Cell Membrane Complex Research Articles

A simplified sum-frequency vibrational imaging setup used for imaging lipid bilayer arrays.

Given the complexity of cell membranes, there is a need for an analytical technique which can explore the physical properties of lipid membranes in a high-throughput and noninvasive manner. A simplified sum-frequency vibrational imaging (SFVI) setup has been developed and characterized using asymmetrically prepared 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC):1,2-distearoyl(d70)-sn-glycero-3-phosphocholine (DSPC-d(70)) lipid bilayer arrays. Exploiting the vibrational selectivity and inherent symmetry constraints of sum-frequency generation, SFVI was successfully used to probe the transition temperature of a patterned DSPC:DSPC-d(70) lipid bilayer array. SFVI was also used to study the phase behavior in a multicomponent micropatterned lipid bilayer array (MLBA) prepared using three different binary lipid mixtures (1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC):DSPC, DOPC:1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC:DSPC). This paper demonstrates that a simplified SFVI setup provides the necessary chemical imaging capabilities with the spatial resolution, sensitivity, and field of view required for exploring lipid membrane properties in a high-throughput array based assay.

The effect of ultrasonication on the micro-splitting of wool fiber

In this study, normal and dichlorodicyanuric acid (DCCA)-treated wool slivers were ultrasonicated in formic acid aqueous solutions. The effect of the ultrasonication condition on the wool fiber splitting was examined and the mechanism of the splitting by ultrasonication in formic acid was elucidated. No wool fiber splitting occurred at formic acid content up to 70 %, but the fiber splitting accelerated as the formic acid content increased from 75 %. Although no fiber splitting occurred up to 450W ultrasonic power, the degree of splitting increased significantly with increasing ultrasonic power above 450W. The wool fiber splitting by ultrasonication was heterogeneous and FE-SEM observations revealed a three-step splitting process: 1) full removal of scale, 2) removal of cell membrane complex (CMC), and 3) fiber splitting. A comparison of the fiber splitting of normal and DCCA-treated wool revealed that the scale of the ultrasonication-treated wool was removed by peeling off rather than by dissolution.

Nanoscale Strain-Hardening of Keratin Fibres

Mammalian appendages such as hair, quill and wool have a unique structure composed of a cuticle, a cortex and a medulla. The cortex, responsible for the mechanical properties of the fibers, is an assemblage of spindle-shaped keratinized cells bound together by a lipid/protein sandwich called the cell membrane complex. Each cell is itself an assembly of macrofibrils around 300 nm in diameter that are paracrystalline arrays of keratin intermediate filaments embedded in a sulfur-rich protein matrix. Each macrofibril is also attached to its neighbors by a cell membrane complex. In this study, we combined atomic force microscopy based nano-indentation with peak-force imaging to study the nanomechanical properties of macrofibrils perpendicular to their axis. For indentation depths in the 200 to 500 nm range we observed a decrease of the dynamic elastic modulus at 1 Hz with increasing depth. This yielded an estimate of 1.6GPa for the lateral modulus at 1 Hz of porcupine quill’s macrofibrils. Using the same data we also estimated the dynamic elastic modulus at 1 Hz of the cell membrane complex surrounding each macrofibril, i.e., 13GPa. A similar estimate was obtained independently through elastic maps of the macrofibrils surface obtained in peak-force mode at 1 kHz. Furthermore, the macrofibrillar texture of the cortical cells was clearly identified on the elasticity maps, with the boundaries between macrofibrils being 40–50% stiffer than the macrofibrils themselves. Elasticity maps after indentation also revealed a local increase in dynamic elastic modulus over time indicative of a relaxation induced strain hardening that could be explained in term of a α-helix to β-sheet transition within the macrofibrils.

