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PubMedDate not supplied

Electrospinning of collagen nanofibers.

Electrospinning is a fabrication process that uses an electric field to control the deposition of polymer fibers onto a target substrate. This electrostatic processing strategy can be used to fabricate fibrous polymer mats composed of fiber diameters ranging from several microns down to 100 nm or less. In this study, we describe how electrospinning can be adapted to produce tissue-engineering scaffolds composed of collagen nanofibers. Optimizing conditions for calfskin type I collagen produced a matrix composed of 100 nm fibers that exhibited the 67 nm banding pattern that is characteristic of native collagen. The structural properties of electrospun collagen varied with the tissue of origin (type I from skin vs type I from placenta), the isotype (type I vs type III), and the concentration of the collagen solution used to spin the fibers. Electrospinning is a rapid and efficient process that can be used to selectively deposit polymers in a random fashion or along a predetermined and defined axis. Toward that end, our experiments demonstrate that it is possible to tailor subtle mechanical properties into a matrix by controlling fiber orientation. The inherent properties of the electrospinning process make it possible to fabricate complex, and seamless, three-dimensional shapes. Electrospun collagen promotes cell growth and the penetration of cells into the engineered matrix. The structural, material, and biological properties of electrospun collagen suggest that this material may represent a nearly ideal tissue engineering scaffold.

Animals↗
PubMed1999

Only multimeric hensin located in the extracellular matrix can induce apical endocytosis and reverse the polarity of intercalated cells.

When an intercalated epithelial cell line was seeded at low density and allowed to reach confluence, it located the anion exchanger band 3 in the apical membrane and an H+-ATPase in the basolateral membrane. The same clonal cells seeded at high density targeted these proteins to the reverse location. Furthermore, high density cells had vigorous apical endocytosis, and low density cells had none. The extracellular matrix of high density cells was capable of inducing apical endocytosis and relocation of band 3 to the basolateral membrane in low density cells. A 230-kDa extracellular matrix (ECM) protein termed hensin, when purified to near-homogeneity, was able to reverse the phenotype of the low density cells. Antibodies to hensin prevented this effect, indicating that hensin is necessary for conversion of polarity. We show here that hensin was synthesized by both low density and high density cells. Whereas both phenotypes secreted soluble hensin into their media, only high density cells localized it in their ECM. Analysis of soluble hensin by sucrose density gradients showed that low density cells secreted monomeric hensin, and high density cells secreted higher order multimers. When 35S-labeled monomeric hensin was added to high density cells, they induced its aggregation suggesting that the multimerization was catalyzed by surface events in the high density cells. Soluble monomeric or multimeric hensin did not induce apical endocytosis in low density cells, whereas the more polymerized hensin isolated from insoluble ECM readily induced it. These multimers could be disaggregated by sulfhydryl reagents and by dimethylmaleic anhydride, and treatment of high density ECM by these reagents prevented the induction of endocytosis. These results demonstrate that hensin, like several ECM proteins, needs to be precipitated in the ECM to be functional.

Anion Exchange Protein 1, Erythrocyte↗
PubMed1988

Immunohistochemical studies on the tissue localization of collagen types I, III, IV, V and VI in schwannomas. Correlation with ultrastructural features of the extracellular matrix.

The distinctive tissue localization of collagen types in typical schwannomas with Antoni type A and B areas was demonstrated immunohistochemically using affinity-purified antibodies against types I, III, IV, V and VI collagen and comparative ultrastructural studies were made on the extracellular matrix components. Antoni type A tissue, which was composed of tightly packed spindle cells with long cytoplasmic processes surrounded by a continuous basement membrane and a few fibrillar components of the extracellular matrix, was almost exclusively immunoreactive for type IV collagen, presumably representing the basement membrane. Verocay bodies, which are organoid structures of Antoni type A tissue, had a variety of more abundant extracellular fibrous components, such as banded collagen fibrils, fibrous long-spacing fibrils and microfibrils. These were positive for type I and III, as well as type IV collagen. In Antoni type B areas, where two types to tumor cells designated Schwann cell-like and fibroblast-like were scattered in large amounts of amorphous extracellular matrix containing microfibrils and thick banded collagen fibrils, type VI collagen as well as types I, III and IV collagen were consistently detected. Type V collagen was localized in dense fibrous tissue areas and around blood vessels. These findings indicate that the differently organized cellular patterns of schwannomas, identified as Antoni types A and B, are characterized not only by the ultrastructural features of the extracellular matrix, but also by the distinctive collagen types produced by neoplastic Schwann cells.

