Good Time for Cell Adhesion and Migration

Our understanding of the molecular and cellular mechanisms controlling cell adhesion and migration has never been so high. The era of “molecular interactions” requires an appropriate tool for exchanging views, presenting outstanding results and discussing new concepts in the field. This is why we decided to launch Cell Adhesion and Migration (CAM), a multi-disciplinary journal publishing original research articles and reviews covering the latest aspects of cellular and molecular mechanisms and their regulation both during physiological and pathological processes.     

神经细胞粘连分子和多聚唾液酸在神经发育和再生中的作用

神经系统的形成依赖于细胞间的互相粘连。本文综述了神经细胞粘连分子（ＮＣＡＭ）及其多聚唾液酸（ＰＳＡ）组份对神经发育和再生的作用。ＮＣＡＭ的基本功能是介导细胞粘连,ＰＳＡ则由于其特殊的分子结构而降低细胞间的粘连。研究表明,鸡胚的发育过程中,ＰＳＡ含量在三个关键时期表达的高低决定了运动神经元能否准确地识别和支配肌肉。成年大鼠周围神经损伤后,肌肉内ＮＣＡＭ含量的高低决定于该肌肉的神经支配状况。成年大鼠脑内,切断内嗅皮层与海马的神经联系,发现齿回外分子层ＰＳＡ含量显著增加,并至少可持续６０天。已有的研究资料提示在去神经靶区域ＰＳＡ的重新表达可能有利于移植神经元轴突的生长并与宿主重建突触联系。     

The Effect of Cortical Elasticity and Active Tension on Cell Adhesion Mechanics

We consider a cell as an elastic, contractile shell surrounding a liquid incompressible cytoplasm and with nonspecific adhesion. We perform numerical simulations of this model to study the mechanics of cell-cell separation. By variation of parameters, we are able to recover well-known limits of the Johnson-Kendall-Roberts theory, the Derjaguin-Muller-Toporov model, adhesive vesicles with surface tension (Brochard-Wyart and de Gennes derivation), and thin elastic shells. We further locate biological cells on this parameter space by comparison to existing experiments on S180 cells. Using this model, we show that mechanical parameters can be obtained that are consistent with both dual pipette aspiration and micropipette aspiration, a problem not successfully tackled so far. We estimate a cortex elastic modulus of E_c ≈ 15 kPa, an effective cortex thickness of t_c ≈ 0.3 μm, and an active tension of γ ≈ 0.4 nN/μm. With these parameters, a Johnson-Kendall-Roberts-like scaling of the separation force is recovered. Finally, the change of contact radius with applied force in a pull-off experiment was investigated. For small forces, a scaling similar to both the Brochard-Wyart and de Gennes derivation and the Derjaguin-Muller-Toporov model is found.     

Characterization of the Topography and Wettability of English Weed Leaves and Biomimetic Replicas

The topography and wettability of the underside of English weed （Oxalis pes-caprae） leaves and of their biomimetic replicas are investigated. Polyvinyl siloxane molds were cast from the leaves and then filled with an epoxy pre-polymer to produce replicas. The particular topographical structures of leaves and replicas were evaluated by optical microscopy and Scanning Electron Microscopy （SEM） analysis. The static wettability of leaves and replicas was assessed by contact angle measurements, while the dynamic wettability was characterized by estimating contact angle hysteresis and studying the dynamic behavior of impacting water droplets. A smooth glass slip and its replica were used as control surfaces. The replica moulding method used was able to transfer the characteristic pattern of irregular 100 μm - 200 μm × 60 μm convex papillae interspersed with stomata of the original leaf to the epoxy replicas. The static contact angle of 143°± 3° and the contact angle hysteresis of 2~ indicate that the underside of the English weed leaf is close to superhydrophobic. The lower contact angles （130° ± 4°） and higher hysteresis （31°） observed for the replica when compared with the original leaves were associated to an inaccurate replication of the chemistry and structures of the three-dimensional wax projections covering the plant surface. Also, trichomes in the original leaves could not be accurately reproduced due to their flexibility and fragility. Differences in wetting behavior were also evident from droplet impact experiments, with rebound regimes prevailing in the original leaves and regimes characterized by higher adhesion and larger dissipation predominating in the replicas. Nevertheless, the morphological features of the leaf transferred to the replica were sufficient to promote a clear hydrophobic behavior of the replica when compared with the smooth epoxy reference surface.     

