成骨细胞在纳米材料表面上粘附特性

在利用静电自组装技术制备的具备不同纳米形貌的S iO2纳米粒子薄膜及空白试片上,进行了SD大鼠成骨细胞的粘附特性考评实验,并对比了成骨细胞分别在纳米粒子基片和空白试片上的粘附数、细胞活性及形态的差异.实验结果表明:利用静电自组装技术获得的纳米结构表面对细胞的粘附特性确实存在良性影响;在所采用的3个纳米尺度下,细胞粘附数随纳米尺度的增加而增加,M TT检测结果表明细胞活性也随之增加.     

Glycan-based interactions involving vertebrate sialic-acid-recognizing proteins

All cells in nature are covered by a dense and complex array of carbohydrates. Given their prominence on cell surfaces, it is not surprising that these glycans mediate and/or modulate many cellular interactions. Proteins that bind sialic acid, a sugar that is found on the surface of the cell and on secreted proteins in vertebrates, are involved in a broad range of biological processes, including intercellular adhesion, signalling and microbial attachment. Studying the roles of such proteins in vertebrates has improved our understanding of normal physiology, disease and human evolution.     

Visual characterization of targeted effect of holo-transferrin-tagged dihydroartemisinin on human breast cancer cells

Targeted drugs could significantly reduce cytotoxic effect and increase therapeutic activity. Dihydroartemisinin (DHA) has been shown to be effective in killing cancer cells. However, it exhibits non-targeted property. Holo-transferrin (TF) is a suitable drug-carrier to target cancer cells, because cancer cells need iron uptake by the TF-mediated mechanism to maintain their uncon- trolled growth. Furthermore, TF receptors (TF-R) are highly expressed on cancer cell surfaces. In this paper, 3-(4,5-dimethylthi- azol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to evaluate the different killing effect of 4-(12-Dihydroart- emisininoxy) Benzoic Acid Hydrozide-transferrin (DBAH-TF) on human breast cancer cells (MCF-7) cells and human normal breast (HNB) cells, and atomic force microscopy (AFM) was used to visually observe the targeted effect of DBAH-TF on MCF-7 cells. MTT results show that DBAH-TF is 172 times more potent than DHA in killing MCF-7 cells, while the cytotoxic effect of DBAH-TF on HNB cells is merely 1/33 to DHA. Also, the killing effect of DBAH-TF on MCF-7 cells is 286 times that on HNB cells, showing targeted effect. Moreover, there are distinct differences in ultrastructures of cellular surfaces after DBAH-TF and DHA treatment. Through AFM imaging, many characteristic holes were observed on the cancer cell surface after being effected by DBAH-TF, which differ from the holes with irregular shapes affected by DHA. These results visually show that the DBAH-TF targeted drug has more potent killing effect on MCF-7 cells compared with DHA.     

Interaction of Protein and Cell with Different Chitosan Membranes

Ｃｈｉｔｉｎ ,ｗｈｉｃｈｍａｉｎｌｙｃｏｎｓｉｓｔｓｏｆ β(1→4 ) 2 ａｃｅｔａｍｉｄｏ 2 ｄｅｏｘｙ Ｄ ｇｌｕｃｏｐｙｒａｎｏｓｅｓｔｒｕｃｔｕｒａｌｕｎｉｔｓ ,ｉｓｏｎｅｏｆｔｈｅｍｏｓｔａｂｕｎｄａｎｔｆｏｒｍｓｏｆｃａｒｂｏｈｙｄｒａｔｅｆｏｕｎｄｉｎｎａｔｕｒｅ[1] .Ｒｅｃｅｎｔｄｅｖｅｌｏｐｍｅｎｔｓｈａｖｅｅｎａｂｌｅｄｔｈｅｕｓｅｏｆｃｈｉｔｉｎａｎｄｃｈｉｔｏｓａｎｉｎｖａｓｔｌｙｄｉｖｅｒｓｅｆｉｅｌｄｓｒａｎｇｉｎｇｆｒｏｍｔｈｅｍｅｄｉｃｉｎｅａｎｄｃｈｅｍｉｃａｌｉ…     

