ABSTRACT
INTRODUCTION: Anxiety sensitivity (AS) as well as negative cognitions about one's ability to quit smoking represent cognitive-affective vulnerabilities implicated in smoking cessation success. However, the extent to which one's perceived sensitivity to anxiety and cessation-related cognitions uniquely and interactively affect acute abstinence outcomes has not been examined. The current study examined the interactive effects of AS and cessation cognitions on percent reductions in cigarettes smoked during the first 24-h of a quit attempt. METHODS: Adult cessation-motivated smokers (n = 64; Mage = 34.21, SD = 11.49) completed a planned quit attempt. AS and cessation cognitions were evaluated prior to quit day. Percent cigarette reduction was assessed by number of cigarettes smoked the day before and during the first 24 h of the quit attempt. RESULTS: Significant interactive effects between AS and cessation cognitions (i.e., expectation of success in quitting, intolerance of withdrawal symptoms, and lack of cognitive coping) were observed. Consistent with hypotheses, individuals reporting higher AS and a greater perceived ability to tolerate withdrawal as well as a greater expectation of success reported larger reductions in cigarettes post quit compared to those who did not endorse these beliefs. Unexpectedly, individuals reporting lower AS who did not endorse the belief that they should be able to tolerate withdrawal discomfort, or a lack of cognitive coping, reported larger reductions compared to those who did endorse this belief. CONCLUSION: AS may interact with specific cessation cognitions. Pre-cessation beliefs that individuals will be successful and be able to tolerate withdrawal symptoms may support cessation efforts.
Subject(s)
Smoking Cessation , Tobacco Products , Adult , Anxiety , Cognition , Humans , SmokingABSTRACT
Pyruvate formate-lyase-activating enzyme (PFL-AE) from Escherichia coli (E. coli) catalyzes the stereospecific abstraction of a hydrogen atom from Gly734 of pyruvate formate-lyase (PFL) in a reaction that is strictly dependent on the cosubstrate S-adenosyl-l-methionine (AdoMet). Although PFL-AE is an iron-dependent enzyme, isolation of the enzyme with its metal center intact has proven difficult due to the oxygen sensitivity and lability of the metal center. We report here the first isolation of PFL-AE under nondenaturing, strictly anaerobic conditions. Iron and sulfide analysis as well as UV-visible, EPR, and resonance Raman data support the presence of a [3Fe-4S](+) cluster in the purified enzyme. The isolated native enzyme, but not apo-enzyme, exhibits a high specific activity (31 U/mg) in the absence of added iron, indicating that the native cluster is necessary and sufficient for enzymatic activity.
Subject(s)
Enzymes/isolation & purification , Iron-Sulfur Proteins/analysis , Acetyltransferases , Electron Spin Resonance Spectroscopy , Enzymes/chemistry , Enzymes/metabolismABSTRACT
A plasmalemmal protein, LBP110, which binds to the alpha1 chain of laminin-1, is acquired by the neural crest-derived precursors of enteric neurons after they colonize the gut. We tested the hypothesis that laminin-1 interacts with LBP110 to promote enteric neuronal development. The effects of laminin-1 on neuronal development were studied in cultures of cells immunoselected from fetal mouse gut (E14-15) with antibodies to LBP110 or p75NTR, a marker for enteric crest-derived cells. No matter which antibody was used, the development of cells expressing neuronal markers was increased three- to fourfold by culturing the cells on a laminin-1-containing substrate. To determine whether this effect of laminin-1 is due to the selective adherence of a neurocompetent subset of precursors, immunoselected cells were permitted to preadhere to poly-D-lysine. Addition of soluble laminin-1 24 h later promoted neuronal but not glial development. The laminin-1-induced increment in neuronal development was abolished both by a peptide containing the sequence of the LBP110-binding domain, IKVAV, and by antibodies to laminin alpha1 that recognize the IKVAV domain. Neither reagent affected the total number of cells. In contrast, the response to laminin-1 was not affected by control peptides, preimmune sera, or antibodies to laminin beta1. Laminin-1 transiently induced the expression of nuclear Fos immunoreactivity; this action was blocked specifically by the IKVAV peptide. These data are consistent with the hypothesis that LBP110 interacts with the IKVAV domain of laminin alpha1 to promote the differentiation of neurons from enteric crest-derived precursors.
