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PubMed1996

Extracellular matrix proteins are potent agonists of human smooth muscle cell migration.

PURPOSE: Extracellular matrix proteins can stimulate smooth muscle cell (SMC) migration by three distinct mechanisms: chemokinesis (nondirected migration in the presence of soluble protein), chemotaxis (directed migration toward soluble protein), and haptotaxis (directed migration toward insoluble, substrate-bound protein). This study investigates the effects of four prevalent extracellular matrix proteins (collagen types I and IV, fibronectin, and laminin), and platelet-derived growth factor (PDGF) on haptotaxis, chemotaxis, and chemokinesis of human SMCs. The role of large guanosine triphosphate-binding proteins (G-proteins) in the signaling mediating these effects is also evaluated. METHODS: Human saphenous vein SMCs were used in all migration studies. Chemokinesis, chemotaxis, and haptotaxis to each of the matrix proteins were measured and compared with PDGF through the use of a 48-well microchemotaxis chamber. The role of G-proteins in matrix-induced SMC migration was studied with the modulators of G-protein function, cholera and pertussis toxins. RESULTS: For all matrix proteins the relative strength of the various stimuli for migration was haptotaxis > chemotaxis > chemokinesis (p < 0.05). For all three stimuli collagen I and IV produced the most significant migration followed by fibronectin > PDGF-AB > laminin (p < 0.05). Pertussis toxin completely inhibited chemotaxis and partially inhibited haptotaxis by laminin but did not affect migration by other matrix proteins, whereas cholera toxin abolished migration in response to all four matrix proteins. CONCLUSION: Matrix proteins, with the exception of laminin, provide a more significant stimulus for SMC locomotion than does the prototypical agonist, PDGF-AB. Of the three mechanisms by which migration can be stimulated, haptotaxis elicits the most profound effect. The importance of G-proteins as second messengers for migration varies with each matrix protein and with the mechanism of stimulation.

Cell Movement↗
PubMed1989

Acellular vascular matrix: a natural endothelial cell substrate.

A preliminary assessment was made of the acellular vascular matrix graft as a substrate for endothelial cell seeding, with respect to surface pretreatment (none versus fibronectin and/or serum) and presence of exogenous growth factor. Arteries were harvested from greyhounds and exposed to a sequential detergent extraction process to produce the acellular vascular matrix. Human umbilical vein endothelial cells were grown in tissue culture, harvested in first passage, then seeded at 10(5) cells/cm2 on sections of acellular vascular matrix and on gel-coated polystyrene positive controls. After 18 hour incubation, endothelial cell-seeded acellular matrices were fixed and processed for histologic and planimetric analysis; control wells were fixed and endothelial cells were counted by planimetry. Pretreatment of the acellular vascular matrix was found to have no effect on the percentage of endothelial cell coverage of the matrix. There was significantly better endothelial cell coverage of the acellular matrix than on matched gel-treated polystyrene control wells. Withdrawal of growth factor resulted in a significant reduction in endothelial cell coverage for all acellular vascular matrix groups. Growth factor withdrawal also significantly reduced attachment of endothelial cells on gel-treated polystyrene. Cell surface area was significantly smaller when growth factor was withdrawn from all groups except from the acellular vascular matrix without pretreatment. We conclude that: (1) the acellular vascular matrix is conductive to endothelial cell adherence and spreading even without pretreatment; and (2) sudden withdrawal of exogenous growth factor may impair early coverage of substrates by endothelial cells due to an effect on their adherence or spreading.

Animals↗
PubMed2000

Monoclonal antibodies to mineralized matrix molecules of the avian eggshell.

The extracellular matrix of the mineralizing eggshell contains molecules hypothesized to be regulators of biomineralization. To study eggshell matrix molecules, a bank of monoclonal antibodies was generated that bound demineralized eggshell matrix or localized to oviduct epithelium. Immunofluorescence staining revealed several staining patterns for antibodies that recognized secretory cells: staining for a majority of columnar lining cells, staining for a minor sub-set of columnar lining cells, intensified staining within epithelial crypts, and staining of the entire tubular gland. Western blotting with the antibody Epi2 on eggshell matrix showed binding to molecules with the apparent molecular weight of eggshell matrix dermatan sulfate proteoglycan (eggshell DSPG). Immunoblots of cyanogen bromide-cleaved eggshell DSPG revealed broad band of reactivity that shifted to 25 kDa after chondroitinase digestion; indicating that the Epi2 binding site is located on a fragment which contains dermatan sulfate side chains. Immunogold labeling showed that Epi2 binds to secretory vesicles within the non-ciliated cells of the columnar epithelium, while the antibodies Tg1 and Tg2 bind to secretory vesicles of tubular gland cells. Immunogold labeling of demineralized shell matrix showed binding of Epi2, Tg1, and Tg2 to the matrix of the palisade layer, and showed little reactivity to other regions of the shell matrix. Quantification of the immunogold particles within the eggshell matrix revealed that antibodies Epi2 and Tg1 bind all calcified regions equally while antibody Tg2 has a greater affinity for the baseplate region of the calcium reserve assembly.

