Laminin Patterns Research Articles

Detection of extracellular matrix protein (laminin) in experimental arterial anastomoses

To investigate the distribution of Laminin in experimental microvascular surgery. Autografts (4 mm) of the left common carotid artery in 20 Wistar rats were harvested after 4 weeks. The specimens were investigated immunohistochemically for the demonstration and distribution of Laminin. The demonstration and distribution pattern of Laminin was a function of the accuracy of each single suture. In the intima, the organisation and amount of Laminin was directed to the lumen and dependent from the distance and vitality of the vessel segments' cells. The media had in its axis direction the lowest reparative potency to regenerate the continuity break. The adventitia formed a thick cuff around the anastomosis, which was circularly grown through with Laminin-positive vasa vasorum. The marked myointimal hyperplasia in the area around the anastomosis and in the entire autograft was constituted of Laminin-positive extracellular matrix. Our results support the demand for a careful and atraumatic suture technique in microvascular surgery.

Immunohistochemical, Ultrastructural, and Molecular Features of Kindler Syndrome Distinguish It From Dystrophic Epidermolysis Bullosa

Kindler syndrome is a rare, inherited skin disease characterized by acral bullae formation, fusion of fingers and toes, and generalized progressive poikiloderma. The purpose of this study was to clarify the nature of the bullous component of Kindler syndrome and to determine whether this inherited skin disorder represents a variant of dystrophic epidermolysis bullosa or a unique independent clinical entity. Two unrelated patients with Kindler syndrome were studied. Electron microscopy demonstrated marked duplication of the lamina densa, and clefts were observed in areas where the lamina densa was destroyed or obscured. Hemidesmosomes and anchoring fibrils showed normal features. Indirect immunofluorescence revealed normal linear labeling with antibodies against hemidesmosomal components (alpha 6 and beta 4 integrins, BPAG1, and BPAG2) and against anchoring filament components such as uncein, as detected by the 19-DEJ-1 monoclonal antibody. However, antibodies against the 3 respective laminin 5 chains, type IV collagen, and various type VII collagen epitopes (the aminoterminal NC1 domain, the central triple helical collagenous domain, and the carboxyterminal end of the triple helical collagenous domain) revealed a broad reticular staining pattern. Molecular screening of the type VII collagen gene (COL7A1) in the patients and their parents by heteroduplex analysis failed to detect any band shifts indicative of pathologic mutations. These results suggest that the bullous component of Kindler syndrome is distinct from dystrophic epidermolysis bullosa caused by mutations in the type VII collagen gene. Additionally, the differential distribution patterns of uncein and laminin 5 in the patients' skin samples support the hypothesis that uncein and laminin 5 are different molecules.

Immunohistochemical Localization of Laminin and Cytokeratin in Embryonic Alligator Gonads

AbstractGonadal sex differentiation is temperature‐dependent in Alligator mississippiensis; testis differentiation occurs in embryos incubated at 33°C and ovary differentiation occurs in embryos incubated at 30°C. Laminin and cytokeratin were examined immunohistochemically in the gonads of alligator embryos incubated at these temperatures. The aim of this study was to determine whether these structural proteins show the same sex‐specific expression patterns reported for mammalian embryos, and to assess their usefulness as early markers of gonadal differentiation in species with temperature‐dependent sex determination. Laminin delineated enlarged seminiferous cords in differentiating testes from developmental stage 23 to hatching. Laminin distribution was more diffuse and revealed smaller cords of cells in differentiating ovaries. Cytokeratin was also detected in developing gonads of both sexes. Cytokeratin became concentrated in the basal cytoplasm of differentiating Sertoli cells in developing testes. In developing ovaries, prefollicular cells of the ovarian cortex and cell cords in the medulla stained strongly for cytokeratin. Cytokeratin did not show the same basal distribution in female medullary cord cells as seen in the Sertoli cells of testes, however. These sex‐specific patterns of laminin and cytokeratin distribution in embryonic alligator gonads may serve as early markers of sexual differentiation.

