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Biomedical subjects

D H Gutmann

Publications and source records attributed to D H Gutmann.

At least 19 recordsLinked to original sources
PubMed2000

Clinic-based study of plexiform neurofibromas in neurofibromatosis 1.

Individuals with neurofibromatosis 1 (NF1) develop both benign and malignant tumors at an increased frequency. One of the most common benign tumors in NF1 is the plexiform neurofibroma. These tumors cause significant morbidity and mortality on account of their propensity to grow and affect adjacent normal tissues. To determine the clinical profile of plexiform neurofibromas in NF1, we conducted a retrospective review of 68 NF1 patients with plexiform neurofibroma. In our series, 44% of tumors were detected by 5 years of age and most were located in the trunk and extremities. Only two patients developed malignant peripheral nerve sheath tumors in their preexisting plexiform neurofibromas. Lastly, we demonstrate that there were no specific clinical features of NF1 associated with the presence of plexiform neurofibroma. These results underscore the importance of careful serial examinations in the evaluation of patients with NF1.

Adolescent↗
PubMed2000

Genetic and cellular defects contributing to benign tumor formation in neurofibromatosis type 1.

Neurofibromatosis type 1 (NF1) is a common inherited cancer predisposition syndrome. The NF1 gene product, neurofibromin, is hypothesized to function as a tumor suppressor and nearly all NF1 patients develop benign peripheral nerve tumors. These neurofibromas presumably arise from NF1 inactivation in S100(+)Schwann cells, but there is no formal proof for this mechanism. We demonstrate that fibro-blasts isolated from neurofibromas carried at least one normal NF1 allele and expressed both NF1 mRNA and protein, whereas the S100(+)cells typically lacked the NF1 transcript. Our findings further indicate that additional molecular events aside from NF1 inactivation in Schwann cells and/or other neural crest derivatives contribute to neurofibroma formation.

Adult↗
PubMed2000

Expression of the tuberous sclerosis complex gene products, hamartin and tuberin, in central nervous system tissues.

Tuberous sclerosis complex (TSC) is a common genetic disorder in which affected individuals can develop mental retardation, developmental brain defects, and seizures. Two genetic loci are responsible for TSC: TSC1 on chromosome 9q and TSC2 on chromosome 16p. Here, we report our analysis of TSC1 (hamartin) and TSC2 (tuberin) protein expression in the central nervous system (CNS). Both tuberin and hamartin are expressed in neurons and astrocytes where they physically interact. In the mouse cerebellum in vivo, tuberin predominantly localizes to the perinuclear region of the Purkinje cell, whereas hamartin is distributed along neuronal or astrocytic processes. In contrast, both hamartin and tuberin demonstrate similar neuronal expression patterns in pure neuronal cultures in vitro. Additionally, hamartin is highly expressed in astrocytes in mixed neuron-glia cultures in vitro, suggesting that hamartin may be important for astrocyte growth control. Unlike tuberin, loss of hamartin expression was not observed in sporadic astrocytomas. These results suggest that tuberin and hamartin may differentially contribute to the CNS pathology in TSC.

Animals↗
PubMed1999

Neurofibromatosis 2 tumor suppressor protein, merlin, forms two functionally important intramolecular associations.

The neurofibromatosis 2 (NF2) tumor suppressor gene product, merlin (schwannomin) forms an intramolecular association that is required for negative growth regulation in vitro and in vivo. In an effort to develop a molecular model for merlin relevant to its tumor suppressor function, we further characterized merlin intramolecular folding. We now demonstrate that merlin forms two intramolecular associations, one between the amino terminal (N-term) domain and the carboxyl terminal (C-term) domain and another within the amino terminal domain (N-term/N-term) itself. The N-term/C-term domain interaction requires contact between residues 302-308 in the N-term and an intact exon 17 (residues 580-595) in the C-term domain. In addition, we demonstrate that the N-term/N-term domain self-interaction is required for N-term/C-term domain association. Lastly, we identify NF2 patient mutations that dramatically reduce each of these interactions in vitro. Based on these findings, we propose a model for merlin folding critical to its ability to function as a tumor suppressor protein.

