Differential properties of attachment of human fibroblasts to various extracellular matrix proteins

Human diploid fibroblasts (TIG-3) were shown to attach and spread onto substrata coated with collagen, fibronectin, laminin and vitronectin. The cell attachment to these proteins required divalent cations. Mg2+ stimulated the cell attachment to all the proteins, while Ca2+ alone was not effective for the attachment to collagen and laminin. A mild trypsin treatment had prevented cells from attaching to the laminin, while it had no effect on the attachment to the other proteins. The fibronectin fragment, which retained cell binding activity, inhibited the cells from attaching and spreading onto fibronectin, but it did not cause any inhibition on the other proteins. The synthetic peptide GRGDSP inhibited the cells from attaching and spreading onto fibronectin and vitronectin, while it did not cause any inhibition on collagen and laminin. In attempts to isolate distinct receptors for these proteins, we were able to purify proteins very similar to the fibronectin and vitronectin receptors of human placenta. Based on the differential properties of the attachment of TIG-3 cells to these proteins and biochemical data, we indicate that human diploid fibroblasts have distinctive binding sites (receptors) for collagen, fibronectin, laminin and vitronectin.     

Embryonic neural retinal cell response to extracellular matrix proteins: developmental changes and effects of the cell substratum attachment antibody (CSAT)

Cell attachment and neurite outgrowth by embryonic neural retinal cells were measured in separate quantitative assays to define differences in substrate preference and to demonstrate developmentally regulated changes in cellular response to different extracellular matrix glycoproteins. Cells attached to laminin, fibronectin, and collagen IV in a concentration-dependent fashion, though fibronectin was less effective for attachment than the other two substrates. Neurite outgrowth was much more extensive on laminin than on fibronectin or collagen IV. These results suggest that different substrates have distinct effects on neuronal differentiation. Neural retinal cell attachment and neurite outgrowth were inhibited on all three substrates by two antibodies, cell substratum attachment antibody (CSAT) and JG22, which recognize a cell surface glycoprotein complex required for cell interactions with several extracellular matrix constituents. In addition, retinal cells grew neurites on substrates coated with the CSAT antibodies. These results suggest that cell surface molecules recognized by this antibody are directly involved in cell attachment and neurite extension. Neural retinal cells from embryos of different ages varied in their capacity to interact with extracellular matrix substrates. Cells of all ages, embryonic day 6 (E6) to E12, attached to collagen IV and CSAT antibody substrates. In contrast, cell attachment to laminin and fibronectin diminished with increasing embryonic age. Age-dependent differences were found in the profile of proteins precipitated by the CSAT antibody, raising the possibility that modifications of these proteins are responsible for the dramatic changes in substrate preference of retinal cells between E6 and E12.     

Attachment of cells to basement membrane collagen type IV

Of ten different cell lines examined, three showed distinct attachment and spreading on collagen IV substrates, and neither attachment nor spreading was enhanced by adding soluble laminin or fibronectin. This reaction was not inhibited by cycloheximide or antibodies to laminin, indicating a direct attachment to collagen IV without the need of mediator proteins. Cell-binding sites were localized to the major triple-helical domain of collagen IV and required an intact triple helical conformation for activity. Fibronectin showed preferential binding to denatured collagen IV necessary to mediate cell binding to the substrate. Fibronectin binding sites of collagen IV were mapped to unfolded structures of the major triple-helical domain and show a similar specificity to fibronectin-binding sites of collagen I. The data extend previous observations on biologically potential binding sites located in the triple helix of basement membrane collagen IV.     