Animals↗
PubMedDate not supplied

Loss of matrix-dependent cytoskeletal tyrosine kinase signals may regulate intestinal epithelial differentiation during mucosal healing.

Intestinal epithelial restitution and the migratory phenotype appear regulated by the extracellular matrix. Since integrin-associated adhesion to matrix triggers tyrosine kinase activity, we hypothesized that matrix-specific tyrosine kinase signals might modulate the intestinal epithelial migratory phenotype, particularly via focal adhesion kinase. Caco-2 cells were seeded at two densities on collagen I, laminin, fibronectin, and tissue culture plastic. Four days later the first cells were confluent, whereas the second cells were not contact inhibited and expressed migratory lamellipodia. Cells were fractionated into membrane/cytoskeletal and cytosolic fractions. Cytoskeletal tyrosine kinase activity in static cells was matrix dependent and, unlike cytoscolic tyrosine kinase, correlated with adhesion, highest on collagen and lowest on plastic. Migrating cells exhibited matrix-dependent increases in cystosolic tyrosine kinase activity. Cytosolic changes in tyrosine kinase activity in motile cells exceeded membrane/cytoskeletal changes. However, matrix-dependent variations in increase in cytosolic tyrosine kinase activity correlated inversely with changes in cytoskeletal tyrosine kinase activity, suggesting cytoskeletal tyrosine kinase translocation to the cytosol during motility. Indeed cytoskeletal focal adhesion kinase activity decreased during migration on collagen. Tyrosine kinase inhibition by genistein both inhibited migration and stimulated expression of brush-border enzymes downregulated during motility. Although enterocyte-matrix interactions alter both cytosolic and cytoskeletal tyrosine kinase activity, matrix-dependent cytoskeletal events are likely to regulate adhesion and differentiation in static cells. Loss of matrix-dependent cytoskeletal tyrosine kinase signals such as focal adhesion kinase during restitution may trigger a phenotypic switch to the "dedifferentiated" migrating intestinal epithelial phenotype.

Caco-2 Cells↗
PubMed2004

2,5-Dihydroxybenzoic acid butylamine and other ionic liquid matrixes for enhanced MALDI-MS analysis of biomolecules.

The performance of the new ionic liquid MALDI-MS matrix 2,5-dihydroxybenzoic acid butylamine (DHBB) was assessed and compared to results obtained with the ionic liquid MALDI-MS matrixes alpha-cyano-4-hydroxycinnamic acid butylamine (CHCAB), 3,5-dimethoxycinnamic acid triethylamine (SinTri), and the frequently used solid MALDI matrixes 2,5-dihydroxybenzoic acid (DHB) and alpha-cyano-4-hydroxycinnamic acid (CHCA). The vacuum-stable, liquid consistency of ionic liquid matrix sample preparations considerably enhanced MALDI-MS analysis in terms of shot-to-shot reproducibility. Consequently, relative standard deviations serving as a measure for reproducibility of intensity-values acquired from 90 different spots on one MALDI-MS preparation were approximately one-half as high when solid DHB was replaced by the ionic liquid DHBB and eight times lower after exchange of solid CHCA by ionic liquid CHCAB. Interestingly, the ionic liquid MALDI matrix DHBB conserved the broad applicability of its solid analogue DHB, reduced MALDI induced fragmentation of monosialylated glycans and gangliosides, and was the superior ionic liquid matrix for MALDI-MS analysis of oligosaccharides and polymers, such as poly(ethylene glycol). It also worked well with glycoconjugates, peptides, and proteins; however, the tendency of DHBB to form multiple alkali adduct ions with peptides and proteins made CHCAB the ionic liquid matrix of choice for peptides. SinTri was the best ionic liquid matrix for proteins of high molecular weight, such as IgG. Furthermore, it was demonstrated for the first time that solvent properties and MALDI matrix properties of ionic liquids, such as DHBB, can be combined to enable fast, direct screening of an enzymatic reaction. This was proven by the desialylation of sialylactose with sialidase from Clostridium perfringens in the presence of diluted aqueous DHBB and subsequent direct MALDI-MS analysis of the reaction mixture.

