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[Structural changes in muscle fiber contractile proteins studied by polarization ultraviolet fluorescence microscopy. VIII. The effect of glutaraldehyde and phalloidine on F-actin conformation].

Conformational changes in F-actin, induced by glutaraldehyde or phalloidin, were found in glycerinated m. psoas rabbit fibres (ghost and reconstructed fibres). It is shown that the conformational changes of F-actin decreased the thin filament flexibility as well as weakened fibre's contractility. It is assumed that the stabilization in F-actin structure may be an important factor involved in the mechanism of muscle contraction regulation.

Actins↗

[Structural changes in muscle fiber contractile proteins studied by polarization ultraviolet fluorescence microscopy. VI. Conformational restructurings of F-actin induced by the binding of heavy meromyosin].

Decrease in tryptophan fluorescence anisotropy of a single ghost muscle fibre was found at the binding of heavy meromyosin (HMM) to actin. Polarized fluorescence revealed its peak changes at a molar ratio of HMM/actin equal to 0.1. The changes observed in polarized fluorescence at F-actin-HMM interaction were found to depend on the state of HMM. The changes in anisotropy fluorescence under the same conditions were assumed to be independent of tryptophane residues in HMM, reflecting cooperative changes in F-actin conformation. Changes in the conformation of F-actin are of great importance in muscular contraction.

Actins↗

Improved staining of extracellular polymer for electron microscopy: examination of Azotobacter, Zoogloea, Leuconostoc, and Bacillus.

Phase contrast, ultraviolet microscopy, and freeze-etching were used to determine the amount of exocellular polymer surrounding unfixed cells of four genera of bacteria: Azotobacter vinelandii, Zoogloea ramigera, Leuconostoc mesenteroides, and an acid-tolerant, floc-forming Bacillus species. Thin-sectional electron microscopy was employed to measure the effectiveness of a modified ruthenium red staining method. The results obtained with this modification of ruthenium red staining technique were compared to results obtained when previously proposed ruthenium red methods of fixation were employed. The results of these relations were then compared to the amounts of exocellular material as determined with phase-contrast microscopy, ultraviolet microscopy, and freeze-etching. The data obtained indicate that improved fixation of exocellular polymer is achieved when cells are pretreated with ruthenium red as described herein. In addition, the modified methods also reveal cytological detail not apparent when other methods of ruthenium fixation are employed.

Azotobacter↗

Studies on conformational changes in F-actin of glycerinated muscle fibers during relaxation by means of polarized ultraviolet fluorescence microscopy.

By means of polarized ultraviolet fluorescence microscopy the conformational changes of F-actin occuring in glycerinated muscle fibers of rabbit and barnacle (Balanus rostratus Hock.) under the influence of adenosine triphosphate in the presence of ethylene glycol bis(beta-amino-ethyl ether)-N,N'-tetraacetic acid were discovered. These changes seem to be located near the surface of the globules thus hampering the penetration of univalent iones and neutral molecules into the F-actin macromolecule. It is suggested that similar changes of F-actin take place in thin myofilaments of living muscle fiber during the contraction-relaxation process.

Actins↗

Studies on structural changes of F-actin and myosin in living, intact and damaged muscle fibres by means of polarized ultraviolet fluorescence microscopy.

By means of polarized ultraviolet fluorescence microscopy the structural changes of F-actin and myosin were discovered at the changing a functional state of a living muscle fibre and during spreading degeneration (Zenker's necrosis). The character of conformational changes of F-actin and myosin at activation, contraction, contracture and rigor is similar, but the number of changed macromolecules depends on a fibre state. At fibre local damage in its morphological intact parts there was found an alternation of zones, reflecting two states unusual for a fibre. During spreading degeneration these states transform into irreversible contracture and then into rigor. Similar changes were observed in muscle fibres obtained from denervated muscles.

Actins↗

[Study of structural changes in muscle fiber contractile proteins using polarization ultraviolet fluorescent microscopy. IV. Several features of the conformational changes in F-actin during muscle fiber relaxation].