Infection-Induced Regulation of Natural Killer Cells by Macrophages and Collagen at the Lymph Node Subcapsular Sinus

Infection leads to heightened activation of natural killer (NK) cells, a process that likely involves direct cell-to-cell contact, but how this occurs in vivo is poorly understood. We have used two-photon laser-scanning microscopy in conjunction with Toxoplasma gondii mouse infection models to address this question. We found that after infection, NK cells accumulated in the subcapsular region of the lymph node, where they formed low-motility contacts with collagen fibers and CD169(+) macrophages. We provide evidence that interactions with collagen regulate NK cell migration, whereas CD169(+) macrophages increase the activation state of NK cells. Interestingly, a subset of CD169(+) macrophages that coexpress the inflammatory monocyte marker Ly6C had the most potent ability to activate NK cells. Our data reveal pathways through which NK cell migration and function are regulated after infection and identify an important accessory cell population for activation of NK cell responses in lymph nodes.

Fractal properties of macrophage membrane studied by AFM

Complexity of cell membrane poses difficulties to quantify corresponding morphology changes during cell proliferation and damage. We suggest using fractal dimension of the cell membrane to quantify its complexity and track changes produced by various treatments. Glutaraldehyde fixed mouse RAW 264.7 macrophage membranes were chosen as model system and imaged in PeakForce QNM (quantitative nanomechanics) mode of AFM (atomic force microscope). The morphology of the membranes was characterized by fractal dimension. The parameter was calculated for set of AFM images by three different methods. The same calculations were done for the AFM images of macrophages treated with colchicine, an inhibitor of the microtubule polymerization, and microtubule stabilizing agent taxol. We conclude that fractal dimension can be additional and useful parameter to characterize the cell membrane complexity and track the morphology changes produced by different treatments.

Nonlinear displacement of ventral stress fibers under externally applied lateral force by an atomic force microscope

Actin-based stress fibers (SFs) have fundamental importance in the maintenance of mechanical stability of living cells. Several in vitro measurements of their elastic properties have therefore been made, but direct mechanical manipulation of individual SFs in vivo for the determination of their mechanical properties has not been attempted. No less important is a search for the possible formation of a global mechanical network involving SFs and other intracellular filamentous components. In this article, we present an application of atomic force microscopy to probe into a live cell and laterally push selected SFs in a fibroblast cells (VNOf 06 fibroblast-like cells derived from rat vomeronasal tissue) transfected with a green fluorescent protein-β-actin gene. The transfected cells were transferred to a serum-depleted medium before the atomic force microscope manipulation. The lateral displacement of the SFs under a point loading condition recorded on a fluorescence microscope revealed both linear and nonlinear displacements against the contour distance from the point of force loading. The nonlinear displacements of the SFs were attributed to their association with a cortical actomyosin-cell membrane complex that effectively pulled them back as a 2D thin plate.

The Combined Effect of Electrostatic Field and Osmotic Pressure on the Stability of Bilayer Lipid Membranes

The issue of cell membrane stability is central in membranology. The extreme complexity of cell membranes urges to study this problem using a model, a bilayer lipid membrane (BLM). The membranes is often impacted both by electric forces and osmotic pressure. Therefore, it is of interest to investigate the joint activity of electric field and osmotic pressure on the stability of BLM. In the given work the joint activity of the osmotic pressure and transmembrane potential difference on the stability of BLM was experimentally investigated. As the capacity parameter a mean lifetime of BLM was taken at the given magnitudes of electric field and the osmotic pressure.At first the change of the mean lifetime of BLM with increasing of potential difference in absence of osmotic pressure was investigated. It was shown that the activity of electrostatic field brings decreasing of BLM mean lifetime. After the influence of potential difference on BLM mean lifetime was experimentally investigated at addition of osmotic pressure. It was shown that a dependence curve of BLM mean lifetime with potential for the second case is lower than in the first case. The loss of stability of BLM at addition of osmotic pressure is conditioned by two factors: either changing linear tension of BLM pore, changing the numbers of defects on BLM. The analysis theoretical formula and its confrontations with obtained data shows that the decreasing of mean lifetime of BLM at addition of osmotic pressure can be connected with the changing of linear tension. It is indicated that the difference in osmotic pressure of BLM does not bring to increasing of an area.