Butylamines↗
PubMed1997

Nuclear matrix, dynamic histone acetylation and transcriptionally active chromatin.

The nuclear matrix, the RNA-protein skeleton of the nucleus, has a role in the organization and function of nuclear DNA. Nuclear processes associated with the nuclear matrix include transcription, replication and dynamic histone acetylation. Nuclear matrix proteins, which are tissue and cell type specific, are altered with transformation and state of differentiation. Transcription factors are associated with the nuclear matrix, with the spectra of nuclear matrix bound factors being cell type specific. There is compelling evidence that the transcription machinery is anchored to the nuclear matrix, and the chromatin fiber is spooled through this complex. Transcriptionally active chromatin domains are associated with dynamically acetylated histones. The energy exhaustive process of dynamic histone acetylation has several functions. Acetylation of the N-terminal tails of the core histones alters nucleosome and higher order chromatin structure, aiding transcriptional elongation and facilitating the binding of transcription factors to nucleosomes associated with regulatory DNA sequences. Histone acetylation can manipulate the interactions of regulatory proteins that bind to the N-terminal tails of the core histones. Lastly, dynamic acetylation may contribute to the transient attachment of transcriptionally active chromatin to the nuclear matrix. Reversible histone acetylation is catalyzed by histone acetyltransferase and deacetylase, enzymes associated with the nuclear matrix. The recent isolation and characterization of histone acetyltransferase and deacetylase reveals that these enzymes are related to transcriptional regulators, providing us with new insights about how these enzymes are targeted to nuclear matrix sites engaged in transcription.

Acetylation↗
PubMed2000

Relationship between cell-associated matrix metalloproteinase 9 and psoriatic keratinocyte growth.

Primary cultures of psoriatic keratinocytes proliferated at a higher rate and produced lower amounts of matrix metalloproteinase 9 than normal keratinocytes cultured under similar conditions. Sup- plementation of psoriatic keratinocyte cell culture medium with batimastat or the use of a matrix metalloproteinase 9 blocking antibody further stimulated psoriatic keratinocyte growth. An increase in intracellular ceramide level enhanced matrix metalloproteinase 9 production and inhibited cell proliferation in parallel. Whether cells were treated with sphingomyelinase or not, however, conditioned media from psoriatic keratinocytes contained higher levels of tissue inhibitor of metalloproteinase-1 compared with matrix metalloproteinase 9 and secreted only the proenzyme form. Pro-matrix metalloproteinase 9, as well as active matrix metalloproteinase 9, was identified in membrane preparations of psoriatic keratinocytes, and enzyme amounts were greatly elevated following sphingomyelinase action. As (i) tissue inhibitor of metalloproteinase-1 antibody nearly totally abrogated keratinocyte growth and (ii) complexes of tissue inhibitor of metalloproteinase-1 and matrix metalloproteinase 9 were recovered in membrane extracts of sphingomyelinase-treated psoriatic keratinocytes, we postulate that an increased level of cell-associated matrix metalloproteinase 9 might compete for tissue inhibitor of metalloproteinase-1 binding to its receptor. As a consequence, the increased levels of matrix metalloproteinase 9 will decrease keratinocyte growth.

Adult↗
PubMed2002

Indirectly formed matrix for multiple composite core restorations: two clinical treatments illustrating an expanded technique.

The technique described in this report is an expansion of a previously reported intraoral procedure for directly developing a matrix for the fabrication of composite cores. The expanded method allows a matrix to be created indirectly after the involved teeth are trimmed or restored on a cast into slightly oversized crown preparations before the treatment visit. The cast may serve for both the matrix and the interim fixed restoration, enabling fabrication by the dental laboratory. A semirigid (occlusal registration) elastomeric material is used to form the matrix. The matrix is used to apply the composite and shape the cores after the abutment teeth have been prepared, pins or posts have been placed, and adhesive procedures have been implemented. The elastomeric matrix can be modified for use as the impression matrix. The described technique allows the simultaneous formation of multiple composite cores. An alternative method in which light-polymerized core composite is used with a vacuum-formed, dual-laminate transparent matrix is described and compared with the semirigid elastomeric matrix.

