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N Ratner

Publications and source records attributed to N Ratner.

At least 37 records · Page 2Linked to original sources
PubMed1997

Schwann cells express NDF and SMDF/n-ARIA mRNAs, secrete neuregulin, and show constitutive activation of erbB3 receptors: evidence for a neuregulin autocrine loop.

Cultured Schwann cells secreted low levels (30 pg/ml/1.5 x 10(6) cells) of a 45-kDa neuregulin protein and showed constitutive activation of a neuregulin receptor, Erb-B3, suggesting the existence of an autocrine loop involving neuregulins in Schwann cells. RT-PCR analyses indicated that Schwann cells and fibroblasts in culture produced SMDF/n-ARIA and NDF but not GGF neuregulin messages. Schwann cell and fibroblast neuregulin messages encoded both beta and alpha domains; Schwann cell transcripts encoded only transmembrane neuregulin forms while fibroblast messages encoded transmembrane and secreted forms. SMDF/n-ARIA and NDF messages were also expressed in early postnatal rat sciatic nerve, suggesting a role for neuregulins in peripheral nerve development. An anti-neuregulin antibody inhibited the mitogenic response of Schwann cells to cultured neurons and to extracts of cultured neurons or embryonic brain, consistent with the accepted paracrine role of neuregulins on Schwann cells. Surprisingly, the same antibody inhibited Schwann cell proliferation stimulated by several unrelated mitogens including bFGF, HGF, and TGF-beta1. These data implicate both paracrine and autocrine pathways involving neuregulin form(s) in Schwann cell mitogenic responses.

Animals↗
PubMed1997

Nf1-deficient mouse Schwann cells are angiogenic and invasive and can be induced to hyperproliferate: reversion of some phenotypes by an inhibitor of farnesyl protein transferase.

We have developed a potential model of Schwann cell tumor formation in neurofibromatosis type 1 (NF1). We show that mouse Schwann cells heterozygous or null at Nf1 display angiogenic and invasive properties, mimicking the behavior of Schwann cells from human neurofibromas. Mutations at Nf1 are insufficient to promote Schwann cell hyperplasia. Here we show that Schwann cell hyperplasia can be induced by protein kinase A activation in mutant cells. Removal of serum from the culture medium also stimulates hyperplasia, but only in some mutant cells. After serum removal, clones of hyperproliferating Schwann cells lose contact with axons in vitro, develop growth factor-independent proliferation, and exhibit decreased expression of the cell differentiation marker P0 protein; hyperproliferating cells develop after a 1-week lag in Schwann cells heterozygous at Nf1. The experiments suggest that events subsequent to Nf1 mutations are required for development of Schwann cell hyperplasia. Finally, an anti-Ras farnesyl protein transferase inhibitor greatly diminished both clone formation and hyperproliferation of null mutant cells, but not invasion; farnesyl transferase inhibitors could be useful in treating benign manifestations of NF1.

Alkyl and Aryl Transferases↗
PubMed1995

Schwann cells from neurofibromin deficient mice exhibit activation of p21ras, inhibition of cell proliferation and morphological changes.

Schwann cells are thought to be abnormal in type 1 neurofibromatosis (NF1) and to contribute to the formation of benign and malignant tumors in this disease. To test the role of the NF1 gene product neurofibromin as a Ras-GTPase activating protein in Schwann cells, and to study the effect of the loss of neurofibromin on Schwann cell proliferation, we isolated Schwann cells from mice with targeted disruption of NF1. The properties of these neurofibromin deficient cells were strikingly similar to those of v-ras expressing rat Schwann cells with normal levels of neurofibromin. The similarities included: growth inhibition, noted as a decrease in cell division in response to glial growth factor 2 (GGF2) and of neuronal contact; morphological changes such as the appearance of elaborated processes; and elevated levels of Ras-GTP. Furthermore, Ras-GTP levels in the neurofibromin deficient Schwann cells were consistently elevated in response to GGF2 treatment. In contrast to these results, introduction of v-ras into a Schwannoma cell line (RN22) led to cell transformation. We conclude that neurofibromin functions as a major regulator of Ras-GTP in Schwann cells; however, mutation in NF1 by itself is unlikely to explain the hyperplasia observed in Schwann cell tumors in NF1 disease.

Animals↗
PubMed1995

Myelin glycoprotein P0 is expressed at early stages of chicken and rat embryogenesis.

