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PubMed2006

Slit2 and netrin 1 act synergistically as adhesive cues to generate tubular bi-layers during ductal morphogenesis.

Development of many organs, including the mammary gland, involves ductal morphogenesis. Mammary ducts are bi-layered tubular structures comprising an outer layer of cap/myoepithelial cells (MECs) and an inner layer of luminal epithelial cells (LECs). Slit2 is expressed by cells in both layers, with secreted SLIT2 broadly distributed throughout the epithelial compartment. By contrast, Robo1 is expressed specifically by cap/MECs. Loss-of-function mutations in Slit2 and Robo1 yield similar phenotypes, characterized by disorganized end buds (EBs) reminiscent of those present in Ntn1(-/-) glands, suggesting that SLIT2 and NTN1 function in concert during mammary development. Analysis of Slit2(-/-);Ntn1(-/-) glands demonstrates an enhanced phenotype that extends through the ducts and is characterized by separated cell layers and occluded lumens. Aggregation assays show that Slit2(-/-);Ntn1(-/-) cells, in contrast to wild-type cells, do not form bi-layered organoids, a defect rescued by addition of SLIT2. NTN1 has no effect alone, but synergistically enhances this rescue. Thus, our data establish a novel role for SLIT2 as an adhesive cue, acting in parallel with NTN1 to generate cell boundaries along ducts during bi-layered tube formation.

Animals↗
PubMed2001

Cortical upper layer neurons derive from the subventricular zone as indicated by Svet1 gene expression.

The cerebral cortex is composed of a large variety of different neuron types. All cortical neurons, except some interneurons, are born in two proliferative zones, the cortical ventricular (VZ) and subventricular (SVZ) zones. The relative contribution of both proliferative zones to the generation of the diversity of the cortical neurons is not well understood. To further dissect the underlying mechanism, molecular markers specific for the SVZ are required. Towards this end we performed a subtraction of cDNA libraries, generated from E15.5 and E18.5 mouse cerebral cortex. A novel cDNA, Svet1, was cloned which was specifically expressed in the proliferating cells of the SVZ but not the VZ. The VZ is marked by the expression of the Otx1 gene. Later in development, Svet1 and Otx1 were expressed in subsets of cells of upper (II-IV) and deep (V-VI) layers, respectively. In the reeler cortex, where the layers are inverted, Svet1 and Otx1 label precursors of the upper and deeper layers, respectively, in their new location. Interestingly, in the Pax6/small eye mutant, Svet1 activity was abolished in the SVZ and in the upper part of the cortical plate while the Otx1 expression domain remained unchanged. Therefore, using Svet1 and Otx1 as cell-type-specific molecular markers for the upper and deep cortical layers we conclude that the Sey mutation affects predominantly the differentiation of the SVZ cells that fail to migrate into the cortical plate. The abnormality of the SVZ coincides with the absence of upper layer cells in the cortex. Taken together our data suggest that while the specification of deep cortical layers occurs in the ventricular zone, the SVZ is important for the proper specification of upper layers.

Animals↗
PubMed2006

Significance of a basal melanin layer to production of non-iridescent structural plumage color: evidence from an amelanotic Steller's jay (Cyanocitta stelleri).

Non-iridescent structural plumage color is typically produced by coherent scattering of light within a matrix of keratin and air (a ;spongy layer') in feather barbs. It remains unclear what role, if any, the basal melanin layer underlying this spongy layer plays in the production of coloration. Amelanism in birds with structural color is a ;natural experiment' in which melanin pigmentation is lost, allowing us to identify the effects of the loss of melanin on structural color production. Here we use full-spectrum spectrometry, transmission electron microscopy and Fourier analysis to compare the color and nanostructure of an amelanotic Steller's jay (Cyanocitta stelleri Gmelin) feather with a normal blue Steller's jay feather. As a control, we also examined a white domestic chicken (Gallus gallus Linnaeus) feather. The pale amelanotic jay feather had a broad reflectance curve with a blue/green peak, while the typical blue feather had a typical distinct bell-shaped blue curve with a UV/violet peak. The white chicken feather had a typical white reflectance curve with no discrete peaks. Electron microscopy revealed that both the amelanotic and blue feather barbs contained well-formed spongy layers that were of the correct size and arrangement to produce their measured peak reflectance values, whereas the chicken feather had no spongy layer. The washed-out color of the amelanotic jay feather was thus most probably caused by the loss of the basal melanin layer, suggesting that melanin functions to absorb incoherently scattered white light from the feather barb thereby increasing the purity of the color produced by the spongy layer.