Subject(s)
Carrier Proteins/metabolism , Laminin/pharmacology , Nerve Tissue Proteins/metabolism , Neural Crest/cytology , Neurons/cytology , Amino Acid Sequence , Animals , Antibody Specificity , Carrier Proteins/analysis , Carrier Proteins/immunology , Cell Adhesion/physiology , Cell Differentiation/drug effects , Cells, Cultured , Enteric Nervous System/cytology , Enteric Nervous System/embryology , Female , Fetus/cytology , Fetus/immunology , Gene Expression/drug effects , Immunomagnetic Separation , Intestines/chemistry , Intestines/cytology , Intestines/innervation , Laminin/chemistry , Laminin/immunology , Mice , Mice, Inbred Strains , Molecular Sequence Data , Nerve Tissue Proteins/analysis , Nerve Tissue Proteins/immunology , Neurofilament Proteins/analysis , Neurofilament Proteins/immunology , Neurons/chemistry , Neurons/immunology , Peptide Fragments/immunology , Peptide Fragments/pharmacology , Pregnancy , Protein Structure, Tertiary , Proto-Oncogene Proteins c-fos/genetics , Receptor, Nerve Growth Factor , Receptors, Nerve Growth Factor/analysis , Receptors, Nerve Growth Factor/immunology , Solubility , Stem Cells/chemistry , Stem Cells/cytologyABSTRACT
Previous studies show that culturing an immortalized human submandibular gland cell line (HSG) on Matrigel, a basement membrane extract, induces cytodifferentiation. We have further defined this model system and identified factors involved in HSG cell acinar development and cyto-differentiation. Acinar development is marked by cell migration into multi-cellular spherical structures, cell proliferation and apoptosis of the centrally localized cells. In addition, functional differentiation was determined by indirect immunofluorescence and immunoblot analysis for cystatin, a salivary gland acinar cell-specific protein found to be produced by differentiated HSG cells. Matrigel contains multiple extracellular matrix proteins, however, laminin-1 was identified as the major matrix component that induced HSG cell acinar development and cytodifferentiation. Antibodies against specific components of Matrigel and against cell surface adhesion molecules were added to cells in culture to identify components important for HSG cell acinar differentiation. Immunostaining of HSG cell acini identified TGF-beta 2 and beta 3 as the predominant isoforms within the cells. Neutralizing antibodies directed against TGF-beta 3 significantly decreased (P < or = 0.0002) the size of acini formed. These results indicate that multiple components, including laminin-1 and TGF-beta 3, contribute to HSG cell acinar development. This model system will be useful to study acinar differentiation and salivary gland-specific protein expression in vitro.
Subject(s)
Laminin/physiology , Submandibular Gland/cytology , Transforming Growth Factor beta/physiology , Apoptosis , Biocompatible Materials , Carcinoma, Acinar Cell/pathology , Cell Differentiation , Cell Division , Cell Line , Collagen , Cystatins/biosynthesis , Drug Combinations , Humans , ProteoglycansABSTRACT
We performed differential cDNA hybridization using RNA from endothelial cells cultured for 4 hours on either plastic or basement membrane matrix (Matrigel), and identified early genes induced during the morphological differentiation into capillary-like tubes. The mRNA for one clone, thymosin beta 4, was increased 5-fold. Immunostaining localized thymosin beta 4 in vivo in both growing and mature vessels as well as in other tissues. Endothelial cells transfected with thymosin beta 4 showed an increased rate of attachment and spreading on matrix components, and an accelerated rate of tube formation on Matrigel. An antisense oligo to thymosin beta 4 inhibited tube formation on Matrigel. The results suggest that thymosin beta 4 is induced and likely involved in differentiating endothelial cells. Thymosin beta 4 may play a role in vessel formation in vivo.