Filopodia initiate choices made by sensory neuron growth cones at laminin/fibronectin borders in vitro

Localized expression of environmental cues is thought to provide directional information to migrating neuronal growth cones by enhancing or suppressing axon outgrowth over limited regions. To investigate how such a mechanism may function in vivo, we observed growth cones of embryonic chick dorsal root ganglion neurons at a substratum border between the extracellular matrix components laminin and fibronectin in vitro using time-lapse phase-contrast and interference reflection microscopy. We found that patterns of laminin and fibronectin could locally promote or suppress the direction of growth cone migration. While migrating on either laminin or fibronectin, at least 79% of growth cones changed their rate and/or direction of outgrowth upon contact with the alternative substratum, in a manner suggesting that growth cones were selecting one substratum over the other. Complex changes in growth cone behavior were initiated by filopodial contact with the alternate substratum, suggesting that filopodia were providing intracellular signals to the growth cone. Using interference reflection microscopy, we have found that selection of a substratum is independent of the degree of close contact to the substratum. We conclude that spatially localized ECM components can direct axon outgrowth by mechanisms based on intracellular signaling through growth cone filopodia.

Histogenetic Consideration of Ovarian Sex Cord-stromal Tumors Analyzed by Expression Pattern of Cytokeratins, Vimentin, and Laminin: Correlation Studies with Human Gonads

A total of 30 sex cord-stromal tumors including 9 adult type and 5 juvenile type granulosa cell tumors (GCTs), 4 Sertoli-Leydig cell tumors (SLTs), 1 gynandroblastoma, 5 thecomas, 2 fibromas and 3 sclerosing stromal tumors were immunohistochemically evaluated by means of cytokeratins of different molecular weight, vimentin and laminin with regard to the histogenesis of these tumors and to the embryogenesis of the sex cord and stroma of developing gonads. For comparison, 7 embryonic gonads, 9 fetal and 9 adult ovaries, 14 fetal and 5 postnatal testes, and 1 gonadoblastoma were also examined. The coelomic epithelium of all gonads were positive for both cytokeratins (CAM 5.2 and AE1) and vimentin. In fetal ovaries, the granulosa cells of primordial follicles express low molecular weight cytokeratins only and those cells of more maturing follicles did not express any cytokeratin or vimentin. In adult ovaries, the granulosa cells of primordial follicles coexpressed low molecular weight cytokeratins and vimentin, but those cells of more maturing follicles expressed vimentin only. In fetal testes before 20 weeks gestational age, the Sertoli and Leydig cells did not express any cytokeratins and vimentin. After that time, both cells expressed vimentin only throughout life. The rete ovarii and rete testis from fetal to adult life coexpressed both low molecular weight cytokeratins and vimentin. The rete ovarii in all ages and rete testis in prenatal and childhood ages were surrounded by the laminin-positive basement membrane, however, the rete testis in adult were not. In neoplasia, the GCTs, thecomas, fibromas, and sclerosing stromal tumors expressed vimentin only.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression of collagen α1(IV), laminin and nidogen genes in the embryonic mouse lung: implications for branching morphogenesis

The patterns of laminin A, B1, B2, nidogen and collagen α1(IV) gene expression in the embryonic mouse lung were determined using in situ hybridization histochemistry at a stage when branching morphogenesis is taking place. Collagen α1(IV), laminin B1 and B2 genes were expressed throughout the mesenchyme and epithelium. Nidogen gene expression was uniform throughout the mesenchyme but was not detected in epithelial cells. Laminin A mRNA was localized to cells closely associated with a basement membrane at the epithelial-mesenchymal interface. However, expression of the laminin A gene was limited to the mesenchymal cells in bronchial regions and to epithelial cells in distal terminal lobules. We propose that the pattern of laminin A gene expression in different regions of the developing lung will influence the structure of the basement membrane at the epithelial-mesenchymal interface and thus have a role in branching morphogenesis.