Using the polarized ultraviolet fluorescent microscopy in the presence of a fluorescence quencher (acrylamide, NO3-) it has been shown that actin tryptophanyls are oriented with their short axes perpendicular to the long axis of the thin filament. Fluorophores in hardly accessible sites of the macromolecule have a more pronounced anisotrophy of orientation in comparison with those in the protein sites easily available by a fluorescence quencher. During the muscle fiber relaxation, conformational changes of F-actin take place which embrace seemingly the sites of subunits, close to the surface of macromolecule, and make difficult the penetration of univalent ions and neutral molecules deep into the protein macromolecule. Some connection between conformational changes of the surface areas of F-actin subunits and the actin incapability of combining with HMM in the presence of ATP is assumed.

Acrylamides↗

[Structural changes in the contractile proteins of muscle fiber studied by polarization ultraviolet fluorescence microscopy. IX. The effect of the pH and ionic strength of the solution on the conformational restructurings of F-actin induced by the binding of heavy meromyosin].

The dependence of F-actin conformational changes induced by the F-actin-HMM complex on pH and ionic strength was found by polarized ultraviolet fluorescence microscopy. It is discovered that pH affects sufficiently the cooperativity of F-actin structural changes, while the ionic strength affects their depth. The actomyosin complex was supposed to be at least in two structural states, differing in their orientation as well as in flexibility of F-actin monomers.

Actins↗

Differential termination times of enamel formation on human deciduous anterior-tooth surfaces revealed by tetracycline labelling.

The positional relationships between the cervical-enamel margin and the intersecting point of a fluorescent (tetracycline) line in dentine, with the outer dentine surface or the dentine-enamel junction, were investigated by ultraviolet microscopy of the human deciduous anterior teeth. The results suggest that enamel formation (1) ended earlier on the mesial than on the distal surface in 18 instances and, nearly simultaneously on both of these surfaces in four, (2) ended earlier on the distal than on the labial surface in all 28 specimens and (3) ended earlier on the labial than on the lingual surface in 14 instances and nearly simultaneously on both of these surfaces in eight instances.

Amelogenesis↗

Protoporphyrin-induced cholestasis in the isolated in situ perfused rat liver.

The pathogenesis of liver disease in protoporphyria has been presumed to result from the hepatic deposition of protoporphyrin. To examine the effects of protoporphyrin on hepatic bile flow and histopathology, studies were performed employing an isolated, in situ, rat liver perfusion system. Rat livers in the control group were perfused with 0-80 mumol sodium taurocholate/h. Rat livers in the experimental group were perfused with sodium taurocholate and (a) sufficient quantities of protoporphyrin to produce maximal canalicular secretion and (b) perfusate protoporphyrin concentrations of 0.01, 0.1, and 1 muM. The administration of protoporphyrin sufficient to achieve maximal canalicular secretion was found to significantly reduce bile flow in rats infused with 0, 40, and 80 mumol sodium taurocholate/h. Linear regression analysis defined the relationship between bile flow and biliary bile acid secretion and showed that the bile acid-independent fraction of bile flow was reduced (P < 0.01). Bile acid-dependent flow was unaffected and there was no significant difference in biliary bile acid secretion rates between control and protoporphyrin-perfused livers. Perfusion of rat livers with varying concentrations of protoporphyrin demonstrated the reduction of bile flow was dose-related. Analysis of perfusate enzyme activity did not reveal abnormalities that could account for the cholestasis. Studies to evaluate the effect of protoporphyrin on regional hepatic hemodynamics were inconclusive. Histopathological studies of control and protoporphyrin-perfused rat livers did not show abnormalities on light microscopy. However, canalicular dilatation, distortion, and loss of microvilli were present in the protoporphyrin-perfused livers examined by transmission electron microscopy. Although ultraviolet microscopy showed diffuse fluorescence of the hepatocytes and canaliculi of protoporphyrin-perfused livers, the deposition of protoporphyrin in amorphous or crystalline forms was notably absent in studies with polarizing and transmission electron microscopy. These studies provide evidence that protoporphyrin has hepatotoxic properties that affect the canalicular secretory apparatus. The mechanism(s) responsible for the injury require further clarification.

Animals↗

The microscopic pattern of tetracycline fluorescence in the cementum of human teeth.