Subject(s)
Collagen/pharmacology , Endothelium, Vascular/drug effects , Extracellular Matrix , Gene Expression Regulation/drug effects , Laminin/pharmacology , Microfilament Proteins/genetics , Proteoglycans/pharmacology , Thymosin/genetics , Base Sequence , Capillaries/metabolism , Cell Cycle/physiology , Cell Differentiation/genetics , Cloning, Molecular , DNA, Complementary/analysis , Drug Combinations , Endothelium, Vascular/cytology , Endothelium, Vascular/metabolism , Humans , Molecular Sequence Data , TransfectionABSTRACT
The basement membrane protein laminin and the IKVAV-containing sequence from the laminin alpha 1 chain have been found to promote the differentiation of primary neurons and a variety of neural cell lines. We previously reported that a 110-kd IKVAV-binding protein (LBP110) isolated from brain appears to be a member of the beta-amyloid precursor protein (APP) family by immunologic and functional studies, which showed that LBP110/APP is also important in neurite outgrowth (Kibbey et al.: Proc Natl Acad Sci USA 90:10150-10153, 1993). In the preparation of this binding protein, a contaminating IKVAV-binding protein of identical molecular weight, nucleolin, was also identified. Here we have studied the relationship between these binding proteins. We find that nucleolin binds specifically to the IKVAV sequence independently of LBP110/ApP. We have also demonstrated significant levels of nucleolin in mature brain and in differentiating neural cells, suggesting that nucleolin functions not only in cell proliferation and in ribosome biogenesis as was previously reported, but also in the differentiation and maintenance of neural tissue. Our identification of cytoplasmic and cell-surface nucleolin, an IKVAV-binding protein, suggests that this protein may function in signalling by extra-cellular matrix.
Subject(s)
Carrier Proteins/metabolism , Laminin/metabolism , Nerve Tissue Proteins/metabolism , Neurites/physiology , Nuclear Proteins/metabolism , Nucleolus Organizer Region/metabolism , Phosphoproteins/metabolism , RNA-Binding Proteins , Amino Acid Sequence , Animals , Binding Sites , Blotting, Western , Carrier Proteins/chemistry , Epitopes , Immunohistochemistry , Laminin/chemistry , Laminin/physiology , Male , Molecular Sequence Data , Nerve Tissue Proteins/chemistry , Nuclear Proteins/isolation & purification , PC12 Cells , Phosphoproteins/isolation & purification , Protein Binding , Rats , Rats, Inbred F344 , Receptors, Cytoplasmic and Nuclear/metabolism , NucleolinABSTRACT
The laminin-derived synthetic peptide containing the SIKVAV (Ser-Ile-Lys-Val-Ala-Val) amino acid sequence has been previously shown to regulate tumor invasion, metastasis, and angiogenesis. Here, we demonstrate that this peptide also modulates human monocyte responses. Moreover, the monocytic responses elicited by this peptide are influenced by the culture conditions. When elutriated monocytes were cultured on SIKVAV substrate or in suspension with this peptide, the synthesis of prostaglandin E2, interstitial collagenase, and gelatinase B was induced and was further enhanced in the presence of concanavalin A (ConA). However, when monocytes were adhered before adding soluble SIKVAV, the peptide alone failed to induce the production of prostaglandin E2 or matrix metalloproteinases. If adherent monocytes were exposed to SIKVAV in the presence of ConA, this peptide enhanced the ConA induced production of these mediators. In contrast to SIKVAV, the intact laminin molecule failed to influence these monocyte responses. This is the first demonstration that a laminin derived peptide is capable of inducing or enhancing monocyte inflammatory responses that may influence a number of biological activities such as wound healing or excessive connective tissue destruction associated with chronic inflammation.
Subject(s)
Collagenases/biosynthesis , Dinoprostone/metabolism , Laminin/chemistry , Macrophages/metabolism , Monocytes/metabolism , Peptide Fragments/pharmacology , Amino Acid Sequence , Cell Adhesion , Humans , Matrix Metalloproteinase 9 , Molecular Sequence DataABSTRACT
Here we review the role of angiogenesis as it pertains to the interactions between the epithelium and the mesenchyme, especially during tumor growth and metastasis. We illustrate and discuss several models of angiogenesis including endothelial tube formation on Matrigel. Finally, we examine angiogenic factors using the Matrigel model and investigate several other matrix molecules for their importance in angiogenesis and epithelial/stromal interactions.