Although previous studies suggest that P0 is expressed only in myelinating Schwann cells, monoclonal antibody 1E8 reacts with P0, yet also stains early Schwann cell precursors and non-myelinating Schwann cells (Bhattacharyya et al.: Neuron 7:831-844, 1991). We therefore characterized the 1E8 epitope and analyzed P0 mRNA expression during development. Immunoblot analyses of P0 fusion proteins and of deglycosylated P0 indicated that the 1E8 epitope is polypeptide. Northern blot and polymerase chain reaction (PCR) analyses revealed that P0 is encoded by a single mRNA that is expressed in chicken embryos as early as E4 and in rat embryos as early as E14. These data indicate that the antigen recognized by 1E8 in early chicken embryos is P0 and that, during development of both chickens and rats, P0 mRNA is expressed long before myelination.

Animals↗
PubMed1995

Effect of transforming growth factor-beta 1 and -beta 2 on Schwann cell proliferation on neurites.

Mechanisms regulating Schwann cell proliferation during development are unclear. Schwann cell division is known to be driven by an unidentified mitogen present on the surface of axons, but it is not known whether other molecules play a role in regulating this proliferation. Transforming growth factor-beta (TGF-beta) which is found in the developing peripheral nervous system (PNS) and is mitogenic for neuron-free Schwann cells in vitro could be involved. We have investigated the effects of TGF-beta 1, TGF-beta 2 and antibodies to TGF-beta 1 and TGF-beta 2 on axon driven Schwann cell proliferation. Rat embryonic dorsal root ganglion neurons (DRG) neurons and Schwann cells from the sciatic nerve were isolated, purified and recombined in vitro. Confirming earlier reports by others, we observed that TGF-beta 1 and TGF-beta 2 added to the culture medium stimulated the proliferation of Schwann cells in the absence of neurons. However, when added to neuron-Schwann cell co-cultures, TGF beta caused a variable response ranging from no effect to moderate inhibition of Schwann cell proliferation in different experiments. A stimulation of Schwann cell proliferation by TGF beta was never observed in neuron-Schwann cell co-cultures. Antibodies to TGF-beta 1 and TGF-beta 2 did not influence axon driven Schwann cell proliferation. To further determine the role of TGF-beta in Schwann cell proliferation and myelination, we studied Schwann cell proliferation in cultures from mice in which the TGF-beta 1 gene was delected by homologous recombination. Neuron-Schwann cell cultures from wild-type, heterozygous and homozygous mice were used. No differences were observed in either Schwann cell proliferation or myelination between cultures obtained from homozygous mutants and their heterozygous and wild-type controls. These findings suggest that TGF-beta does not function as a part of the mitogenic mechanism presented by neurons to Schwann cells, but that the presence of active TGF beta in the cellular environment might regulate the degree of proliferation induced by neuronal contact.

Animals↗
PubMed1995

Regulation of neurofibromin expression in rat sciatic nerve and cultured Schwann cells.

Loss of function mutations at the NF1 locus may act intrinsically in Schwann cells to cause the formation of benign Schwann cell tumors (neurofibromas) in patients with type 1 neurofibromatosis. To identify contexts in Schwann cells in which such mutations may play an important role, we measured the levels of NF1 mRNA and neurofibromin in rat sciatic nerve during development, after axotomy, and in cultured rat Schwann cells. NF1 mRNA was present in developing sciatic nerve throughout the period of active Schwann cell proliferation and myelination. After nerve transection, no alteration in NF1 message level was detected, but neurofibromin levels increased, as assessed by immunohistochemistry and Western blotting, suggesting that, in vivo, neurofibromin expression in Schwann cells is post-transcriptionally induced during Wallerian degeneration. Cultured rat Schwann cells constitutively expressed NF1 mRNA and neurofibromin. Schwann cell proliferation induced by exposure to serum and forskolin was not associated with changes in NF1 mRNA or neurofibromin expression, whereas Schwann cell proliferation induced by extracts of embryonic brain membranes was associated with increased NF1 message and neurofibromin expression. Thus, Schwann cells, both in vivo and in vitro, express NF1 mRNA constitutively; the expression of NF1 mRNA and neurofibromin is modulated by only some mitogenic stimuli in Schwann cells.

Animals↗
PubMed1995

Neurofibromin expression and astrogliosis in neurofibromatosis (type 1) brains.

