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Biomedical subjects

David H Gutmann

Publications and source records attributed to David H Gutmann.

At least 55 records · Page 3Linked to original sources
PubMed2004

Loss of tuberous sclerosis complex 1 (Tsc1) expression results in increased Rheb/S6K pathway signaling important for astrocyte cell size regulation.

Individuals with tuberous sclerosis complex (TSC) develop central nervous system abnormalities that may reflect astrocyte dysfunction. In an effort to model astrocyte dysfunction in TSC, we generated mice lacking Tsc1 (hamartin) expression in astrocytes and demonstrated that Tsc1-null astrocytes exhibit abnormalities in contact inhibition growth arrest. In this study, we demonstrate that hamartin-deficient astrocytes are also defective in cell size regulation. We show that the increase in Tsc1-null astrocyte size is associated with increased activation of the S6-kinase pathway. In keeping with recent reports that the hamartin/tuberin complex may regulate Rheb and downstream S6K activation, we demonstrate that expression of either Rheb or S6K in primary astrocytes results in increased S6 pathway activation, and that inhibition of Rheb activity in Tsc1-deficient astrocytes using either pharmacologic or genetic strategies markedly reduces S6 activation. Collectively, these observations suggest that TSC inactivation in astrocytes results in defective cell size regulation associated with dysregulated Rheb/mTOR/S6K pathway activity.

Animals↗
PubMed2004

Disruption of 14-3-3 binding does not impair Protein 4.1B growth suppression.

Meningiomas are common central nervous system tumors; however, the mechanisms underlying their pathogenesis are largely unknown. Collaborative studies from our laboratory demonstrated a direct association of 14-3-3 with the meningioma tumor suppressor Protein 4.1B, which was not observed with other members of the Protein 4.1 family, including the NF2 meningioma tumor suppressor, merlin/schwannomin. Given the role of 14-3-3 in the regulation of cell proliferation and apoptosis, we sought to determine the functional significance of 14-3-3 binding to Protein 4.1B growth suppression. Based on comparative binding studies performed with additional members of the Protein 4.1 family, we generated specific missense mutations within the minimal growth suppressor fragment of Protein 4.1B (DAL-1, differentially expressed in adenocarcinoma of the lung). Complementary in vitro GST affinity chromatography and in vivo interaction experiments demonstrated that the F359Y mutation abrogated binding to 14-3-3, but did not impair DAL-1 binding to other known Protein 4.1B interacting proteins. Similar to wild-type DAL-1, the expression of the F359Y DAL-1 14-3-3-binding mutant resulted in reduced Protein 4.1B-deficient IOMM-Lee and CH157-MN meningioma cell line colony formation. Moreover, similar to wild-type DAL-1, the stable expression of the DAL-1 F359Y mutant significantly reduced cell proliferation in independently isolated IOMM-Lee clones, as assessed by thymidine incorporation. Collectively, these results suggest that binding to 14-3-3 is not essential for the growth suppressor function of Protein 4.1B in meningiomas.

14-3-3 Proteins↗
PubMed2004

Expression profiling in tuberous sclerosis complex (TSC) knockout mouse astrocytes to characterize human TSC brain pathology.

Individuals with tuberous sclerosis complex (TSC) exhibit a variety of neurologic abnormalities, including mental retardation, epilepsy, and autism. Examination of human TSC brains demonstrate dysplastic astrocytes and neurons, areas of abnormal neuronal migration (tubers), and hamartomatous growths, termed subependymal nodules, which can progress to subependymal giant cell astrocytomas (SEGA). Previous studies have suggested that these neuropathologic features may result from abnormal neuroglial cell differentiation. In an effort to provide support for this hypothesis and to identify specific markers of aberrant neuroglial cell differentiation in TSC, we employed gene expression profiling on Tsc1 conditional knockout (Tsc1(GFAP)CKO) mouse astrocytes. We identified several transcripts implicated in central nervous system development that are differentially expressed in Tsc1(-/-) astrocytes compared to wild-type astrocytes. We validated the differential expression of select transcripts on the protein level both in primary cultures of Tsc1(-/-) astrocytes and in Tsc1(GFAP)CKO mouse brains. Moreover, we show that these markers are also differentially expressed within cortical tubers, but not in adjacent normal tissue from TSC patient brains. This study provides supportive evidence for a developmental defect in neuroglial cell differentiation relevant to the genesis of TSC nervous system pathology and underscores the utility of mouse modeling for understanding the molecular pathogenesis of human disease.