Animals↗
PubMed1991

Filtering centrifugation through two layers of silicone oil: a method for the kinetic analysis of rapid metabolite transport in organelles.

Kinetic studies of ATP uptake in amyloplasts from sycamore (Acer pseudoplatanus L.) have been performed with a newly developed method of centrifugation through a double layer of silicone oil; the results are compared with the frequently used method of centrifugation through a single layer. The present technique employs two separate silicone layers: the upper one prevents mixing of the organelle preparation with the incubation layer, which contains the metabolites, and the lower one prevents mixing of the incubation and pelleting layers. Incubation of the organelles takes place in the incubation layer after the upper layer of silicone inverts during centrifugation. Depending on the speed of centrifugation, incubation periods as short as 1 sec and as long as 2 min can be achieved. High reproducibility and accuracy, acquisition of multiple data in a single centrifugation, and maintenance of structural integrity and metabolic activity of the organelles make the technique advantageous for the analysis of metabolite transport in amyloplasts and other kinds of organelles.

Adenosine Triphosphate↗
PubMed1996

Polarity of cultured newborn rat skin basal cell layer.

Basal cells were prepared from the newborn rat skin epidermis and cultured on a type I collagen-coated filter. This cultured basal cell layer showed an amiloride-sensitive polarity, and the apical surface of the cell layer was more acidic than the basal surface. When the basal cells were cultured with ascorbic acid, a polarity reverse of the cultured cell layer was observed. In this case, synthesized and accumulated type IV collagen was detected on the whole apical surface of the cultured cell layer. An inhibitor of collagen synthesis, cis-hydroxyproline (cis-OH-Pro), suppressed the ascorbic acid-induced polarity reversal. Furthermore, when basal cells were cultured on a type IV collagen-coated filter, the amiloride-sensitive polarity of the cultured cell layer resembled the cultivation on a type I collagen-coated filter. In this case, the polarity was reversed by the addition of ascorbic acid, and cis-OH-Pro suppressed this reversal. Based on these results, we concluded that cultured skin basal cell layer showed an amiloride-sensitive polarity. Moreover, newly synthesized and accumulated type IV collagen was deduced to affect the polarity reversal in the cultured basal cell layer.

Amiloride↗
PubMed1976

[Respiratory function of mitochondria in the inner and outer layers of the dog myocardium].

The respiratory effect of dog myocardial mitochondria was studied by measuring oxygen content in a reaction chamber using Garbani electrode. 1) In normal dog mitochondria, oxygen consumption rate (QO2) in state 3 and state 4, respiratory control ratio (RCR) and ADP/O ratio were not different between the inner and outer layers when glutamate or succinate was used as substrate. 2) Thirty min after the occlusion of one of the small branches of the anterior descending artery, there were no differences in the respiratory effects between the inner and outer layers when glutamate or succinate was used as substrate. HOowever, compared with the normal dog mitochondria, ADP/O ratio, QO2 in state 3 and state 4 were lower in the outer layer and QO2 in state 4 was lower in the inner layer when succinate was used as substrate. 3) When the anterior descending branch was narrowed to decrease the blood flow to about a half, the respiratory effects were not different between the inner and outer layers when succinate was used as substrate. However, ADP/O ratio in the outer layer was lower than that of the normal dog. 4) Nitroglycerin had no effect on the respiratory effects either in vitro (1.63 X 10(-6) M, succinate as substrate) or in vivo (20 mug/kg i.v., glutamate as substrate.) 5) Dipyridamole had no effect on the respiratory effects in vivo (250 mug/kg, i.v., glutamate as substrate) but, ADP/O ratio in the outer layer was diminished in vitro (7.34 X 10(-6) M, succinate as substrate).

Animals↗
PubMed1999

Rat gastric mucous gel layer contains sialomucin not produced by the stomach.