Dependence on pH of polarized sorting of secreted proteins

The plasma membranes of epithelial cells are divided into apical and basolateral domains. These two surfaces are characterized by markedly different protein compositions, reflecting the ability of the cell to target newly synthesized membrane proteins to specific regions of the cell surface. This targeting capability is also apparent in the polarized release of secretory products. Recent studies using canine renal tubule (MDCK) cells have suggested that distinct sets of secretory proteins are released from their apical and basolateral poles. We report experiments designed to examine secretory protein sorting by MDCK cells. We have shown that secretion of basement membrane components (laminin and heparan sulphate proteoglycan (HSPG] takes place from the basolateral cell surface and that this polarized release results from active sorting. The sorting process which mediates this polarized secretion requires an acidic intracellular compartment. MDCK cells treated with NH4Cl to raise the pH of their intracellular compartments, secrete laminin and HSPG by a default pathway which leads to their release in roughly equal quantities into the medium of both the apical and basolateral compartments.     

Hepatic Bel-7402 Cell Proliferation on Different Phospholipid Surfaces

ＩｎｔｒｏｄｕｃｔｉｏｎＣｙｔｏｃｏｍｐａｔｉｂｉｌｉｔｙｓｔｕｄｉｅｓａｒｅａｎｉｍｐｏｒｔａｎｔｔｏｏｌｔｏｅｖａｌｕａｔｅｂｉｏｌｏｇｉｃａｌｒｅｓｐｏｎｓｅｓｔｏｂｉｏｍａｔｅｒｉａｌｓ .Ａｇｒｅａｔｄｅａｌｏｆｒｅｓｅａｒｃｈｈａｓｂｅｅｎｒｅｐｏｒｔｅｄｆｒｏｍｖａｒｉｏｕｓｐｏｉｎｔｓｏｆｖｉｅｗ ,ｓｕｃｈａｓｃｅｌｌａｄｈｅｓｉｏｎ ,ｃｅｌｌｍｏｒｐｈｏｌｏｇｙｏｂｓｅｒｖａｔｉｏｎｓ ,ｐｒｏｌｉｆｅｒａｔｉｏｎ ｑｕａｎｔｉｆｉｃａｔｉｏｎ ,ａｎｄｅｖａｌｕａｔｉｏｎｏｆｃｅｌ…     

接种密度对Vero细胞在微载体表面生长的影响

研究了接种密度对Ｖｅｒｏ细胞在微载体表面生长行为的影响,发现培养过程中所获得的细胞最高密度随接种密度的增加而有所提高,当细胞的表面接种密度高于１．０×１０＾５ｃｅｌｌｓ／ｍｇＭＣ时,因贴壁后细胞扩展受到限制而不利于进入对数生长期。就整个Ｖｅｒｏ细胞培养过程,适宜的表面接种密度为３×１０＾－４－７×１０＾４ｃｅｌｌｓ／ｍｇＭＣ,此时可以获得较高的比生长速率和细胞增殖倍数。     

Guidance of optic nerve fibres by N-cadherin adhesion molecules

The dendritic branches (neurites) of developing neurons migrate along specific pathways to reach their targets. It has been suggested that this migration is guided by factors present on the surface of other neurons or glial cells. The molecular nature of such factors, however, remains to be elucidated. N-cadherin is a cell-surface glycoprotein which belongs to the cadherin family of cell-cell adhesion molecules. This adhesion molecule is expressed in various neuronal cells as well as in glial cells of the central and peripheral nervous systems in vertebrate embryos and recent immunological studies suggested that N-cadherin may play a role in guiding the migration of neurites on myotubes or astrocytes. To further examine this possibility, we used a molecular-genetic approach; that is, we examined the outgrowth of chicken embryonic optic axons on monolayer cultures of Neuro 2a or L cells transfected with the complementary DNA encoding chicken N-cadherin. The data indicate that N-cadherin is used as a guide molecule for the migration of optic axons on cell surfaces.     