Animals↗
PubMed2004

Effect of merlin phosphorylation on neurofibromatosis 2 (NF2) gene function.

The neurofibromatosis 2 (NF2) tumor suppressor gene product, merlin, belongs to the ezrin-radixin-moesin (ERM) subgroup of the Protein 4.1 family, which links cell surface glycoproteins to the actin cytoskeleton. Previous studies have suggested that phosphorylation of merlin, similar to other ERM proteins, may regulate its function. To determine whether merlin phosphorylation has functional consequences for merlin suppression of cell growth and motility, we generated doxycycline-regulatable RT4 schwannoma cell lines that inducibly express full-length merlin with mutations at two potential phosphorylation sites (amino-acid residues S518 and T576). Whereas a mutation at S518 that mimics constitutive phosphorylation (S518D) abrogates the ability of merlin to suppress cell growth and motility, the S518A merlin mutant, which mimics nonphosphorylated merlin, functions equivalently to wild-type merlin. Similar mutations involving T576, the analogous phosphorylation site in ERM proteins important for regulating their function, had no effect. In contrast to other functionally inactive missense merlin mutants, the regulated overexpression of S518D merlin resulted in dramatic changes in cell shape and the elaboration of filopodial extensions. These results provide the first direct demonstration that the S518D merlin mutation, which mimics merlin phosphorylation, impairs not only merlin growth and motility suppression but also leads to an acquisition of a novel phenotype previously ascribed to ERM proteins.

Amino Acid Sequence↗
PubMed2004

Functional significance of S6K overexpression in meningioma progression.

One common genetic change in anaplastic meningiomas is amplification of chromosome 17q23 containing the S6 kinase (S6K) gene. We show, for the first time to our knowledge, increased S6K mRNA expression in anaplastic meningiomas compared with benign tumors. To evaluate S6K as a candidate meningioma progression gene, we generated IOMM-Lee human meningioma cell lines overexpressing S6K. Whereas no effect of S6K overexpression on meningioma cell growth, motility, or adhesion was observed in vitro, S6K overexpression resulted in increased tumor size in vivo. Collectively, these results suggest that S6K is functionally important for meningioma progression and may represent a target for future meningioma therapy.

Animals↗
PubMed2004

CD44-independent hepatocyte growth factor/c-Met autocrine loop promotes malignant peripheral nerve sheath tumor cell invasion in vitro.

Malignant peripheral nerve sheath tumors (MPNSTs) are invasive peripheral nerve neoplasms that express both the receptor tyrosine kinase c-Met and its ligand hepatocyte growth factor (HGF). The combined expression of these proteins has been implicated in tumor cell growth and metastasis. However, HGF/c-Met autocrine activity requires the presence of a serine protease, the HGF activator (HGFA), and, in some cells, the CD44 transmembrane glycoprotein. Here, we found that HGFA, HGF, c-Met, and CD44 are coexpressed in MPNSTs but their localization did not correlate with increased cell proliferation. The ST8814 MPNST cell line also expresses all of these proteins, can convert pro-HGF to active HGF, and exhibits constitutive c-Met phosphorylation. Blocking c-Met activity or expression inhibits the invasive behavior of these cells but not their proliferation. Interestingly, although a CD44 splice variant contributes to MPNST cell invasion and interacts with c-Met and HGF in ST8814 cells, it is not required for c-Met activation. These data indicate that an HGF/c-Met autocrine loop can promote MPNST invasion through a CD44-independent mechanism and suggest that c-Met, HGFA, and HGF are potential molecular targets to inhibit MPNST metastasis.

Autocrine Communication↗
PubMed2004

Molecular pathogenesis of meningiomas.

