The development of epidermal feet in preparation for metamorphosis in an insect

Insect epidermal cell surfaces can be seen by scanning electron microscopy after removal of the basal lamina. This let us study surface changes in the 5th larval stage of Calpodes ethlius (Lepidoptera, Hesperiidae) in preparation for metamorphosis at the end of the stadium, in particular changes in the basal cell processes or feet, intercellular lymph spaces, filopodia and hemidesmosomes. The feet develop in three phases, initiation, elongation and contraction. Initial growth begins immediately after ecdysis and continues until commitment to pupation 66 hr later. During this phase the feet are randomly oriented. Elongation and orientation begin after commitment to pupation. Orientation is probably achieved by selective survival and growth of those feet that are axially oriented rather than by reorientation. As the larva shortens to the pupal form late in the stadium, contraction of the feet occurs and the cells become columnar. The feet finally disappear as the cells rearrange themselves into new positions in the pupal epidermis. The lateral margins of the feet are united by adhesions even when their interdigitations are most complex. The adhesions separate an intercellular lymph space from the haemolymph. The lymph space remains small through most of the stadium, but enlarges with the loss of lateral junctions as the feet contract and eventually extends along most of the length of the columnar cells. Filopodia then form and span the gaps between the cells as though they have been induced by the separation and loss of lateral cell to cell contact. Scanning electron microscopy also shows that hemidesmosomes reflect the axial alignment of the cells even before the orientation of the feet. The hemidesmosome plaques are linear structures having a constant width of 0.15 - 0.2 mum and variable length. They arise in short sections and lengthen by the linear addition of more sections with the same width. Late in the stadium they lose their axial alignment and may become branched.     

Formation of the Primitive Streak and Mesoderm Cells in Mouse Embryos—Detailed Scanning Electron Microscopical Study

We describe morphological events of the mammalian gastrulation in pre- to middle-primitive-streakstage mouse embryos by using scanning electron microscopy. The first sign of the ingression of the mesodermal cells was disruption of the epithelial structure of ectoderm and the underlying basal lamina, thus forming a semicircular area of the presumptive primitive streak. Then, cells at periphery of the semicircular region spread on the basal lamina by extending many filopodia to it. The majority of the migrating cells formed a loosely arranged cell sheet. We found solitary cells and isolated small groups of cells migrating away from the periphery of the cell sheet. These cells were well spread on the basal lamina, and had large cell processes and many filopodia in the direction of cell migration. Filopodia of these cells were attached to the basal lamina or a meshwork of the extracellular fibrils. These observations suggest that the extracellular matrix serves as the substratum for cell adhesion and migration, and plays an important role in the mammalian gastrulation.     

Fine structure of the down feather during its early development

The down feather of the chick embryo has been examined by electron microscopy during three distinct stages of its early development; the presumptive stage, represented by dorsal skin of an area from which the feather organ will arise; the thickening stage, during which areas of the basal epidermis form spurs projecting into the mesenchyme, and the latter condenses under a thickened area of the epidermis; the elevation stage, at which time the basal epidermis flattens, the entire epidermis increases in thickness, and the underlying mesenchyme becomes more compact. As development proceeds the rough endoplasmic reticulum of the epidermal cells dilates, but during the elevation stage begins to flatten, and Golgi is observed with increasing frequency. The mitochondria do not appear to differ except for those in the periderm during the presumptive stage, in which case they reveal a vacant matrix and irregular cristae. Evidence is presented for actual contact between basal epidermal spurs and filopodia of cells within the mesenchyme, some of which contain numerous vesicles. The basal epidermal spurs are also seen in intimate association with collagen and anchor filaments and a network of reticulin. Evidence is also presented for the presence of neuronal elements within the mesenchyme during the thickening stage. Cross sections of cell processes within the condensations of the mesenchyme resemble unmyelinated nerve fibers, and cross sections of filopodia similar to arborizing axons abound at and within the basal lamina of both the thickening and elevation stages. Further support for the presence of nerve fibers within the mesenchyme comes from positive staining results with Bodian's and Ungewitter's methods. This comparative study of three stages of early development of the feather organ serves as a basis for more detailed investigations of each stage.     