Blotting, Western↗
PubMed1999

Differential effects of cAMP in neurons and astrocytes. Role of B-raf.

Mitogen-activated protein kinase (MAPK) activation provides cell type-specific signals important for cellular differentiation, proliferation, and survival. Cyclic AMP (cAMP) has divergent effects on MAPK activity depending on whether signaling is through Ras/Raf-1 or Rap1/B-raf. We found that central nervous system-derived neurons, but not astrocytes, express B-raf. In neurons, cAMP activated MAPK in a Rap1/B-raf-dependent manner, while in astrocytes, cAMP decreased MAPK activity. Inhibition of MAPK in neurons decreased neuronal growth factor-mediated survival, and activation of MAPK by cAMP analogues rescued neurons from death. Furthermore, constitutive expression of B-raf in astrocytoma cells increased MAPK activation, as seen in neurons, and enhanced proliferation. These data provide the first experimental evidence that B-raf is the molecular switch which dominantly permits differential cAMP-dependent regulation of MAPK in neurons versus astrocytes, with important implications for both survival and proliferation.

Animals↗
PubMed1999

Haploinsufficiency for the neurofibromatosis 1 (NF1) tumor suppressor results in increased astrocyte proliferation.

Individuals affected with neurofibromatosis 1 (NF1) harbor increased numbers of GFAP-immunoreactive cerebral astrocytes and develop astrocytomas that can lead to blindness and death. Mice heterozygous for a targeted Nf1 mutation (Nf1+/-) were employed as a model for the human disease to evaluate the hypothesis that reduced NF1 protein (neurofibromin) expression may confer a growth advantage for astrocytes, such that inactivation of only one NF1 allele is sufficient for abnormal astrocyte proliferation. Here, we report that Nf17+/- mice have increased numbers of cerebral astrocytes and increased astrocyte proliferation compared to wild-type littermates. Intriguingly, primary Nf1+/- astrocyte cultures failed to demonstrate a cell-autonomous growth advantage unless they were cocultured with C17 neuronal cells. This C17 neuronal cell-induced Nf1+/- increase in proliferation was blocked by MEK inhibition (PD98059), suggesting a p21-ras-dependent effect. Furthermore, mice heterozygous for a targeted mutation in another GAP molecule, p120-GAP, demonstrated no increases in cerebral astrocyte number. These findings suggest that reduced NF1 expression results in a cell context-dependent increase in astrocyte proliferation that may be sufficient for the development of astrocytic growth abnormalities in patients with NF1.

Animals↗
PubMed1999

Intracranial gliomas in neurofibromatosis type 1.

Optic pathway gliomas and brainstem gliomas are the predominant intracranial neoplasms associated with neurofibromatosis type 1 (NF1). Before the past 15 years, studies of optic pathway gliomas in NF1 were hampered by the inaccurate diagnosis of NF1, the unavailability of noninvasive neuroimaging techniques, and the frequent rendering of what would now be considered unnecessary, overly aggressive therapy. When studied systematically, these tumors behave in a much more benign fashion than their counterparts in children who do not have NF1. While they may cause symptoms in as many of 50% of cases, progression to the point where specific intervention is deemed necessary is unusual. Consequently, screening neuroimaging of asymptomatic patients is unwarranted. Because optic pathway tumors universally arise in children younger than 7 years of age, all such children should undergo yearly ophthalmologic evaluations and annual assessments of growth to monitor for signs of precocious puberty. Am. J. Med. Genet. (Semin. Med. Genet.) 89:38-44, 1999.

Brain Neoplasms↗
PubMed1999

Developmental regulation of a neuron-specific neurofibromatosis 1 isoform.

