Topical treatments with acylceramide dispersions restored stratum corneum lipid lamellar structures in a reconstructed human epidermis model.

Long-periodicity phase (LPP) lamellar structures in intercellular lipid matrixes of the stratum corneum (SC) are considered important for maintenance of skin permeability barriers. Acylceramides are essential components of LPP structures, and their absence influences skin barriers under physiological and pathological conditions, such as atopic dermatitis and dry skin. Although topical applications of acylceramide have been shown to facilitate maintenance of the skin barrier, it is unknown whether topically applied acylceramides are incorporated into intercellular lipids to form LPP structures. Thus, we assessed the effects of topical treatments with monomodal acylceramides on the formation of LPP structures in a surfactant-insulted reconstructed human epidermis model using small-angle X-ray scattering (SAXS) and transmission electron microscopy (TEM) analyses. In SAXS experiments, LPP structures give rise to a diffraction peak which indicates the presence of a structure with a 13 nm real space repeat distance. LPP patterns of intercellular lipid matrixes in the SC were disrupted' by surfactant treatments and were recovered by topical acylceramide treatments. TEM images also showed specific repeating patterns of LPP structures, indicating that topical acylceramide treatments facilitate recovery of LPP structures in the SC. The present data show that the application of acylceramides might temporarily modify the lipid structure to resemble that of normal skin although the underlying cause of dry or diseased skin is not fully clarified.

Label-free imaging by stimulated parametric emission microscopy reveals a difference in hemoglobin distribution between live and fixed erythrocytes.

We demonstrate that stimulated parametric emission (SPE) microscopy enables label-free, 3-D visualization of internal hemoglobin distribution of live mouse and chicken erythrocytes with high sensitivity. Change in hemoglobin distribution in chicken erythrocytes before and after ethanol fixation is clearly visualized.

Broadband group delay dispersion compensation for a microscope objective lens with a specially-designed mechanical deformable mirror.

We propose that the bending profile of a mechanical deformable mirror can be designed by shaping its form, realizing a simple, compact, and broadband group delay dispersion compensator in a 4-f pulse shaper arrangement. By using the proposed compensator, spectral phase distortion of a microscope objective lens is successfully pre-compensated for to generate a sub-8 fs pulse at the focus of the lens.

Axopodial degradation in the heliozoon Raphidiophrys contractilis: a novel bioassay system for detecting heavy metal toxicity in an aquatic environment.

We observed the physiological effects of zinc, lead, mercury, copper, cadmium, and arsenic on the axopodia of the centrohelid heliozoon Raphidiophrys contractilis. In the presence of these heavy metal ions, the axopodial length of the heliozoon decreased in a concentration-dependent manner. When the heavy metal ions were examined at the same concentration, mercury produced the strongest effect on axopodia. At a high concentration (> 10-3 M) of any of the heavy metal ions examined, axopodia disappeared and cells became disrupted. Axopodia were also degraded by the addition of solutions with an acidic (< or = 6) or basic (> or = 8) pH. These observations indicate the toxic effects of heavy metal ions and non-neutral pHs on axopodial length, and also signify that R. contractilis can be used as an effective biological tool for the study of metal poisoning in eukaryotic cells.

Ca2+-dependent in vitro contractility of a precipitate isolated from an extract of the heliozoon Actinophrys sol.