Expression of Laminin and Nidogen Genes during the Postimplantation Development of the Mouse Placenta1

The expression patterns of laminin A, B1, B2, and nidogen genes were identified by in situ hybridization in postimplantation mouse extraembryonic tissues and maternal decidua during the period when the chorioallantoic placenta is established. Laminin and nidogen genes were not coordinately expressed either in the decidua or in trophoblast cells, indicating that these genes are regulated independently in these cell types during the establishment of the placenta. Laminin A mRNA was absent from the decidua except in the outer layer of cells adjacent to the myometrium and in the central decidual zone adjacent to the remnant of the uterine epithelium on Day 9. At this stage laminin B1, B2, and nidogen genes were strongly expressed in these cells and also in other regions of the decidua. Laminin B1 mRNA was present at higher levels in the decidua capsularis than in the decidua basalis, while nidogen mRNA showed highest expression in the decidua basalis. Laminin B2 mRNA was produced uniformly throughout the decidua at very high levels, suggesting that laminin B2 chains may be an important component of the decidual matrix. By Day 11, the nidogen gene was expressed only in endothelial cells lining the maternal blood spaces within the decidua. Laminin B1 and nidogen mRNAs were found at high levels within trophoblast giant cells at all stages, while laminin A mRNA was detected in trophoblast giant cells at later stages and laminin B2 mRNA was not produced in high levels by these cells. The patterns of gene expression show a very high degree of regional specialization, suggesting that the extracellular matrices in different regions of the decidua and extraembryonic membranes are likely to be composed of quite different ratios of laminin and nidogen polypeptides.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential expression of laminin, nidogen and collagen IV genes in the midgestation mouse placenta

The distribution of laminin A, B1, B2, nidogen and collagen alpha 1(IV) mRNA was studied in the 12.5-day mouse placenta and uterus. This was compared to the pattern of laminin, nidogen and collagen IV immunoreactivity in the placenta at this time. High levels of B2 mRNA were distributed throughout the decidual layer, compared to lower levels of laminin B1, nidogen and collagen IV. In contrast, laminin B1 and nidogen mRNA were found at very high levels in the trophoblast giant cells and cytotrophoblast. Laminin B2 was much lower in trophoblast cells than in the decidua. Nidogen mRNA levels were low in the decidual cells, but high in endothelial cells lining the placental blood spaces in the decidual layer. Immunofluorescence staining of the placenta showed colocalization of laminin, nidogen and collagen IV in fetal and maternal layers of the placenta. In the antimesometrial area where the uterine epithelium was reforming laminin B1 and B2 mRNA were uniformly distributed between the epithelium and stroma, whereas nidogen and collagen alpha 1(IV) mRNA were only produced by the stromal cells. In all cell types, apart from parietal endoderm cells forming Reichert's membrane, laminin A chain mRNA was very low or absent. These results demonstrate that laminin and nidogen genes are not coordinately expressed in the midgestation mouse placenta. Comparison of the distribution of these mRNAs with collagen alpha 1(IV) mRNA suggests that different regions of the placenta produce specialized extracellular matrices which may contain different ratios of these polypeptides.

Genes Coding for Basement Membrane Glycoproteins Laminin, Nidogen, and Collagen IV Are Differentially Expressed in the Nervous System and by Epithelial, Endothelial, and Mesenchymal Cells of the Mouse Embryo

The pattern of laminin A, B1, B2, nidogen, and collagen alpha;1(IV) gene expression in the 12.5-day mouse embryo was determined by in situ hybridization. Laminin B1, B2, and collagen α1(IV) mRNAs were present in many epithelial and mesenchymal compartments. Laminin A mRNA had a more restricted distribution, being present in cells closely associated with basement membranes and also in the ependymal layer of the neural tube. Nidogen was not produced by any epithelium, but was abundant in mesenchymal and endothelial cells. These results demonstrate that mesenchymal cells contribute significantly to basement membrane production, and that many cells not associated with typical basement membranes produced high levels of mRNAs coding for basement membrane components. Very few cell types produced all five gene products, and some tissues preferentially expressed only one or two of the five genes. This study shows that basement membranes at the epithelial-mesenchymal interface in the majority of mouse embryonic tissues are assembled from components derived from both cell types, and that heterogeneous matrix structures containing different laminin subunits and/or nidogen are likely to be present in the central nervous system and other tissues of the midgestation mouse embryo.