Composite Resins↗
PubMed1990

Progressive changes in the protein composition of the nuclear matrix during rat osteoblast differentiation.

Primary cultures of fetal rat calvarial osteoblasts undergo a developmental sequence with respect to the temporal expression of genes encoding osteoblast phenotypic markers. Based on previous suggestions that gene-nuclear matrix associations are involved in regulating cell- and tissue-specific gene expression, we investigated the protein composition of the nuclear matrix during this developmental sequence by using high-resolution two-dimensional gel electrophoresis. The nuclear matrix was isolated at times during a 4-week culture period that represent the three principal osteoblast phenotypic stages: proliferation, extracellular matrix (ECM) maturation, and mineralization. The most dramatic changes in the nuclear matrix protein patterns occurred during transitions from the proliferation to the ECM maturation stage and from ECM maturation to the mineralization period, with only minor variations in the profiles within each period. These stage-specific changes, corresponding to the major transition points in gene expression, indicate that the nuclear matrix proteins reflect the progressive differentiation of the bone cell phenotype. Subcultivation of primary cells delays mineralization, and a corresponding delay was observed for the nuclear matrix protein patterns. Thus, the sequential changes in protein composition of the nuclear matrix that occur during osteoblast differentiation represent distinct stage-specific markers for maturation of the osteoblast to an osteocytic cell in a bone-like mineralized ECM. These changes are consistent with a functional involvement of the nuclear matrix in mediating modifications of developmental gene expression.

Animals↗
PubMed1993

Osteocalcin gene promoter-binding factors are tissue-specific nuclear matrix components.

The nuclear matrix appears to play an important role in developmental gene expression during osteoblast differentiation. To better understand this role, we examined nuclear matrix DNA-binding proteins that are sequence-specific and interact with the osteocalcin gene promoter. Multiple protein-DNA interactions involving two distinct nuclear matrix proteins occur within the 5' regulatory sequences (nt -640 to -430). One of these proteins, NMP-1, is a ubiquitous, cell growth-regulated protein that is related to the transcription factor ATF and resides in both the nuclear matrix and the nonmatrix nuclear compartment. The other protein, NMP-2, is a cell type-specific, 38-kDa promoter factor that recognizes binding sites resembling the consensus site for the CCAAT/enhancer-binding protein C/EBP and is localized exclusively on the nuclear matrix. NMP-1 and NMP-2 each interact with two nuclear matrix protein-binding elements. These elements are present near key regulatory sites of the osteocalcin gene promoter, such as the principal steroid hormone (vitamin D)-responsive sequences. Binding in this region of the osteocalcin gene promoter suggests transient associations with the nuclear matrix that are distinct from the stable interactions of matrix attachment regions. Our results are consistent with involvement of the nuclear matrix in concentrating and/or localizing transcription factors that mediate the basal and steroid hormone responsiveness of osteocalcin gene transcription.

Animals↗
PubMed1998

Integrin-linked protein kinase regulates fibronectin matrix assembly, E-cadherin expression, and tumorigenicity.

Fibronectin (Fn) matrix plays important roles in many biological processes including morphogenesis and tumorigenesis. Recent studies have demonstrated a critical role of integrin cytoplasmic domains in regulating Fn matrix assembly, implying that intracellular integrin-binding proteins may be involved in controlling extracellular Fn matrix assembly. We report here that overexpression of integrin-linked kinase (ILK), a newly identified serine/threonine kinase that binds to the integrin beta1 cytoplasmic domain, dramatically stimulated Fn matrix assembly in epithelial cells. The integrin-linked kinase activity is involved in transducing signals leading to the up-regulation of Fn matrix assembly, as overexpression of a kinase-inactive ILK mutant failed to enhance the matrix assembly. Moreover, the increase in Fn matrix assembly induced by ILK overexpression was accompanied by a substantial reduction in the cellular E-cadherin. Finally, we show that ILK-overexpressing epithelial cells readily formed tumors in nude mice, despite forming an extensive Fn matrix. These results identify ILK as an important regulator of pericellular Fn matrix assembly, and suggest a novel critical role of this integrin-linked kinase in cell growth, cell survival, and tumorigenesis.