Cell orientation on a stripe-micropatterned surface

Stripe-micropatterned surfaces have recently been a unique tool to study cell orientation. In this paper, we prepared, by the photolithography transfer technique, stable gold （Au） micropatterns on PEG hydrogel surfaces with defined cell-resistant （PEG hydrogel） and cell-adhesive （gold microstripes） properties. 3T3 fibroblasts were cultured on Au-microstripe surfaces to observe cell adhesion and orientation. Five statistical parameters were defined and used to describe cell orientation on micropatterns. With the increase of inter-stripe distance, the orientational order parameter, the ratio of long and short axes of a cell, and the occupation fraction of cells on stripes increased gradually, whereas the spreading area of a single cell decreased. The abrupt changes of these four parameters did not happen at the same inter-distance. The adhesion ratio of a cell on Au stripes over cell spreading area did not change monotonically as a function of inter-stripe distance. The combination of the 5 statistical parameters represented well the cell orientation behaviors semi-quantitatively.     

A functional barrier to movement of lipids in polarized neurons.

In polarized neurons, axons and dendrites perform different functions, which are reflected in their different molecular organization. Studies on the sorting of viral and endogenous glycoproteins in epithelial cells and hippocampal neurons suggest that there may be similarities in the mechanism of sorting in these two cell types. The mechanisms that maintain the distinct composition of the two plasma membrane domains in these two cell types must, however, be different. We have proposed the existence of a functional barrier at the axonal hillock/initial segment which prevents the intermixing of membrane constituents. Here we test this hypothesis by fusing liposomes containing fluorescent phospholipids into the plasma membrane of polarized hippocampal cells in culture. Fusion was induced by lowering the pH and mediated by influenza virus haemagglutinin expressed on the axonal surface of neurons infected with fowl plague virus. Labelling was found exclusively on axons after fusion. Although the fused lipids were mobile on the axonal membrane, no labelling was detected on the cell body and dendritic surfaces. These results suggest that there is a diffusion barrier at the axonal hillock/initial segment which maintains the compositional differences between the axonal and somatodendritic domains.     

Surface mechanics mediate pattern formation in the developing retina

Pattern formation of biological structures involves organizing different types of cells into a spatial configuration. In this study, we investigate the physical basis of biological patterning of the Drosophila retina in vivo. We demonstrate that E- and N-cadherins mediate apical adhesion between retina epithelial cells. Differential expression of N-cadherin within a sub-group of retinal cells (cone cells) causes them to form an overall shape that minimizes their surface contact with surrounding cells. The cells within this group, in both normal and experimentally manipulated conditions, pack together in the same way as soap bubbles do. The shaping of the cone cell group and packing of its components precisely imitate the physical tendency for surfaces to be minimized. Thus, simple patterned expression of N-cadherin results in a complex spatial pattern of cells owing to cellular surface mechanics.     

High resolution imaging using scanning ion conductance microscopy with improved distance feedback control

Microscopy is an essential technique for observation on living cells. There is currently great interest in applying scanning probe microscopy to image-living biological cells in their natural environment at the nanometer scale. Scanning ion conductance microscopy is a new form of scanning probe microscopy, which enables non-contact high-resolution imaging of living biological cells. Based on a scanned nanopipette in physiological buffer, the distance feedback control uses the ion current to control the distance between the pipette tip and the sample surface. However, this feedback control has difficulties over slopes on convoluted cell surfaces, which limits its resolution. In this study, we present an improved form of feedback control that removes the contribution of up to the third-order slope from the ion current signal, hence providing a more accurate signal for controlling the distance. We show that this allows faster and lower noise topographic high-resolution imaging.     