Gradient Distribution of LFA-1 cluster Size in the Immunological Synapse

Coordinated rearrangement of membrane receptors into clusters upon ligand binding has emerged as an important regulator of cellular signaling. However, due to cell membrane complexity and constant change in membrane morphology, probing protein clusters that are below optical resolution in live cells has long been a challenging task. Here, we use a hybrid live cell-supported membrane interface to enable high resolution fluorescence fluctuation measurements. We explore the cluster distribution of LFA-1:ICAM-1 complexes that form a ring that spans ∼10µm in diameter in a mature immunological synapse(IS). IS formation is triggered by exposing primary T cells to a supported membrane that is functionalized with activating peptide-MHC and ICAM-1-YFP. We scan across the IS with a focused laser beam to monitor the photobleaching and fluorescence fluctuations of LFA-1 bound ICAM-1-YFP molecules within the optical area. By changing the excitation laser from continuous to a pulsed form and applying photon counting histogram analysis, we are able to distinguish between the fluorescent signals of large clusters from those of small clusters. Instead of an even distribution, our results indicate that LFA-1:ICAM-1 complexes at the IS exhibit two differently clustered populations. One population shows a strong association to the T cell cytoskeleton and thus displays low lateral mobility. Interestingly, the fraction and the size of clusters of the cytoskeleton-associated complexes increase toward the center of the IS. The second population of LFA-1:ICAM-1 complexes exhibits random diffusion and a small cluster number that is no more than 3 on average. Therefore, more compartmentalized domains exist beyond the classical picture of three concentric zones at the IS. This supports the frictional force coupling model as the mechanism of IS formation, i.e. larger clusters experience larger transport force toward the center of the IS.

Hair Shaft Damage from Heat and Drying Time of Hair Dryer

BackgroundHair dryers are commonly used and can cause hair damage such as roughness, dryness and loss of hair color. It is important to understand the best way to dry hair without causing damage.ObjectiveThe study assessed changes in the ultra-structure, morphology, moisture content, and color of hair after repeated shampooing and drying with a hair dryer at a range of temperatures.MethodsA standardized drying time was used to completely dry each hair tress, and each tress was treated a total of 30 times. Air flow was set on the hair dryer. The tresses were divided into the following five test groups: (a) no treatment, (b) drying without using a hair dryer (room temperature, 20℃), (c) drying with a hair dryer for 60 seconds at a distance of 15 cm (47℃), (d) drying with a hair dryer for 30 seconds at a distance of 10 cm (61℃), (e) drying with a hair dryer for 15 seconds at a distance of 5 cm (95℃). Scanning and transmission electron microscopy (TEM) and lipid TEM were performed. Water content was analyzed by a halogen moisture analyzer and hair color was measured with a spectrophotometer.ResultsHair surfaces tended to become more damaged as the temperature increased. No cortex damage was ever noted, suggesting that the surface of hair might play a role as a barrier to prevent cortex damage. Cell membrane complex was damaged only in the naturally dried group without hair dryer. Moisture content decreased in all treated groups compared to the untreated control group. However, the differences in moisture content among the groups were not statistically significant. Drying under the ambient and 95℃ conditions appeared to change hair color, especially into lightness, after just 10 treatments.ConclusionAlthough using a hair dryer causes more surface damage than natural drying, using a hair dryer at a distance of 15 cm with continuous motion causes less damage than drying hair naturally.