The sialylated mucus components of the normal gastric mucosa and mucous gel layer of rats were studied by using various histochemical staining methods including Maackia amurensis II (MAL-II) and Sambucus nigra (SNA) lectins, alcian blue (AB) pH 2.5 -- periodic acid Schiff (PAS) and high iron diamine (HID) -- AB pH 2.5. The acidic and neutral mucins characterized by the AB-PAS staining were abundantly present in the mucous gel layer as well as in the gastric mucosa. The sialomucin characterized by HID-AB was barely found in either the mucous gel layer or the mucosa. The sialomucin positive to MAL-II and SNA, which react with the N-acetyl neuraminic acid residue linked to galactose via an alpha-linkage, was moderately detected only in the mucous gel layer, but not in the entire mucosal layer. Furthermore, in animals given surgery to form an esophageal fistula through which saliva was excluded or in animals subjected to salivectomy, the mucous gel layer stained with MAL-II and SNA lectins was markedly decreased. These results indicate that a part of the sialomucin containing-mucous gel layer covering normal rat gastric mucosa originates from the saliva and that MAL-II and SNA lectins are useful for detecting this specific sialomucin.

Animals↗
PubMedDate not supplied

The four-layer bandage system from a nursing perspective.

This article describes an evaluation that took place in 20 health centres, community clinics and hospitals throughout the UK. The aim was to explore nurses' opinions of three different four-layer bandage systems: kit A (System 4); kit B (Charing Cross); and kit C (Profore). Fifty-seven pairs of nurses were involved; within each pair, one was the applier and one the wearer. Each nurse was asked to complete a questionnaire which highlighted their opinions relating to ease of application, conformability, comfort and preferred bandage length. Responses were analysed using simple frequency counts, means, standard errors, Fisher's exact test and Kruskal-Wallis' one-way analysis of variance. The nurses involved were regular users of either the Charing Cross or Profore systems. Kit A was significantly easier to apply than kit B for layer 1 (P < 0.001), layer 2 (P < 0.05) and layer 3 (P < 0.01). There was no significant difference in relation to layer 4. With regard to ease of application there were no statistically significant differences between kits A and C or B and C. Layer 1 of kit A was significantly more conformable than layer 1 of kit B (P < 0.001). There were no significant differences between kits B and C, or A and C or in relation to comfort for all bandage kits. Overall preference by the applier was: 25 for kit A, nine for kit B and 15 for kit C. Overall preference by the wearer was: 23 for kit A, 10 for kit B and 14 for kit C.

Attitude of Health Personnel↗
PubMed2003

Determination of the optical properties of a two-layer tissue model by detecting photons migrating at progressively increasing depths.

We have investigated a method for solving the inverse problem of determining the optical properties of a two-layer turbid model. The method is based on deducing the optical properties (OPs) of the top layer from the absolute spatially resolved reflectance that results from photon migration within only the top layer by use of a multivariate calibration model. Then the OPs of the bottom layer are deduced from relative frequency-domain (FD) reflectance measurements by use of the two-layer FD diffusion model. The method was validated with Monte Carlo FD reflectance profiles and experimental measurements of two-layer phantoms. The results showed that the method is useful for two-layer models with interface depths of >5 mm; the OPs were estimated, within a relatively short time (<1 min), with a mean error of <10% for the Monte Carlo reflectance profiles and with errors of <25% for the phantom measurements.

Algorithms↗
PubMed2001

Recovery of optical parameters in multiple-layered diffusive media: theory and experiments.

Diffuse photon density waves have lately been used both to characterize diffusive media and to locate and characterize hidden objects, such as tumors, in soft tissue. In practice, most biological media of medical interest consist of various layers with different optical properties, such as the fat layer in the breast or the different layers present in the skin. Also, most experimental setups consist of a multilayered system, where the medium to be characterized (i.e., the patient's organ) is usually bounded by optically diffusive plates. Incorrect modeling of interfaces may induce errors comparable to the weak signals obtained from tumors embedded deep in highly heterogeneous tissue and lead to significant reconstruction artifacts. To provide a means to analyze the data acquired in these configurations, the basic expressions for the reflection and transmission coefficients for diffusive-diffusive and diffusive-nondiffusive interfaces are presented. A comparison is made between a diffusive slab and an ordinary dielectric slab, thus establishing the limiting distance between the two interfaces of the slab for multiple reflections between them to be considered important. A rigorous formulation for multiple-layered (M-layered) diffusive media is put forward, and a method for solving any M-layered medium is shown. The theory presented is used to characterize a two-layered medium from transmission measurements, showing that the coefficients of scattering, mu'(s) , and absorption, mu(a) , are retrieved with great accuracy. Finally, we demonstrate the simultaneous retrieval of both mu;(s) and mu(a).

