The equatorial streak in the small-angle X-ray scattering (SAXS) pattern from nylon-6 fibers, normally assigned to microvoids or to internal fibrillar structure, is due to scattering from the fiber ...surfaces. This scattering is best thought of as refraction by the cylindrical fibers. The streak intensity increases by 50 times during the early stages of drying a wet bundle of nylon-6 fibers, when superficial water, water between the fibers in the bundle, is lost. Separate X-ray experiments using a nonpenetrating fluid, a large diameter monofilament, and microbeam SAXS confirmed this assignment. There are many fibers where internal voids are known to be the source of a strong equatorial streak in SAXS, but in general, a check for surface scattering is advised before interpreting this feature. Simultaneous small- and wide-angle X-ray diffraction (WAXD) was obtained during drying. Following the initial loss of superficial water, there are changes in both the WAXD reflections and the SAXS lamellar reflection. In the later stages of drying, WAXD shows that the crystals decrease in perfection, in apparent lateral size and in degree of misorientation. Lamellar structure changes are slower to start and include a 1.3% decrease in axial lamellar spacing and a 40% increase in the contrast between the crystalline lamellae and the interlamellar regions. All these changes are explained as effects of the removal of absorbed water from the interlamellar amorphous regions and their reduction in volume and density. Interpretation of these experimental results thus does not require a fibrillar structure in nylon-6.
The fiber morphology of the dragline silk of Nephila clavipes has been investigated by the detailed analysis of wide-angle X-ray diffraction (WAXD) patterns. WAXD gives the crystal lattice ...dimensions, the orientation distribution, the crystalline fraction, and an estimate of the crystal size. It is found that the crystals are very small and well oriented. The mean (minimum) crystal dimensions are 2 × 5 × 7 nm, and the angle, φ, between the molecular chains in the crystals and the fiber axis has a full width at half-maximum (fwhm) of 15.7° and an orientation function f = 0.981. The X-ray crystallinity is in the range 10−15%, and the amorphous diffraction is divided 60:40 between an isotropic ring and an oriented halo with fwhm 30°. This means one-third of the material is in the oriented amorphous state, with a chain orientation of fwhm 43° and f = 0.87. When the fiber is extended up to 10%, the orientation of the crystals increases as predicted for affine deformation at constant volume. There is no observable change in crystallinity and apparently a small reduction in the lateral crystal size on deformation.
Successful gene therapy will require that the therapeutic gene be expressed at a sufficient level in the correct cell type(s). To improve the specificity of gene transfer for cystic fibrosis (CF) and ...other airway diseases, we have begun to develop cell-type specific promoters to target the expression of transgenes to specific airway cell types. Using a FOXJ1 promoter construct previously shown to direct transgene expression specifically to ciliated cells, we have generated transgenic mice expressing human cystic fibrosis transmembrane conductance regulator (CFTR) in the murine tracheal and nasal epithelia. RNA analysis demonstrated levels of CFTR expression is greater than or equal to the level of endogenous mouse CFTR. Immunoprecipitation and western blotting demonstrated the production of human CFTR protein, and immunochemistry confirmed that CFTR was expressed in the apical region of ciliated cells. The transgenic animals were bred to CFTR null mice (Cftr(tm1Unc)) to determine if expression of CFTR from the FOXJ1 promoter is capable of correcting the airway defects in Cl(-) secretion and Na(+) absorption that accompany CF. Isolated trachea from neonatal CF mice expressing the FOXJ1/CFTR transgene demonstrated a correction of forskolin-stimulated Cl(-) secretion. However, expression of human CFTR in ciliated cells of the nasal epithelia failed to significantly change the nasal bioelectrics of the CF mice.
Cystic Fibrosis/Pulmonary Research and Treatment Center, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina
Submitted 10 November 2008
; accepted in final form 24 March 2009
...The nasal epithelium of the cystic fibrosis (CF) mouse has been used extensively in CF research because it exhibits ion transport defects similar to those of human CF airways. This tissue is composed of 50% olfactory (OE) and 50% ciliated epithelium (CE), and on the basis of previous observations, we hypothesized that a significant fraction of the bioelectric signals from murine nasal tissue may arise from OE rather than CE, while CE is the target tissue for CF gene therapy. We compared the bioelectric properties of isolated OE from the nasal cavity and CE from the nasopharynx in Ussing chamber studies. Hyperabsorption of Na + amiloride response; CF vs. wild type (WT) was 7.5-fold greater in the OE compared with the CE. The forskolin response in native tissues did not reliably distinguish genotypes, likely due to a cyclic nucleotide-gated cation conductance in OE and a calcium-mediated Cl – conductance in CE. By potential difference assay, hyperabsorption of Na + (CF vs. WT) and the difference in response to apical 0 Cl – buffer (CF vs. WT) were 2-fold greater in the nasal cavity compared with the nasopharynx. Our studies demonstrate that in the CF mouse, both the hyperabsorption of Na + and the Cl – transport defect are of larger magnitude in the OE than in the CE. Thus, while the murine CF nasal epithelium is a valuable model for CF studies, the bioelectrics are likely dominated by the signals from the OE, and assays of the nasopharynx may be more specific for studying the ciliated epithelium.