Identification of asparagine-linked oligosaccharides involved in tumor cell adhesion to laminin and type IV collagen

MDW4, a wheat germ agglutinin-resistant nonmetastatic mutant of the highly metastatic murine tumor cell line called MDAY-D2 has previously been shown to attach to fibronectin and type IV collagen, whereas MDAY- D2 and phenotypic revertants of MDW4 attached poorly to these substrates. The increased adhesiveness of the mutant cells appeared to be closely related to a lesion in cell surface carbohydrate structures. In an effort to identify the carbohydrates involved in cell attachment, glycopeptides isolated from mutant and wild-type cells as well as from purified glycoproteins were tested for their ability to inhibit the attachment of MDW4 cells to plastic surfaces coated with fibronectin, laminin, or type IV collagen. The addition of mannose-terminating glycopeptide to the adhesion assay inhibited MDW4 cell attachment to type IV collagen. In contrast, a sialylated poly N-acetyllactosamine- containing glycopeptide, isolated from wheat germ agglutinin-sensitive MDAY-D2 cells but absent in MDW4 cells, inhibited MDW4 attachment to laminin. None of the glycopeptides used in this study inhibited attachment of MDW4 cells to fibronectin-coated plastic. Peptide N- glycosidase treatment of the cells to remove surface asparagine-linked oligosaccharides inhibited MDW4 adhesion to type IV collagen, but not to laminin, and the same treatment of the wheat germ agglutinin- sensitive cells enhanced attachment to laminin. Tumor cell attachment to, and detachment from, the sublaminal matrix protein laminin and type IV collagen are thought to be important events in the metastatic process. Our results indicate that tumor cell attachment to these proteins may be partially modulated by the expression of specific oligosaccharide structures associated with the cell surface.     

Interactions of rat hepatocytes with type IV collagen, fibronectin and laminin matrices. Distinct matrix-controlled modes of attachment and spreading

We have examined the interaction of adult rat hepatocytes in primary culture, to type IV collagen, fibronectin, and laminin, the major basement membrane proteins of normal rat liver. Culture substrata consisted of glass coverslips, which were covalently derivatized with individual purified basement membrane constituents at varying densities of protein. The attachment of freshly prepared hepatocytes was examined after incubation at 37 degrees C for 30 min as a function of the amount of protein on the coverslips. For each of the three types of substratum under study, distinct modes of cell attachment were observed, with the apparent affinity of hepatocytes for type IV collagen being three-fold greater than for fibronectin and ten-fold greater than for laminin. Cell attachment exhibited saturation on all substrata. Hepatocyte spreading was measured by scanning electron microscopy of cells incubated at 37 degrees for 2 h on similarly prepared coverslips. A five-fold greater surface density of type IV collagen was required for maximal spreading compared with attachment. For cells on fibronectin or laminin the maximal cell spreading reached on type IV collagen did not occur even at coverslip protein densities 10 to 20 times those providing for maximal cell attachment. A very similar qualitative pattern of cell proteins was secreted within a few hours of plating on the various substrata and further studies failed to reveal any evidence that attachment and spreading was mediated by endogenously produced matrix molecules.(ABSTRACT TRUNCATED AT 250 WORDS)     

Concurrent changes in sinusoidal expression of laminin and affinity of hepatocytes to laminin during rat liver regeneration.

Distribution of fibronectin, laminin, and collagens type I, III, IV, and V in the lobular regions of regenerating rat liver was studied by indirect immunofluorescence. Little or no laminin was detected in sham-operated controls throughout the experimental period, while it was detected in sinusoids of regenerating liver as early as 6 h after partial hepatectomy (PH). After reaching a maximum at 24 h, it decreased and was barely detectable 6 days after PH. Changes in the other extracellular matrix (ECM) proteins were evident 3 days after PH, but not earlier than 24 h. Hepatocytes isolated from regenerating rat livers were tested in a short term assay for attachment to the substrates coated with the ECM proteins. The attachment of hepatocytes to laminin substrates increased 12 h after PH, reached a maximum at 24 h, and decreased to the control level 6 days after PH, while that of the control remained constant. The attachment to fibronectin substrates was not different between regenerating livers and controls at any time point. The attachment to collagen did not change earlier than 24 h after PH, but increased slightly 3 days after PH. Primary rat hepatocytes cultured on the substrates coated with the ECM proteins were determined for replicative DNA synthesis in response to epidermal growth factor. Both in normal liver and in regenerating liver 24 h after PH, laminin was one of the most effective substrates in supporting the responsiveness of hepatocytes to the growth stimulus. Taken together, these results suggest the importance of hepatocyte-laminin interaction during the early stage of liver regeneration possibly in growth stimulation of hepatocytes and/or maintenance of hepatocyte-specific functions.     