Effects of heat shock on the pattern of fibronectin and laminin during somitogenesis in Xenopus laevis.

Heat shock causes partial disruption of the segmentation pattern during somitogenesis in Xenopus as well as in other vertebrates. However, Xenopus undergoes a different type of somite formation than that of most vertebrates: Somites are formed by rotation of cell blocks out of the paraxial mesoderm. We attempted to determine whether or not the segmentation disorder following heat shock is caused by an altered pattern of fibronectin and/or laminin, that could then effect the rotation of cell blocks. Therefore, we carried out heat shock experiments and analyzed the distribution of both ECM proteins in correlation to the position of somitic cells. Our results reveal that heat shock causes an incorrect deposition of fibronectin as well as laminin during somite formation. This leads to an intermingling of cells from different segments and to an anchorage of cells at the lateral matrix. Immunoblots show that the defects in the pattern of these ECM proteins do not correlate with a decrease of both proteins. However, immunohistological staining patterns demonstrate that oversized blocks of 20-cell width, instead of the normal ones of about 9-cell width, are separated out of the paraxial mesoderm following heat shock treatment. This indicates that the altered pattern of fibronectin and laminin might be a secondary effect caused by incorrect segregation and detachment of cell blocks during somitogenesis. Since anchorage of somitic cells is mostly affected by the altered distribution of fibronectin and laminin, it is more likely that both ECM proteins function in anchorage of migrating presomitic cells and in maintaining of segment borders rather than in stimulating cell rotation movements.

Laminin and basement membrane-associated microfilaments in wild-type and mutant Drosophila ovarian follicles

The localization of the extracellular matrix glycoprotein laminin was studied using polyclonal anti-laminin antibodies. The laminin patterns of the basement membranes of the muscular epithelial sheath that envelops the ovariole were conspicuously different from those of the basement membrane of the follicular epithelium. In the latter structure laminin was stained in a pattern of parallel stripes oriented perpendicular to the long axis of the follicle; microfilament bundles at the adjacent basal side of the follicle cells have the same orientation. At late vitellogenic stages the orientation of the microfilaments remained the same while the laminin stripes were no longer visible. The orientation of laminin and F-actin was abnormal in follicles of the egg-shape mutant kugel, which produces shorter and thicker eggs than wild-type flies. This phenotype might result from the disturbance of the normal circular microfilament and/or laminin pattern.