The cell membrane complex: Three related but different cellular cohesion components of mammalian hair fibers

The structure, chemistry and physical properties of the cell membrane complex (CMC) of keratin fibers are reviewed, highlighting differences in the three types of CMC. Starting with Rogers' initial description of the CMC in animal hairs, several important developments have occurred that will be described, adding new details to this important structure in mammalian hair fibers. These developments show that essentially all of the covalently bound fatty acids of the beta layers are in the cuticle and exist as monolayers. The beta layers of the cortex are bilayers that are not covalently bonded but are attached by ionic and polar linkages on one side to the cortical cell membranes and on the other side to the delta layer. The delta layer between cortical cells consists of five sublayers; its proteins are clearly different from the delta layer that exists between cuticle cells. The cell membranes of cuticle cells are also markedly different from the cell membranes of cortical cells. Models with supporting evidence are presented for the three different types of cell membrane complex: cuticle-cuticle CMC, cuticle-cortex CMC, and cortex-cortex CMC.

Effects of potential calcium sensing receptor inducers on promoting chemosensitivity of human colon carcinoma cells

We have previously reported that by inducing calcium sensing receptor (CaSR), calcitriol, the active form of vitamin D, promoted the sensitivity of the human colon carcinoma cells to anticancer drugs. In the current study we tested several other potential CaSR modulators, calcipotriol and the effective components of Chinese herbal medicine lentinan, for their functions in inducing CaSR expression in human colon carcinoma cell line CBS, Moser, Fet, and SW480 cells and subsequently promoting sensitivity of the cells to anticancer drugs. Calcipotriol and lentinan suppressed invasion of the colon carcinoma cells and enhanced the cytotoxicity of anticancer regimen FOLFIRI to cells in culture or in anchorage-independent growth. In the mechanism study we found that calcipotriol and lentinan suppressed protein expression and gene transcriptional activities of survivin and thymidylate synthase, increased E-cadherin cell membrane localization and complex formation of E-cadherin and beta-catenin, and repressed TCF4 transcriptional activation. These effects were attenuated, however, in CaSR knocked-down cells, indicating that CaSR was required in the pathway. We concluded that calcipotriol and lentinan are efficient in promoting chemosensitivity in colon carcinoma cells. Since both compounds have much less side effects than calcitriol in clinic, they have greater potential to be applied as supplements of colon cancer therapy.

Immunocamouflage: The biophysical basis of immunoprotection by grafted methoxypoly(ethylene glycol) (mPEG)

Development of novel approaches for the immunomodulation of donor cells would have significant utility in transfusion and transplantation medicine. Immunocamouflage of cell surfaces by covalently grafted methoxypoly(ethylene glycol) (mPEG) (PEGylation) has emerged as a promising approach. While previous studies demonstrated the in vitro and in vivo efficacy of immunocamouflaged allogeneic blood cells, the biophysical mechanisms of immunoprotection have not been well-defined due to the fragility of intact cells. To overcome this limitation, polystyrene beads (1.2 and 8.0 μm) were used to elucidate the biophysical effects of polymer size, density and linker chemistry on charge camouflage and protein adsorption. These findings were correlated with biological studies using red blood cells and lymphocytes. Charge camouflage of both beads and cells was best achieved with long polymers. However, protein adsorption studies demonstrated an unexpected effect of target size. For 1.2 μm beads, decreased protein adsorption was best achieved with short (2 kDa) polymers whereas long chain (20 kDa) polymers were optimal for 8.0 μm particles. The biophysical findings correlated well with biological immunocamouflage as measured by particle electrophoresis and the inhibition of antibody–antigen (CD3, CD4 and CD28) recognition. Moreover, it was observed that antigen topography (CD28 vs. CD4) was of significance in selecting the appropriate polymer size. The biophysical interactions of PEGylated surfaces and macromolecules involve complex mechanisms dependent on the molecular weight, grafting concentration, target size and surface complexity. Cellular PEGylation strategies must be customized to account for target cell size, membrane complexity and antigen density and height.