Adult↗
PubMed1997

Pharmacological modulation of pressure-overload cardiac hypertrophy: changes in ventricular function, extracellular matrix, and gene expression.

BACKGROUND: Appropriate cardiac hypertrophy (CH) is necessary in several clinical settings, such as pulmonary artery banding in the two-stage arterial switch operation for transposition of the great arteries. Pressure-overload CH, however, produces ventricular dysfunction due to structural and molecular changes. The beta2-adrenergic receptor agonist clenbuterol has been shown to induce CH without such adverse effects to the rat heart. This study was performed to determine its effects on left ventricular (LV) function, structure, and gene expression in pressure-overload CH. METHODS AND RESULTS: Sprague-Dawley rats were assigned to one of four groups: 1, sham-operated (n=15); 2, banding of ascending aorta (n=22); 3, banding+clenbuterol (n=18); and 4, banding+thyroxine (n= 17). At the end of 3 weeks, groups 2, 3, and 4 showed an increase in LV mass index of 49.7+/-5.1%, 66.1+/-3.8%, and 47.6+/-4.6%, respectively, relative to group 1. A subgroup with severe CH (>50%) in group 2 was found to have significantly impaired developed pressure and diastolic relaxation and an increase in passive stiffness, with significantly reduced LV expression of sarcoplasmic reticulum Ca2+-ATPase2a (SERCA2a) mRNA and increased LV collagen concentration. In comparison, similarly hypertrophied animals in groups 3 and 4 demonstrated improved developed pressure, normal relaxation and diastolic stiffness with normal collagen concentration, and a greater abundance of SERCA2a mRNA. CONCLUSIONS: Clenbuterol administration in conjunction with pressure overload produces a specific type of CH with preserved LV function. In addition, an increase in LV mass was associated with less fibrosis and greater expression of SERCA2a mRNA than banding alone.

Analysis of Variance↗
PubMed2006

Photogeneration of heptacene in a polymer matrix.

Heptacene (1) was generated by the photodecarbonylation of 7,16-dihydro-7,16-ethanoheptacene-19,20-dione (2) in a polymer matrix using a UV-LED lamp (395 +/- 25 nm). Compound 1 showed a long wavelength absorption band extending from 600 to 825 nm (lambdamax approximately 760 nm) and was found to be stable up to 4 h in the polymer matrix. However, irradiation of a solution of 2 in toluene produced only oxygen adducts.

Journal Article↗
PubMed1981

Lateral mobility of erythrocyte membrane proteins studied by the fluorescence photobleaching recovery technique.

Erythrocyte membrane peripheral and integral proteins have been isolated and purified, and the lateral diffusion of these proteins in a well-defined phospholipid bilayer matrix (dimyristoylphosphatidylcholine) has been studied by fluorescence photobleaching recovery measurements. Our own instrument for the recovery measurements is described and some data for lipid diffusions are compared with those previously reported by other investigators. The peripheral proteins (spectrin and band 4.1) diffuse rapidly on the lipid membrane in its fluid phase. The diffusion constant of approximately 5 x 10(-8) cm2 . s-1 (30 degrees C) was only a little smaller than that for lipid diffusion. The diffusion was greatly slowed down when the host lipid matrix became solid. The integral protein band 3 also diffuses rapidly in the fluid membrane. The diffusion constant of 1.6 x 10(-8) cm2 . s-1 (30 degrees C) was smaller than those for lipids and for the peripheral proteins. The lateral motion is compatible with diffusion of a cylinder with radius 3 nm in a two dimensional matrix with an inner viscosity of 2 poises and an inner thickness of 4 nm. The band 3 lateral motion was restricted by binding of the cytoskeletal component proteins (ankyrin, spectrin, actin, and band 4.1) to the reconstituted membranes. The diffusion constant decreased to half. The results provide a basis for the elucidation of transmembrane control mechanisms in more complex cellular systems.

Anion Exchange Protein 1, Erythrocyte↗
PubMed2002

In silico chromosome staining: reconstruction of Giemsa bands from the whole human genome sequence.

Giemsa staining has been used for identifying individual human chromosomes. Giemsa-dark and -light bands generally are thought to correspond to GC-poor and GC-rich regions; however, several experiments showed that the correspondence is quite poor. To elucidate the precise relationship between GC content and Giemsa banding patterns, we developed an "in silico chromosome staining" method for reconstructing Giemsa bands computationally from the whole human genome sequence. Here we show that 850-level Giemsa bands are best correlated with the difference in GC content between a local window of 2.5 megabases and a regional window of 9.3 megabases along a chromosome. The correlations are of strong statistical significance for almost all 43 chromosomal arms. Our results clearly show that Giemsa-dark bands are locally GC-poor regions compared with the flanking regions. These findings are consistent with the model that matrix-associated regions, which are known to be AT-rich, are present more densely in Giemsa-dark bands than in -light bands.