Four patterns of laminin-immunoreactive structure in developing rat brain

Laminin has been known to promote neurite outgrowth in culture. We have recently demonstrated that laminin facilitates and guides fiber growth of grafted neurons in the brain. In this study, I report that 4 distinct patterns of laminin immunoreactivity — small and large punctiform laminin, sheath laminin, and somal laminin — were expressed in the brain, each having unique spatial and temporal distributions. They are extensively produced in the developing brain while nerve fibers and vessels are actively growing. Two patterns of laminin, small and large puncta, disappear when the brain matures; the other two, sheath laminin and somal laminin, reduce in intensity but persist through adult life. The small puncta, size less than 1 μm in diameter, frequently accumulated along acellular spaces where future fiber bundles will form, interneuronal spaces and surfaces of neurons where terminal fibers dwell, and around the microvessels where neomicrovessels form. The small puncta, which reportedly appear at the eight-cell stage, were observed in our study at stage E10. They increased in expression from E14 to 19 during the stage when many nerve fibers are navigating to their targets, and subsided in the early postnatal days (P6) when less fiber growth occurs. Large puncta, size about 2 μm in diameter, existed exclusively in the hippocampus. They appeared about E16, and ended P16. The sheath laminin is known to form wrappings on microvessels, the ependymal layer, the choroid plexus, and the surface of the brain. This pattern of laminin attaches to non-neuronal structures which associate with CNS fluid. It was detected at E10 (our youngest group), increased its distribution along with expansion of the associated structure, and persisted throughout the adult life. The somal laminin, associated mainly with the soma of neurons, was the latest appearing laminin pattern (not until E16). It also persisted through adult life in the brain. The somal laminin, which was highly associated with the appearance of neuronal groups in brain nuclei, appeared group by group along with neurons. In summary, laminin attaches with distinct groups of cells in the central nervous system with specific temporal association. This close temporal and spatial association may allow laminin to actively affect nerve growth during brain development. The detailed mapping of laminin distribution is in progress in our lab.

Dendritic reticulum cell-related immunostaining for laminin in follicular and diffuse B-cell lymphomas

We performed a comparative immunohistocytochemical study of the distribution patterns of laminin and follicular dendritic reticulum cells (DRCs) within their follicular microenvironment in both nodular or diffuse B-cell non Hodgkin's lymphomas (NHLs). Twenty nine cases of immunophenotypically diagnosed B-cell NHLs (19 of follicular center cell origin-FCCL- and 10 of the diffuse well differentiated lymphocytic type-WDLL-) and five reactive lymph nodes with follicular hyperplasia were analyzed by immunoperoxidase and immunofluorescence techniques. Serial frozen sections and cytospin preparations were tested either with single antibodies anti laminin and DRC-1, or paired reagents in double labeling immunofluorescence. Our results indicated consistently that within both the reactive germinal centers and the neoplastic nodules of FCCL laminin immunostaining visualized a punctate-granular pattern apart from the linear vascular basement membrane positivity. Double immunofluorescence assay demonstrated that there was a close parallelism between this laminin staining pattern and DRC-1 distribution showing a well developed DRCs meshwork; in the diffuse tumour areas of both FCCL and WDLL, laminin immunoreactivity was found only in those cases in which nests of DRCs were observed. Double immunofluorescence studies performed on cytospin preparations demonstrated that the groups of cells containing DRC-1 positive cells, contained a positivity for laminin, although within the cell the staining for DRC-1 was intense and diffuse, while that for laminin was granular and more sparse. Our results suggested that these laminin and DRC-1 positive reactive sites may be present on the same cells. Since the reduction in number or loss of both DRCs and their related immunostaining for laminin within the microenvironment was consistently associated with a loss of nodularity by lymphoma cells, whereas nodularity in reactive and neoplastic conditions was associated with a rich DRCs meshwork and the related laminin immunostaining, a trapping function of DRCs exercised in the presence of laminin should be considered.

Prognostic significance of laminin in adenocarcinoma of the lung

The distribution of laminin in tumor-associated basement membrane was immunohistochemically investigated in 115 cases of adenocarcinoma of the lung. The distribution of laminin was classified into continuous and discontinuous staining patterns. The incidence of the discontinuous pattern was less in early-stage disease than that in advanced stages (P less than 0.01). In patients with stage I, the incidence of discontinuous patterns was greater in short-term survivors than in long-term survivors (P less than 0.05). By contrast, in patients with stage III, the discontinuous pattern of laminin was frequently seen in both long-term survivors and short-term survivors, with no difference between the two groups. These data suggest that the discontinuous pattern of laminin in tumor-associated basement membrane reflects the spread and dissemination of tumor, hence a close relationship to the prognosis.