Time Course of Cell Sheet Adhesion to Porcine Heart Tissue after Transplantation

Multilayered cell sheets have been produced from bone marrow-derived mesenchymal stem cells (MSCs) for investigating their adhesion properties onto native porcine heart tissue. Once MSCs reached confluence after a 7-day culture on a temperature-responsive culture dish, a MSCs monolayer spontaneously detached itself from the dish, when the culture temperature was reduced from 37 to 20°C. The basal extracellular matrix (ECM) proteins of the single cell sheet are preserved, because this technique requires no proteolytic enzymes for harvesting cell sheet, which become a basic building block for assembling a multilayer cell sheet. The thickness of multilayered cell sheets made from three MSC sheets was found to be approximately 60 μm. For investigating the adhesion properties of the basal and apical sides, the multilayered cell sheets were transplanted onto the surface of the heart’s left ventricle. Multilayered cell sheets were histological investigated at 15, 30, 45 and 60 minutes after transplantation by hematoxylin eosin (HE) and azan dyes to determine required time for the adhesion of the multilayered sheets following cell-sheet transplantation. The results showed that only the basal side of multilayered cell sheets significantly enhanced the sheets adhesion onto the surface of heart 30 minutes after transplantation. This study concluded that (1) cell sheets had to be transplanted with its basal side onto the surface of heart tissue and (2) at least 30 minutes were necessary for obtaining the histological adhesion of the sheets to the heart tissue. This study provided clinical evidence and parameters for the successful application of MSC sheets to the myocardium and allowed cell sheet technology to be adapted clinical cell-therapy for myocardial diseases.     

Influence of the Surface Topography of Stainless Steel on Bacterial Adhesion

Bacterial adhesion on stainless steel may cause problems such as microbially induced corrosion or represent a chronic source of microbial contamination. The investigation focussed on how the extent and patterns of four bacterial species comprising three different phyla and a broad variety of physicochemical characteristics was influenced by the surface topography of AISI 304 stainless steel. Five types of surface finish corresponding to roughness values R a between 0.03 and 0.89 w m were produced. Adhesion of all four bacteria was minimal at R a =0.16 w m, whereas smoother and rougher surfaces gave rise to more adhesion. This surface exhibited parallel scratches of 0.7 w m, in which a high proportion of bacteria of three of the strains aligned. Reduced overall adhesion was attributed to unfavorable interactions between this surface and bacteria oriented other than parallel to the scratches. Interaction energy calculations and considerations of micro-geometry confirmed this mechanism. Rougher surfaces exhibiting wider scratches allowed a higher fraction of bacteria to adhere in other orientations, whereas the orientation of cells adhered to the smoothest surface was completely random.     

Chemistry of Abscisic Acid, Abscisic Acid Catabolites and Analogs

Recently there have been breakthroughs on a number of fronts in abscisic acid (ABA) biology research that have advanced the field significantly, including discovery of genes involved in ABA metabolism, along with progress in understanding of ABA signaling (Finkelstein and others 2002; Kushiro and others 2004; Lim and others 2005; Saito and others 2004). At the same time, the chemistry of ABA has advanced. New analytical methods have been developed for profiling ABA and catabolites (Ross and others 2004; Zaharia and others 2005). Novel bioactive catabolites have been discovered from feeding studies with deuterated ABA and catabolites (Zaharia and others 2004; Zhou and others 2004). This review covers recent advances and prospects in natural products chemistry, analysis of ABA catabolism, and applications of ABA analogs for biochemical studies and horticultural uses.     