Meningiomas are common central nervous system tumors that originate from the meningeal coverings of the brain and the spinal cord. Most meningiomas are slowly growing benign tumors that histologically correspond to World Health Organization (WHO) grade I. However, certain rare histological variants (clear cell, chordoid, papillary, and rhabdoid), as well as atypical (WHO grade II) and anaplastic (WHO grade III) meningiomas show a more aggressive biological behavior and are clinically associated with a high risk of local recurrence and a less favorable prognosis. This review summarizes the most important features of meningioma pathology and provides an up-to-date overview about the molecular mechanisms involved in meningioma initiation and progression. Current data indicate that meningioma initiation is closely linked to the inactivation of one or more members of the highly conserved protein 4.1 superfamily, including the neurofibromatosis type 2 gene product merlin/schwannomin, protein 4.IB (DAL-1) and protein 4.1R. The genetic alterations in atypical meningiomas are complex and involve losses on 1p, 6q, 10, 14q and 18q, as well as gains on multiple chromosomes. The relevant genes are still unknown. Anaplastic meningiomas show even more complex genetic alterations, including frequent alteration of the CDKN2A, p14ARF, and CDKN2B tumor suppressor genes at 9p21, as well as gene amplification on 17q23. A better understanding of the molecular mechanisms involved in meningioma pathogenesis may not only lead to the identification of novel diagnostic and prognostic marker but will also facilitate the development of new pathogenesis-based therapeutic strategies.

Animals↗
PubMed2004

Glioneuronal tumours in neurofibromatosis type 1: MRI-pathological study.

Neurofibromatosis type 1 (NF1) is an inherited disorder in which affected individuals develop both benign and malignant tumours at an increased frequency. Glioneuronal tumours, such as ganglioglioma and dysembryoplastic neuroepithelial tumour, have been previously reported in patients with NF1. We describe two patients with glioneuronal tumours and typical clinical features of NF1. Molecular analysis of these tumours did not demonstrate loss of the NF1 gene by fluorescence in situ hybridization (FISH) or immunohistochemistry analysis, suggesting they might not be causally associated with gross defects in NF1 expression. Because of the excellent prognosis following the resection of these tumours, it is important to distinguish them from other NF1-associated tumours.

Adult↗
PubMed2004

Loss of tumor suppressor in lung cancer-1 (TSLC1) expression in meningioma correlates with increased malignancy grade and reduced patient survival.

Meningiomas represent the second most common central nervous system tumor affecting adults. Two of the most frequent early events in meningioma tumorigenesis involve loss of expression of the neurofibromatosis 2 (NF2) and 4.1B genes. Recently, 4.1B was shown to interact with the tumor suppressor in lung cancer-1 (TSLC1) protein, prompting us to examine the expression of TSLC1 in meningiomas. We developed specific anti-TSLC1 antibodies to examine TSLC1 expression in normal human leptomeninges, human meningioma cell lines, and human meningiomas of different pathological grades by Western blot (n = 10) and immunohistochemistry (n = 123). Whereas TSLC1 was expressed in normal human leptomeninges by immunohistochemistry, TSLC1 expression was absent in 3 human malignant meningioma cell lines and markedly reduced or absent in 30% of benign meningiomas by Western blot. Restoration of TSLC1 expression in a TSLC1-deficient human meningioma cell line resulted in reduced cell proliferation. In a series of 123 meningiomas (98 adult and 25 pediatric), TSLC1 expression was absent in 48% of benign (WHO grade I), 69% of atypical (grade II), and 85% of anaplastic (grade III) meningiomas. Moreover, TSLC1 loss was associated with decreased patient survival, within the overall group, and in the atypical meningiomas. Collectively, these results suggest that TSLC1 plays an important role in meningioma pathogenesis.

Animals↗
PubMed2004

Recent advances in neurofibromatosis type 1.