Basement membrane components in lymphoid follicles: Immunohistochemical demonstration and retationship to the follicular dendritic cell network

We analyzed the distribution of two basement membrane components, laminin and type IV collagen, in human reactive and neoplastic lymphoid follicles by using immunoenzymatic and immunofluorescence methods on serial frozen sections from 11 reactive lymph nodes, three palatine tonsils, and 11 immunophenotypically defined B cell non-Hodgkin's lymphomas. The patterns of distribution of these two antigens were compared with that of follicular dendritic reticulum cells (DRCs) as visualized by anti-DRC-1 and anti-desmoplakin 1 and 2 antibodies. Immunostaining for laminin and type IV collagen produced overlapping results. Germinal centers contained some vascular and fiber-like linear staining, but the most striking feature was the presence of a large amount of punctate-granular staining. This staining was present throughout the entire area of most of the germinal centers, although often more densely in the central areas. The stronger punctate-granular staining for laminin, type IV collagen, and desmoplakin 1 and 2 corresponded to the more densely stained areas for DRC-1. Double immunofluorescence assay demonstrated that there was a close similarity between the punctate-granular staining pattern of laminin and the dendritic network of DRC-1—positive cells. In the nine B cell lymphomas of follicular center origin in which DRCs were present, although distributed in different patterns, immunostaining for laminin and type IV collagen could be observed only in those areas showing immunoreactivity for DRC-1. The results of this study suggest that the punctate-granular staining pattern for laminin and type IV collagen is specific for the follicle. This finding may be strictly related to the follicular microenvironment because the pattern of distribution displayed by laminin and type IV collagen seems to be spatially related to the staining pattern of DRCs, as visualized by the anti-DRC-1 and desmoplakins antibodies, both in reactive and neoplastic conditions.

Destruction of Basement Membrane and Cell Infiltrates in Oral Lichen Planus

Immunohistochemical staining analysis of oral lichen planus (OLP) using monoclonal antibodies (MoAbs) against the components of basement membrane (BM) and infiltrated cells was performed in 27 cases. Integrity of lining epithelial BM was more strictly elucidated by staining for laminin and type IV collagen than by fibronectin. Staining patterns of laminin were similar to type IV collagen. In the cases examined, continuity of BM was proven by laminin staining in 10 cases, partially destructive BM was seen in 9 cases and complete destruction of BM in 8 cases. Dividing OLP into 20 symptomatic (ulcerative/erosive) and 7 asymptomatic (non-ulcerative/-erosive) cases, intact BM could be observed in only 5 cases of the symptomatic group, while 5 cases of the other group possessed undamaged BM. The difference between the two groups was significant. Stromal and epidermal cell infiltrates, which were mainly composed of T cell subsets with accompanying Leu-M5 and 6-positive cells, were correlated with the condition of BM. Stromal Leu-M5+ and Leu-6+ cell infiltrates were clearly more dominant in laminin-negative (BM-destructed) cases than infiltrates in BM-intact cases. This relation was also found in epidermal infiltrates with the same manner of Leu-4 infiltration. From the above, it seems that the integrity of BM is concerned with cell infiltrates and clinical manifestations of OLP.

Changes in distribution of extracellular matrix proteins during wound repair in corneal endothelium.

The distribution of fibronectin (FN) and laminin (LM) in non-injured and injured rat corneal endothelium in vivo was investigated by light microscopy using immunoperoxidase cytochemistry. In non-injured tissues, both FN and LM have distinct pericellular staining patterns and exhibit some diffuse cytoplasmic staining. After a circular freeze injury, cells migrating into the wound area at 24 hr lack the characteristic pericellular staining observed in non-injured cells but show cytoplasmic staining for both extracellular matrix glycoproteins. Endothelial cells on the periphery of such preparations do not partake in wound repair and retain their pericellular staining patterns. Forty-eight hours after injury, cells have filled in the wound area but are disorganized. They display intracellular FN and LM staining but do not demonstrate any pericellular staining. When observed 10 days after injury, a uniform monolayer has formed but neither FN nor LM is detected pericellularly. By 14 days post injury, endothelial cells in the wound area display pericellular FN patterns but not LM patterns. This may reflect differences in the function of each glycoprotein in maintaining the attachment of the endothelium to Descemet's membrane.