Effect of the Cell Membrane Complex on Dyeing of Wool Fibers with Hematein

We have found the difference on the color between the silk fiber and the wool fiber dyed with hematein and then investigated the effect of the cell membrane complex (CMC) on the K/S-wavelength (l) curve of the dyed wool fiber to clarify the origin of their difference. First we determined the dye uptake of the cortex cells dyed with hematein and found that the uptake of cortex cells was low apparently compared to that of the wool fiber, indicating that hematein adsorbed mainly to the CMC region of the wool fiber. Further, we examined the dyeing behavior of the wool fiber pretreated by ethylene diaminetetraacetic acid (EDTA) with hematein and found that the K/S-l curve of the EDTA pretreated wool fiber is compatible with that of the dyed silk fiber. We assumed from these results that the difference on the color between the silk fiber and the wool fiber dyed with hematein can be ascribed in part to the complex between hematein and the trace metal ions included in the CMC regions.

Morphological Analysis of Shrinkproof Wool Fibers by SEM Combined with Alkaline and Enzymatic Etching Techniques: Microstructural Differences of DCCA- and Kroy-Processed Fibers

Morphologies of the shrinkproof wool fibers were analyzed by scanning electron microscopy (SEM) combined with enzymatic and alkaline etchings of the cross-sectional fiber specimens. The enzymatic etching revealed the microstructural changes of the fiber components produced by after the DCCA (dichloroisocyanuric acid)- and Kroy-processes. It was suggested that the inner-fiber reactions had extended to cell membrane complex (CMC), intermacrofibril material (int-MF), and nuclear remnants (NRs), as well as to the components in their vicinity of present in the orthocortex and paracortex of the both shrinkproof wool fibers. On the other hand, the alkaline etching gave different patterns between the DCCA- and Kroy-processed fibers; in the former fibers the paracortex and orthocortex gave weak and almost disintegrated patterns, respectively, while in the latter fibers both cortexes gave weak patterns. It was therefore supported that the inner-fiber reactions are more homogeneous in the Kroy-process than in the DCCA-process although the degrees of inner-fiber reaction are almost comparable in both processes. Based on these data, the difference in laundering effect on the Kroy- and DCCA-processed fibers was discussed.

Macrophage- and Dendritic-Cell-Derived Interleukin-15 Receptor Alpha Supports Homeostasis of Distinct CD8+ T Cell Subsets

Interleukin-15 receptor alpha (IL-15R alpha) is a pleiotropically expressed molecule that chaperones and trans-presents IL-15 to NK and T cells. To investigate whether IL-15R alpha presented by different cells perform distinct physiological functions, we have generated four lines of mice lacking IL-15R alpha in various cell types. We find that IL-15R alpha expression on macrophages but not dendritic cells (DCs) supports the early transition of antigen specific effector CD8(+) T cells to memory cells. After memory CD8(+) T cell differentiation, IL-15R alpha expression on DCs selectively supports central memory CD8(+) T cells, whereas IL-15R alpha expression on macrophages supports both central and effector memory CD8(+) T cells. By contrast, mice lacking IL-15R alpha on macrophages, DCs, or both, exhibit equivalent defects in NK cell homeostasis and activation. These studies define unique roles for macrophage expression of IL-15R alpha and show that NK cells rely upon distinct IL-15R alpha dependent IL-15 signals than memory CD8(+) T cells. Moreover, they demonstrate the diversity, specification, and geographic restriction of cytokine signals.

Physico-chemical and Dyeing Properties of Raw Wool Extracted By Pressurized CO2/Modifiers

Raw wool fibers were extracted with CO2 with different organic modifiers (acetone, ethanol and methanol) at constant pressure and temperature. The influence of experimental extraction conditions on the properties of extracted wools was studied to determine the suitability of the fibers for textile applications. Suitable wool wax removal from raw wool fibers was qualitatively evaluated by optical microscopy and scanning electron microscopy observation as well as by whiteness, yellowness and brightness measurements. Moreover, the possible bulk fiber modification as a consequence of internal wool lipid extraction belonging to the cell membrane complex was assessed by determining the bursting resistance and dyeing behavior. The results suggest that both CO2/ethanol and CO2/methanol extraction methods provide well-scoured wool fibers with an acceptable dyeing behavior, the bulk modification being negligible.