Amino Acid Sequence↗
PubMed2001

A-kinase-anchoring protein AKAP95 is targeted to the nuclear matrix and associates with p68 RNA helicase.

The cell nucleus is structurally and functionally organized by the nuclear matrix. We have examined whether the nuclear cAMP-dependent protein kinase-anchoring protein AKAP95 contains specific signals for targeting to the subnuclear compartment and for interaction with other proteins. AKAP95 was expressed in mammalian cells and found to localize exclusively to the nuclear matrix. Mutational analysis was used to identify determinants for nuclear localization and nuclear matrix targeting of AKAP95. These sites were found to be distinct from previously identified DNA and protein kinase A binding domains. The nuclear matrix-targeting site is unique but conserved among members of the AKAP95 family. Direct binding of AKAP95 to isolated nuclear matrix was demonstrated in situ and found to be dependent on the nuclear matrix-targeting site. Moreover, Far Western blot analysis identified at least three AKAP95-binding proteins in nuclear matrix isolated from rat brain. Yeast two-hybrid cloning identified one binding partner as p68 RNA helicase. The helicase and AKAP95 co-localized in the nuclear matrix of mammalian cells, associated in vitro, and were precipitated as a complex from solubilized cell extracts. The results define novel protein-protein interactions among nuclear matrix proteins and suggest a potential role of AKAP95 as a scaffold for coordinating assembly of hormonally responsive transcription complexes.

A Kinase Anchor Proteins↗
PubMedDate not supplied

Accessibility to the fibronectin synergy site in a 3D matrix regulates engagement of alpha5beta1 versus alphavbeta3 integrin receptors.

Cell adhesion and migration on fibronectin (FN) extracellular matrix are mediated by integrin receptors. Integrins alpha5beta1 and alphavbeta3 require the RGD cell-binding sequence in FN, but alpha5beta1 also requires the nearby synergy site for maximal binding. In this study, we investigated how differences in the numbers of RGD or synergy sites within a three-dimensional (3D) FN-rich matrix influence cell adhesion and migration. CHO cell adhesion, spreading, and migration were reduced on 3D chimeric matrix containing FN lacking RGD (FN(RGD-)). Incorporation of FN with mutation of the synergy site (FN(syn-)), however, resulted in selective usage of integrins. CHO cells expressing alpha5beta1 showed decreased interactions with FN(syn-) chimeric matrix. In contrast, the presence of FN(syn-) had no effect on CHOalphavbeta3 cell migration. Interestingly, CHOalpha5/alphavbeta3 cells expressing both integrins selectively used alpha5beta1 for migration on wild type FN matrix but preferred alphavbeta3 for migration on FN(syn-) chimeric matrix. Thus sequestration or exposure of the FN synergy site within a 3D matrix may represent a novel mechanism for regulating cell functions through differential usage of integrin receptors. [Supplementary materials are available for this article. Go to the publisher's online edition of Cell Communication and Adhesion for the following free supplemental resource: a video recording shows migration of HT1080 cells on 3D matrix. HT1080 cells were allowed to attach to the matrix in serum-free DMEM for 2 h. FBS was then added to the medium to a final concentration of 10% and video recording was started. Images were taken every 5 min for 2 h. The video plays at 6 frames/s.].

Animals↗
PubMed1986

Schwann cell myelination: induction by exogenous basement membrane-like extracellular matrix.

Exposing rat Schwann cells co-cultured with nerve cells to a reconstituted basement membrane induced the formation of myelin segments by Schwann cells. This occurred in a serum-free culture medium in which, in the absence of this matrix, Schwann cells proliferate but fail to differentiate. This reconstituted basement membrane was prepared from solubilized extracellular matrix proteins synthesized by a basement membrane-producing murine tumor. The major constituents of this reconstituted matrix are collagen type IV, laminin, heparan sulfate proteoglycan, entactin, and nidogen. The matrix also elicited striking morphological changes in Schwann cells, inducing them to spread longitudinally along the nerve fibers (a necessary early step in the process of ensheathment of nerve fibers). Several observations indicated that the effect of the matrix was exerted directly on Schwann cells and not indirectly through an effect on nerve cells. First, the matrix-induced cell spreading occurred only in areas in which Schwann cells directly contacted the matrix; Schwann cells that were associated with the same nerve fibers but that did not themselves directly contact the matrix did not exhibit spreading. Second, the matrix-induced alteration in Schwann cell morphology was observed in cultures in which the nerve cells were removed. These results provide direct evidence that basement membrane contact induces normal Schwann cell differentiation, and support the idea that Schwann cell differentiation in vivo may be regulated by the appearance of the basement membrane, which normally envelops terminally differentiating Schwann cells.