Evidence that the serological determinant of H-Y antigen is carbohydrate

The histocompatibility-Y (H-Y) antigen is a minor histocompatibility antigen which has been detected on cell surfaces from the heterogametic sexes of mammalian, bird, amphibian, teleost and invertebrate species. H-Y is thought to be a male-determining substance in mammals because of its almost perfect correlation with maleness among a variety of mammalian species. To characterize the molecular determinant responsible for H-Y specific serological activity, H-Y positive immunoabsorbent cells were first subjected to various treatments which alter protein or carbohydrate structure and then tested for their ability to absorb H-Y antisera. We present here evidence that the serological determinant of H-Y antigen is carbohydrate.     

Regenerative medicine of cornea by cell sheet engineering using temperature-responsive culture surfaces

Recently, regenerative medicine has been focused on as next-generation definitive therapies for several diseases or injuries for which there are currently no effective treatments. These therapies have been rapidly developed through the effective fusion be- tween different fields such as stem cell biology and biomaterials. So far, we have proposed ＂cell sheet engineering＂ through our core technology that simply applies alterations of the temperature which allows regulation of the attachment or detachment of living cells on the culture surfaces grafted with the temperature-responsive polymer ＂poly（N-isoproplyacrylamide）＂. This tech- nology enables us to construct bioengineered sheet-like tissues, termed ＂cell sheets＂, without the need of using biodegradable scaffolds and protease treatments that are traditionally used. Therefore, our carrier-free cell sheets not only are independent of the issues observed in conventional methods, but also showed further advantages in the reconstruction of the corneal epithelium or endothelium （e.g. improvement of optical transparency and cell-cell interactions to host stroma in reconstructed tissues）. Moreo- ver, our approach does not have issues such as immune rejection or limited number of donors, due to the use of cell sheets derived from autologous limbal （in patients with unilateral disease） or oral mucosal epithelial cells （in patients with bilateral disorders）. Indeed, we have successfully performed the clinical application for the reconstruction of ocular surfaces through the transplanta- tion of our carrier-free corneal epithelial cell sheets, as evidenced by improved visual acuity as well as long-term maintenance of postoperative health conditions on ocular surfaces in all patients. We have also proposed a novel approach for the reconstruction of the corneal endothelium using corneal endothelial cell sheets by demonstrating its successful application. Thus, our cell sheet engineering technique provides a breakthrough in the field of regenerative medicine applied to the cornea.     

Targeting of transmembrane and GPI-anchored forms of N-CAM to opposite domains of a polarized epithelial cell.

The calcium-independent neural cell adhesion molecule N-CAM is expressed transiently during development in many tissues, including epithelia. The three naturally occurring principal isoforms of N-CAM differ in the way in which they associate with the membrane and in their cytoplasmic domains. These isoforms are generated by developmentally regulated alternative splicing of a single gene: the large cytoplasmic domain (ld) form (relative molecular mass 180,000 (Mr 180K] is specific for post-mitotic neurons; the 120K small cytoplasmic domain (ssd) and 140K small surface domain (sd) forms also occur on other cell types. One function of the different isoforms could be to specify cellular localization; for example, glycosyl phosphatidyl inositol (GPI)-membrane anchoring acts as a targeting signal for expression on the apical surface of polarized epithelial cells. Neurons and epithelial cells may use similar mechanisms for polarizing their plasma membrane proteins. We have therefore investigated the targeting of GPI-anchored (ssd N-CAM, 120K) and transmembrane forms of N-CAM (sd N-CAM, 140K; ld N-CAM, 180K) by comparing the expression of each after transfection of the appropriate complementary DNAs into polarized epithelial cells. We find that isoforms with alternative modes of membrane association are targeted to different surfaces of polarized epithelial cells: ssd N-CAM is expressed on the apical surface, whereas sd and ld N-CAM are expressed on the basolateral surface. These results suggest that the different isoforms of N-CAM determine their own diverse cellular destinations. They also support the hypothesis that the GPI anchor acts as an apical targeting signal in epithelia.     