Azure Stains↗
PubMed1998

The muscleblind gene participates in the organization of Z-bands and epidermal attachments of Drosophila muscles and is regulated by Dmef2.

We report the embryonic phenotype of muscleblind (mbl), a recently described Drosophila gene involved in terminal differentiation of adult ommatidia. mbl is a nuclear protein expressed late in the embryo in pharyngeal, visceral, and somatic muscles, the ventral nerve cord, and the larval photoreceptor system. All three mbl alleles studied exhibit a lethal phenotype and die as stage 17 embryos or first instar larvae. These larvae are partially paralyzed, show a characteristically contracted abdomen, and lack striation of muscles. Our analysis of the somatic musculature shows that the pattern of muscles is established correctly, and they form morphologically normal synapses. Ultrastructural analysis, however, reveals two defects in the terminal differentiation of the muscles: inability to differentiate Z-bands in the sarcomeric apparatus and reduction of extracellular tendon matrix at attachment sites to the epidermis. Failure to differentiate both structures could explain the partial paralysis and contracted abdomen phenotype. Analysis of mbl expression in embryos that are either mutant for Dmef2 or ectopically express Dmef2 places mbl downstream of Dmef2 function in the myogenic differentiation program. mbl, therefore, may act as a critical element in the execution of two Dmef2-dependent processes in the terminal differentiation of muscles.

Animals↗
PubMed1983

A discontinuous electrophoretic system for separating peptides on polyacrylamide gels.

An electrophoretic system for separating, with high resolution, peptides 25-250 residues in length is described. The peptides are stacked by discontinuous electrophoresis to form very sharp bands at the origin. They are then separated on a matrix of 20% polyacrylamide, 8 M urea, and 0.1% dodecyl sulfate. Through this combination, high resolution and clean separation, based on polymer length, are achieved.

Animals↗
PubMed1991

Rapid estimation of chromosomal damage in yeast due to the effects of environmental chemicals using pulsed field gel electrophoresis.

We present a procedure to rapidly estimate the damage to yeast chromosomes by toxic chemicals. This procedure employs the following steps: incubation of yeast cells with the chemicals, DNA preparation in an agarose matrix, separation of chromosome-sized DNA molecules into reproducible band patterns by pulsed field gel electrophoresis, and quantification of the intensity of chromosomal bands by densitometry. Saccharomyces cerevisiae cells have been treated with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and cis-Platinum(II) diamminedichloride (cisPT), both of which are known to interact with DNA, and trichlorethylen (TCE), for which such an effect has not been shown in yeast. Treatment of cells with MNNG and cisPt led to an impairment of the intensity of the band pattern to an extent dependent on the concentration of the chemicals applied. For TCE a similar effect could not be discerned. This procedure will be useful as a screening test for the estimation of the biological hazards of toxic chemicals.

Chromosomes, Fungal↗
PubMed1988

Lattices of high histochemical activity occur in the human, monkey, and cat superior colliculus.

A lattice of high oxidative metabolic activity occurs in the intermediate gray layer of the human, monkey, and cat superior colliculus. It is composed of a matrix of high enzyme activity that surrounds pale islands or bands of lower activity. In the human the pale bands are 300-400 micron wide while in the smaller colliculi of the monkey and cat they are 100-200 micron wide. The lattice was demonstrated by studying either cytochrome oxidase or succinate dehydrogenase. In the cat and monkey the lattice occurs at the same depth as the lattice of intense acetylcholinesterase activity, but the two lattices are not in spatial register. In the human the lattice of high oxidative metabolic activity is in the middle of the intermediate gray layer, whereas the lattice of intensely stained cholinesterase activity is at the base of this layer, but again the two lattices are not in spatial register. However, in the middle of the intermediate gray layer of the human, there are elongated islands and bands of very low acetylcholinesterase activity that coincide with the pale islands and bands of low cytochrome oxidase activity. An additional lattice of high enzyme activity occurs based on the enzyme nicotinamide dinucleotide phosphate (reduced form)-diaphorase. This lattice is prominent in the cat, occurs more faintly in the monkey, but did not appear to be present in the human. In the intermediate gray layer it had a high degree of overlap with the acetylcholinesterase lattice. The lattice of high oxidative metabolism contains loosely knit clusters of large multipolar cells containing high cytochrome oxidase activity and these cells do not occur in the pale islands. By contrast the cell bodies in the intermediate gray layer that contain either acetylcholinesterase or the diaphorase occur both between and within the patches of corresponding, high enzyme activity. It is suggested that the acetylcholinesterase and diaphorase lattices are mainly associated with afferent fibers while the lattice of high oxidative metabolism is mainly associated with intrinsic cells. The lattices occur in all mammals studied to date and appear to represent a fundamental principle in the organization of the mammalian colliculus. It is concluded that the lattices will provide a useful basis for further studies of the relationship between the many afferent and efferent modules thought to exist in this structure.