Subject(s)
Mesoderm/physiology , Neoplasms/blood supply , Neoplasms/pathology , Neovascularization, Pathologic/physiopathology , Animals , Cell Differentiation , Cell Division , Cells, Cultured , Collagen , Drug Combinations , Epithelial Cells , Epithelium/pathology , Epithelium/physiology , Extracellular Matrix/physiology , Growth Substances/physiology , Humans , Laminin/physiology , Mesoderm/pathology , Models, Biological , Neoplasm Metastasis , Neoplasms/physiopathology , Proteoglycans , Tumor Cells, CulturedABSTRACT
Expansion of the tumor-cell mass is dependent on both the degree of tumor vascularization and the rate of angiogenesis. Blood vessel growth is controlled, in part, by the matrix surrounding it, in particular, the basement membrane underlying the endothelium. Here we illustrate that laminin, a major component of basement membrane, has several biologically active sites that can bind to endothelial and tumor cells, and have the ability to regulate angiogenesis and tumor growth. We show that synthetic peptides at two sites in the laminin B1 chain (the RGD and YIGSR sequences) inhibit angiogenesis, whereas a third site in the A chain, designated SIK-VAV, stimulates vessel and tumor cell growth. By developing strategies that promote or inhibit the activities of these sites in laminin, we may obtain methods to inhibit angiogenesis and subsequent tumor growth.
Subject(s)
Basement Membrane/chemistry , Laminin/physiology , Neoplasm Invasiveness/physiopathology , Neovascularization, Pathologic/metabolism , Amino Acid Sequence , Animals , Basement Membrane/physiology , Humans , Laminin/analysis , Molecular Sequence DataABSTRACT
Angiogenesis has been investigated in vivo using subcutaneously injected reconstituted basement membrane (Matrigel) supplemented with angiogenic factors. Previously we found that the laminin-derived synthetic peptide containing SIKVAV (ser-ile-lys-val-ala-val) promoted angiogenesis in vivo. In parallel studies, it was observed that new vessel formation in response to this peptide occurred several days after basic fibroblast growth factor-induced angiogenesis. Since this delay suggested that SIKVAV-induced angiogenesis may be secondary to other events, we investigated here earlier time points to determine if both indirect and direct mechanisms of angiogenesis are involved. We found that neutrophils are continuously recruited to the SIKVAV-containing plugs between 4 hours to 3 days following the initial injection. By day 7, columns of endothelial cells begin to migrate into the plug and form small blood vessels. In contrast, neutropenic mice had a 62% reduction in SIKVAV-induced angiogenesis when compared to control mice. Freshly isolated neutrophils also degraded laminin, the major component of the basement membrane Matrigel. These cells also produced factors in response to SIKVAV peptide which induced proliferation of human umbilical vein endothelial cells relative to a control peptide. In vitro experiments utilizing human neutrophils demonstrated that these cells migrate to the SIKVAV peptide and possess a specific cell surface SIKVAV binding protein of approximately 56 kD. These data suggest that neutrophils are induced to migrate to the Matrigel plugs, at least in part, by SIKVAV peptide, where they may release their own angiogenic factors and degrade the matrix, thus physically facilitating cell migration and liberating additional angiogenic matrix fragments and/or cytokines.
Subject(s)
Laminin/pharmacology , Neovascularization, Pathologic/chemically induced , Neutrophils/cytology , Neutrophils/physiology , Amino Acid Sequence , Cell Division/drug effects , Cell Division/physiology , Cell Movement/drug effects , Cell Movement/physiology , Cells, Cultured , Endothelium, Vascular/cytology , Humans , Laminin/analysis , Molecular Sequence Data , Time FactorsABSTRACT
A large family of bZIP proteins, containing a basic DNA-binding domain and a leucine zipper, have been described that recognize the CRE and AP-1 elements. Here, we have identified two new members, designated LZIP-1 and LZIP-2. The murine cDNA for LZIP-1 coded for a 379-amino-acid (aa) residue protein containing several distinct domains, including a Ser-rich region, a basic DNA-binding region, and an unusually long leucine zipper. A second form, LZIP-2, contained an additional 25 aa in the N-terminal region. Western immunoblotting revealed that antibody raised against part of recombinant LZIP-1 detected both forms in a variety of tissues. Gel mobility shift assays demonstrated that the recombinant protein possessed specific DNA-binding activity for both the CRE AP-1 sites. The present identification of two more ubiquitous members of the bZIP family emphasizes the complex nature of transcription factor interactions at the CRE and AP-1 sites.