An ultrastructural analysis of cell and matrix differentiation during early limb development in Xenopus laevis

Early development of the hind limb of Xenopus (stages 44–48) has been analyzed at the level of ultrastructure with emphasis on differentiation of extracellular matrix components and intercellular contacts. By stages 44–45, mesenchyme is separated from prospective bud epithelium by numerous adepidermal granules in a subepithelial compartment (the lamina lucida), a continuous basal lamina and several layers of collagen (the basement lamella). Tricomplex stabilization of amphoteric phospholipid demonstrates that each adepidermal granule consists of several membranelike layers (electron-lucent band 25–30 Å; electron-dense band 20–40 Å), which are usually parallel to the basal surface of adjacent epithelial cells. Collagen fibrils are interconnected by filaments (35 Å in diameter) which stain with ruthenium red. Epithelial cells possess junctional complexes at their superficial borders, numerous desmosomes at apposing cell membranes and hemidesmosomes at their basal surface. Mesenchymal cells predominantly exhibit close contacts (100–150 Å separation) with few focal tight junctions at various areas of their surface. By stages 47–48, adepidermal granules are absent beneath bud epithelium and layers of collagen in the basement lamella lose filamentous cross-linking elements. Filopodia of mesenchymal cells penetrate the disorganized matrix and abut the basal lamina. Hemidesmosomes disappear at the basal surface of the epidermis and mesenchymal cells immediately subjacent to epithelium exhibit focal tight junctions and gap junctions at their lateral borders. These structural changes may be instrumental in the epitheliomesenchymal interactions of early limb development. Degradation of oriented collagenous lamellae permits direct association of mesenchymal cell surfaces (filopodia) with surface-associated products of epithelial cells (organized into the basal lamina). Development of structural pathways for intercellular ion and metabolite transport in mesenchyme may coordinate events specific to limb morphogenesis.     

Filopodia and adhesion in cancer cell motility

Slender bundled actin containing plasma membrane protrusions, called filopodia, are important for many essential cellular processes like cell adhesion, migration, angiogenesis and the formation of cell-cell contacts. In migrating cells, filopodia are the pioneers at the leading edge which probe the environment for cues. Integrins are cell surface adhesion receptors critically implicated in cell migration and they are transported actively to filopodia tips by an unconventional myosin, myosin-X. Integrin mediated adhesion stabilizes filopodia and promotes cell migration even though integrins are not essential for filopodia initiation. Myosin-X binds also PIP3 and this regulates its activation and localization to filopodia. Filopodia stimulate cell migration in many cell types and increased filopodia density has been described in cancer. Furthermore, several proteins implicated in filopodia formation, like fascin, are also relevant for cancer progression. To investigate this further, we performed a meta-analysis of the expression profiles of 10 filopodia-linked genes in human breast cancer. These data implicated that several different filopodia inducing genes may contribute in a collective manner to cancer progression and the high metastasis rates associated with basal-type breast carcinomas.     

Pioneer neurones use basal lamina as a substratum for outgrowth in the embryonic grasshopper limb

During axonogenesis, contacts made by the growth cone with its substratum are important in guiding the direction of neurone outgrowth. This study examines the contacts made by the growth cones of pioneer neurones in the embryonic grasshopper limb. Individual pioneer neurones at different stages of development were injected with horseradish peroxidase and the contacts made by the filopodia at the tip of their growth cones were examined by electron microscopy. Filopodia made few contacts with mesodermal cells, some contacts with ectodermal cells and very frequent contacts with basal lamina underlying the ectoderm. Components of the basal lamina may therefore play a role in guiding pioneer axon outgrowth.     

Filopodia and adhesion in cancer cell motility

Slender bundled actin containing plasma membrane protrusions, called filopodia, are important for many essential cellular processes like cell adhesion, migration, angiogenesis and the formation of cell-cell contacts. In migrating cells, filopodia are the pioneers at the leading edge which probe the environment for cues. Integrins are cell surface adhesion receptors critically implicated in cell migration and they are transported actively to filopodia tips by an unconventional myosin, myosin-X. Integrin mediated adhesion stabilizes filopodia and promotes cell migration even though integrins are not essential for filopodia initiation. Myosin-X binds also PtdIns(3,4,5)P₃ and this regulates its activation and localization to filopodia. Filopodia stimulate cell migration in many cell types and increased filopodia density has been described in cancer. Furthermore, several proteins implicated in filopodia formation, like fascin, are also relevant for cancer progression. To investigate this further, we performed a meta-analysis of the expression profiles of 10 filopodia-linked genes in human breast cancer. These data implicated that several different filopodia-inducing genes may contribute in a collective manner to cancer progression and the high metastasis rates associated with basal-type breast carcinomas.Key words: filopodia, integrins, migration, cancer     