Phorbol esters enhance attachment of NIH/3T3 cells to laminin and type IV collagen substrates

The effect of phorbol esters on the adhesive properties of NIH/3T3 mouse fibroblasts was investigated using plastic substrates precoated with the extracellular matrix proteins fibronectin, collagen, and laminin. Treatment with phorbol 12-myristate 13-acetate (PMA) enhanced NIH/3T3 cell attachment to laminin and type IV collagen substrates but had little or no effect on attachment to fibronectin and type I collagen substrates. The effect of PMA in enhancing cell attachment to laminin and type IV collagen substrates was dose dependent between 10(-9) and 10(-7) M. PMA was effective as early as 30 min; the effect reached a maximum at 2 h and decreased gradually. Phorbol 12, 13-dibenzoate and phorbol 12, 13-diacetate were effective but to a lesser extent and phorbol 12-myristate and phorbol 13-acetate showed little or no effect. These results suggest that PMA may enhance NIH/3T3 cell adhesion through effects on laminin and type IV collagen receptors. Retinoic acid, which itself requires at least 6 h to show an effect on attachment, did not have any effect on cell attachment in 2 h and, if anything, slightly inhibited PMA-enhanced cell attachment to laminin and type IV collagen substrates.     

Functionally distinct laminin receptors mediate cell adhesion and spreading: the requirement for surface galactosyltransferase in cell spreading

The molecular mechanisms underlying cell attachment and subsequent cell spreading on laminin are shown to be distinct form one another. Cell spreading is dependent upon the binding of cell surface galactosyltransferase (GalTase) to laminin oligosaccharides, while initial cell attachment to laminin occurs independent of GalTase activity. Anti-GalTase IgG, as well as the GalTase modifier protein, alpha-lactalbumin, both block GalTase activity and inhibited B16-F10 melanoma cell spreading on laminin, but not initial attachment. On the other hand, the addition of UDP galactose, which increases the catalytic turnover of GalTase, slightly increased cell spreading. None of these reagents had any effect on cell spreading on fibronectin. When GalTase substrates within laminin were either blocked by affinity- purified GalTase or eliminated by prior galactosylation, cell attachment appeared normal, but subsequent cell spreading was totally inhibited. The laminin substrate for GalTase was identified as N-linked oligosaccharides primarily on the A chain, and to a lesser extent on B chains. That N-linked oligosaccharides are necessary for cell spreading was shown by the inability of cells to spread on laminin surfaces pretreated with N-glycanase, even though cell attachment was normal. Cell surface GalTase was distinguished from other reported laminin binding proteins, most notably the 68-kD receptor, since they were differentially eluted from laminin affinity columns. These data show that surface GalTase does not participate during initial cell adhesion to laminin, but mediates subsequent cell spreading by binding to its appropriate N-linked oligosaccharide substrate. These results also emphasize that some of laminin's biological properties can be attributed to its oligosaccharide residues.     