We have identified a protein isoform of the neurofibromatosis 1 (NF1) gene (neurofibromin) containing the alternatively spliced exon 9a that is expressed in forebrain neurons. Exon 9a neurofibromin is localized in the cytoplasm, sediments in a P100 fraction, and is expressed throughout the soma and processes in cortical neurons in vitro. Expression of exon 9a neurofibromin is developmentally regulated, with expression first detected after postnatal day 2. The identification of this novel protein isoform with restricted neuronal expression suggests novel functions for neurofibromin in the postmitotic brain that are perhaps relevant to the learning disabilities observed in children with NF1.

Alternative Splicing↗
PubMed1999

Molecular analysis of malignant triton tumors.

Triton tumors are rare variants of malignant peripheral nerve sheath tumor (MPNST) with muscle differentiation, often seen in patients with neurofibromatosis 1 (NF1). Individuals affected with NF1 harbor mutations in the NF1 tumor suppressor gene and develop neurofibromas and MPNSTs. The NF1 gene is expressed in Schwann cells and its expression is lost in schwannian neoplasms, suggesting a role in malignant development. Separately, there is evidence that p53 suppressor gene mutations are involved in MPNSTs. To determine the role of the NF1 and p53 genes in the development of the malignant Triton tumor we examined 2 such tumors, 1 from a 3-year-old boy without clinical manifestations of NF1 and another from a 24-year-old man with NF1. Histological analysis of these tumors showed both neural and muscle differentiation with S-100 and desmin immunoreactivity, respectively. Reverse transcribed RNA polymerase chain reaction (RT-PCR) of NF1 mRNA showed NF1 expression in the sporadic tumor. Strong nuclear immunoreactivity for p53 was observed throughout the malignant population in both tumors. This was confirmed by loss of heterozygosity for p53 in the non-NF1 patient, suggesting that p53 is involved in both hereditary and sporadic Triton tumors. The finding of preserved NF1 gene expression in the non-NF1-related Triton tumor suggests that different genetic events predispose to the development of this rare neoplasm in sporadic cases.

Adult↗
PubMed1999

Increased expression of the NF2 tumor suppressor gene product, merlin, impairs cell motility, adhesionand spreading.

The neurofibromatosis 2 ( NF2 ) gene product, merlin, is a tumor suppressor protein mutated in schwanno-mas and several other tumors. Merlin, which shares significant homology with the actin-associated proteins ezrin, radixin and moesin (ERM proteins), inhibits cell growth when overexpressed in cell lines. The similarities between merlin and ERM proteins suggest that merlin's growth-regulatory capabilities may be due to alterations in cytoskeletal function. We examined this possibility in rat schwannoma cell lines overexpressing wild-type merlin isoforms and mutant merlin proteins. We found that overexpression of wild-type merlin resulted in transient alterations in F-actin organization, cell spreading and cell attachment. Merlin overexpression also impaired cell motility as measured in an in vitro motility assay. These effects were only observed in cells overexpressing a merlin isoform capable of inhibiting cell growth and not with mutant merlin molecules (NF2 patient mutations) or a merlin splice variant (isoform II) lacking growth-inhibitory activity. These data indicate that merlin may function to maintain normal cytoskeletal organization, and suggest that merlin's influence on cell growth depends on specific cytoskeletal rearrangements.

Animals↗
PubMed1999

Overexpression of bax in human glioma cell lines.