FORMATION AND ORIGIN OF BASAL LAMINA AND ANCHORING FIBRILS IN ADULT HUMAN SKIN

The purpose of this investigation was to study the formation and origin of basal lamina and anchoring fibrils in adult human skin. Epidermis and dermis were separated by "cold trypsinization." Viable epidermis and viable, inverted dermis were recombined and grafted to the chorioallantoic membrane of embryonated chicken eggs for varying periods up to 10 days. Basal lamina and anchoring fibrils were absent from the freshly trypsinized epidermis before grafting although hemidesmosomes and tonofilaments of the basal cells remained intact. Basal lamina and anchoring fibrils were absent from freshly cut, inverted surface of the dermis. Beginning 3 days after grafting, basal lamina was noted to form immediately subjacent to hemidesmosomes of epidermal basal cells at the epidermal-dermal interface. From the fifth to the seventh day after grafting, basal lamina became progressively more dense and extended to become continuous in many areas at the epidermal-dermal interface. Anchoring fibrils appeared first in grafts consisting of epidermis and viable dermis at five day cultivation and became progressively more numerous thereafter. In order to determine the epidermal versus dermal origin of basal lamina and anchoring fibrils, dermis was rendered nonviable by repeated freezing and thawing 10 times followed by recombination with viable epidermis. Formation of basal lamina occurred as readily in these recombinants of epidermis with freeze-thawed, nonviable dermis as with viable dermis, indicating that dermal viability was not essential for synthesis of basal lamina. This observation supports the concept of epidermal origin for basal lamina. Anchoring fibrils did not form in recombinants containing freeze-thawed dermis, indicating that dermal viability was required for anchoring fibrils formation. This observation supports the concept of dermal origin of anchoring fibrils.     

A preformed basal lamina alters the metabolism and distribution of hyaluronan in epidermal keratinocyte "organotypic" cultures grown on collagen matrices

A rat epidermal keratinocyte (REK) line which exhibits histodifferentiation nearly identical to the native epidermis when cultured at an air-liquid interface was used to study the metabolism of hyaluronan, the major intercellular macromolecule present in basal and spinous cell layers. Two different support matrices were used: reconstituted collagen fibrils with and without a covering basal lamina previously deposited by canine kidney cells. REKs formed a stratified squamous, keratinized epithelium on both support matrices. Hyaluronan and its receptor, CD44, colocalized in the basal and spinous layers similar to their distribution in the native epidermis. Most (approximately 75%) of the hyaluronan was retained in the epithelium when a basal lamina was present while most (approximately 80%) diffused out of the epithelium in its absence. While REKs on the two matrices synthesized hyaluronan at essentially the same rate, catabolism of this macromolecule was much higher in the epithelium on the basal lamina (half-life approximately 1 day, similar to its half-life in native human epidermis). The formation of a true epidermal compartment in culture bounded by the cornified layer on the surface and the basal lamina subjacent to the basal cells provides a good model within which to study epidermal metabolism.     

Filopodial initiation and a novel filament-organizing center, the focal ring

This study examines filopodial initiation and implicates a putative actin filament organizer, the focal ring. Filopodia were optically recorded as they emerged from veils, the active lamellar extensions of growth cones. Motile histories revealed three events that consistently preceded filopodial emergence: an influx of cytoplasm into adjacent filopodia, a focal increase in phase density at veil margins, and protrusion of nubs that transform into filopodia. The cytoplasmic influx probably supplies materials needed for initiation. In correlated time lapse-immunocytochemistry, these focal phase densities corresponded to adhesions. These adhesions persisted at filopodial bases, regardless of subsequent movements. In correlated time lapse-electron microscopy, these adhesion sites contained a focal ring (an oblate, donut-shaped structure approximately 120 nm in diameter) with radiating actin filaments. Filament geometry may explain filopodial emergence at 30 degree angles relative to adjacent filopodia. A model is proposed in which focal rings play a vital role in initiating and stabilizing filopodia: 1) they anchor actin filaments at adhesions, thereby facilitating tension development and filopodial emergence; 2) "axial" filaments connect focal rings to nub tips, thereby organizing filament bundling and ensuring the bundle intersects an adhesion; and 3) "lateral" filaments interconnect focal rings and filament bundles, thereby helping stabilize lamellar margins and filopodia.     