Analysis of fibronectin receptor function with monoclonal antibodies: roles in cell adhesion, migration, matrix assembly, and cytoskeletal organization

We have developed two rat mAbs that recognize different subunits of the human fibroblast fibronectin receptor complex and have used them to probe the function of this cell surface heterodimer. mAb 13 recognizes the integrin class 1 beta polypeptide and mAb 16 recognizes the fibronectin receptor alpha polypeptide. We tested these mAbs for their inhibitory activities in cell adhesion, spreading, migration, and matrix assembly assays using WI38 human lung fibroblasts. mAb 13 inhibited the initial attachment as well as the spreading of WI38 cells on fibronectin and laminin substrates but not on vitronectin. Laminin-mediated adhesion was particularly sensitive to mAb 13. In contrast, mAb 16 inhibited initial cell attachment to fibronectin substrates but had no effect on attachment to either laminin or vitronectin substrates. When coated on plastic, both mAbs promoted WI38 cell spreading. However, mAb 13 (but not mAb 16) inhibited the radial outgrowth of cells from an explant on fibronectin substrates. mAb 16 also did not inhibit the motility of individual fibroblasts on fibronectin in low density culture and, in fact, substantially accelerated migration rates. In assays of the assembly of an extracellular fibronectin matrix by WI38 fibroblasts, both mAbs produced substantial inhibition in a concentration-dependent manner. The inhibition of matrix assembly resulted from impaired retention of fibronectin on the cell surface. Treatment of cells with mAb 16 also resulted in a striking redistribution of cell surface fibronectin receptors from a streak-like pattern to a relatively diffuse distribution. Concomitant morphological changes included decreases in thick microfilament bundle formation and reduced adhesive contacts of the streak-like and focal contact type. Our results indicate that the fibroblast fibronectin receptor (a) functions in initial fibroblast attachment and in certain types of adhesive contact, but not in the later steps of cell spreading; (b) is not required for fibroblast motility but instead retards migration; and (c) is critically involved in fibronectin retention and matrix assembly. These findings suggest a central role for the fibronectin receptor in regulating cell adhesion and migration.     

Antagonistic effects of laminin and fibronectin in cell-to-cell and cell-to-matrix interactions in MCF-7 cultures

Summary During morphogenesis, tumor progression and metastasis, cell adhesion, dissociation, and migration result from a complex balance between cell-to-cell and cell-to-matrix interactions. Two different organization patterns of MCF-7 cells were induced by different extracellular matrix proteins. When plated on plastic or polymeric type I collagen gel used as a model of interstitial matrix, MCF-7 cells spread and grew in monolayer. When cultured on a solid gel of basement membrane (BM) proteins (85% laminin) used as a model of BM, cells formed clusters attached to the matrix. Matrix proteins regulated these two types of cell organization by preferentially promoting cell-to-cell or cell-support interactions. On plastic in the presence of soluble laminin or on laminin-coated dishes, cells also formed clusters. Addition of soluble fibronectin induced spreading of the cells, suggesting that laminin and fibronectin have competitive antagonistic effects on MCF-7 cell morphology. Antilaminin antibodies inhibited cluster formation and attachment, emphasizing the important role of this glycoprotein not only in promoting cluster attachment but also in cell-to-cell contact formation. Such effects of extracellular matrix proteins could play significant roles in tumor progression and metastasis. This work was supported by grants 3.4512.85 and 3.4514.85 from the Belgian Fonds de la Recherche Scientifique Médicale and the Fonds Cancérologique de la CGER.     

Retinoic acid modulates attachment of mouse fibroblasts to laminin substrates

The effect of retinoic acid treatment on cell attachment to plastic substrates precoated with fibronectin, gelatin, laminin, and type IV collagen was investigated. Both retinoic acid-treated and control cells attached efficiently to fibronectin or gelatin substrates without any significant difference. In contrast, retinoic acid-treated cells attached to laminin or type IV collagen substrates, while control cells showed little or no attachment. The minimal effective concentration of retinoic acid for pretreatment to yield a significant increase in the attachment assay was higher than 10(-8) M. The attachment of retinoic acid-treated cells to laminin substrates reached a maximum at 60 min, while that to type IV collagen substrates had a time lag and did not reach a maximum by 60 min. The effect of retinoic acid treatment reached a maximum at 2 days and was partly reversible. These results suggest that retinoic acid may increase NIH/3T3 cell adhesion through an effect on laminin receptors. Other mouse fibroblast lines, 3T3-Swiss, 3T6-Swiss, Balb/3T3, and Balb/3T12-3 (spontaneously transformed Balb/3T3), responded to retinoic acid treatment in a manner similar to that of NIH/3T3 cells. However, the virus-transformed Balb/3T3 lines, SV-T2 and M-MSV, showed significant attachment to laminin substrates without retinoic acid treatment, and retinoic acid did not affect or slightly decreased the cell attachment to laminin substrates.     