OBJECT: Cells that lose their ability to undergo apoptosis may promote the development of neoplasms and result in resistance to clinical treatment with DNA-damaging modalities such as radio- and chemotherapy. Four established human glioma cell lines that are resistant to apoptosis were transfected with the proapoptotic gene bax and assessed for their sensitivity to a proapoptotic stimulus. METHODS: Two cell lines had a wild-type p53 genotype (U87 and D247MG) and two had mutant p53 genotypes (U138 and U373). Constitutive overexpression of murine bax was achieved in U138 and U373 only, which resulted in an increased sensitivity of these lines to the apoptosis-inducing effect of cytosine arabinoside (ara-C). Multiple attempts to produce constitutive overexpression of bax in U87 and D247MG cells resulted in spontaneous, near-complete cell loss. Vector-only control transfections were successful in all four cell lines. Inducible overexpression of bax was achieved in the U87 cells and elevated levels of BAX were observed as early as 6 hours after gene induction. This overexpression of BAX resulted in the spontaneous induction of apoptosis in these cells. CONCLUSIONS: Overexpression of BAX in four human glioma cell lines resulted in increased sensitivity to apoptosis. In the two lines that had a wild-type p53 genotype, overexpression of BAX produced spontaneous apoptosis. In contrast, the lines that had mutant, nonfunctional P53 did not undergo spontaneous apoptosis, but they were rendered more sensitive to the apoptosis-inducing effect of ara-C. Modulation of BAX expression may be a useful therapeutic modality for gliomas, regardless of p53 genotype.

Animals↗
PubMed1999

Establishment of primary vestibular schwannoma cultures from neurofibromatosis type-2 patients.

Primary cultures were established from vestibular schwannomas of NF2 patients. The cultured tumor cells were selectively amplified by growth factor supplemented medium and characterized by immunocytochemistry. NF2 cDNA was amplified by RT-PCR and mutations were detected by both the non-isotopic RNase cleavage assay and direct DNA sequencing, no detectable wild-type NF2 transcript was found in cDNA from the cultured cells. Distinguishable morphology and growth rate differences have been observed in different passages of the primary cells. The data suggest that a pure schwannoma primary culture can be established and could be very useful in vitro model for further understanding the NF2 gene function in Schwann cells.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1998

Merlin differentially associates with the microtubule and actin cytoskeleton.

The neurofibromatosis 2 (NF2) suppressor gene encodes a protein termed merlin (or schwannomin) with sequence similarity to a family of proteins that link the actin cytoskeleton to cell surface glycoproteins. Members of this ERM family of proteins include ezrin, radixin, and moesin. These proteins contain a carboxyl (C-) terminus actin binding site. In contrast to the ERM proteins, merlin lacks the conventional C-terminal actin binding site, but still localizes to the ruffling edge of plasma membranes. In this study, we investigate the ability of merlin to interact with actin through a nonconventional actin binding domain. We demonstrate for the first time that merlin can associate with polymerized actin in vitro by virtue of an amino (N-) terminal actin binding domain including residues 178-367. Merlin actin binding is not affected by several naturally-occurring NF2 patient mutations or alternatively spliced isoforms. These results suggest that merlin, like other ERM proteins, can directly interact with the actin cytoskeleton. In addition, merlin associates with polymerized microtubules in vitro using a novel microtubule binding region in the N-terminal region of merlin that is masked in the full-length merlin molecule, such that wild-type functional merlin in the "closed" conformation fails to bind polymerized microtubules. These microtubule association results confirm the notion that merlin exists in "open" and "closed" conformations relevant to its function as a negative growth regulator.

Actins↗
PubMed1998

Parallels between tuberous sclerosis complex and neurofibromatosis 1: common threads in the same tapestry.

Neurofibromatosis type 1 (NF1) and tuberous sclerosis complex (TSC) represent two neurocutaneous disorders in which affected individuals develop tumors at an increased frequency. Although the clinical manifestations of these disorders are distinctive, the identification of the genes responsible for these disorders has demonstrated remarkable similarities on a molecular level between the NF1 and TSC tumor suppressor gene products. The NF1 and TSC2 gene products are hypothesized to function as growth regulators by modulating the activities of small GTPase molecules. The overlap between the functions of these tumor suppressor genes has yielded important insights into the molecular pathogenesis underlying each of these disorders and suggested possible pharmacological therapies specifically targeted for affected individuals.

Calcium-Calmodulin-Dependent Protein Kinases↗