Ultrastructural studies of regenerating spines of the sea urchin Strongylocentrotus purpuratus I. Cell types without spherules

The fine structure of regenerating tips of spines of the sea urchin Strongylocentrotus purpuratus was investigated. Each conical tip consisted of an inner dermis, which deposits and contains the calcite skeleton, and an external layer of epidermis. Although cell types termed spherulecytes containing large, intracellular membrane bound spherules were also present in spine tissues, only epidermal and dermal cell types lacking such spherules are described in this paper. The epidermis was composed largely of free cells representing several functional types. Over the apical portion of the tip these cells occurred in groups, while proximally they were distributed within longitudinal grooves present along the periphery of the spine from the base to the tip. The terminal portions of apical processes extending from some of the epidermal cells formed a thin, contiguous outer layer consisting of small individual islands of cytoplasm bearing microvilli. Adjacent islands were connected around the periphery by a junctional complex extending roughly 200 Å in depth in which the opposing plasma membranes were separated by a narrow gap about 145 Å in width bridged by amorphous material. Other epidermal cells were closely associated with the basal lamina, which was 900 Å in thickness and delineated the dermoepidermal junction; some of these cells appeared to synthesize the lamina, while others may be sensory nerve cells. The dermis at the spine tip also consisted of several functional types of free cells; the most interesting of these was the calcoblast, which deposits the skeleton. Calcoblasts extended a thin, cytoplasmic skeletal sheath which surrounded the tips and adjacent proximal portions of each of the longitudinally oriented microspines comprising the regenerating skeleton, and distally, formed a conical extracellular channel ahead of the mineralizing tip. The intimate relationship between calcoblasts and the growing mineral surface strongly suggests that these cells directly control both the kinetics of mineral deposition and morphogenesis of the skeleton. Other cell types in the dermis were precalcoblasts and phagocytes. Precalcoblasts may function as fibroblasts and are possible precursors of calcoblasts. Closely associated with the basal lamina at the dermoepidermal junction were extracellular unbanded anchoring fibrils 150 Å to 200 Å in diameter. Scattered proximally among dermal cells were other extracellular fibrils, presumably collagenous, about 300 Å in diameter with a banding periodicity of 210 Å.     

Stimulation of human epidermal differentiation by delta-notch signalling at the boundaries of stem-cell clusters

BACKGROUND: Human epidermis is renewed throughout life from stem cells in the basal layer of the epidermis. Signals from the surrounding keratinocytes influence the differentiation of the stem cells, but the nature of the signals is unknown. In many developing tissues, signalling mediated by the transmembrane protein Delta1 and its receptor Notch1 inhibits differentiation. Here, we investigated the role of Delta-Notch signalling in postnatal human epidermis. RESULTS: Notch1 expression was found in all living epidermal layers, but Delta1 expression was confined to the basal layer of the epidermis, with highest expression in those regions where stem cells reside. By overexpressing Delta1 or Delta(T), a truncated form of Delta1, in primary human keratinocytes and reconstituting epidermal sheets containing mixtures of Delta-overexpressing cells and wild-type cells, we found that cells expressing high levels of Delta1 or Delta(T) failed to respond to Delta signals from their neighbours. In contrast, wild-type keratinocytes that were in contact with neighbouring cells expressing Delta1 were stimulated to leave the stem-cell compartment and initiate terminal differentiation after a few rounds of division. Delta1 promoted keratinocyte cohesiveness, whereas Delta(T) did not. CONCLUSIONS: We propose that high Delta1 expression by epidermal stem cells has three effects: a protective effect on stem cells by blocking Notch signalling; enhanced cohesiveness of stem-cell clusters, which may discourage intermingling with neighbouring cells; and signalling to cells at the edges of the clusters to differentiate. Notch signalling in epidermal stem cells thus differs from other progenitor cell populations in promoting, rather than suppressing, differentiation.     