Diffusion↗
PubMed2004

Compositional changes of the muscular layers of the stomach with aging.

To elucidate compositional changes of the stomach with aging, the authors investigated age-related changes of the elements and relationships among the elements in the muscular layers of the pylorus, cardia, lesser curvature, and greater curvature by inductively coupled plasma-atomic emission spectrometry. After ordinary dissection by medical students, the pylori, cardias, lesser curvatures, and greater curvatures were removed from the subjects, consisting of 19 men and 1 woman, ranging in age from 65 to 95 yr. The muscular layers were isolated and the element contents were determined. The calcium content increased progressively with aging in the muscular layers of the pylorus, cardia, and lesser curvature, whereas it tended to increase in the muscular layer of the greater curvature with aging. Regarding sulfur, the content increased significantly in the muscular layer of the pylorus with aging, but not significantly in the muscular layers of the cardia, lesser curvature, and greater curvature with aging. Regarding the relationships among the elements, significant direct correlations were found among calcium, phosphorus, sulfur, and magnesium in both the muscular layers of the pylori and cardias, with some exceptions.

Aged↗
PubMed2006

Radial migration of superficial layer cortical neurons controlled by novel Ig cell adhesion molecule MDGA1.

MAM (meprin/A5 protein/receptor protein tyrosine phosphatase mu) domain glycosylphosphatidylinositol anchor 1 (MDGA1), a unique cell surface glycoprotein, is similar to Ig-containing cell adhesion molecules that influence neuronal migration and process outgrowth. We show in postnatal mice that MDGA1 is expressed by layer 2/3 neurons throughout the neocortex. During development, MDGA1 is expressed in patterns consistent with its expression by migrating layer 2/3 neurons, suggesting a role for MDGA1 in controlling their migration and settling in the superficial cortical plate. To test this hypothesis, we performed loss-of-function studies using RNA interference (RNAi) targeting different sequences of mouse MDGA1. RNAi or empty vectors were coelectroporated with an enhanced green fluorescent protein reporter in utero into the lateral ventricle at embryonic day 15.5 to transfect progenitors of superficial layer neurons; the distributions of transfected neurons were analyzed late on postnatal day 0. We found a direct correlation between effectiveness of an RNAi in suppressing MDGA1 expression and disrupting migration of superficial layer neurons. An RNAi with no effect on MDGA1 expression has no effect on the migration. In contrast, an RNAi that suppresses MDGA1 expression also blocks proper migration of transfected superficial layer neurons, with essentially all transfected cells found deep in the cortical plate or beneath it. This migration defect is rescued by cotransfection of a rat MDGA1 expression construct along with the effective RNAi, confirming that the RNAi effect is specific to diminishing mouse MDGA1 expression. RNAi transfections of deep layer neurons that do not express MDGA1 do not significantly affect their migration. We conclude that MDGA1 acts cell autonomously to control the migration of MDGA1-expressing superficial layer cortical neurons.

Animals↗
PubMed2004

The density and refractive index of adsorbing protein layers.

The structure of the adsorbing layers of native and denatured proteins (fibrinogen, gamma-immunoglobulin, albumin, and lysozyme) was studied on hydrophilic TiO(2) and hydrophobic Teflon-AF surfaces using the quartz crystal microbalance with dissipation and optical waveguide lightmode spectroscopy techniques. The density and the refractive index of the adsorbing protein layers could be determined from the complementary information provided by the two in situ instruments. The observed density and refractive index changes during the protein-adsorption process indicated the presence of conformational changes (e.g., partial unfolding) in general, especially upon contact with the hydrophobic surface. The structure of the formed layers was found to depend on the size of the proteins and on the experimental conditions. On the TiO(2) surface smaller proteins formed a denser layer than larger ones and the layer of unfolded proteins was less dense than that adsorbed from the native conformation. The hydrophobic surface induced denaturation and resulted in the formation of thin compact protein films of albumin and lysozyme. A linear correlation was found between the quartz crystal microbalance measured dissipation factor and the total water content of the layer, suggesting the existence of a dissipative process that is related to the solvent molecules present inside the adsorbed protein layer. Our measurements indicated that water and solvent molecules not only influence the 3D structure of proteins in solution but also play a crucial role in their adsorption onto surfaces.