chloride transport; nasal potential difference; sodium transport; cystic fibrosis
Address for reprint requests and other correspondence: B. R. Grubb, Cystic Fibrosis/Pulmonary Research and Treatment Center, 7011 Thurston-Bowles Bldg., CB 7248, The Univ. of North Carolina, Chapel Hill, NC 27599-7248 ( bgrubb{at}med.unc.edu )
We have previously shown that most small-angle scattering (SAS) patterns from a range of polymer fibers can be best fit using elliptical coordinates, where the minor axis of the ellipse is parallel ...to the fiber axis. Analysis of small-angle X-ray scattering patterns obtained during elongation of poly(ethylene-
co-1-octene) films now shows that elliptical coordinates also provide a natural way to fit four-point radiating-out patterns, also called “butterfly” or “eyebrow” patterns. To fit this type of pattern the minor axis of the ellipse is tilted away from the fiber draw direction by some angle
α, and as the films have rotational symmetry, the pattern is fitted with the sum of two ellipses, at +
α and −
α. As a result, the reflections appear to fall on a hyperbolic arc – the butterfly pattern. The Matlab code to carry out this analysis on full 2D data is available on request. The elliptical analysis is primarily an empirical fit, much strengthened by its ability to be applied to all observed types of SAS patterns and by its relevance to the mechanical behavior of polymers.
A poly(hydroxybutyrate-
co-hydroxyvalerate) with 9% hydroxyvalerate content has been thermally degraded to give a set of materials of different molecular weights. The effect of molecular weight on ...the lamellar structure, thermal and mechanical properties was investigated. The long period, lamellar and amorphous thickness all increase as molecular weight increases; their values vary linearly with 1/(molecular weight). Observed melting temperatures increase with molecular weight, following the same functional form, while melting enthalpy and non-isothermal crystallization temperature decrease. Young's modulus varies by 13% with molecular weight; changes in crystallinity cannot explain this effect in detail. Ultimate tensile strength increases rapidly with molecular weight and then levels off at 28.5
MPa above 10
5
g/mol. This can also be seen as a linear variation with 1/(molecular weight). The strain at the point of ultimate tensile strength also increases rapidly up to 10
5
g/mol but then continues to increase at a slower rate.
Cystic Fibrosis/Pulmonary Research and Treatment Center, University of North Carolina, Chapel Hill, North Carolina
Submitted 8 June 2005
; accepted in final form 7 September 2005
The ion transport ...defects reported for human cystic fibrosis (CF) airways are reproduced in nasal epithelia of the CF mouse. Although this tissue has been studied in vivo using the nasal potential difference technique and as a native tissue mounted in the Ussing chamber, little information is available on cultured murine nasal epithelia. We have developed a polarized cell culture model of primary murine nasal epithelia in which the CF tissue exhibits not only a defect in cAMP-mediated Cl secretion but also the Na + hyperabsorption and upregulation of the Ca 2+ -activated Cl conductance observed in human airways. Both the wild-type and CF cultures were constituted predominantly of undifferentiated cuboidal columnar cells, with most cultures exhibiting a small number of ciliated cells. Although no goblet cells were observed, RT-PCR demonstrated the expression of Muc5ac RNA after 22 days in culture. The CF tissue exhibited an adherent layer of mucus similar to the mucus plaques reported in the distal airways of human CF patients. Furthermore, we found that treatment of CF preparations with a Na + channel blocker for 7 days prevented formation of mucus adherent to epithelial surfaces. The cultured murine nasal epithelial preparation should be an excellent model tissue for gene transfer studies and pharmacological studies of Na + channel blockers and mucolytic agents as well as for further characterization of CF ion transport defects. Culture of nasal epithelia from F508 mice will be particularly useful in testing drugs that allow F508 CFTR to traffic to the membrane.
F508 mice; cystic transmembrane conductance regulator; epithelial sodium channel; mucus; Na + hyperabsorption
Address for reprint requests and other correspondence: B. R. Grubb, Cystic Fibrosis/Pulmonary Research and Treatment Center, 7011 Thurston-Bowles Bldg., CB#7248, Univ. of North Carolina, Chapel Hill, NC 27599-7248 (e-mail: bgrubb{at}med.unc.edu )
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