Influence of Internal Lipid on Dyeing of Wool Fibers

Wool is a natural fiber that is mainly composed of protein. It has an external lipid content widely used in cosmetics (lanoline) and minor internal lipid content (1.5%), which could be of added value given its high content of ceramides. Internal wool lipids (IWL) were mostly extracted at laboratory level and the lipid composition was quantitatively analyzed. The hydrophobic character of both extracted and non-extracted fibers was evaluated. These wool fibers were dyed with two acid dyes with a different degree of hydrophilic character. Dyeing behavior and washing fastness tests were assessed to study whether the lipid extraction could affect fiber dyeing. The results obtained showed the different behaviors of extracted and non-extracted wool fibers when dyes with different hydrophilic character were used. Given the absence of surface hydrophobic differences between the two fibers, the different dyeing behavior may be attributed to the interaction between IWL and the dyestuff, highlighting the importance of dye penetration through the cell membrane complex. A lipid extraction could therefore modify wool dyeing strategies and reduce the final dyeing temperature without impairing the washing fastness of the fibers.

No. 1 Vol. 42, No. 3, pp. 185–200, 2008 A tentative mechanism of oxidative dyeing for keratin fibers

A number of papers have been published on the oxidation coupling reactions of the active intermediate of a dye precursor with an electron‐rich dye coupler. At the present stage, however, little information is available about the oxidative reaction mechanism to colored oxidation products and the dye distribution inside the keratin fibers. From the results of dyeing in the presence of both reducing agent and chelate agent, we found that the coupling reaction developing oxidation dyes occurred on the outer surface in the cuticle cell phase, and that not only the cell membrane complex (CMC) regions play an important role as accumulation regions of the finished dye, but also the components of CMC contribute directly to the oxidation coupling reactions. Furthermore, we suggested from the other results that the metal ions and disulphide bond presented primarily within the intercellular materials play an important role in developing the colored oxidation dye. Then, on the basis of the general scheme of the oxidation dye process, we proposed a tentative mechanistic scheme of oxidative dyeing to account for the oxidation dying phenomenon of keratin fibers. The scheme is made up of four step‐reactions.Keywords: oxidation dye, oxidative dyeing, precursor, coupler, active intermediate, keratin fiber, chelate agent, cell membrane complex, intercellular materials, metal ion, disulphide bond

Absorption of nanoparticles by wool

Wool was treated with inorganic and polymer‐based nanoparticles. The diffusion of nanoparticles into wool appears to be dependent on electrostatic interactions. In particular, it is optimised at low pH in which there are very few anionic groups on the wool fibre; the nanoparticles also need to have sufficient charge to maintain their stability as dispersion. The findings support the view that the cell membrane complex and other low sulphur regions are the main route of entry for both molecular and macromolecular treatment chemicals. The use of this technique to treat wool may lead to new coloration effects and other functions such as antimicrobial action.

Scaffolded Vesicles as a Model Membrane System

Biological cell membranes are complex systems that consist primarily of a phospholipid bilayer, into which cholesterol, proteins etc. may be integrated. Due to the complexity of biological cell membranes, it is desirable to develop model membrane systems that can be more easily studied. Scaffolded vesicles are an example of such a system. A scaffolded vesicle model membrane system offers a number of advantages with respect to other model systems. By using a porous material as the scaffold, one can achieve an aqueous environment on both sides of the model membrane. This allows for the study of membrane transport processes. The scaffolds porosity may also allow one to more easily integrate transmembrane proteins into the bilayer. Importantly, such a system would remain accessible to both electrochemical and surface analytical techniques. Using FTIR-ATR spectroscopy, the orientation of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) coated on the porous scaffold (as a 70:30 DMPC:cholesterol bilayer) will be determined. Proteins may then be incorporated into the bilayer of the scaffolded model membrane system for study.