High-resolution freeze-etching replica images of the disk and the plasma membrane surfaces in purified bovine rod outer segments

Molecular organization of the photoreceptor disk membrane was revealed by the freeze-deep etching replica method using purified and successively rinsed bovine rod outer segment (ROS). Various membrane particles with different shape and sizes were found on cytoplasmic surface (PS face) as well as on both P and E fracture faces, which are presumed to be peripheral membrane proteins such as transducin, phosphodiesterase, guanylate cyclase and so on. Membrane particles seen on PS face were catalogued in size. The histogram on their number and size showed that they were classified at least into two major groups, the group of particles about 50 nm2 in size and the group of particles about 115 nm2 in size. The distribution density of the 115 nm2 particle was 1200 microm(-2) in native state, but it decreased to 125 microm(-2) after washing with hypotonic buffer solution. Namely, the group of the 115 nm2-particle seems to be mainly composed of peripheral membrane proteins. Rinsing with the sucrose free buffer at the final step of the purification procedure enabled us to observe three types of filaments localized in ROS (filaments connecting disk to disk at the margin, filaments connecting disk to the plasma membrane, filaments associated with PS face of disk membrane); and also to find characteristic domains with crystal arrangement of particles on the external surface (ES face) of ROS plasma membrane.     

On the Origin of Ion Selectivity in Ultrathin Nanopores: Insights for Membrane‐Scale Osmotic Energy Conversion

Nanopores in ultrathin or atomically thin membranes attract broad interest because the infinitesimal pore depth allows selective transport of ions and molecules with ultimate permeability. Toward large‐scale osmotic energy conversion, great challenges remain in extrapolating the promising single‐pore demonstration to really powerful macroscopic applications. Herein, the origin of the selective ion transport in ultrathin nanopores is systematically investigated. Based on a precise Poisson and Nernst–Planck model calculation, it is found that the generation of net diffusion current and membrane potential stems from the charge separation within the electric double layer on the outer membrane surface, rather than that on the inner pore wall. To keep the charge selectivity of the entire membrane, a critical surface charged area surrounding each pore orifice is therefore highly demanded. Otherwise, at high pore density, the membrane selectivity and the overall power density would fall down instead, which explains the giant gap between the actual experimental achievements and the single‐pore estimation. To maximize the power generation, smaller nanopores (pore diameter ≈1–2 nm) are appropriate for large‐scale osmotic energy conversion. With a porosity of ≈10%, the total power density approaches more than 200 W m^‐2, anticipating a substantial advance toward high‐performance large‐scale nanofluidic power sources.     

Electron microscopy of pathogenic yeasts Cryptococcus neoformans and Exophiala dermatitidis by high-pressure freezing

A high-pressure freezing method was used to observe the ultrastructure of pathogenic yeasts, Cryptococcus neoformans and Exophiala dermatitidis, after freeze-substitution and ultrathin sectioning. The method well preserved the cell structure in its natural state, since the capsule, cell wall, plasma membrane, nucleus, outer and inner nuclear membranes, nuclear pores, nucleolus, mitochondria, mitochondrial membrane and cristae, vacuoles, endoplasmic reticulum, Golgi apparatus, spindle pole body, ribosomes, lipid droplets, microtubules, actin filaments, and glycogen granules were clearly visible. The method was shown to freeze cells as deep as 0.1 mm by sectioning the sample perpendicular to specimen surface. The quality of the cell image was similar to that obtained by a rapid freezing method when compared using the same materials. Thus, high-pressure freezing would be useful for making serial ultrathin sections for three-dimensional analysis of cells, which should give basic information of structure and function of pathogenic yeast cells necessary for finding an effective therapy for mycoses.     

Ultrastructural verification of anchoring role of lamina fibroreticularis of dental basement membrane in odontogenesis