PURPOSE OF REVIEW: The past decade, since the identification of the neurofibromatosis type 1 (NF1) gene, has witnessed great advances in our understanding of the role of the NF1 gene in the molecular pathogenesis of NF1-associated clinical abnormalities. The purpose of this review is to highlight recent advances in defining the molecular etiology of nervous system tumors and learning disabilities. RECENT FINDINGS: Neurofibromas and optic pathway gliomas result from NF1 inactivation in Schwann cells and astrocytes, respectively, but other cellular factors contribute to tumorigenesis. In addition, malignant progression of plexiform neurofibromas to malignant peripheral nerve sheath tumors requires additional genetic changes, including increased expression of growth factor receptors, molecules that are involved in tumor invasion and metastasis, and inactivation of critical cell cycle regulators. In addition, specific types of NF1 gene mutation may be associated with an increased risk for malignancy in individuals with NF1. SUMMARY: Research over the past few years has resulted in a detailed understanding of the molecular genetics of benign and malignant tumors affecting individuals with NF1 as well as the development of refined small animal models for these tumors. In addition, clinical studies have begun to define specific subpopulations of patients at risk for cancer and have identified targeted therapies for NF1-associated tumors, based on basic science research advances.

Adult↗
PubMed2004

Meningioma: an update.

PURPOSE OF REVIEW: Recent clinical and molecular research has shed new light on the biology of meningiomas--a common but understudied CNS neoplasm. This review will focus on recent advances and their significance for future research and treatment. RECENT FINDINGS: Meningiomas represent the second most common brain tumor in adults, and while improved diagnostic modalities are available, these tumors remain underreported. Radiosurgery is an effective adjuvant therapy against meningioma; however, no effective chemotherapy exists. In addition to histologic grading and estimates of the extent of resection, biomarkers, such as progesterone receptor, cyclooxygenase 2, S100A5 and ornithine decarboxylase may be useful in predicting tumor recurrence and/or progression potential in patients with meningioma. On the genetic level, cytogenetic losses on chromosomes 1, 7, 10 and 14 and telomerase activation are observed in clinically aggressive meningioma, whereas monosomy 22 is a common early molecular event in tumor formation. Several candidate growth regulatory genes have been identified, including the Neurofibromatosis 2 (NF2), Tumor Suppressor in Lung Cancer-1 (TSLC1), Protein 4.1B, p53/MDM2 and S6-Kinase genes. The roles of these genes in meningioma formation and progression, as well as the clinical implications of these genetic changes, are discussed. SUMMARY: The recent insights into the molecular biology and genetics of meningioma provide new avenues for basic science research aimed at understanding the mechanisms underlying meningioma formation and malignant progression. These advances may be useful in improving our ability to predict clinical outcome and developing targeted therapies to improve outcomes in patients with clinically aggressive meningiomas.

Animals↗
PubMed2004

Gene expression profiling reveals unique molecular subtypes of Neurofibromatosis Type I-associated and sporadic malignant peripheral nerve sheath tumors.

Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive Schwann cell neoplasms that are frequently associated with Type I Neurofibromatosis (NF1) and respond poorly to current therapeutic regimens. To better understand the molecular heterogeneity of these tumors, we performed gene expression profiling on 25 NF1-associated and 17 sporadic MPNSTs using oligonucleotide microarrays representing approximately 8100 unique human gene transcripts. Using several previously reported statistical approaches, we were unable to identify a molecular signature that could reliably distinguish between NF1-associated and sporadic MPNSTs in independent training and test sample sets. However, using an unsupervised clustering approach, we identified an extensive gene expression signature that distinguished 9 of the 42 tumors analyzed. This signature corresponded to relative overexpression of transcripts associated with neuroglial differentiation (NCAM, MBP, L1CAM, P1P) and relative down-regulation of proliferation and growth factor associated transcripts (IGF2, FGFR1, MDK, Ki67). All tumors with this gene expression signature lacked expression of EGFR and all but one tumor were derived from patients with NF1. However, there were no other obvious associations with histological grade, tumor site, metastasis, recurrence, age, or patient survival. We conclude that distinct molecular classes of MPNST exist and that the ability to stratify these tumors based on unique and biologically relevant gene expression profiles may be important for future targeted therapeutics.

Adolescent↗
PubMed2004

Mouse models of tuberous sclerosis complex.

