Migration Tracks Research Articles

Keratinocyte Migration Is Partially Supported by the Cell-Binding Domain of Fibronectin and Is RGDS-Dependent

Fibronectin (FN) plays a key role in cell attachment, embryonic development, and wound healing. In this respect, it is known that FN promotes keratinocyte migration. The aim of this study was to examine specific FN domains (120-kD cell-binding fragment, 45-kD collagen fragment, and 40-kD heparin fragment) and a biologically active peptide within the molecule (RGDS) for their ability to influence human keratinocyte (HK) locomotion. HKs were plated on gold salts coated with different substrates (type IV collagen, FN with or without the RGDS peptide, and the three FN fragments). After 20 h, locomotion tracks were quantified by computer-assisted image analysis that determines the area of each microscopic field occupied by migration tracks, a so-called migration index (MI). MIs on type IV collagen and FN were 39.14 +/- 2.8% and 30 +/- 0.4%, respectively. The maximal MIs on the collagen-binding domain and heparin-binding domain of FN were similar to our negative controls (plastic and albumin): 3 +/- 1%. In contrast, the maximal MI on the cell-binding fragment of FN was 18.45 +/- 2.1%. The effect of the cell-binding domain on keratinocyte motility was found to be dose dependent. Moreover, we could specifically inhibit the FN-driven locomotion using the RGDS sequence contained in the cell-binding fragment. We did not observe a synergistic effect (i.e., a higher MI) when we added the three fragments in a same dish. These results suggest i) that the cell-binding fragment of FN partially supports HK locomotion, ii) that other untested FN domain(s) should act in synergy with the cell-binding fragment to promote keratinocyte locomotion, or alternatively iii) that the FN function of promoting cell migration resides within the FN cell-binding domain, but the proper presentation of this domain to the cell requires an intact, native FN molecule, and iv) that the RGDS sequence is essential for HK movement.

Pleuropulmonary paragonimiasis: radiologic findings in 71 patients.

Pleuropulmonary paragonimiasis is a disease caused by lung flukes characterized by migration of a juvenile worm in the early stage and by formation of cysts around the worm later on. The purpose of this study was to describe the radiologic manifestations of pleuropulmonary paragonimiasis, with special emphasis on the worm cyst and worm migration track. We retrospectively studied 71 patients who had evidence of pleuropulmonary paragonimiasis on chest radiographs (n = 71) and CT scans (n = 17). The diagnosis was based on the detection of eggs or on positive antibody tests. On chest radiographs, 59 patients (83%) had pulmonary lesions and 43 patients (61%) had pleural lesions. Pulmonary findings included patchy air-space consolidation (n = 37) with or without cystic changes; ring shadows (n = 16); and peripheral linear opacities (n = 29), which were more prominent in patients with pleural effusion. Twelve patients (17%) had bilateral pleural effusions or pneumothoraces. On CT scans, round low-attenuation cystic lesions (5-15 mm), filled either with fluid (n = 5) or gas (n = 5), were characteristically seen within the consolidation. Peripheral linear opacities seen on radiographs were suggestive of worm migration tracks on CT scans. CT may reveal an intracystic worm. Although the findings vary depending on the stage of the disease, findings on chest radiographs are usually typical of paragonimiasis. CT provides more specific information about the worm cyst and the worm migration track.

The epidermal growth factor-induced migration of rat liver epithelial cells is associated with a transient inhibition of DNA synthesis

Epidermal growth factor (EGF) is a potent mitogen for most cultured cells and has previously been shown to induce the migration of rat liver epithelial cells. We have now demonstrated that under migration-inducing conditions EGF does not stimulate cell proliferation, but causes instead a transient inhibition of DNA synthesis. Analysis at the single-cell level by [3H]thymidine autoradiography indicated that in 40-50% of the EGF-treated cell population the entry into S phase is delayed. The simultaneous demonstration of migration tracks by laminin immunofluorescence revealed that the transient inhibition of DNA synthesis is not restricted to the migratory cells. The effect is also observed with the stationary subpopulation and appears, therefore, to be independent of the induction of migration. The independence of both processes was further supported by showing that induction of migration by EGF proceeds undisturbed in cells blocked in S phase by aphidicolin. These results indicated that for rat liver epithelial cells the induction of migration by EGF has priority over cell proliferation. The data also emphasize the need for a time-course analysis when studying factors that stimulate or inhibit DNA synthesis or cell proliferation.

Binding and endocytosis of thrombospondin and thrombospondin fragments in endothelial cell cultures analyzed by cuprolinic blue staining, colloidal gold labeling, and silver enhancement techniques.