Integrins beta1, alpha6, and alpha3 contribute to mechanical strain-induced differentiation of fetal lung type II epithelial cells via distinct mechanisms

Mechanical forces regulate lung maturation in the fetus by promoting type II epithelial differentiation. However, the cell surface receptors that transduce these mechanical cues into cellular responses remain largely unknown. When distal lung type II epithelial cells isolated from embryonic day 19 rat fetuses were cultured on flexible plates coated with laminin, fibronectin, vitronectin, collagen, or elastin and exposed to a level of mechanical strain (5%) similar to that observed in utero, transmembrane signaling responses were induced under all conditions, as measured by ERK activation. However, mechanical stress maximally increased expression of the type II cell differentiation marker surfactant protein C when cells were cultured on laminin substrates. Strain-induced alveolar epithelial differentiation was inhibited by interfering with cell binding to laminin using soluble laminin peptides (IKVIV or YIGSR) or blocking antibodies against integrin beta1, alpha3, or alpha6. Additional studies were carried out with substrates coated directly with different nonactivating anti-integrin antibodies. Blocking integrin beta1 and alpha6 binding sites inhibited both cell adhesion and differentiation, whereas inhibition of alpha3 prevented differentiation without altering cell attachment. These data demonstrate that various integrins contribute to mechanical control of type II lung epithelial cell differentiation on laminin substrates. However, they may act via distinct mechanisms, including some that are independent of their cell anchoring role.     

Regulation of cell attachment and cell number by fibronectin and laminin

We have examined the effect of laminin and fibronectin on the attachment and growth on type IV collagen of a line of mouse epithelial cells and a strain of adult human fibroblasts. Laminin stimulated attachment of the epidermal cells and fibronectin stimulated fibroblast attachment. At high concentrations (100 micrograms/ml), the attachment proteins altered the growth of cells in culture. The epidermal cells grew better in media containing fibronectin-free serum supplemented with laminin. Fibroblasts, on the other hand, grew best in media containing serum supplemented with fibronectin. These data suggest that laminin promotes epithelial cell growth whereas fibronectin promotes fibroblast growth. This observation was confirmed when these cells were cocultured in the presence of the attachment proteins or of their respective antibodies. The mouse epidermal cells grew best when laminin was added to cocultures of fibroblasts and epithelial cells. Fibroblasts grew best in the presence of antibody to laminin and poorly in the presence of antibody to fibronectin. Thus, fibronectin and laminin may participate in the regulation of cell populations in vivo and may be involved in epithelial-mesenchymal interactions.     

层粘连蛋白促进实体型瘤细胞粘着、铺展及~3H-TdR参人

 恶性肿瘤细胞或粘着于基膜以及在一定的基质上增殖对于肿瘤的浸润转移至关重要。层粘连蛋白(1aminin,LN)是基膜所特有的非胶原糖蛋白。我们研究了LN在肿瘤细胞粘着、铺展及增殖中的作用。通过测定粘着细胞所释放的LDH活性定量分析细胞粘着率。LN可显著增加体外培养的小鼠S180-V肉瘤及B16-MBK黑色素瘤细胞在玻璃及Ⅳ型胶原基质上粘着及铺展。纤维粘连蛋白(fibronectin,FN)亦有类似作用。而非糖蛋白(牛血清清蛋白)或其他糖蛋白(鸡卵清清蛋白或免疫球蛋白)均无类似作用。此外,LN或FN抗体可分别抑制细胞粘着于LN或FN。这些结果表明LN或FN促细胞粘着作用是专一性的。扫描电镜观察表明,粘着在LN敷盖的玻璃表面的瘤细胞呈多突扁平形、膜皱襞及微绒毛十分稀少,而粘着在裸露的玻璃表面者为球形,膜皱襞及微绒毛甚多。再者,~3H-TdR向粘着于LN基质上的瘤细胞群的参入量显著高于粘着在裸露玻璃面者。     