Adsorption↗
PubMed1999

Multi-layered periodontal pocket epithelium reconstituted in vitro: histology and cytokeratin profiles.

BACKGROUND: In order to study inter-individual differences in bacterial adhesion/invasion of periodontal tissues, an in vitro model for culturing multi-layered pocket epithelium without feeder layers or stromal equivalents (including the evaluation of their cytokeratin profiles) was developed. METHODS: Pocket epithelium was collected and grown until confluent in Falcon flasks using keratinocyte-serum free medium (KSFM), without a feeder layer. In the second passage, oral keratinocytes were re-grown in a 2 compartment system using either a clear polyester (transwell-clear [TCL]) or a collagen (transwell-col [TCO]) membrane as culture surface. After the first week, the calcium concentration was raised to 1.2 mM and in half the wells, the KSFM was supplemented with 10% fetal calf serum (FCS). Histology and immunohistochemistry were performed after 1, 2, and 3 weeks of additional growth. RESULTS: In general, all conditions resulted in a structured epithelium consisting of 3 to 5 layers, but important differences were observed between the membrane types and between the media. CK4 was rarely and only lightly expressed while CK18 and 19 (characteristic of junctional epithelium) were very strongly expressed in the older (2 and 3 weeks) cultures. CK13 and 14 (characteristic of any stratifiable epithelial cell) also tended to increase over time; CK13 seemed to be stronger in KSFM with FCS while the contrary was true for CK14. The multi-layer created by the combination TCL/KSFM + 10% FCS resembled a junctional epithelium most, while that grown on TCO without FCS mimicked the sulcular epithelium. CONCLUSIONS: It seems possible to create a histiotypic culture resembling either periodontal pocket or junctional epithelium without the use of stromal equivalents or feeder layers which make this approach more cumbersome. This multi-layered culture offers a model to investigate the permeability of pocket epithelium and the adhesion and penetration of bacteria under well-defined environmental conditions.

Animals↗
PubMedDate not supplied

Effect of practical layered dielectric loads on SAR patterns from dual concentric conductor microstrip antennas.

Radiation patterns of 2 and 4cm square Dual Concentric Conductor (DCC) microstrip antennas were studied theoretically with Finite Difference Time Domain (FDTD) analysis and compared with experimental measurements of power deposition (SAR) in layered lossy dielectric loads. Single and array configurations were investigated with 915 MHz excitation applied across either one, two or four sides, or four corners of the square apertures. FDTD simulations were carried out for realistic models of a muscle tissue load coupled to the DCC antennas with a 5 mm thick bolus of either distilled water or low loss Silicone Oil. This study characterizes the effect on SAR of adding three additional thin dielectric layers which are necessary for clinical use of the applicator. These layers consist of a 0.1 mm thick dielectric coating on the array surface to provide electrical isolation of DCC apertures, and 0.15 mm thick plastic layers above and below the bolus to contain the liquid. Experimental measurements of SAR in a plane 1 cm deep in muscle phantom agree well with theoretical FDTD simulations in the multi-layered tissue models. These studies reveal significant changes in SAR for applicator configurations involving low dielectric constant (Er) layers on either side of a high Er water bolus layer. Prominent changes include a broadening and centring of the SAR under each aperture as well as increased SAR penetration in muscle. No significant differences are noted between the simple and complete load configurations for the low Er Silicone Oil bolus. Both theoretical and measured data demonstrate relatively uniform SAR distributions with > 50% of maximum SAR extending to the perimeter of single and multi-aperture array configurations of DCC applicators when using a thin 5 mm water or Silicone Oil bolus.

Hyperthermia, Induced↗
PubMed1995

Evidence for synchronous activation of neurons located in different layers of primary somatosensory cortex.