Structures in focus--filopodia

Filopodia are thin cell surface extensions filled with tight parallel bundles of actin filaments. They are highly dynamic structures which rapidly extend and retract as well as sweep up and down and from side to side, and can be found at the leading edge of many types of motile cells such as fibroblasts and keratinocytes, as well as the growth cone tips of migrating axons. Cells appear to use filopodia to explore the extracellular matrix (ECM) and surfaces of other cells, identifying appropriate targets for adhesion or in the case of a migrating growth cone, for sensing guidance cues that enable the axon to navigate to it's appropriate target. As well as this sensory role, filopodia have also recently been shown to play an important mechanical role in epithelial adhesion, and are likely to be key players in developmental processes that require migrating epithelial sheets to zipper and fuse to one another. Their dynamic properties as well as their tendency to be damaged or lost after fixation mean they are best analysed using live imaging techniques. As this field improves, the number of tissues in which filopodia are seen to be playing key roles is fast increasing.     

The unc-5, unc-6, and unc-40 genes guide circumferential migrations of pioneer axons and mesodermal cells on the epidermis in C. elegans

Three known genes guide circumferential migrations of pioneer axons and mesodermal cells on the nematode body wall. unc-5 affects dorsal migrations, unc-40 primarily affects ventral migrations, and unc-6 affects migrations in both directions. Circumferential movements still occur, but are misdirected whereas longitudinal movements are normal in these mutants. Pioneer growth cones migrating directly on the epidermis are affected; growth cones migrating along established axon fascicles are normal. Thus these genes affect cell guidance and not cell motility per se. We propose that two opposite, adhesive gradients guide circumferential migrations on the epidermis. unc-5, unc-6, and unc-40 may encode these adhesion molecules or their cellular receptors. Neurons have access to the basal lamina and the basolateral surfaces of the epidermis, but mesodermal cells contact only the basal lamina. These genes probably identify molecular cues on the basal lamina that guide mesodermal migrations. The same basal lamina cues, or perhaps related molecules on the epidermal cell surfaces, guide pioneer neurons.     

三种国产阴地蕨科植物叶的表皮构造

运用光学显微镜和扫描电镜对三种国产阴地蕨科植物：劲直蕨萁(Botrypus strictus)、小阴地蕨(Botrychium lunaria)和薄叶阴地蕨(Sceptridium daucifolium)〖WTBZ〗叶的成熟表皮构造进行了详细观察和研究。它们具有共同的特征：气孔散生,气孔类型无规则型,气孔长轴方向多与叶脉延伸方向一致。但也存在明显区别,特别是劲直蕨萁与另外二个种的区别更为明显：前者表皮细胞垂周壁直,相邻气孔不接触,保卫细胞平周壁具细条纹。但后二者之间亦有一定区别：薄叶阴地蕨的气孔为下生式,不下陷,而小阴地蕨的气孔为两面气孔型,气孔下陷。本文是国内首次对阴地蕨科叶的表皮构造进行研究,其研究结果表明,表皮构造在阴地蕨科植物的鉴定上具有重要意义,并且在研究阴地蕨科的分类以及起源和演化上也有一定参考价值。     

Organization of actin networks in intact filopodia

Filopodia are finger-like extensions of the cell surface that are involved in sensing the environment, in attachment of particles for phagocytosis, in anchorage of cells on a substratum, and in the response to chemoattractants or other guidance cues. Filopodia present an excellent model for actin-driven membrane protrusion. They grow at their tips by the assembly of actin and are stabilized along their length by a core of bundled actin filaments. To visualize actin networks in their native membrane-anchored state, filopodia of Dictyostelium cells were subjected to cryo-electron tomography. At the site of actin polymerization, a peculiar structure, the "terminal cone," is built of short filaments fixed with their distal end to the filopod's tip and with their proximal end to the flank of the filopod. The backbone of the filopodia consists of actin filaments that are shorter than the entire filopod and aligned in parallel or obliquely to the filopod's axis. We hypothesize that growth of the highly dynamic filopodia of Dictyostelium is accompanied by repetitive nucleation of actin polymerization at the filopod tip, followed by the rearrangement of filaments within the shaft.     

Regulation of dendritic branching and filopodia formation in hippocampal neurons by specific acylated protein motifs

Although neuronal axons and dendrites with their associated filopodia and spines exhibit a profound cell polarity, the mechanism by which they develop is largely unknown. Here, we demonstrate that specific palmitoylated protein motifs, characterized by two adjacent cysteines and nearby basic residues, are sufficient to induce filopodial extensions in heterologous cells and to increase the number of filopodia and the branching of dendrites and axons in neurons. Such motifs are present at the N-terminus of GAP-43 and the C-terminus of paralemmin, two neuronal proteins implicated in cytoskeletal organization and filopodial outgrowth. Filopodia induction is blocked by mutations of the palmitoylated sites or by treatment with 2-bromopalmitate, an agent that inhibits protein palmitoylation. Moreover, overexpression of a constitutively active form of ARF6, a GTPase that regulates membrane cycling and dendritic branching reversed the effects of the acylated protein motifs. Filopodia induction by the specific palmitoylated motifs was also reduced upon overexpression of a dominant negative form of the GTPase cdc42. These results demonstrate that select dually lipidated protein motifs trigger changes in the development and growth of neuronal processes.     