Endothelial Cell Adhesion Molecule Expression and Lymphocyte Adhesion to Endothelial Cells: Effect of Nitric Oxide

Interactions between circulating leukocytes and vascular endothelial cells are of fundamental importance in controlling normal recirculation and migration of cells into sites of inflammation. Nitric oxide (NO), which is synthesized by vascular endothelial cells, has been reported to decrease the binding of platelets, monocytes, macrophages, and neutrophils to endothelial cells. Using NO donors and inhibitors of the enzyme NO synthase, we found no evidence that physiologically relevant levels of NO alter adhesion of purified lymphocytes to an endothelial cell line derived from human umbilical vein endothelial cells (SGHEC-7). In addition, NO donors did not alter the cell surface expression of VCAM-1, ICAM-1, or E-selectin on SGHEC-7 cells.     

整合素β2促进人乳腺癌细胞MCF-7的迁移,侵袭与粘附

本研究主要目标为探讨整合素β2 (ITGB2)的高表达对人乳腺癌细胞MCF-7迁移,侵袭与粘附能力的影响。本研究首先构建了ITGB2过表达质粒,实验设阴性对照组(pcDNA-3.1+)与ITGB2基因过表达组(pcDNA-3.1+/ITGB2)。ITGB2过表达质粒转染MCF-7细胞后,采用逆转录PCR与Western blotting方法分别检测ITGB2 mRNA转录水平与蛋白翻译水平;流式细胞术检测细胞周期的改变;划痕实验检测细胞横向迁移能力;Transwell小室实验检测细胞纵向迁移能力及侵袭能力;人脐静脉血管内皮细胞(HUVEC)粘附实验检测癌症细胞与血管内皮细胞之间的粘附能力,Western blotting实验检测侵袭相关指标MMP9,整合素经典通路中FAK蛋白磷酸化水平的改变。研究结果表明:转染ITGB2过表达质粒后,MCF-7细胞中ITGB2的m RNA水平(p<0.01)与蛋白水平(p<0.05)均显著增高;流式细胞术实验中,实验组S期的细胞所占比例与对照组无明显差异;划痕实验与Transwell小室实验中,实验组的迁移侵袭能力显著性增强;人脐静脉血管内皮细胞粘附实验中,实验组乳腺癌细胞与血管内皮细胞的粘附能力强于对照组(p<0.05);且Western blotting结果显示MMP9和p-FAK蛋白水平明显上升。由以上结果可得出结论,过表达ITGB2后会增强人乳腺癌细胞MCF-7的迁移、侵袭与粘附能力,而对其增殖能力无明显影响。     

Inhibition of Rac and ROCK Signalling Influence Osteoblast Adhesion,Differentiation and Mineralization on Titanium Topographies

Reducing the time required for initial integration of bone-contacting implants with host tissues would be of great clinical significance. Changes in osteoblast adhesion formation and reorganization of the F-actin cytoskeleton in response to altered topography are known to be upstream of osteoblast differentiation, and these processes are regulated by the Rho GTPases. Rac and RhoA (through Rho Kinase (ROCK)). Using pharmacological inhibitors, we tested how inhibition of Rac and ROCK influenced osteoblast adhesion, differentiation and mineralization on PT (Pre-treated) and SLA (sandblasted large grit, acid etched) topographies. Inhibition of ROCK, but not Rac, significantly reduced adhesion number and size on PT, with adhesion size consistent with focal complexes. After 1 day, ROCK, but not Rac inhibition increased osteocalcin mRNA levels on SLA and PT, with levels further increasing at 7 days post seeding. ROCK inhibition also significantly increased bone sialoprotein expression at 7 days, but not BMP-2 levels. Rac inhibition significantly reduced BMP-2 mRNA levels. ROCK inhibition increased nuclear translocation of Runx2 independent of surface roughness. Mineralization of osteoblast cultures was greater on SLA than on PT, but was increased by ROCK inhibition and attenuated by Rac inhibition on both topographies. In conclusion, inhibition of ROCK signalling significantly increases osteoblast differentiation and biomineralization in a topographic dependent manner, and its pharmacological inhibition could represent a new therapeutic to speed bone formation around implanted metals and in regenerative medicine applications.     