Subject(s)
Leucine Zippers/genetics , Transcription Factors/genetics , Animals , Base Sequence , Cell Line , DNA-Binding Proteins/genetics , DNA-Binding Proteins/isolation & purification , DNA-Binding Proteins/metabolism , Fibroblasts , Melanoma, Experimental/chemistry , Melanoma, Experimental/genetics , Mice , Molecular Sequence Data , Transcription Factors/isolation & purification , Transcription Factors/metabolismSubject(s)
Laminin/chemistry , Neoplasms, Experimental/drug therapy , Neoplasms/drug therapy , Neovascularization, Pathologic/prevention & control , Amino Acid Sequence , Animals , Basement Membrane/physiology , Cell Division/drug effects , Cell Division/physiology , Disease Models, Animal , Humans , Molecular Sequence Data , Neoplasms/blood supply , Neoplasms/pathology , Neoplasms, Experimental/blood supply , Neoplasms, Experimental/pathologyABSTRACT
Laminin has been shown to promote the malignant phenotype and the expression of certain laminin receptors has been correlated with the malignant character of the tumors. Here new cell lines were isolated from a human colon cancer cell line (LCC-C1) based on their adhesiveness to laminin. The laminin-adherent subclone formed large tumors in nude mice, whereas the laminin-nonadherent subclone failed to form sizable tumors. Only the laminin-adherent subclone adhered to laminin and invaded through Matrigel-coated filters. The adhesive and invasive ability of the cells was almost completely blocked by low concentrations (1.0 microgram/ml) of anti-beta 1 integrin antibody. The amounts of total cellular beta 1 integrin protein were similar in the two subclones when compared by Western blot, and the mRNA levels also did not differ. The localization of beta 1 integrin laminin receptor varied in the two subclones; the laminin-adherent subclone showed a linear distribution along the cell-cell junctions, while the laminin-nonadherent subclone did not stain between the cells. Using laminin-Sepharose affinity chromatography, more beta 1 integrin was obtained from the laminin-adherent subclone. These findings suggest that alterations in the affinity of beta 1 integrin for laminin and in its membrane distribution might be involved in the increased tumorigenicity observed in colon cancer cells.
Subject(s)
Colonic Neoplasms/pathology , Integrins/physiology , Laminin/metabolism , Amino Acid Sequence , Animals , Blotting, Western , Cell Adhesion/drug effects , Cell Adhesion/physiology , Cell Communication/physiology , Chromatography, Affinity , Collagen , Colonic Neoplasms/metabolism , Drug Combinations , Extracellular Matrix/physiology , Humans , Immunohistochemistry , Integrin beta1 , Integrins/analysis , Laminin/pharmacology , Mice , Mice, Nude , Molecular Sequence Data , Neoplasm Invasiveness , Neoplasm Transplantation , Proteoglycans , Tumor Cells, CulturedABSTRACT
Laminin has been shown to promote the malignant phenotype and the level of the 32/67 Kd laminin receptor has been found to correlate with Dukes' staging of colon cancer. A biopsy of a Dukes' stage B2 human colon carcinoma formed a tumor in a nude mouse after coinjection with Matrigel. The parental tumor and the murine tumor appeared identical at the histological level. A cell line LCC-C1 was established from the murine tumor. The cell line appeared moderately differentiated although it did not produce mucin in vitro; however, the xenograft in vivo did produce low levels of mucin. Laminin adherent and non-adherent cell lines were selected. The parental and the laminin-selected cell subclones adhered equally well to plastic and to fibronectin and showed similar growth rates on plastic. When injected subcutaneously into nude mice, the laminin-adherent cells formed relatively undifferentiated tumors that were twice as large as the parental cell tumors whereas the laminin non adherent cells formed very small, but highly differentiated tumors. These data demonstrate that subpopulations of tumor cells which differ in their tumorigenic properties can be selected based on their adhesion to laminin and thus provide models for studying the mechanisms of tumor growth.