Acetylcholinesterase↗
PubMed1977

Photo-induced charge transfer. A critical test of the mechanism and range of biological electron transfer processes.

The vibronic coupling theory of electron tunneling between biomolecules requires that all such tunnelings involve vibronic coupling, finds temperature dependence to tunneling at finite temperatures, and predicts relatively short tunneling distances. This theory might be expected to apply to most electron transfers involved in the membrane-bound electron transfer reactions of photosynthesis and oxidative phosphorylation. This paper calculates the properties of a weak charge-transfer optical absorption band, whose predicted characteristics are a direct and simple consequence of the model that describes vibronically coupled tunneling. The new absorption band provides the basis for a critical experimental test of the constructs and parameters of the tunneling theory. If the tunneling theory is valid, the oscillator strength of such bands will be the most reliable measure of the tunneling matrix element and of the distance between the sites exchanging an electron.

Electron Transport↗
PubMed1996

Structural fluctuations of myoglobin from normal-modes, Mössbauer, Raman, and absorption spectroscopy.

A normal-mode analysis of carbon monoxymyoglobin (MbCO) and deoxymyoglobin (Mb) with 170 water molecules is performed for (54)Fe and (57)Fe. A projection is defined that extracts iron out-of-plane vibrational modes and is used to calculate spectra that can be compared with those from resonance Raman scattering. The calculated spectra and the isotopic shift (57)Fe versus (54)Fe agree with the experimental data. At low temperatures the average mean square fluctuations (MSFs) of the protein backbone atoms agree with molecular dynamics simulation. Below 180 K the MSFs of the heme iron agree with the data from Mossbauer spectroscopy. The MSFs of the iron atom relative to the heme are an order of magnitude smaller than the total MSFs of the iron atom. They agree with the data from optical absorption spectroscopy. Thus the MSFs of the iron atom as measured by Mossbauer spectroscopy can be used to probe the overall motion of the heme within the protein matrix, whereas the Gaussian thermal line broadening of the Soret band and the resonance Raman bands can be used to detect local intramolecular iron-porphyrin motions.

Heme↗
PubMed2003

Variable-frequency EPR study of Mn(2+)-doped NH(4)Cl(0.9)I(0.1) single crystal at 9.6, 36, and 249.9 GHz: structural phase transition.

Multifrequency electron paramagnetic resonance studies on the Mn(2+) impurity ion in a mixed single crystal NH(4)Cl(0.9)I(0.1) were carried out at 9.62 (X-band) in the range 120-295 K, at 35.87 (Q-band) at 77 and 295 K, and at 249.9 GHz (far-infrared band) at 253 K. The high-field EPR spectra at 249.9 GHz are well into the high-field limit leading to a considerable simplification of the spectra and their interpretation. Three magnetically inequivalent, but physically equivalent, Mn(2+) ions with their respective magnetic Z-axes oriented along the crystallographic [100], [010], [001] axes were observed. Simultaneous fitting of EPR line positions observed at X-, Q-, and far infra-red bands was performed using a least-squares procedure and matrix diagonalization to estimate accurately the Mn(2+) spin-Hamiltonian parameters. The temperature variation of the linewidth and peak-to-peak intensities of the EPR lines indicate the presence of lambda-transitions in the mixed NH(4)Cl(0.9)I(0.1) crystal at 242 and 228 K consistent with those observed in the pure NH(4)Cl and NH(4)I crystals, respectively. A superposition-model analysis of the spin-Hamiltonian parameters reveals that the local environment of the Mn(2+) ion is considerably reorganized to produce axially symmetric crystal fields about the respective Z-axes of the three magnetically inequivalent ions as a consequence of the vacancy created due to charge-compensation when the divalent Mn(2+) ion substitutes for a monovalent NH(4)(+) ion in the NH(4)Cl(0.9)I(0.1) crystal. This reorganization is almost the same as that observed in NH(4)Cl and NH(4)I single crystals, although the latter two are characterized by different, simple cubic and face-centered cubic, structures.