Patients with type 1 neurofibromatosis (NF1) have mutations in the gene encoding the protein neurofibromin. Immunocytochemistry on sections of cortex and cerebellum of unaffected and NF1 individuals and wild-type and NF1-deficient mice showed that the distribution of neurofibromin was similar to that reported for rat. However, dystrophic neurofibromin-expressing neurons were found in human but not rodent brain. Intensity of anti-neurofibromin reactivity was reduced in NF1-deficient mice but not in human brains. GFAP was upregulated in three NF1 brains studied by immunocytochemistry; a 4-18-fold increase in GFAP levels was documented by Western blot analysis in three brains. GFAP content/cell and the number of GFAP-immunoreactive astrocytes was increased in NF1 brains as compared to the controls. These results suggest that mutations in the NF1 gene do not grossly alter the pattern of neurofibromin expression, but activation of astrocytes may be common in NF1. Presence of degenerative debris in one of two brains using the cupric silver method suggests that degeneration is not always detectable in NF1 brains.

Adult↗
PubMed1995

Neurofibromin-deficient fibroblasts fail to form perineurium in vitro.

To identify cell type(s) that might contribute to nerve sheath tumors (neurofibromas) in patients with neurofibromatosis type 1, we generated cell cultures containing neurons. Schwann cells and fibroblasts from transgenic mouse embryos in which the type 1 neurofibromatosis gene was disrupted by homologous recombination (Brannan et al. (1994) Genes Development, 8,1019-1029). Normal fascicle formation by perineurial cells failed to occur in the absence of neurofibromin. Fascicles were reduced in number and showed abnormal morphology when normal neurons and Schwann cells were cultured up to 37 days with fibroblasts lacking neurofibromin. Proliferation was increased in a majority of fibroblast cell strains analyzed from embryos lacking neurofibromin. These observations suggest that mutations in the neurofibromatosis type I gene affect fibroblast behavior that might contribute to neurofibroma formation in patients with neurofibromatosis type 1.

Animals↗
PubMed1994

Targeted disruption of the neurofibromatosis type-1 gene leads to developmental abnormalities in heart and various neural crest-derived tissues.

The neurofibromatosis (NF1) gene shows significant homology to mammalian GAP and is an important regulator of the ras signal transduction pathway. To study the function of NF1 in normal development and to try and develop a mouse model of NF1 disease, we have used gene targeting in ES cells to generate mice carrying a null mutation at the mouse Nf1 locus. Although heterozygous mutant mice, aged up to 10 months, likely attributable to a severe malformation of the heart. Interestingly, mutant embryos also display hyperplasia of neural crest-derived sympathetic ganglia. These results identify new roles for NF1 in development and indicate that some of the abnormal growth phenomena observed in NF1 patients can be recapitulated in neurofibromin-deficient mice.

Alleles↗
PubMed1994

Amphoterin (P30, HMG-1) and RIP are early markers of oligodendrocytes in the developing rat spinal cord.

We have determined the cellular localization of the neurite outgrowth-promoting protein, amphoterin (p30), within the developing rat spinal cord in order to gain insight into possible function(s). Both neurons and oligodendrocytes are labelled by anti-amphoterin antibodies in tissue sections. Double labelling confirmed that labelled non-neuronal cells were oligodendrocytes and not astrocytes. White matter cells labelled by anti-amphoterin were first apparent in the spinal cord at embryonic day 18 (E18). The spinal cord white matter was most densely populated by amphoterin immunoreactive oligodendrocytes during the first postnatal week. In the adult spinal cord anti-amphoterin-labelled oligodendrocytes were infrequent. Neuronal staining was not diminished in adult animals. We have identified multiple populations of developing oligodendrocytes in the rat spinal cord. Oligodendrocytes (or precursors) in the presumptive white matter at E20 are elongated, radially oriented cells. Some are amphoterin+ and Rip-, while others are positive for both antigens. At the same age, a small number of process-bearing Rip+ cells are present in the grey matter. These morphologically and antigenically defined populations of cells could reflect different oligodendrocyte lineages, development stages, or different responses to environmental cues. While the function of amphoterin is unknown, the finding that amphoterin is expressed in numerous oligodendrocytes from late embryonic development suggests that amphoterin expression may be important in the early stages of oligodendrocyte maturation. The onset of amphoterin expression, prior to the expression of myelin-specific antigens, makes amphoterin a useful marker for studying oligodendrocyte development.

Animals↗
PubMed1994

Localization of neurofibromin to keratinocytes and melanocytes in developing rat and human skin.