Contraction of axopodia in actinophrid heliozoons (protozoa) is induced by a unique contractile structure, the "contractile tubules structure (CTS)". We have previously shown that a cell homogenate of the heliozoon Actinophrys sol yields a precipitate on addition of Ca2+ that is mainly composed of filamentous structures morphologically identical to the CTS. In this study, to further characterize the nature of the CTS in vitro, biochemical and physiological properties of the precipitate were examined. SDS-PAGE analysis showed that the Ca2+-induced precipitate was composed of many proteins, and that no proteins in the precipitate showed any detectable changes in electrophoretic mobility on addition of Ca2+. Addition of extraneous proteins such as bovine serum albumin to the cell homogenate resulted in cosedimentation of the proteins with the Ca2+-induced precipitate, suggesting that the CTS has a high affinity for other proteins that are not related to precipitate formation. Appearance and disappearance of the precipitate were repeatedly induced by alternating addition of Ca2+ and EGTA, and its protein composition remained unchanged even after repeated cycles. When adhered to a glass surface, the precipitate showed Ca2+-dependent contractility with a threshold of 10-100 nM, and this contractility was not inhibited by colchicine or cytochalasin B. The precipitate repeatedly contracted and relaxed with successive addition and removal of Ca2+, indicating that the contraction was controlled by Ca2+ alone with no need for any other energy supply. From our characterization of the precipitate, we concluded that its Ca2+-dependent formation and contraction are associated with the unique contractile organelle, the "contractile tubules structure".

Ca2+-dependent nuclear contraction in the heliozoon Actinophrys sol.

Ca2+-dependent contractility was found to exist in the nucleus of the heliozoon protozoan Actinophrys sol. Upon addition of Ca2+ ([Ca2+]free = 2.0 x 10(-3) M), diameters of isolated and detergent-extracted nuclei became reduced from 16.5+/-1.7 microm to 11.0+/-1.3 microm. The threshold level of [Ca2+]free for the nuclear contraction was 2.9 x 10(-7) M. The nuclear contraction was not induced by Mg2+, and was not inhibited by colchicine or cytochalasin B. Contracted nuclei became expanded when Ca2+ was removed by EGTA; thus cycles of contraction and expansion could be repeated many times by alternating addition of Ca2+ and EGTA. The Ca2+-dependent nuclear contractility remained even after high salt treatment, suggesting a possible involvement of nucleoskeletal components in the nuclear contraction. Electron microscopy showed that, in the relaxed state, filamentous structures were observed to spread in the nucleus to form a network. After addition of Ca2+, they became aggregated and constructed a mass of thicker filaments, followed by re-distribution of the filaments spread around inside of the nucleus when Ca2+ was removed. These results suggest that the nuclear contraction is induced by Ca2+-dependent transformation of the filamentous structures in the nucleus.

Axopodial contraction in the heliozoon Raphidiophrys contractilis requires extracellular Ca2+.

Axopodial contraction of the centrohelid heliozoon Raphidiophrys contractilis was induced by mechanical or electrical stimulation. For inducing contraction, extracellular Ca(2+) was required. The threshold level of extracellular Ca(2+) was between 10(-6)-10(-7) M. The speed of axopodial contraction was faster than 3.0 mm/sec. Re-elongation of axopodia started just after contraction, and its initial velocity was approximately 0.30 microm/sec. Electron microscopic observations were carried out using an improved fixative that contained 1 mg/ml ruthenium red and 15 microM Taxol. This fixative prevented artificial retraction of axopodia and resulted in better fixation. A bundle of hexagonally-arranged microtubules was observed in each axopodium, but no other filamentous structures were detected, suggesting that the contractile machinery of axopodia in R. contractilis may be different from that in actinophryid heliozoons in which Ca(2+)-dependent contractile filaments are employed for contraction.

Gliding movement in Peranema trichophorum is powered by flagellar surface motility.

A colorless euglenoid flagellate Peranema trichophorum shows unique unidirectional gliding cell locomotion on the substratum at velocities up to 30 micro m/s by an as yet unexplained mechanism. In this study, we found that (1) treatment with NiCl(2) inhibited flagellar beating without any effect on gliding movement; (2) water currents applied to a gliding cell from opposite sides caused detachment of the cell body from the substratum. With only the anterior flagellum adhering to the substratum, gliding movement continued along the direction of the anterior flagellum; (3) gentle pipetting induced flagellar severance into various lengths. In these cells, gliding velocity was proportional to the flagellar length; and (4) Polystyrene beads were translocated along the surface of the anterior flagellum. All of these results indicate that a cell surface motility system is present on the anterior flagellum, which is responsible for cell gliding in P. trichophorum.