Comparative structure of the labellum in Ophrys fusca and O. lutea (Orchidaceae)

The morphology and anatomy of the labellar epidermal cells and the way in which they are arranged are described in an attempt to locate and characterize the osmophore in Ophrys fusca and O. lutea. The micromorphology of the labellum of these two species is similar. Four types of epidermal cells are present on the adaxial surface of the labellum. Long unicellular trichomes with straight tips cover the basal region of the labellum, whereas short unicellular trichomes with polygonal flattened bases form the reflective median speculum. The apical region of the labellum possesses a villous indumentum of long acuminate trichomes with bent or sinuate tips. Large smooth-walled, dome-shaped papillae occur on the margins and on the distal region of the abaxial surface of the labellum. These remarkable papillae have high polarity; the protoplasm at the apex of each cell contains several small vacuoles, while a prominent nucleus surrounded by numerous hypertrophied amyloplasts occurs at the opposite end of the cell. Positive reactions to Vogel's staining test and to Sudan black B enabled us to conclude that the osmophores of both species are composed of these peculiar secretory epidermal cells and by two or three subsecretory layers of parenchyma cells.     

Building the actin cytoskeleton: filopodia contribute to the construction of contractile bundles in the lamella

Filopodia are rodlike extensions generally attributed with a guidance role in cell migration. We now show in fish fibroblasts that filopodia play a major role in generating contractile bundles in the lamella region behind the migrating front. Filopodia that developed adhesion to the substrate via paxillin containing focal complexes contributed their proximal part to stress fiber assembly, and filopodia that folded laterally contributed to the construction of contractile bundles parallel to the cell edge. Correlated light and electron microscopy of cells labeled for actin and fascin confirmed integration of filopodia bundles into the lamella network. Inhibition of myosin II did not subdue the waving and folding motions of filopodia or their entry into the lamella, but filopodia were not then integrated into contractile arrays. Comparable results were obtained with B16 melanoma cells. These and other findings support the idea that filaments generated in filopodia and lamellipodia for protrusion are recycled for seeding actomyosin arrays for use in retraction.     

Maturation of Filopodia Shaft Adhesions Is Upregulated by Local Cycles of Lamellipodia Advancements and Retractions

While cell-substrate adhesions that form between the protruding edge of a spreading cell and flat surfaces have been studied extensively, processes that regulate the maturation of filopodia adhesions are far less characterized. Since little is known about how the kinetics of formation or disassembly of filopodia adhesions is regulated upon integration into the lamellum, a kinetic analysis of the formation and disassembly of filopodia adhesions was conducted at the leading edge of β3-integrin-EGFP-expressing rat embryonic fibroblasts spreading on fibronectin-coated glass or on soft polyacrylamide gels. Filopodia β3-integrin adhesions matured only if the lamellipodium in their immediate vicinity showed cyclic protrusions and retractions. Filopodia β3-integrin shaft adhesions elongated rapidly when they were overrun by the advancing lamellipodium. Subsequently and once the lamellipodium stopped its advancement at the distal end of the filopodia β3-integrin adhesion, these β3-integrin shaft adhesions started to grow sidewise and colocalize with the newly assembled circumferential actin stress fibers. In contrast, the suppression of the cyclic protrusions and retractions of the lamellipodium by blocking myosin light chain kinase suppressed the growth of filopodia adhesion and resulted in the premature disassembly of filopodia adhesions. The same failure to stabilize those adhesions was found for the advancing lamellipodium that rapidly overran filopodia shaft adhesions without pausing as seen often during fast cell spreading. In turn, plating cells on soft polyacrylamide gels resulted in a reduction of lamellipodia activity, which was partially restored locally by the presence of filopodia adhesions. Thus filopodia adhesions could also mature and be integrated into the lamellum for fibroblasts on soft polyacrylamide substrates.