In a previous study of the developing tooth a characteristic fibrillar layer associated with the basement membrane of the inner enamel epithelium was found to be a highly specialized lamina fibroreticularis of the basement membrane which is unusually rich in basotubules, 10 nm wide microfibril-like structures. In this study this layer was further examined in detail in the hope of ultrastructurally elucidating its role in odontogenesis. Tooth germs of the monkey (Macaca fuscata) were processed for thin section observations. Dental papilla cell processes were inserted into the lamina fibroreticularis and their surface was closely associated with numerous parallel basotubules. With high-resolution observations the space between the surface and nearest basotubules as well as the spaces between neighbouring basotubules were bridged by 1.5-3 nm wide filaments running perpendicular to the axis of basotubules. These results indicate that the processes of dental papilla cells are linked to groups of basotubules by means of 1.5-3 nm wide filaments. Immunoperoxidase staining showed the presence of fibronectin along basotubules as well as within the space between the process and basotubule. This result, together with the comparison of these filaments with microfibril-associated 1.2-3 nm wide fibronectin filaments and the reported abundance of fibronectin in the basement membrane area during odontogenesis, indicates that these 1.5-3 nm wide filaments are composed of fibronectin. After immunostaining for amyloid P component, done with the rat tissue because of the nature of an available antiserum, basotubules in the lamina fibroreticularis were positively stained, as has been shown in basotubules/microfibrils in other locations. Microfibrils function as anchoring rods by interlinking connective tissue components to one another and to the cells. Basotubules, thought to be basement membrane-incorporated microfibrils, in the lamina fibroreticularis in this study are also likely to function as an anchoring device to immobilize dental papilla cells along the basement membrane. Such an arrangement of mesenchymal cells is known to be crucial for the successful differentiation of odontoblasts in the developing tooth.     

Correspondence between the location of evoked potential generators and sites of maximal sensitivity to stimulation

The potential recorded by a set of electrodes as an action potential traverses a small axonal segment is proportional to the transmembrane potential produced during stimulation of that axon segment by the same set of recording electrodes, under certain circumstances. First, the membrane must have a constant thickness which is so small that the difference between the surface area of the inner and outer surfaces is minimal. Second, all media must be linear. Third, there must be a monotonically increasing relation between the mean transmembrane potential induced by a stimulus and the maximum transmembrane potential. Fourth, as each axon segment depolarizes, the transmembrane current and change in membrane potential during this time are same. This principle remains true for magnetic stimulation and recording as long as currents generated at the boundaries between regions of differing conductivity outside the axon contribute minimally to the field at the axon. This allows the identification of the point at which an action potential generates a maximal extracellular potential as the point that is stimulated with the lowest threshold.     

Rhodopsin and retinitis pigmentosa: shedding light on structure and function

Rhodopsin is the dim-light activated photoreceptor located in the rod cells of the eye. It belongs to the large superfamily of G-protein-coupled receptors (GPCRs). Many consider it the proto-typical GPCR as numerous studies since its cloning in 1983 (Nathans and Hogness 1983) have established many fundamental principles of seven transmembrane-spanning GPCRs. Abundant expression in the rod's outer segment, constituting about 90% of the total membrane protein in the discs, and the development of techniques to purify large quantities of functional protein has facilitated this process. Another distinct feature is rhodopsin's ligand, 11-cis-retinal, which is covalently bound via a Schiff base to transmembrane seven (TM VII), allowing extensive spectroscopic studies. Exciting recent developments include the discovery of naturally occurring mutations that lead to retinal degeneration, the determination of transmembrane movements using electron paramagnetic resonance (EPR) and biochemical techniques, and the discovery of its 3D X-ray crystal structure, the first among GPCRs. The impact of these major advances will be discussed in this review.     

Ultracytochemistry of enzymes in cell biology of the retina.

In matured chick retina, alkaline phosphatase activity is specifically localized in the outer plexiform layer and in the horizontal and Müller cells. In developing chick retina, the activity is recognized in growing neurites from horizontal cells, when synaptogenesis begins in the outer plexiform layer. Using levamisole, a potent inhibitor of alkaline phosphatase, on chick retina in vivo and in vitro, the enzyme was shown to play a significant role in retinal cell differentiation. 5'-Nucleotidase is used as a marker for the rod photoreceptors. It became apparent that the 'displaced' rod cells are localized in the inner nuclear layer of postnatal retina. High activity of glucose-6-phosphatase is confirmed in the cisternae of the smooth and rough endoplasmic reticulum, together with the space of the perinuclear envelope in the pigment epithelium of rat. The process of disc membrane recycling in the rod outer segment was investigated cytochemically to reveal sequential changes in lysosomal digestion both by conventional enzyme cytochemistry and by high voltage electron microscopy. With conventional enzyme histochemistry as well as with rapid freeze substitution enzyme cytochemistry, all enzyme for cGMP metabolism were observed to be on the cytoplasmic side of the disc membranes.     

局部包络片控制误差简化三角网格模型

利用内外包络控制三角网格模型简化误差是一种可行的方法.但构造整个模型的内外包络需要花费较多的时间,且每次简化操作时也会进行大量重复的三角形相交检测,使算法效率下降.针对这些问题,本文提出了局部包络片控制误差的思想,即每次简化操作仅利用相关部分的包络来控制误差.算法采用三角形收缩操作简化模型,当原始模型上的一个三角形被选中,并尝试进行收缩操作时,先构造其对应的局部包络片;在该三角形收缩后,原来与之相邻的三角形将发生形变,若这些形变后的三角形位于局部包络片之间,则认为此次三角形收缩操作可以接受,否则予以取消.实验结果表明,局部包络片保持了良好的误差控制效果,且由于构造简单,很少发生自交现象,减少了三角形相交检测次数,提高了计算效率.     