Animals↗
PubMed1987

Perturbation of human endothelial cells by thrombin or PMA changes the reactivity of their extracellular matrix towards platelets.

In this study we have examined the influence of perturbation of endothelial cells on the amounts of fibronectin and von Willebrand factor in their extracellular matrix and the consequences of a changed composition of the matrix on platelet adhesion. For this purpose, we have used an in vitro perfusion system with which we can investigate the interactions of platelets in flowing blood with cultured endothelial cells and their extracellular matrix (Sakariassen, K. S., P. A. M. M. Aarts, P. G. de Groot, W. P. M. Houdgk, and J. J. Sixma, 1983, J. Lab. Clin Med. 102:522-535). Treatment of endothelial cells with 0.1-1.0 U/ml thrombin for 2 h increased the reactivity of the extracellular matrix, isolated after the thrombin treatment, towards platelets by approximately 50%. The increased reactivity did not depend on de novo protein synthesis but was inhibited by 3-deazaadenosine, an inhibitor of phospholipid methylation, which also inhibits the stimulus-induced instantaneous release of von Willebrand factor from endothelial cells. However, no changes in the amounts of von Willebrand factor and fibronectin in the matrix were detected. Thrombin may change the organization of the matrix proteins, not the composition. When endothelial cells were perturbed with the phorbol ester PMA or thrombin for 3 d, the adhesion of platelets to the extracellular matrix of treated cells was strongly impaired. This impairment coincided with a decrease in the amounts of von Willebrand factor and fibronectin present in the matrix. These results indicate that, after perturbation, endothelial cells regulate the composition of their matrix, and that this regulation has consequences for the adhesion of platelets.

Blood Platelets↗
PubMed1988

Metabolism of receptor-bound and matrix-bound basic fibroblast growth factor by bovine capillary endothelial cells.

Bovine capillary endothelial (BCE) cells were incubated at 4 degrees C with 5 ng/ml 125I-basic fibroblast growth factor (bFGF) to equilibrate 125I-bFGF with high affinity cell surface receptors and low affinity matrix binding sites. 67% of the added 125I-bFGF bound to the matrix and 7% bound to receptors. The fate of bound bFGF was followed after cells were incubated in bFGF-free medium and were shifted to 37 degrees C to restore cell metabolism. 125I-bFGF bound to receptors decreased rapidly while the amount of 125I-bFGF bound to matrix was reduced more slowly. The rapid decrease in receptor-bound 125I-bFGF appeared to be due to a down-regulation of bFGF receptors; cells that had been treated for 5 h with bFGF had 60% fewer high affinity receptors than untreated cells. Despite the initial high level of 125I-bFGF binding to matrix, most of this 125I-bFGF was mobilized and metabolized by the cells. 125I-bFGF was internalized by the cells at 37 degrees C, leading to a constant accumulation of 125I-bFGF within the cell. Internalized bFGF was rapidly cleaved from an 18-kD form to a 16-kD form. The 16-kD form was more slowly degraded with a half-life of approximately 8 h. Degradation of internalized 125I-bFGF was inhibited by chloroquine, suggesting that the digestion occurred in a lysosomal compartment. The role of matrix binding sites in the internalization process was investigated. Binding to matrix sites seemed not to be directly involved in the internalization process, since addition of heparin at a concentration that blocked 95% of the binding to matrix had no effect on the initial rate of internalization of bFGF. BCE cells also released a substance that competed for the binding of bFGF to matrix but not to receptors. This substance bound to DEAE-cellulose and was sensitive to heparinase treatment, suggesting that it was a heparinlike molecule. Thus, heparinlike molecules produced by BCE cells can modulate the cellular interaction with bFGF. Matrix-associated heparinlike molecules bind bFGF which can later be metabolized by the cell, and secreted heparinlike molecules release bFGF from matrices.

Animals↗
PubMed1989

The fibronectin receptor is organized by extracellular matrix fibronectin: implications for oncogenic transformation and for cell recognition of fibronectin matrices.