Journal Article↗
PubMed2002

Anomeric and mesomeric effects in methoxycarbonylsulfenyl chloride, CH(3)OC(O)SCl: an experimental and theoretical study.

The molecular structure and conformational properties of methoxycarbonylsulfenyl chloride, CH(3)OC(O)SCl, were determinated in the gas and solid phases by gas electron diffraction, low-temperature X-ray diffraction, and vibrational spectroscopy. Furthermore, quantum chemical calculations were performed. Experimental and theoretical methods result in structures with a planar C-O-C(O)-S-Cl skeleton. The electron diffraction intensities are reproduced best with a mixture of 72(8)% syn and 28(8)% anti conformers (S-Cl bond synperiplanar/antiperiplanar with respect to C=O bond) and the O-CH(3) bond synperiplanar with respect to the C=O bond. The syn form is the preferred form and becomes the exclusive form in the crystalline solid at low temperature. This experimental result is reproduced very well by Hartree-Fock approximation and by density functional theory at different levels of theory but not by the MP2/6-311G method, which overestimates the value of DeltaG between the syn and anti conformers. The results are discussed in terms of anomeric effects and a natural bond orbital (NBO) calculation. Photolysis of matrix-isolated CH(3)OC(O)SCl with broad-band UV-visible irradiation produces an interconversion of the conformers, and the concomitant decomposition leads to formation of OCS and CO molecules.

Journal Article↗
PubMed2006

Optical characterization of eu-doped and undoped gd(2)o(3) nanoparticles synthesized by the hydrogen flame pyrolysis method.

Rare-earth-doped nanoparticles are promising materials for fluorescent labeling, as they are characterized by a high Stokes shift, narrow emission spectra, long lifetimes, minimized photobleaching, and low toxicity. We examined the structural and optical properties of europium-doped gadolinium oxide nanoparticles synthesized by the flame pyrolysis method, with specific emphasis on full spectral characterization and fluorescence kinetics. The emission-excitation characterization revealed the presence of predominantly monoclinic but also highly luminescent cubic phases with a prominent oxygen-to-europium charge-transfer band in the 230-260 nm range. A broad emission band in the visible region, corresponding to a similar band in undoped Gd(2)O(3), related to the matrix surface defects, was observed in time-gated spectroscopy of doped nanopowders. All of the examined nanopowders showed very short decay components, on the order of 2 ns, and much longer millisecond decay times characteristic of lanthanide ions. At intermediate times, on the order of 20-100 ns, a complex behavior of the decay was observed, indicative of progressive energy transfer to the lanthanide ion, which varied with different intrashell transitions. Structural characterization data by means of XRD measurements allowed for unambiguous determination of the Eu:Gd(2)O(3) crystallographic structure and cell dimensions to be consistent with a predominantly monoclinic phase.

Journal Article↗
PubMed2005

Intermolecular vibrations of the water trimer, a matrix isolation study.

Infrared spectra from 25 to 4000 cm(-1) have been recorded of water (H2O, D2O and H218O) matrix isolated in neon, argon, and krypton matrices. Intermolecular absorption bands of different isotopologues of the water trimer and tetramer have been assigned from concentration dependencies and diffusion behavior, using the well-known mid-infrared trimer and tetramer absorption bands as measures of the trimer and tetramer concentrations. The results are compared to ab initio calculations.

Journal Article↗
PubMed2005

Vibrational and electronic absorption spectroscopy of dibenzo[b,def]chrysene and its ions.

The vibrational and electronic absorption spectra of dibenzo[b,def]chrysene (DBC) and its ions in argon matrixes have been recorded. Assignment of the observed infrared (IR) bands has been made by comparison with the density functional theory (DFT) computations of harmonic vibrational frequencies (with 6-31G(d,p) or 6-311+G(d,p) basis sets). Extensive time-dependent (TD) DFT calculations of vertical excitation energies have aided in the assignment of the experimental electronic absorption transitions. In general, the theoretical predictions are in good agreement with the observed ultraviolet and visible bands. By correlating IR and UV-visible band intensities (after UV photolysis), it has been shown that both DBC cations and anions are formed. The IR band intensity distributions of the DBC ions differ markedly from neutral DBC. A synthetic spectrum composed of neutral, cationic, and anionic DBC contributions compares reasonably well with the interstellar features of the "unidentified infrared" (UIR) bands from the reflection nebula NGC 7023. Finally, it is shown that the electronic absorption bands of the DBC ions lie in close proximity to several of the diffuse interstellar visible absorption bands (DIBs).

Chrysenes↗