Ultraviolet microscopy of longitudinal non-decalcified sections revealed fluorescence characteristic of in vivo tetracycline deposition in the cementum of 55 out of 378 human permanent teeth. Patterns of fluorescence could be related to various stages in root development and to pathological events, as well as to the approximate duration of drug intake. Cementum lines formed during apposition of primary root dentin were associated with dentin bands and met the latter at the cemento-dentinal junction: other lines corresponded to incremental cementum apposition after completion of the root. Label was also visualized in localized areas of cementum hyperplasia and in repaired resorption lacunae. Discontinuities in broad fluorescent lines at or near the cementum surface were related to lack of connective tissue attachment when drug exposure occurred, or with subsequent root caries. Observations suggest that typical short, single periods of antibiotic therapy are recorded in the cementum, although marginally or close to the limit of resolution in some cases. Longer, chronic administration or multiple short regimens closely spaced in time are clearly registered. Study of these patterns may be useful in clinical periodontal research.

Dental Cementum↗

[Structural changes in contractile proteins of muscle fibers studied by polarization ultraviolet fluorescence microscopy. X. The effect of ATP, Ca2+, pH change and ionic strength of the washing solution on the structural state of thick filaments].

By the method of polarized ultraviolet fluorescence microscopy, effects of ATP, Ca2+, changes in pH and ionic strength of washing solution of the structural state of thick filaments in both actin-free muscle fibers of rabbit and anisotropic discs (A-discs) of glycerinated fibers of crab were studied. The dependence of tryptophan fluorescence anisotropy of thin filaments upon physico-chemical parameters (compounds) of washing solution has been found. The structural state of thick filaments was suggested to be influenced by ATP, Ca2+, changes in pH and ionic strength of washing solution.

Adenosine Triphosphate↗

[Study of structural changes of contractile muscle proteins with the aid of polarization ultraviolet fluorescence microscopy. 1. Conformational changes of F-actin in the muscle fiber caused by ATP and its analogs].

Increase of anisotropy of F-actin fluorescence of balanus and rabbit muscle fibers under the influence of ATP, AMP and pyrophosphate in EGTA presence was detected by means of the polarized ultraviolet (UV) fluorescent microscopy methods. The fluorescence anisotropy changes are assumed to be associated with the conformational changes in the actin. ATP cause more noticeable changes of actin structure, than pyrophosphate and AMP. The conformational changes in the actin of balanus and rabbit muscle fibres were similar. ATP and its analogs induced also decrease of UV fluorescence anisotropy of A-band which appears to be associated with conformational changes in myosin. It was siggested that the changes in fluorescence of anisotropy of A-bands are due to structural changes in both HMM and LMM parts of myosin molecule.

Actins↗

Repair process of external root resorption subsequent to palatal expansion treatment.

The repair process of external root resorption (ERR) and the role of retention mechanics in enhancing ERR repair were studied on eight Macaca fascicularis monkeys that were divided equally into short- and long-term groups. Six monkeys received palatal expansion appliances, and two received sham appliances. The short-term group received active treatment. The long-term group received additional retention (4 months) and relapse (2 months) treatment periods with biweekly injections of individual vital dye per phase, i.e., procion red H-8B and violet H-3R (80 mg/kg B.W.), respectively. Histomorphologic examinations included scanning electron microscopy, light, and ultraviolet microscopy. The short-term group demonstrated penetrated resorption with pulp exposure at sites with initial deficiency of the protecting odontoblastic layer (apical zone, nutrition canal). The long-term group showed two forms of ERR repair: (1) Nonfunctional retarded repair cementum, comprised of overlapped incremental lines and deprived of extrinsic fibers, was delineated in severe pulp exposure. The pulp/dentin complex showed intense incorporation of procion dye in the dentinal tubuli, conceivably related to a defense response in the form of sclerotic dentin. (2) Functional rapid repair cementum, comprised of discriminated incremental lines mainly of mixed cellular cementum, with a consistent pattern of five sequential phases: the lag phase (14 to 28 days), the incipient phase (14 days), the peak phase (14 to 28 days), the steady phase (42 to 56 days) and the retreating phase (70 days). Sharpey's fibers at functional ERR sites were scarce, never emerging from the dentinocemental junction, and not developing into principal fibers. The pulp/dentin complex showed an increase in pulp stones but no formation of tertiary dentin. The apical area responded by hypercementosis in the form of apical occlusion and a displaced pulp canal. The application of a fixed retention device is recommended in light of accelerated apposition of repair cementum during the retention period. However, increased formation of Sharpey's fibers during the relapse period might suggest a restricted duration in splinting therapy.

Animals↗