ABSTRACT
We previously characterized a 110-kDa membrane-associated laminin-binding protein (LBP110) from brain which binds the laminin A chain -Ile-Lys-Val-Ala-Val-(IKVAV) site and increases in injury. Here we demonstrate that antisera directed against different epitopes of beta-amyloid precursor protein (APP) recognize LBP110 and that APP is recognized by LBP110 antiserum. APP specifically binds IKVAV and not another biologically active laminin-derived peptide containing the amino acid sequence -Tyr-Ile-Gly-Ser-Arg-. PC-12 cells transfected with antisense APP RNA produce less APP and LBP110, and they form fewer processes when cultured on either laminin or the IKVAV peptide. Thus, LBP110 is a member of the APP family and a function for APP in neurite outgrowth is now defined.
Subject(s)
Amyloid beta-Protein Precursor/metabolism , Laminin/chemistry , Laminin/metabolism , Neurites/physiology , Amino Acid Sequence , Amyloid beta-Protein Precursor/biosynthesis , Animals , Binding Sites , Genetic Vectors , Molecular Sequence Data , Nerve Growth Factors/pharmacology , PC12 Cells , Peptide Fragments/chemistry , Peptide Fragments/metabolism , RNA, Antisense , TransfectionABSTRACT
A 110,000 mol.wt laminin-binding protein from newborn mouse brain recognizes a neurite promoting laminin A chain site and is related to the beta-amyloid precursor protein. In the present study, we examined the expression of 110,000 mol.wt laminin-binding protein in brains of adult mice, rats, and non-human primates. Essentially identical immunoreactivities were observed across species with distinct staining of cortical pyramidal neurons with apical dendrites, cerebellar basket cell axons, hippocampal mossy fibers, and fine labeling of processes throughout the brain. Colocalization of immunoreactivities to 110,000 mol.wt laminin-binding protein and to laminin in neurons of the adult rat brain was observed. Electron microscopy demonstrated that 110,000 mol.wt laminin-binding protein-like immunoreactivity is intracellular and is possibly associated with the neuronal cytoskeleton. Western blot analysis revealed that anti-110,000 mol.wt laminin-binding protein also recognizes a 140,000 mol.wt protein in the pellet, in addition to the 110,000 mol.wt protein in the Triton soluble extract. Antibody fractions specific to the two reactive protein species (110,000 mol.wt and 140,000 mol.wt) exhibited cross-reactivity on immunoblots and revealed similar immunohistochemical staining in adult brain. Results suggest a significant interaction between laminin-like molecules and 110,000 mol.wt laminin-binding protein-like molecules in normal brain function, in response to CNS injury and possibly in the pathogenesis of Alzheimer's disease.
Subject(s)
Brain Chemistry , Brain/ultrastructure , Carrier Proteins/analysis , Nerve Tissue Proteins/analysis , Amino Acid Sequence , Amyloid beta-Protein Precursor , Animals , Animals, Newborn , Blotting, Western , Carrier Proteins/chemistry , Cytoskeleton/chemistry , Female , Fixatives/pharmacology , Macaca mulatta , Male , Mice , Mice, Inbred C57BL , Microscopy, Immunoelectron , Molecular Sequence Data , Molecular Weight , Neoplasm Proteins/chemistry , Nerve Tissue Proteins/chemistry , Neuroglia/chemistry , Neurons/chemistry , Primates , Protein Structure, Tertiary , Rats , Rats, Inbred F344 , Sarcoma, Experimental/chemistry , Species SpecificityABSTRACT
Basement membrane has a variety of effects on tumor cells and promotes malignant behavior. Tumor cell growth is enhanced both in vitro and in vivo in mice in the presence of basement membrane. This has led to the ability to grow various tumors including human biopsy specimens in nude mice. Furthermore, low cell numbers can be used when coinjected with Matrigel, a basement membrane extract. The basement membrane glycoprotein laminin is important in promoting invasive behavior and the level of a 32/67 kDa laminin receptor has been shown to correlate with malignancy. A sequence of five amino acids, tyrosine-isoleucine-glycine-serine-arginine (YIGSR) has been shown to recognize this receptor and to reduce experimental metastases (tail vein injection resulting in colonization