Demonstration of high affinity fibronectin receptors on rat hepatocytes in suspension

A cell-binding peptide (Mr = 85,000) which lacks the gelatin- and heparin-binding domains, was purified from trypsin-digested fibronectin. Preincubation of rat hepatocytes in suspension with the peptide, inhibited initial attachment of the cells to immobilized fibronectin while attachment to immobilized laminin and collagen was unaffected. 125I-labeled 85-kDa peptide bound to the cells in suspension at 4 degrees C in a time-dependent, saturable, and partially reversible reaction. Scatchard analysis of the binding data indicated a single class of receptors with a Kd of 1.7 X 10(-8) M. The number of binding-sites was approximately 2.8 X 10(5)/cell. Unlabeled 85-kDa peptide inhibited the binding of 125I-labeled 85-kDa peptide 30-fold more effectively than intact fibronectin. These results provide direct evidence for the presence of a domain in the fibronectin molecule which has or may acquire a high affinity for receptors involved in adhesion of hepatocytes to immobilized fibronectin.     

Modulation of growth factor induced fiber outgrowth in rat pheochromocytoma (PC12) cells by a fibronectin receptor antibody

Rat pheochromocytoma PC12 cells respond to the binding of nerve growth factor (NGF) and basic fibroblast growth factor (bFGF) by extending neurites in a manner resembling sympathetic neurons. This response requires cell attachment to an appropriate substratum (Fujii et al., J. Neurosci., 2:1157, 1982); attachment factors which function in this capacity include the adhesive proteins fibronectin and laminin. Incubating PC12 cells with a polyclonal antiserum directed against a putative 140-kDa fibroblast cell surface fibronectin receptor (anti-gp140) perturbed spreading but not attachment of the cells to fibronectin and laminin substrates. However, in the presence of anti-gp 140 or its Fab fragments, NGF-stimulated neurite outgrowth was dramatically reduced. The antibody also caused a retraction of previously extended neurites. SDS-PAGE analysis of immunoprecipitates of PC12 cells surface labeled with 125I identified a prominent 120-140-kDa band, suggesting that the site of anti-gp140 action in PC12 cells is also through a fibronectin receptor.     

Sphingosine inhibits attachment of murine Lewis lung carcinoma cells to laminin and type IV collagen

The effect of sphingosine (SPH) on the adhesive properties of Lewis lung carcinoma (3LL) cells was investigated using plastic precoated with the extracellular matrix proteins, laminin, fibronectin, or type IV collagen. Treatment of 3LL cells with SPH (0.5-10 microM) resulted in a dose-dependent decrease in the ability to bind to laminin and type IV collagen but had little or no effect on attachment to fibronectin. Phorbol 12-myristate 13-acetate (PMA) selectively enhanced attachment of 3LL cells to laminin and collagen. The inhibitory effect of SPH on attachment to both proteins was competitively antagonized by PMA. These results suggest that SPH acts as a negative effector for cell attachment to laminin and collagen, and that the cell attachment process to both proteins might be regulated in part by protein kinase C.     

