Asynchrony in Peritumoral Resting-State Blood Oxygen Level-Dependent fMRI Predicts Meningioma Grade and Invasion.

BACKGROUND AND PURPOSE: Meningioma grade is determined by histologic analysis, with detectable brain invasion resulting in a diagnosis of grade II or III tumor. However, tissue undersampling is a common problem, and invasive parts of the tumor can be missed, resulting in the incorrect assignment of a lower grade. Radiographic biomarkers may be able to improve the diagnosis of grade and identify targets for biopsy. Prior work in patients with gliomas has shown that the resting-state blood oxygen level-dependent fMRI signal within these tumors is not synchronous with normal brain. We hypothesized that blood oxygen level-dependent asynchrony, a functional marker of vascular dysregulation, could predict meningioma grade. MATERIALS AND METHODS: We identified 25 patients with grade I and 11 patients with grade II or III meningiomas. Blood oxygen level-dependent time-series were extracted from the tumor and the radiographically normal control hemisphere and were included as predictors in a multiple linear regression to generate a blood oxygen level-dependent asynchrony map, in which negative values signify synchronous and positive values signify asynchronous activity relative to healthy brain. Masks of blood oxygen level-dependent asynchrony were created for each patient, and the fraction of the mask that extended beyond the contrast-enhancing tumor was computed. RESULTS: The spatial extent of blood oxygen level-dependent asynchrony was greater in high (grades II and III) than in low (I) grade tumors (P < 0.001) and could discriminate grade with high accuracy (area under the curve = 0.88). CONCLUSIONS: Blood oxygen level-dependent asynchrony radiographically discriminates meningioma grade and may provide targets for biopsy collection to aid in histologic diagnosis.

Local Glioma Cells Are Associated with Vascular Dysregulation.

BACKGROUND AND PURPOSE: Malignant glioma is a highly infiltrative malignancy that causes variable disruptions to the structure and function of the cerebrovasculature. While many of these structural disruptions have known correlative histopathologic alterations, the mechanisms underlying vascular dysfunction identified by resting-state blood oxygen level-dependent imaging are not yet known. The purpose of this study was to characterize the alterations that correlate with a blood oxygen level-dependent biomarker of vascular dysregulation. MATERIALS AND METHODS: Thirty-two stereotactically localized biopsies were obtained from contrast-enhancing (n = 16) and nonenhancing (n = 16) regions during open surgical resection of malignant glioma in 17 patients. Preoperative resting-state blood oxygen level-dependent fMRI was used to evaluate the relationships between radiographic and histopathologic characteristics. Signal intensity for a blood oxygen level-dependent biomarker was compared with scores of tumor infiltration and microvascular proliferation as well as total cell and neuronal density. RESULTS: Biopsies corresponded to a range of blood oxygen level-dependent signals, ranging from relatively normal (z = -4.79) to markedly abnormal (z = 8.84). Total cell density was directly related to blood oxygen level-dependent signal abnormality (P = .013, R2 = 0.19), while the neuronal labeling index was inversely related to blood oxygen level-dependent signal abnormality (P = .016, R2 = 0.21). The blood oxygen level-dependent signal abnormality was also related to tumor infiltration (P = .014) and microvascular proliferation (P = .045). CONCLUSIONS: The relationship between local, neoplastic characteristics and a blood oxygen level-dependent biomarker of vascular function suggests that local effects of glioma cell infiltration contribute to vascular dysregulation.

A Multiparametric Model for Mapping Cellularity in Glioblastoma Using Radiographically Localized Biopsies.

BACKGROUND AND PURPOSE: The complex MR imaging appearance of glioblastoma is a function of underlying histopathologic heterogeneity. A better understanding of these correlations, particularly the influence of infiltrating glioma cells and vasogenic edema on T2 and diffusivity signal in nonenhancing areas, has important implications in the management of these patients. With localized biopsies, the objective of this study was to generate a model capable of predicting cellularity at each voxel within an entire tumor volume as a function of signal intensity, thus providing a means of quantifying tumor infiltration into surrounding brain tissue. MATERIALS AND METHODS: Ninety-one localized biopsies were obtained from 36 patients with glioblastoma. Signal intensities corresponding to these samples were derived from T1-postcontrast subtraction, T2-FLAIR, and ADC sequences by using an automated coregistration algorithm. Cell density was calculated for each specimen by using an automated cell-counting algorithm. Signal intensity was plotted against cell density for each MR image. RESULTS: T2-FLAIR (r = -0.61) and ADC (r = -0.63) sequences were inversely correlated with cell density. T1-postcontrast (r = 0.69) subtraction was directly correlated with cell density. Combining these relationships yielded a multiparametric model with improved correlation (r = 0.74), suggesting that each sequence offers different and complementary information. CONCLUSIONS: Using localized biopsies, we have generated a model that illustrates a quantitative and significant relationship between MR signal and cell density. Projecting this relationship over the entire tumor volume allows mapping of the intratumoral heterogeneity in both the contrast-enhancing tumor core and nonenhancing margins of glioblastoma and may be used to guide extended surgical resection, localized biopsies, and radiation field mapping.