Pigmentation defects are common in the inherited disease type 1 neurofibromatosis (NF1), predicting a role for the NF1 gene product, neurofibromin, in the skin. We used immunohistochemistry to determine the distribution of neurofibromin in normal developing and adult rat skin, normal neonatal and adult human skin, and skin from patients affected with NF1. The distribution of NF1 mRNA in the epidermis was also analyzed by in situ hybridization. NF1 mRNA and neurofibromin are highly enriched in the keratinocytes in the stratum spinosum of the epidermis in the embryonic, but not adult, rat and in both neonatal and adult humans. Immunoelectron microscopic analysis confirmed that neurofibromin is associated with the keratinocyte plasma membrane, particularly adjacent to desmosomes. Neurofibromin is also detectable in human melanocytes. Analysis of skin from NF1 patients showed normal neurofibromin expression in nine of ten hyperpigmented cafe-au-lait macules and in adjacent unaffected skin. We conclude that 1) neurofibromin expression in the perinatal rat closely parallels expression in the human; 2) reduction in detectable neurofibromin cannot be used to distinguish NF1 and normal human skin; and 3) neurofibromin might function in keratinocytes, as well as melanocytes.

Adult↗
PubMed1994

Axons arrest the migration of Schwann cell precursors.

The neural crest gives rise to a variety of cell types including Schwann cells of the peripheral nervous system. Schwann cell precursors begin to differentiate early and migrate along specific pathways in the embryo before associating with nerve trunks. To determine whether motor axons direct the migration of Schwann cell precursors along specific pathways, we tested the effect of ablating the ventral half of the neural tube, which contains motor neuron cell bodies. The ventral neural tube was removed unilaterally from lumbar regions of chicken embryos at stage 17, when neural crest cells are just beginning to migrate and before motor axons have extended out of the neural tube. At several stages after ventral tube ablation, sections of the lumbar region of these embryos were stained with anti-acetylated tubulin to label developing axons, HNK-1 to label migrating neural crest cells and 1E8 to label Schwann cell precursors. In many embryos the ablation of motor neurons was incomplete. The staining patterns in these embryos support the idea that some Schwann cells are derived from the neural tube. In embryos with complete motor neuron ablation, at stage 18, HNK-1-positive neural crest cells had migrated to normal locations in both control and ablated sides of the embryo, suggesting that motor axons or the ventral neural tube are not required for proper migration of neural crest cells. However, by stage 19, cells that were positive for HNK-1 or 1E8 were no longer seen in the region of the ventral root, nor ventral to the ventral root region. Because Schwann cell precursors require neural-derived factors for their survival in vitro, we tested whether neural crest cells that migrate to the region of the ventral root in ventral neural tube-ablated embryos then die. Nile Blue staining for dead and dying cells in ventral neural tube-ablated embryos provided no evidence for cell death at stage 18. These results suggest that motor axons arrest the migration of Schwann cell precursors during neural crest migration.

Animals↗
PubMed1994

Hepatocyte growth factor is a mitogen for Schwann cells and is present in neurofibromas.

To characterize mitogens that might contribute to Schwann cell proliferation during development or in tumors, we tested the ability of hepatocyte growth factor (HGF) to stimulate Schwann cell division in vitro. HGF is a potent mitogen for purified rat Schwann cells; DNA synthesis in rat Schwann cells was stimulated 20-40-fold by 3-10 ng/ml HGF. Rat Schwann cells express c-met mRNA, encoding the HGF receptor, but not HGF mRNA, implying that HGF might act as a paracrine Schwann cell growth factor. HGF-stimulated Schwann cell proliferation differs from that of previously described Schwann cell mitogens in that its activity is abolished by forskolin and is not inhibited or potentiated by addition of transforming growth factor beta (TGF beta) or fibroblast growth factor (FGF). HGF is probably not a component of the axonal signal thought to cause Schwann cell division during development, as anti-HGF neutralizing antibodies failed to block neuron-stimulated Schwann cell proliferation. In contrast, mitogenic activity present in normal human adult nerves and in neurofibromas from patients with type 1 neurofibromatosis analyzed in the absence of forskolin is largely inhibitable by anti-HGF. Thus, HGF is a novel mitogen for Schwann cells in vitro and it is present in Schwann cell tumors, suggesting a potential role for HGF after wounding of peripheral nerves or in tumor growth.

Adolescent↗
PubMed1993

Neuron-Schwann cell signals are conserved across species: purification and characterization of embryonic chicken Schwann cells.