The most devastating complications of tuberous sclerosis complex affect the central nervous system and include epilepsy, mental retardation, autism, and glial tumors. Mutations in one of two genes, TSC1 and TSC2, result in a similar disease phenotype by disrupting the normal interaction of their protein products, hamartin and tuberin, which form a functional signaling complex. Disruption of these genes in the brain results in abnormal cellular differentiation, migration, and proliferation, giving rise to characteristic brain lesions called cortical tubers. Relevant animal models, including conventional and conditional knockout mice, are valuable tools for studying the normal functions of tuberin and hamartin and how disruption of their expression gives rise to the variety of clinical features that characterize tuberous sclerosis complex. In the future, these animals will be invaluable preclinical models for the development of highly specific and efficacious treatments for children affected with tuberous sclerosis complex.

Animals↗
PubMed2003

Optic nerve glioma in mice requires astrocyte Nf1 gene inactivation and Nf1 brain heterozygosity.

Whereas biallelic neurofibromatosis 1 (NF1) inactivation is observed in NF1-associated gliomas, astrocyte-restricted Nf1 conditional knockout mice do not develop gliomas. These observations suggest that NF1 glioma formation requires additional cellular or genetic conditions. To determine the effect of an Nf1 heterozygous brain environment on NF1 glioma formation, we generated Nf1+/- mice lacking Nf1 expression in astrocytes. In contrast to astrocyte-restricted Nf1 conditional knockout mice, Nf1+/- mice lacking Nf1 in astrocytes develop optic nerve gliomas. This mouse model demonstrates that Nf1+/- cells contribute to the pathogenesis of gliomas in NF1 and provides a tool for the preclinical evaluation of potential therapeutic interventions for these tumors.

Animals↗
PubMed2003

Genetic heterogeneity of stably transfected cell lines revealed by expression profiling with oligonucleotide microarrays.

Large-scale gene expression measurements with oligonucleotide microarrays have contributed tremendously to biological research. However, to distinguish between relevant expression changes and falsely identified positives, the source and magnitude of errors must be understood. Here, we report a source of biological variability in microarray experiments with stably transfected cell lines. Mouse embryonic fibroblast (MEF/3T3) and rat schwannoma (RT4) cell lines were generated to provide regulatable schwannomin expression. The expression levels of 29 samples from five different mouse embryonic fibroblast clonal cell lines and 18 samples from 3 RT4 cell lines were monitored with oligonucleotide microarrays. Using hierarchical clustering, we determined that the changes in gene expression induced by schwannomin overexpression were subtle when compared with those detected as a consequence of clonal selection during generation of the cell lines. The hierarchical clustering implies that significant alterations of gene expression were introduced during the transfection and selection processes. A total of 28 genes were identified by Kruskal-Wallis rank test that showed significant variation between clonal lines. Most of them were related to cytoskeletal function and signaling pathways. Based on these analyses, we recommend that replications of experiments with several selected cell lines are necessary to assess biological effects of induced gene expression.

Animals↗
PubMed2003

Optic pathway gliomas in neurofibromatosis type 1: the effect of presenting symptoms on outcome.

Children with neurofibromatosis type 1 (NF1) may present with optic pathway gliomas (OPG) that can progress to visual loss or other neurologic symptoms. These tumors may become evident either as a result of patient signs or symptoms or as an incidental finding on "baseline" neuroimaging studies. In an attempt to determine if there were differences between symptomatic and asymptomatic children with OPG, a retrospective cohort study of ninety children with NF1 and OPG was performed using data from two large NF1 referral centers. Age at diagnosis, presenting symptoms, tumor location, associated features, and clinical response were assessed for children who were initially symptomatic from their OPG (n = 51) and compared to similar data of asymptomatic children whose tumors were incidentally discovered by MRI (n = 39). There were no differences in age at presentation, tumor location, NF1-associated symptoms, or clinical response between the groups. Initially symptomatic children were much more likely to require treatment (OR: 14.8, 95% CI [1.9-116.7]) than those with incidentally discovered, asymptomatic OPG. Although 36% of OPG were diagnosed in children over the age of 6 years, none received prior neuroimaging and only two children had previously normal eye examinations, suggesting that the vast majority of OPG in this group were longstanding, undiagnosed tumors. Based on these findings, we do not advocate "baseline" MRI in children with NF1, but strongly recommend that all children of the age 10 years and younger with NF1 have complete annual ophthalmologic evaluations.

Child↗