Cells interact with extracellular fibronectin (FN) via adhesive fibronectin receptors (FNRs) that are members of the very late antigens (VLAs) subgroup of the integrin family. In stationary fibroblasts, the FNR is highly organized and distributed identically to extracellular FN fibrils. However, in highly migratory neural crest cells and embryonic somatic fibroblasts, this organization is lost and the FNR appears diffuse. Similarly, oncogenic transformation typically leads to disorganization of the FN receptor and loss of matrix FN. Two models can account for these observations. First, the FN matrix may organize the FN receptor at extracellular matrix contacts on the cell surface. Motile cells not depositing FN matrices thus lack organized receptors. Alternatively, as the FNR is required for optimal FN matrix assembly, (McDonald, J. A., B. J. Quade, T. J. Broekelmann, R. LaChance, K. Forseman, K. Hasegawa, and S. Akiyama. 1987. J. Biol. Chem. 272:2957-2967; Roman, J. R. M. LaChance, T. J. Broekelmann, C. J. R. Kennedy, E. A. Wayner, W. G. Carter, J. A. McDonald. 1989. J. Cell Biol. 108:2529-2543) and has putative cytoskeletal links, it could be organized from within the cell helping to position newly forming FN fibrils. To study this question, we developed peptide antibodies specifically recognizing the alpha 5 subunit of the FNR. Using these antibodies, we examined the organization of FN and of the FNR in normal, matrix assembly inhibited, and SV40-transformed human fibroblasts. On FN-coated substrates, the FNR is found in focal contacts rather than diffusely on the basal cell surface, suggesting FNR interaction with intracellular components. However, when FN fibrils are deposited, the FNR is co-distributed with these fibrils. Preventing FN matrix assembly prevents organization of the FNR. Moreover, when fibroblasts with well established FN matrices and co-distributed FNR are incubated briefly with monoclonal antibodies that block FNR binding to FN, the FNR is no longer co-distributed with the FN matrix. Thus, the FN receptor is organized in fibrils on the cell surface in response to extracellular FN. Because exogenous FN restores a FN matrix and receptor organization to SV40-transformed cells, the diffuse FN receptor phenotype appears to be related to loss of the FN matrix rather than to impaired FNR function. These results explain diffusely distributed FNRs in migratory neural crest and embryonic fibroblasts lacking well organized FN matrices and emphasize the existence of separate but related systems controlling FN deposition and recognition by receptor-armed cells.

Cell Adhesion↗
PubMed1990

An instructive role for the interstitial matrix in tissue patterning: tissue segregation and intercellular invasion.

Intercellular invasion is the intrusion of the cells of one tissue into space occupied by a second tissue. The alternative situation to invasion, one characteristic of most coherent tissues, is segregation, with identifiable boundaries existing between contiguous tissues. The interfaces between mesenchymal and myocardial tissues in the developing avian heart show a profoundly different character in different regions of the heart: the interface between epicardial mesenchyme and heart wall myocardium is planar, without intermingling of the two cell types, whereas the interface between endocardial cushion mesenchyme and myocardium is diffuse, with extensive invasion of both tissue types across the border to produce intermingling of the two tissues. Thus, invasion and tissue segregation coexist in different regions of the mesenchyme-myocardium contact zone. Investigation of the involvement of the interstitial matrix in invasion and segregation has been conducted by maintaining the two tissues in mutual contact in organ culture. Investigation of the mechanisms by which the two cell types sort out in randomized chimeric tissue reaggregates has provided insight into the conditions for tissue segregation. We have modeled invasion in organ culture by fusing aggregates of myocardial cells with aggregates of cardiac mesenchymal cells. Cells of both tissues invaded the partner aggregate during a period of 1-3 d of coculture. Both invasion and segregation in the aggregates appear to depend on the presence or absence of a fibronectin-rich interstitial matrix elaborated by the cardiac mesenchyme. During sorting, the matrix appears selectively in regions occupied by the mesenchyme. Under conditions of culture that are nonpermissive for matrix deposition, sorting fails to occur. Stimulation of matrix deposition by addition of serum, transforming growth factor beta, or isolated matrix itself is accompanied by sorting out of the two tissues. Sorting out is blocked reversibly by inclusion of the fibronectin adhesion site peptide, GRGDSP. Invasion of fused aggregates is preceded by a redistribution of the fibronectin-containing matrix of the mesenchymal aggregate such that matrix-poor regions come to occupy the interface with the myocardial partner aggregate. The invasion that ensues involves mesenchymal cells emigrating from, and myocardial cells intruding into, matrix-poor regions of the mesenchymal aggregate.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