A monoclonal antibody, 1E8, which recognizes the peripheral myelin protein, P0, specific for chicken Schwann cells and their precursors (Bhattacharyya et al., Neuron 7:831-844, 1991), was used to immunoselect Schwann cells from embryonic day 14 (E14) chicken sciatic nerve. When cultured, these immunoselected cells displayed properties characteristic of perinatal rodent Schwann cells, including S100-immunoreactivity and O4 antigen-immunoreactivity. In addition, the purified chicken Schwann cells divided slowly when cultured alone, but when co-cultured with chicken or rat sensory neurons, they bound to axons and proliferated. Proliferation was also stimulated by the addition of bovine brain membrane extracts or chicken brain membranes. The 1E8 monoclonal antibody was also used to test the effect of axonal contact on P0 expression. Chicken Schwann cells purified using the 1E8 monoclonal antibody gradually lost P0 when cultured alone. These cells remained 1E8-negative even after prolonged co-culture with embryonic rat dorsal root ganglion neurons or chicken sensory ganglia. These results demonstrate that chicken Schwann cells behave like rodent Schwann cells in their expression of specific antigens, interactions with axons, and regulation of P0 expression. In addition, chicken Schwann cells respond to neuronal signals from the rat and cow, illustrating the cross-species conservation of these signals.

Animals↗
PubMed1993

Neurofibromin is enriched in the endoplasmic reticulum of CNS neurons.

NF1 patients display CNS abnormalities including learning disabilities, clumsiness, astrocytomas, and abnormalities on magnetic resonance imaging exams. To determine whether the cellular and neuroanatomical distribution of neurofibromin reveals possible function for neurofibromin in the brain, we stained rat brain tissue sections with anti-neurofibromin antibodies. Neurofibromin is highly enriched in large projection neurons, such as cortical and hippocampal pyramidal cells and cerebellar Purkinje cells. Neurofibromin is present in cell bodies and in axons, but is highly enriched in dendrites. Immunoelectron microscopic analysis demonstrates that NF1 is associated with smooth vesiculotubular elements and cisternal stacks and with multivesicular bodies in the cell body and dendrites, but not with the plasma membrane, nucleus, nuclear envelope, Golgi apparatus, mitochondria, or rough endoplasmic reticulum. The preferential localization of neurofibromin to the smooth endoplasmic reticulum, together with evidence that neurofibromin modulates ras GTPase activity, suggests that some, if not all, of the CNS manifestations of NF1 might result from the altered expression of neurofibromin in neurons, perhaps through disruption of Ca2+ signaling, translocation of organelles, or endocytic pathways.

Animals↗
PubMed1992

Abnormal regulation of mammalian p21ras contributes to malignant tumor growth in von Recklinghausen (type 1) neurofibromatosis.

Tumor cell lines derived from malignant schwannomas removed from patients with neurofibromatosis type 1 (NF1) have been examined for the level of expression of NF1 protein. All three NF1 lines examined expressed lower levels of NF1 protein than control cells, and the level in one line was barely detectable. The tumor lines expressed normal levels of p120GAP and p21ras. Although the p21ras proteins isolated from the tumor cells had normal (nonmutant) biochemical properties in vitro, they displayed elevated levels of bound GTP in vivo. The level of total cellular GAP-like activity was reduced in extracts from the tumor line that expresses very little NF1 protein. Introduction of the catalytic region of GAP into this line resulted in morphological reversion and lower in vivo GTP binding by endogenous p21ras. These data implicate NF1 protein as a tumor suppressor gene product that negatively regulates p21ras and define a "positive" growth role for ras activity in NF1 malignancies.

Animals↗
PubMed1992

Development of olfactory nerve glia defined by a monoclonal antibody specific for Schwann cells.

Although there is considerable interest in the possible role of olfactory glia in the pathfinding abilities of olfactory nerve axons, the complete development of these glia in vivo has not been described. Using a specific Schwann cell marker, the 1E8 antibody, we have found that olfactory nerve glia can be identified throughout development. These glia appear to originate in the olfactory placode and migrate initially into the periphery of the olfactory nerve, and later into the center of the nerve. Olfactory nerve glia enter the presumptive olfactory bulb with the olfactory receptor neuron axons and distribute themselves along the edge of the olfactory nerve layer. The fact that olfactory nerve glia are specifically immunostained by the 1E8 monoclonal antibody, which recognizes the Schwann cell-specific protein P0, suggests that these cells more closely resemble Schwann cells than astrocytes or enteric glia. These results support and extend previous findings suggesting that olfactory nerve glia have distinctive developmental and anatomical features which may be important to the regenerative capacity of the olfactory system.

Aging↗