Modeling of the internal fields distribution in human inner hearing system exposed to 900 and 1800 MHz

This paper investigates the internal electric and magnetic field distribution and the specific absorption rate (SAR) values in a magnetic resonance imaging-based model of the inner hearing system exposed to 900 and 1800 MHz. The internal fields distributions were calculated using the Finite Integration Technique. The estimation of the field values was evaluated along lines passing through that target organ, specifically from the vestibular to the cochlear region and from the apex to the base of the cochlea. The specific findings are: 1) higher internal fields strength and SAR value in the vestibular region rather than in the auditory region, especially for the inner ear closer to the external source; 2) higher internal fields strength in the basal and apical region of the cochlea than in the middle one; 3) local differences in the internal fields distribution and SAR value, comparing the head models including or not the inner auditory system model; 4) results' variability evaluated by changing the head-source mutual position and the dielectric properties of the inner hearing system.     

Three-dimensional studies of cytoskeletal organizations in cultured thyroid cells by quick-freezing and deep-etching method

Isolated porcine thyroid cells were cultured on collagen gels (control group, TSH-stimulated group, and double-layered culture). They were split or cut to remove cytoplasmic soluble proteins for replica preparations. Some specimens were immunostained with anti-actin antibody or decorated with S1 myosin fragments to identify actin filaments. The basal cell membranes of thyroid cells of monolayer culture were in contact with collagen gels and the apical cell membranes faced the culture medium. Networks of actin filaments were attached to the cytoplasmic sides of the apical cell membranes, while intermediate filaments were localized along the basal ones. The thyroid-stimulating hormone (TSH) treatment induced the formation of microvilli only on the apical cell membranes and the accumulation of actin filaments under the apical cell membranes, indicating the apical-basal polarity of the cells. In double-layered culture, the primitive follicular lumens with microvilli appeared between two adjacent cells. The interaction of cell membranes with collagen gels is a determinant factor in the orientation of apical-basal polarity. Moreover, the TSH treatment and cell-cell contact further intensify the polarization through reorganizing the cytoskeletons.     

High‐Efficiency Water‐Transport Channels using the Synergistic Effect of a Hydrophilic Polymer and Graphene Oxide Laminates

Graphene oxide (GO) laminates possess unprecedented fast water‐transport channels. However, how to fully utilize these unique channels in order to maximize the separation properties of GO laminates remains a challenge. Here, a bio‐inspired membrane that couples an ultrathin surface water‐capturing polymeric layer (<10 nm) and GO laminates is designed. The proposed synergistic effect of highly enhanced water sorption from the polymeric layer and molecular channels from the GO laminates realizes fast and selective water transport through the integrated membrane. The prepared membrane exhibits highly selective water permeation with an excellent water flux of over 10 000 g m^?2 h^?1, which exceeds the performance upper bound of state‐of‐the‐art membranes for butanol dehydration. This bio‐inspired strategy demonstrated here opens the door to explore fast and selective channels derived from 2D or 3D materials for highly efficient molecular separation.     

An osmium-ferricyanide staining method for the intercellular space in the small intestinal epithelium.

The post-fixation with osmium tetroxide and potassium ferricyanide (OsFeCN) produced dense deposits on the outer surface of the lateral plasma membrane in the mouse small intestinal epithelium. The deposits also filled up the intercellular space. No deposits were found on the surface of apical and basal plasma membranes. This staining pattern was highly reproducible when pH of the OsFeCN solution was adjusted to 7.0 by cacodylate buffer without calcium ion. Thus, our modified OsFeCN method is useful to selectively stain the intercellular space in epithelial tissues.     

Crystal structure and mechanism of GlpT, the glycerol-3-phosphate transporter from E. coli

The major facilitator superfamily represents the largest group of secondary active membrane transporters in prokaryotic and eukaryotic cells. They transport a vast variety of substrates, presumably via similar mechanisms, yet the details of these mechanisms remain unclear. Here we report the 3.3 A resolution structure of a member of this superfamily--GlpT, the glycerol-3-phosphate transporter from the E. coli inner membrane, in the absence of a substrate. The antiporter mediates the exchange of glycerol-3-phosphate for inorganic phosphate across the membrane. Its N- and C-terminal domains exhibit a pseudo 2-fold symmetry along an axis perpendicular to the membrane. Eight of the twelve transmembrane alpha-helices are arranged around a centrally located substrate translocation pore that is closed off at the periplasmic surface. Present at the beginning of the pore are two arginine residues that presumably comprise the substrate-binding site which is accessible only from the cytosol, suggesting an inward-facing conformation for the transporter. The central loop connecting the N- and C-terminal domains is partially disordered and exhibits reduced susceptibility to trypsin in the presence of substrate, indicating conformational changes. We propose that GlpT operates via a single binding-site, alternating-access mechanism.     