of the lung) and subcutaneous tumor growth. This peptide is active in both models either when coinjected or when daily intraperitoneal injections are given after tumor growth has initiated. YIGSR does not effect cell arrest but does inhibit angiogenesis which is necessary for tumor growth. YIGSR also appears to have an additional antitumor effect via its interaction with a specific receptor. YIGSR-adherent cells established after 30 successive selections on YIGSR-coated dishes in vitro formed more lung colonies after intravenous injection and larger tumors after subcutaneous injection than the parent B16F10 melanoma cells. The YIGSR-non-adherent cells formed fewer lung colonies and smaller subcutaneous tumors. These data demonstrate the importance of laminin-tumor cell interactions in malignancies and suggest that a short sequence from laminin has multiple effects in reducing tumor growth and spread.(ABSTRACT TRUNCATED AT 250 WORDS)
Subject(s)
Collagen/physiology , Laminin/physiology , Neoplasms, Experimental/pathology , Oligopeptides/physiology , Proteoglycans/physiology , Amino Acid Sequence , Animals , Basement Membrane/physiology , Cell Division/drug effects , Collagen/chemistry , Drug Combinations , Humans , Laminin/chemistry , Mice , Molecular Sequence Data , Neoplasm Metastasis , Oligopeptides/chemistry , Proteoglycans/chemistry , Receptors, Laminin/physiology , Tumor Cells, CulturedABSTRACT
The distribution of 110/140 laminin-binding protein (110/140 LBP) in the spinal dorsal root ganglia (DRG) and its regulation by partial constriction of the sciatic nerve was studied in adult rats. The cross-sectional area of neurons with 110/140 LBP-immunoreactivity (-I) showed an approximately normal frequency distribution. The 110/140 LBP-I was observed in neuronal cell bodies exclusive of the nucleus. Following sciatic nerve constriction, the 110/140 LBP-I was downregulated in the ipsilateral L4-5 DRG. DRG neurons with a cross-sectional area > or = 1600 microns 2 were preferentially affected. Neonatal capsaicin-treatment, a procedure that selectively destroys a subpopulation of DRG neurons with fine unmyelinated axons, had no effect on the reduction of 110/140 LBP in the DRG induced by sciatic nerve constriction. Western immunoblot analysis confirmed a reduction of 110/140 LBP on the side ipsilateral to the constriction. These results demonstrate a LBP within primary sensory neurons and its suppression by peripheral nerve injury. The data support a role for LBP in the adult nervous system.
Subject(s)
Capsaicin/pharmacology , Carrier Proteins/metabolism , Ganglia, Spinal/metabolism , Receptors, Laminin/metabolism , Sciatic Nerve/injuries , Animals , Animals, Newborn/physiology , Blotting, Western , Carrier Proteins/immunology , Down-Regulation/drug effects , Ganglia, Spinal/anatomy & histology , Ganglia, Spinal/drug effects , Immunohistochemistry , Neurons/drug effects , Neurons/ultrastructure , Rats , Rats, Sprague-Dawley , Receptors, Laminin/immunologyABSTRACT
Culture of the human neoplastic submandibular gland intercalated duct cell line, HSG, on the basement membrane extract Matrigel induces dramatic morphologic changes and cytodifferentiation. Transmission electron microscopy demonstrated an acinar cell phenotype with polarized cells containing a well-developed Golgi apparatus, multiple microvilli-like projections from the apical surfaces into a lumenal-like area, and numerous granule-like organelles. Amylase, an acinar cell marker, was detected by both immunocytochemical and Northern blot analyses. A 50% reduction in [3H]thymidine incorporation by cells cultured on Matrigel, as compared to cells cultured on tissue culture plates, confirmed the differentiated phenotype of the cells. Multiple components of Matrigel appear to contribute to the morphologic differentiation of the HSG cells since antibodies to both laminin and collagen IV, as well as the laminin-derived bioactive peptide containing SIKVAV, have potent inhibitory effects on HSG cell organization on Matrigel. Collectively, these data indicate that culture of HSG cells on Matrigel is a useful model to study salivary gland acinar development.