Although many studies have examined the columnar organization of primary somatosensory (SI) cortex, the functional relationship among neurons in different layers remains unclear. To understand how activity is coordinated among different cortical layers, the present investigation tested the hypothesis that the initial part of a peripheral stimulus produces a serial pattern of laminar activation in SI cortex. Extracellular discharges of 334 histologically recovered neurons were recorded from the medial bank of the coronal sulcus in nine anesthetized cats during electrical or cutaneous stimulation of the distal forelimb. Mean responses during the initial 50-msec period following stimulus onset were largest in layers IIIb or IV for both types of stimulation, but laminar differences in the magnitude of onset responses were not statistically significant. Among 175 neurons with responses exceeding 0.5 spikes per stimulus, electrical stimulation consistently produced shorter response latencies than mechanical indentation in the extragranular (II, IIIa, V, VI), but not in the middle (IIIb, IV), cortical layers. The average minimum latencies for different cortical layers ranged from 7.4 to 10.1 msec for responses to electrical stimulation and from 10.3 to 11.6 msec for responses to mechanical indentations, but these laminar differences were not statistically significant. In some experiments, neurons in different layers of a cortical column were recorded simultaneously with dual-electrode assemblies; among 37 neuron pairs in which both neurons responded with more than 0.5 spikes per stimulus, response latencies were similar, even though the neurons were separated by several hundred microns. Cross-correlation analysis of the onset responses for neurons recorded simultaneously from different layers also indicated that many cells throughout a cortical column were activated nearly simultaneously by the initial phase of a peripheral stimulus. Results from the present study are compared with previous reports examining laminar patterns of activation.

Afferent Pathways↗
PubMedDate not supplied

In vitro electro-mechanical activity of the human colon. Simultaneous recording of the electrical patterns of the two muscle layers.

Electrical and mechanical activity on longitudinal and circular layers of the human sigmoid colon were simultaneously studied. Recordings were obtained from two electrode sites spaced 3 cm apart in a piece of colon which had been resected surgically and perfused in an organ bath. Spontaneous electrical activity of the colon showed slow waves and spikes. Slow waves were present for only 24.5% and 12% of the recording time on the longitudinal and circular layers, respectively, and they appeared as localized activity which was irregular in amplitude and varying in frequency. Electrical coupling between the two muscle layers was rarely seen and slow waves were not associated with pressure changes. Spiking activity were recorded as short and long spike bursts on both muscle layers. Short spike bursts were localized activity superimposed on slow waves. The associated mechanical activity, which consisted of single weak pressure changes or prolonged contractions with summation, was determined by slow wave frequency. Long spike bursts were seen at irregular intervals and were either propagated or not propagated activity associated with electrical oscillations ranging from 24 to 46 cpm. Mechanical activity consisted of sustained tonic contractions propagated or not propagated in the same way as the electrical pattern. Coordinated electrical activity of the two muscle layers seldom occurred when spontaneous activity was being recorded. Electrical activity on both muscle layers was very sensitive to stretching and could be initiated or modulated by pharmacological agents. In particular, our findings showed that stimulation induced coordinated spiking activity on the two muscle layers and caused mechanical activity, propagated orally or aborally, which consisted of long lasting, high amplitude contractions.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗
PubMed1991

Relationship between bile tolerance and the presence of a ruthenium red staining layer on strains of Lactobacillus acidophilus.

Seventeen strains of Lactobacillus acidophilus were evaluated to determine the relationship between bile tolerance and the presence of an outer polysaccharide layer exterior to the cell wall when viewed by transmission electron microscopy. Bile tolerance is necessary for survival of lactobacilli in the intestinal tract, and the polysaccharide layer may be responsible for adherence to human intestinal tissue. These two factors may be the basis for use of L. acidophilus as a dietary adjunct. Ten strains exhibited a ruthenium red-stained outer polysaccharide layer. Three of the 10 strains had extremely dense layers, which may indicate stronger adherence properties. Seven strains did not contain a ruthenium red-stained outer layer; however, six strains that did not have the stained layer were resistant to 1.0% bile concentration. Fourteen strains were tolerant to 1% bile, one strain was tolerant to 6% bile, and two strains were sensitive to bile. No relationship between bile tolerance and the presence of the ruthenium red-stained outer polysaccharide layer was apparent.

Bacterial Adhesion↗