水稻胚乳细胞淀粉质体被膜与其增殖的关系

水稻胚乳细胞经高锰酸钾固定法固定后，显示出十分清晰的内膜结构，尤其展现了淀粉质体被膜与淀粉质体增殖在结构上的密切关系。常见的淀粉质体出芽、缢缩和形成中间隔板的增殖方式分别与淀粉质体外层被膜、内外层被膜、内层被膜的活动有关。另外，淀粉质体外层被膜也能出芽，形成双层膜小泡，再积累淀粉形成淀粉质体；而淀粉质体内层被膜向内出泡或内外层被膜同时内陷，在淀粉体内形成新淀粉质体，这是淀粉质体增殖的两种新方式。对淀粉质体被膜在淀粉质体增殖中的作用进行了讨论。     

基于统计特征的非理想虹膜图像分割方法

由于退化条 件的存在,非理想虹膜识别的关键在于正确 分割虹膜区域,这一区域包含能 够用于个体识别的纹理。本文提出了一种基于统计特性的非理想虹膜图像分割方法,包括内 边界定位、外边界定位和眼睑检 测3个阶段。在内边界定位阶段,通过高斯混合(GMM)模型及多弦长均衡策略,实现对瞳 孔及虹膜中心的精确定位;在外边界定 位阶段,利用简化的基于区域信息的曲线演化方法,将其与序统计滤波(OSF)结合,以确保 曲线收敛至虹膜外边界;在 眼睑检测阶段,利用二次曲线对眼睑进行建模。对多个数据库进行实验的结果表明,本 文 方法能够有效克服反光、睫毛和 眼睑遮挡、外边界模糊等不利因素的影响,精确实现了非理想虹膜图像的分割。     

QoS Guaranteed Resource Allocation Scheme for Cognitive Femtocells in LTE Heterogeneous Networks with Universal Frequency Reuse

In this paper, we propose a novel resource allocation scheme for co-channel interference avoidance in LTE heterogeneous networks with universal spectrum reuse where both macro users (MUs) and cognitive femto base stations (FBSs) within the same macrocell coverage can dynamically reuse whole spectrum. Specifically, resource blocks (RBs) are shared between cognitive FBSs in underlay mode while the resource sharing among FBSs and MUs is in overlay mode. The macrocell is divided into inner and outer regions with the inner region further divided into three sectors. The proposed scheme addresses co-channel interference (CCI) by employing fractional frequency reuse (FFR) for RB allocation in the outer region of the macrocell and increase the distance of users that reuse the same RB within the macrocell. Part of RBs are allocated to the outer region of the macrocell with a FFR factor of 1/3, while the remaining RBs are dynamically allocated to each sector in the inner region of macrocell based on MUs demand to efficiently utilize the available spectrum. A basic macro base station (MBS) assistance is required by the FBS in selection of suitable RB to avoid interference with MU in each sector. With the proposed solution, both macro and femto users can dynamically access the whole spectrum while having minimum bandwidth guarantee even under fully congested scenarios. Moreover, the proposed scheme practically eliminates the cross-tier interference and the CCI problem in heterogeneous network reduces to inter-femtocell interference. The throughput and outage performances of the proposed scheme are validated through extensive simulations under LTE network parameters. Simulation results show that the proposed scheme achieves a performance gain of more than 1.5 dB in terms of SINRs of both macro user and femto user compared to traditional cognitive and non-cognitive schemes without bandwidth guarantee for femtocells.     

A Hierarchical Janus Nanofibrous Membrane Combining Direct Osteogenesis and Osteoimmunomodulatory Functions for Advanced Bone Regeneration