We investigated the distribution of thrombospondin-specific binding sites and the uptake of thrombospondin-gold conjugates in cultured porcine endothelial cells by light and electron microscopy. Colloidal gold marker and silver enhancement techniques were applied for cytochemical detection of monomeric thrombospondin and fragments of thrombospondin. Thrombospondin binds to granular and fibrillar structures and to sites of cell-cell contact on the cell surface, as indicated by many proteoglycan-cuprolinic blue precipitates. Cell migration tracks on the culture dish bottom are most heavily stained. Labeling of intact thrombospondin and of proteolytic fragments of thrombospondin with colloidal gold followed by silver intensification enables one to detect its binding and uptake in endothelial cells. Binding to the cell surface and uptake of thrombospondin-gold particles was inhibited by heparin but not by hyaluronic acid or chondroitin sulfate. The heparin binding region at the N-terminal end of the thrombospondin molecule proved to be essential for cell surface binding. Gold-conjugated thrombospondin fragments devoid of the heparin binding region were not internalized. After 60 min incubation at 37 degrees C, thrombospondin-gold particles accumulated in the lysosomal compartment close to the nucleus. In the presence of monensin and ammonium chloride, vesicles in this area are swollen and the concentration of particulate marker is reduced. Binding and uptake of thrombospondin by vascular endothelial cells appears to require linkage of the heparin binding region of the thrombospondin molecule to coated pits and heparan sulfate-rich molecules as receptors. Colloidal gold conjugation of thrombospondin fragments proved to be useful for cytochemical characterization of molecular domains.

Immunofluorescent localization of eosinophil granule major basic protein in fatal human cases of Baylisascaris procyonis infection.

We examined eosinophil degranulation in tissues from patients infected with Baylisascaris procyonis as shown by the extracellular deposition of granule major basic protein (MBP). We utilized immunofluorescence to localize MBP in eosinophils and at sites of degranulation to study specimens from 2 fatal cases of B. procyonis infection. Large numbers of intact eosinophils were present in the brain around blood vessels and necrotic migration tracks and in mesenteric granulomata. Extensive extracellular MBP deposition was present in the necrotic migration tracks in the brain and around larvae in the mesenteric granulomata in association with the radiating eosinophilic deposits characteristic of the "Splendore-Hoeppli phenomenon." The Splendore-Hoeppli deposits consist in part of eosinophil granule MBP. Release of the cytotoxic MBP in response to invading larvae may cause tissue damage. Central nervous system tissue damage by cytotoxic eosinophil granule proteins may contribute to the neurologic symptoms of B. procyonis infection.

Liver epithelial cell migration induced by epidermal growth factor or transforming growth factor alpha is associated with changes in the gene expression of secreted proteins.

Rat liver epithelial cells are induced to migrate by epidermal growth factor (EGF) or transforming growth factor alpha (TGF-alpha) in serum-free medium supplemented with insulin. Immunohistological staining of the migration tracks containing laminin and fibronectin has allowed a quantitative analysis of the process. The growth factor-induced migration is relatively slow, but very efficient. Between 24 and 48 h after exposure to EGF (or TGF-alpha), 50 to 70% of the cells have migrated away from their site of initial attachment and spreading. This delayed effect of the interaction of the receptor with its ligands is associated with changes in gene expression, but is not associated with a stimulation of cell proliferation. In serum-free medium supplemented with insulin, the cells secrete six major proteins, as revealed by SDS-polyacrylamide gel electrophoresis. The media of cultures supplemented with insulin plus EGF (or TGF-alpha) contain in addition two new proteins and an increased amount of fibronectin. One secreted protein is synthesized in significantly reduced amounts. The most conspicuously EGF-induced protein (EIP-1; Mr 47,000) is detected within 2 h, depends on the continued presence of the growth factor, and has not been detected as bound to the substratum. The stringent regulation of EIP-1 suggests that this gene product might participate in the modulation of the changes induced by the growth factor. The system is being used for the further analysis of the regulation of gene expression by EGF and of the migration of normal and neoplastically transformed epithelial cells.

Extracellular matrix (ECM) modulates the EGF-induced migration of liver epithelial cells in serum-free, hormone-supplemented medium.

The influence of the extracellular matrix (ECM) glycoproteins collagen IV, laminin (LN), and fibronectin (FN) on the in vitro migration of epithelial cells was studied using the ECM migration track method (4) with preparations immunostained for LN and FN. The locomotion of rat liver epithelial cells stimulated to migrate in serum-free medium by epidermal growth factor (EGF) in the presence of insulin is inhibited by substratum-bound FN. The inhibition is concentration-dependent up to 0.7 microgram of the protein per cm2. Neither LN nor collagen IV decreased the number of migrating cells, indicating that the inhibition is a specific effect of fibronectin. The data also indicate that the FN-mediated inhibition of migration is an additional and not alternative mechanism to the well-established contact inhibition of locomotion (1) which also occurs in liver epithelial cell cultures. The system is being used for a further analysis of the factors that influence migration of normal and neoplastic epithelial cells and the biochemical mechanisms underlying the migration reaction.

Stimulation of growth and migration of liver epithelial cells by insulin and epidermal growth factor in serum-free medium. immunohistological detection of migration tracks

Stimulation of growth and migration of liver epithelial cells by insulin and epidermal growth factor in serum-free medium. immunohistological detection of migration tracks