Colocalization of laminin and fibronectin in bovine lens epithelial cells in vitro.

The distribution and organization of the extracellular matrix (ECM) proteins laminin, fibronectin, entactin, and type IV collagen were investigated in primary colonies and secondary cultures of bovine lens epithelial cells using species-specific antisera and indirect immunofluorescence microscopy. Primary cell colonies fixed in formaldehyde and permeabilized with Triton X-100 displayed diffuse intracellular patterns for type IV collagen and entactin, observed to a similar degree throughout the colonies. In contrast, thick bundles of laminin and fibronectin were located on the basal cell surfaces and in between cells in the densely packed center of the colonies, and as "adhesive plaques" and fine extracellular matrix cords in the sparsely populated (migratory) outer edge of the colonies. The distribution of ECM proteins observed in secondary lens epithelial cell cultures was similar to that observed at the periphery of the primary colony. Extraction of the secondary cell cultures with sodium deoxycholate confirmed that laminin and fibronectin were deposited on the basal cell surface. Indeed, the patterns of laminin and fibronectin deposition suggested that these proteins codistribute. These results establish that lens epithelial cells in culture can be used as a model system to study the synthesis and extracellular deposition of the basement membrane proteins, laminin and fibronectin.

Response of sensory neurites and growth cones to patterned substrata of laminin and fibronectin in vitro

During neurite elongation in the developing peripheral nervous system, the distribution of laminin and fibronectin may provide preferred substrates for neurite elongation. In this study, the response of sensory neurites and growth cones to patterns of laminin or fibronectin applied to a background substrate of Type IV collagen was studied to determine any possible substrate preference. Neurites exhibited elongation restricted to a laminin pattern, but not a fibronectin pattern, indicating that sensory neurites prefer to elongate on laminin compared to Type IV collagen. When polylysine is included in the background substrate, neurite preference for laminin is decreased. Laminin also enhances neurite elongation and defasciculation and stabilizes growth cone protrusions. These results suggest an adhesive as well as a cytoskeletal involvement in the response to laminin, but direct adhesion estimates indicate that laminin decreases overall adhesion, arguing against an adhesive involvement. Regardless of the mechanism involved, the observed neurite preference for laminin is consistent with the hypothesis that spatial and temporal laminin distributions provide preferred pathways for peripheral neurite elongation.

Neurofilament phosphorylation in peripheral nerve regeneration

A monoclonal antibody to the 200 kdalton neurofilament (NF) polypeptide selectively decorated axons in tissue sections. Neuronal perikarya and dendrites, including motor and sensory neurons reacting to axotomy, were not stained. Axonal staining was abolished by dilution of the monoclonal supernatants with phosphate buffer and by digestion of tissue sections with phosphatase, thus suggesting that the antibody reacted with a phosphorylated epitope 24. Conventional monoclonal and polyclonal antibodies, i.e. antibodies decorating NF regardless of their location (axons, perikarya and dendrites) were not affected by these procedures. Compared to conventional NF antibodies, staining with the axon-specific monoclonal antibody was a late event in peripheral nerve regeneration. One week after operation, the whole distal stump of crushed rat sciatic nerve was invaded by bundles of axons strongly reacting with conventional NF antibodies. Axon-specific NF immunoreactivity was confined to the proximal segment of the stump at this time and progressively extended distally in the following week. Furthermore, NF phosphorylation appeared to coincide with the return of a normal nerve structure as evidenced by the distribution of laminin immunoreactivity. Bundles of axons growing within columns of laminin-positive Schwann cells did not stain with the axon-specific NF antibody. Immunoreactivity with this antibody coincided with the return of a normal laminin pattern, i.e. selective decoration of the endoneurial basal membranes surrounding the axons.