An ideal guided bone regeneration membrane (GBRM) is expected not only to perform barrier function, but also to enhance osteogenesis and resist bacteria infection. However, currently available membranes have limited bioactivities. To address this challenge, a Janus GBRM (JGM) is designed and fabricated by sequential fractional electrospinning here. The random gelatin fibers loaded with hydroxyapatite (HAP) are designed as the inner face to promote the osteoblasts’ adhesion, proliferation, and osteogenic differentiation, meanwhile the aligned poly(caprolactone) (PCL) nanofibers loaded with poly(methacryloxyethyltrimethyl ammonium chloride-co-2-Aminoethyl 2-methylacrylate hydrochloride) (P(DMC-AMA)) are designed as the outer layer to resist epithelia invasion and bacterial infection. In vitro assays reveal that the inner face displays enhanced osteogenic effects, meanwhile the outer surface can regulate the epithelia to spread along the aligned direction and kill the contacted bacteria. Interestingly, the outer face can induce macrophages to polarize toward the M2 phenotype, thus manipulating a favorable osteoimmune environment. These results suggest that the JGM simultaneously meets the critical requirements of barrier, osteogenic, antibacterial, and osteoimmunomodulatory functions. Consequently, the JGM shows better in vivo bone tissue regeneration performance than the commercial Bio-Gide membrane. This work provides a novel platform to design multi-functional membranes/scaffolds, displaying great potential applications in tissue engineering.     

Genetically Modifiable Flagella as Templates for Silica Fibers: From Hybrid Nanotubes to 1D Periodic Nanohole Arrays

Bacterial flagellum is a protein nanotube that is helically self‐assembled from thousands of a protein subunit called flagellin. The solvent‐exposed domain of each flagellin on the flagella is genetically modifiable, in that a foreign peptide can be genetically inserted into this domain, leading to the high‐density display of this foreign peptide on the surface of flagella. In this work, wild‐type and genetically engineered flagella (inner diameter of ～2 nm and outer diameter of ～14 nm) detached from the surface of Salmonella bacterial cells are used as templates to site‐specifically form silica sheaths on the flagellar surface, resulting in the formation of double‐layered silica/flagella nanotubes. The flagella templates inside the silica/flagella nanotubes can be removed to obtain silica nanotubes by calcining the nanotubes at high temperature (550°C). Further calcination of the silica nanotubes at a higher temperature (800 °C) leads to the formation of a periodic nanohole array along the silica fibers with a center‐to‐center nanohole spacing of ～79 nm. It is demonstrated that the double‐layered silica‐flagella nanotubes can be used for selective CdTe quantum dot uptake into the inner channels or selective Au nanoparticle coating on the outer wall of the nanotubes due to the different chemistry between inner flagellum core (protein) and outer silica wall of the nanotubes. It is also found that flagella displaying different peptides result in different morphologies of the silica nanotubes. This work suggests that the monodisperse diameter and genetically tunable surface chemistry of the flagella can be exploited for the fabrication of silica nanotubes with uniform diameter and controllable morphologies as well as silica nanofibers decorated with periodic nanohole arrays.     

Scanning electron microscopy of proteoglycans in metaphyseal cartilage.

Alcian blue (AB) was used for scanning electron microscope investigations on metaphyseal cartilage. In the pericellular area three-dimensional network connecting the cell membrane surface to the lacunar wall is evident. The network is formed by very long rod-like filaments about 50 nm thick. The segments may be interpreted as the proteoglycans (PGs) of the pericellular area. In the pericellular area in hypertrophic and degenerative zones, the rod-like segments are closely connected to "chain granules" which are the morphological expression of Ca-P non crystalline compounds. The rod-like segments are not at all evident either in glutaraldehyde-osmium fixed fragments or in predigested-(streptococcal hyaluronidase and chondroitinase) AB stained ones. It is concluded that AB is a good method to detect the three-dimensional spatial disposition of cartilage PGs.     

Rate Regions for Relay Broadcast Channels

A partially cooperative relay broadcast channel (RBC) is a three-node network with one source node and two destination nodes (destinations 1 and 2) where destination 1 can act as a relay to assist destination 2. Inner and outer bounds on the capacity region of the discrete memoryless partially cooperative RBC are obtained. When the relay function is disabled, the inner bound reduces to an inner bound on the capacity region of broadcast channels that includes an inner bound of Marton, and GePfand and Pinsker. The outer bound reduces to a new outer bound on the capacity region of broadcast channels that generalizes an outer bound of Marton to include a common message, and that generalizes an outer bound of GePfand and Pinsker to apply to general discrete memoryless broadcast channels. The proof for the outer bound simplifies the proof of GePfand and Pinsker that was based on a recursive approach. Four classes of RBCs are studied in detail. For the partially cooperative RBC with degraded message sets, inner and outer bounds are obtained. For the semideterministic partially cooperative RBC and the orthogonal partially cooperative RBC, the capacity regions are established. For the parallel partially cooperative RBC with unmatched degraded subchannels, the capacity region is established for the case of degraded message sets. The capacity is also established when the source node has only a private message for destination 2, i.e., the channel reduces to a parallel relay channel with unmatched degraded subchannels.