimal hyperplasia was confined to the outermost graft layer. CONCLUSIONS: Cryopreservation modified the reparative response of the grafts. Owing to faster degeneration of the medial layer and a delay in the appearance of intimal hyperplasia, arterial wall thickness was reduced relative to that of the non-cryopreserved autografts. This thinning, at least in the short-term (90 days), does not seem to give rise to aneurysms owing to the generation of a neointima that stabilizes the vessel wall.

Animals↗
PubMedDate not supplied

[The structural changes in the lungs in the acute period of a myocardial infarct in the wake of ventricular fibrillation].

Time-related course was studied of structural changes in the lung tissue in acute myocardial infarction having developed in the wake of secondary ventricular fibrillation, using investigational methods common in biology, histochemistry, and electron-microscopy. It has been found out that development in patients with acute myocardial infarction of ventricular fibrillation greatly aggravates the course of the illness inducing origination of profound and even irreversible alternative changes in all constituents of the aerohematic barrier. The above patients developed structural as well as functional pulmonary insufficiency which is at its greatest within 2 to 4 days of the onset of ventricular fibrillation.

Acute Disease↗