Surface Wettability and Chemistry of Ozone Perfusion Processed Porous Collagen Scaffold

Crosslinking treatment of collagen has often been used to improve the biological stability and mechanical properties of 3D porous collagen scaffolds. However, accompanying these improvements, the collagen fibril surface becomes hydrophobic nature resulting in a reduced surface wettability. The wetting of the collagen fibril by culture medium is reduced and it is difficult for the medium to diffuse into the 3D structure of a porous collagen scaffold. This paper reports a “perfusion processing” strategy using ozone to improve the surface wettability of chemical crosslinked collagen scaffolds. Surface wettability, surface composition and biological stability were analyzed to evaluate the effectiveness of this surface processing strategy. It was observed that ozone perfusion processing improved surface wettability for both exterior and interior surfaces of the porous 3D collagen scaffold. The improvement in wettability is attributed to the incorporation of oxygen-containing functional groups onto the surface of the collagen fibrils, as confirmed by X-ray Photoelectron Spectroscopy (XPS) analysis. This leads to a significant improvement in water taking capability without compromising the bulk biological stability and mechanical properties, and confirms that ozone perfusion processing is an effective tool to modify the wettability both for interior and exterior surfaces throughout the scaffold.     

Effect of Poxvirus Infection on Host Cell Deoxyribonucleic Acid Synthesis

Deoxyribonucleic acid (DNA) synthesis was studied in poxvirus-infected cells by measuring (14)C-thymidine incorporation into viral and host cell DNA. A complete separation of the two species of DNA was achieved by combining the previously used "Dounce method" with a separation method based on different reannealing properties of viral and vertebrate DNA. Shortly after infection of HeLa cells with poxviruses, a burst of viral DNA synthesis occurred in the cytoplasm, but a rapid inhibition of host-cell DNA synthesis in the nucleus was observed. This inhibition of cellular DNA synthesis was also found if an accumulation of viral DNA was prevented. At high multiplicites, ultraviolet-irradiated virus inhibited host-cell DNA synthesis to the same extent as fully infectious poxvirus. Under the same conditions, heating at 60 C for 15 min caused a decrease in the ability of cowpox virus to inhibit host-cell DNA synthesis, but did not produce the same effect on vaccinia virus strain WR.     

不同诱导子对枇杷悬浮细胞生长及熊果酸、齐墩果酸含量的影响

在已建立的稳定枇杷细胞悬浮培养系的试验中,研究加入诱导子水杨酸（SA）、茉莉酸甲酯（MJ）、水解酪蛋白（CH）和酵母提取物（YE）对枇杷悬浮细胞生长及次生代谢产物熊果酸（UA）、齐墩果酸（OA）含量的影响。结果表明,从细胞生长、UA及OA含量三方面综合考虑,以MJ为100.0 mg.L^-1时培养最佳,此时细胞生长、UA及OA含量分别为对照的1.081、2.540及2.590倍。     

Enhanced Cell Adhesion and Alignment on Micro-Wavy Patterned Surfaces

Various micropatterns have been fabricated and used to regulate cell adhesion, morphology and function. Micropatterns created by standard photolithography process are usually rectangular channels with sharp corners (microgrooves) which provide limited control over cells and are not favorable for cell-cell interaction and communication. This paper proposes a new micropattern with smooth wavy surfaces (micro-waves) to control the position and orientation of cells. To characterize cell growth and responses on the micro-patterned substrates, bovine aortic endothelial cells were seeded onto surfaces with micro-grooves and micro-waves for 24 h. As a result, the cells on the micro-wavy pattern appeared to have a lower death rate and better alignment compared to those on the micro-grooved pattern. In addition, flow-induced shear stress was applied to examine the adhesion strength of cells on the micro-wavy pattern. Results showed that cells adhered to the wavy surface displayed both improved alignment and adhesion strength compared to those on the flat surface. The combination of increased alignment, lower death rate and enhanced adhesion strength of cells on the micro-wavy patterns will offer advantages in potential applications for cell phenotype, proliferation and tissue engineering.     