PC12 adhesion and neurite formation on selected substrates are inhibited by some glycosaminoglycans and a fibronectin-derived tetrapeptide

The effects of added soluble glycosaminoglycans (GAGs) on adhesion and neurite formation by cultured PC12 pheochromocytoma cells on several substrates were tested. PC12 cells adhere more rapidly to Petri plastic coated with fibronectin, laminin, poly-L-lysine, or conA, than to either uncoated Petri plastic or tissue culture plastic. Adhesion to poly-L-lysine, fibronectin- and laminin-coated dishes was significantly inhibited by added dextran sulfate and to a lesser extent heparin--but not by chondroitin sulfate. PC12 adhesion to fibronectin could also be totally inhibited by the putative fibronectin cell binding tetrapeptide L-arginyl-glycyl-L-aspartyl-L-serine (Pierschbacher, MD & Ruoslahti, E, Nature 309 (1984) 30). The inhibitory effects of combinations of this tetrapeptide and heparin or dextran sulfate (but not chondroitin sulfate or hyaluronic acid) were additive. Nerve growth factor (NGF) pretreatment increased the percentage of PC12 cells adherent to all substrates and reduced the GAG inhibition of adhesion. PC12 cells previously treated with NGF to induce morphologic differentiation will rapidly re-extend neurites when plated on all four substrates. On fibronectin and poly-L-lysine-coated dishes this neurite growth is inhibited by added heparin and dextran sulfate, while on laminin it is not. Neurite formation on fibronectin-coated dishes was also inhibited by low concentrations of fibronectin tetrapeptide. In summary, PC12 adhesion and neurite formation can be inhibited by sulfated GAGs on some substrates, including fibronectin, but not other substrates, suggesting that these cells have at least two independent molecular adhesion mechanisms.     

Effect of serum,fibronectin, and laminin on adhesion of rabbit intestinal epithelial cells in culture

Rabbit intestinal epithelial cells, obtained after a limited hyaluronidase digestion, were incubated in medium with or without calf serum, on bacteriological plastic dishes. The dishes, either plain or coated with an air-dried type I collagen film, were pretreated with medium alone or with medium containing purified laminin or purified fibronectin. Cells did not attach in significant numbers to untreated bacteriological plastic, even in the presence of serum. Cells did attach to collagen-coated dishes, and were judged viable on the basis of their incorporation of radiolabeled leucine into cell protein. Cell adhesion to the collagen substrate increased in proportion to the concentration of serum in the medium, with maximal attachment at 5% serum or greater. Pretreatment of plain or collagen-coated dishes with increasing amounts of fibronectin enhanced cell adhesion in a concentration-dependent manner. Either serum, or fibronectin-free serum in the medium enhanced cell attachment to substrates pretreated with cither fibronectin or laminin. Thus, intestinal epithelial cells appear to possess surface receptors for both laminin and fibronectin. The evidence further suggests that calf serum may contain factors, other than fibronectin, capable of enhancing intestinal epithelial cell attachment to collagen substrates.     

Collagen can modulate cell interactions with fibronectin

We have examined the effects of soluble collagen on the function of fibronectin in baby hamster kidney (BHK) cells. Collagen and its purified alpha1(l) chain noncompetitively inhibited cell spreading on substrates precoated with fibronectin or a 75,000-D cell-binding fragment of fibronectin. Neither preincubation of cells with collagen followed by washing nor the addition of collagen to previously spread cells had any inhibitory effect on cell spreading, which indicates a requirement for the concurrent presence of collagen during the process of spreading. Treatment of collagen or alpha1(l) chain with collagenase abolished the inhibitory effect on fibronectin-mediated cell spreading. However, direct attachment of BHK cells to fibronectin-coated or 75,000-D fragment-coated substrates was not inhibited by collagen or by the alpha1(l) chain. Moreover, the binding of [3H]fibronectin or the 3'-75,000-D fragment to cell surfaces was not inhibited by the presence of soluble collagen, whereas soluble fibronectin inhibited binding. Although the binding of [3H]fibronectin-coated beads to BHK cell surfaces was also not inhibited by collagen, the phagocytosis of such beads was inhibited by the presence of collagen. On the other hand, soluble fibronectin partially inhibited the binding of fibronectin-coated beads but did not inhibit phagocytosis of the beads that did bind. The mechanism of the inhibition of fibronectin function by collagen and the possible interactions of two different kinds of receptors on the cell surface are discussed.