High resolution imaging using scanning ion conductance microscopy with improved distance feedback control

Microscopy is an essential technique for observation on living cells. There is currently great interest in apply scanning probe microscopy to image living biological cells in their natural environment at the nanometer scale. Scanning ion conductance microscopy is a new form of scanning probe microscopy, which enables non-contact high resolution imaging of living biological cells. Based on a scanned nanopipette in physiological buffer, the distance feedback control uses the ion current to control the distance between the pipette tip and the sample surface. However, this feedback control has difficulties over slopes on convoluted cell surfaces, which limits its resolution. In this study, we present an improved form of feedback control that removes the contribution of up to the third order slope from the ion current signal, hence providing a more accurate signal for controlling the distance. We show that this allows faster and lower noise topographic high resolution imaging.     

Tetherin inhibits retrovirus release and is antagonized by HIV-1 Vpu

Human cells possess an antiviral activity that inhibits the release of retrovirus particles, and other enveloped virus particles, and is antagonized by the HIV-1 accessory protein, Vpu. This antiviral activity can be constitutively expressed or induced by interferon-alpha, and it consists of protein-based tethers, which we term 'tetherins', that cause retention of fully formed virions on infected cell surfaces. Using deductive constraints and gene expression analyses, we identify CD317 (also called BST2 or HM1.24), a membrane protein of previously unknown function, as a tetherin. Specifically, CD317 expression correlated with, and induced, a requirement for Vpu during HIV-1 and murine leukaemia virus particle release. Furthermore, in cells where HIV-1 virion release requires Vpu expression, depletion of CD317 abolished this requirement. CD317 caused retention of virions on cell surfaces and, after endocytosis, in CD317-positive compartments. Vpu co-localized with CD317 and inhibited these effects. Inhibition of Vpu function and consequent mobilization of tetherin's antiviral activity is a potential therapeutic strategy in HIV/AIDS.     

Escherichia coli swim on the right-hand side

The motion of peritrichously flagellated bacteria close to surfaces is relevant to understanding the early stages of biofilm formation and of pathogenic infection. This motion differs from the random-walk trajectories of cells in free solution. Individual Escherichia coli cells swim in clockwise, circular trajectories near planar glass surfaces. On a semi-solid agar substrate, cells differentiate into an elongated, hyperflagellated phenotype and migrate cooperatively over the surface, a phenomenon called swarming. We have developed a technique for observing isolated E. coli swarmer cells moving on an agar substrate and confined in shallow, oxidized poly(dimethylsiloxane) (PDMS) microchannels. Here we show that cells in these microchannels preferentially 'drive on the right', swimming preferentially along the right wall of the microchannel (viewed from behind the moving cell, with the agar on the bottom). We propose that when cells are confined between two interfaces--one an agar gel and the second PDMS--they swim closer to the agar surface than to the PDMS surface (and for much longer periods of time), leading to the preferential movement on the right of the microchannel. Thus, the choice of materials guides the motion of cells in microchannels.     

L1 mono- and polyclonal antibodies modify cell migration in early postnatal mouse cerebellum

A major event of nervous system development is the migration of granule cell neurones, during the early postnatal development of the cerebellar cortex, from their germinating zone in the external granular layer to their final location in the internal granular layer. During migration, many granule cells are seen in direct cell-surface contact with processes of Bergmann glia, a subclass of astrocytes. In the neurological mutant mouse weaver, however, migration of granule cells is impaired, probably due to a deficit in cell-cell interactions. To gain insight into the cellular and molecular mechanisms involved in granule cell migration, we have used a modification of an in vitro assay system, previously described by Moonen et al., which displays migratory behaviour in small tissue explants during several days of suspension culture. The aim of this study was to investigate the process of granule cell migration by using antibodies directed against cell-surface components of developing neural cells. We report here that migration of 3H-thymidine-labelled granule cell neurones can be modified by Fab fragments of both mono- and polyclonal L1 antibodies, but not by Fab fragments of polyclonal antibodies prepared against mouse liver membranes, which also react with cerebellar cell surfaces.