牛蛙皮胶原蛋白酸提取条件优化及其对细胞生长和粘附性的影响

应用羟脯氨酸法测定了牛蛙皮胶原蛋白含量,通过正交实验对牛蛙皮胶原蛋白酸法提取条件进行了优化,并结合MTT法测定了胶原蛋白对细胞生长和粘附性的影响。结果显示,牛蛙皮胶原蛋白的含量约为45．1％;温度条件对提取率影响最大,其次依次为酸种、时间和浓度,最优组合为乙酸、1．5mol／L、37℃、24h;在低浓度时,牛蛙皮胶原蛋白对正常人类肝细胞的生长和粘附性无显著影响,当浓度升高到一定值时,对细胞生长和粘附性有显著促进作用。     

Effect of Cell Appendages on the Adhesion Properties of a Highly Adhesive Bacterium,Acinetobacter sp. Tol 5

A toluene-degrading bacterium, Acinetobacter sp. Tol 5, shows noteworthy adhesiveness mediated by two types of cell appendages. In this study, we obtained a less-adhesive mutant, T1, which lost both types of appendages, and investigated how the cell appendages affect the adhesion properties of this useful bacterium for environmental technology. Wild-type cells attained irreversible adhesion to polyurethane carriers within 30 s, while adhesion of T1 cells was still reversible at that time. While T1 showed decreased adhesion with decreasing ionic strength and did not adhere at all at 0.015 mM, adhesion of the wild type was fully independent of ionic strength. Acinetobacter sp. Tol 5 was also found to be not motile. Our results suggest that through the long distant interaction mediated by the appendages between the cells and surfaces, Tol 5 cells can attain irreversible adhesion very quickly without approaching the vicinity of the substratum.     

G-helix of Maspin Mediates Effects on Cell Migration and Adhesion

Maspin is a member of the serine protease inhibitor (serpin) superfamily that lacks protease inhibitory ability, although displaying tumor metastasis-suppressing activity resulting from its influence on cell migration, invasion, proliferation, apoptosis, and adhesion. The molecular mechanisms of these actions of maspin are as yet undefined. Here, we sought to identify critical functional motifs by the expression of maspin with point mutations at sites potentially involved in protein-protein interactions: the G α-helix (G-helix), an internal salt bridge or the P1 position of the reactive center loop. Our findings indicate that only mutations in the G-helix attenuated inhibition of cell migration by maspin and that this structural element is also involved in the effect of maspin on cell adhesion. The action of maspin on cell migration could be mimicked by a 15-mer G-helix peptide, indicating that the G-helix is both essential and sufficient for this effect. In addition, we provide evidence that the effects of the G-helix of maspin are dependent on β1 integrins. These data reveal that the major extracellular functions associated with the tumor suppressive action of maspin likely involve interactions in which the G-helix plays a key role.     

Fine Structural Analysis of Animal Cell Surfaces: Membranes and Cell Surface Topography

Current research on the mechanism of transmembrane regulationof topographic modulation at the cell surface is described forthe sea urchin egg and Sarcoma 180 ascites tumor cells of SwissWebster white mice. The transmembrane system is characterizedin terms of three components: glycocalyx, membrane, and cytofibrillarstructures. The importance of membrane molecular architectureper se relative to the other surface components is assessedin terms of freeze fracture analysis of both cell types as wellas concanavalin A (ConA)-mediated long term agglutination, cytochalasinB effects, and other drug-induced changes at Sarcoma 180 cellsurfaces. A quantitative and qualitative assessment of intramembranousparticle (IMP) sizes and density distributions reveals intrinsicstructural changes of the fusing membranes at cortical reactionduring sea urchin egg fertilization and also with the post-fertilizedaccumulation of surface microvilli. Comparable changes in IMPare noted for microvillus retraction and membrane smoothingin Sarcoma 180 cells under a variety of experimental conditions.On the other hand, chemical perturbation of S-180 cell surfacesreveals a rather non-ubiquitous, though identifiable, involvementof microfilaments and no microtubule involvement in these topographicchanges. These observations suggest that the plasma membraneis a dynamic structure poised between "restrictive" and "lessrestrictive" states of fluidity or deformability and, hence,is a determinant component in topographic change.