Atypical pleomorphic neoplasms of the pineal gland: Case report and review of the literature.

BACKGROUND: Pineal region tumors are rare and diverse. Among them exist reports of pleomorphic xanthroastrocytoma (PXA) and pleomorphic granular cell astrocytoma (PGCA) of the pineal gland. These related tumors are remarkably similar sharing pleomorphic histologic features with only minor immunohistochemical and ultrastructural differences. CASE DESCRIPTION: We present a case of a 42-year old right-handed woman presented with a longstanding history of migraine headaches which had worsened over the two months leading up to her hospitalization. MRI revealed a 1.7 × 1.3 × 1.6 cm intensely enhancing lesion originating in the pineal gland. The tumor closely resembled PGCA but did not strictly fit the diagnostic requirements of either PGCA or PXA. CONCLUSION: The present case highlights the exotic nature of pineal region tumors with pleomorphic cell histology. Given the diverse range of tumors encountered in the pineal region, pathological confirmation is mandatory. Favorable clinical outcomes demonstrate that surgical resection alone can yield excellent long-term results for tumors falling within the spectrum of pleomorphic lesions of the pineal gland.

Roles of HAUSP-mediated p53 regulation in central nervous system development.

The deubiquitinase HAUSP (herpesvirus-associated ubiquitin-specific protease; also called USP7) has a critical role in regulating the p53-Mdm2 (murine double minute 2) pathway. By using the conventional knockout approach, we previously showed that hausp inactivation leads to early embryonic lethality. To fully understand the physiological functions of hausp, we have generated mice lacking hausp specifically in the brain and examined the impacts of this manipulation on brain development. We found that deletion of hausp in neural cells resulted in neonatal lethality. The brains from these mice displayed hypoplasia and deficiencies in development, which were mainly caused by p53-mediated apoptosis. Detailed analysis also showed an increase of both p53 levels and p53-dependent transcriptional activation in hausp knockout brains. Notably, neural cell survival and brain development of hausp-mutant mice can largely be restored in the p53-null background. Nevertheless, in contrast to the case of mdm2- and mdm4 (murine double minute 4)-mutant mice, inactivation of p53 failed to completely rescue the neonatal lethality of these hausp-mutant mice. These results indicate that HAUSP-mediated p53 regulation is crucial for brain development, and also suggest that both the p53-dependent and the p53-independent functions of HAUSP contribute to the neonatal lethality of hausp-mutant mice.

PDGF stimulates the massive expansion of glial progenitors in the neonatal forebrain.

Platelet-derived growth factor (PDGF) plays a major role in regulating migration, proliferation, and differentiation of glial progenitors during normal brain development and in the abnormal proliferation and dispersion that drives the formation of malignant gliomas. To further explore the relationship between PDGF's effects on normal glial progenitors and its role in the formation of gliomas, we infected progenitor cells in the subventricular zone (SVZ) of the lateral ventricle of neonatal rat pups with a retrovirus that expresses PDGF and green fluorescent protein (GFP). At 3 days post-injection (dpi), a proliferation of PDGFRalpha+ progenitors was seen in the SVZ and white matter around the injection site and by 10 dpi the animals had large diffusely infiltrating tumors that resembled glioblastomas. The tumors contained a massive proliferation of both infected and uninfected PDGFRalpha+ progenitors, suggesting that PDGF was driving tumor formation via both autocrine and paracrine signaling. Rats co-injected with two retroviruses (one that expresses PDGF-IRES-DSRED and one that expresses only GFP) formed tumors that contained a mixture of DSRED+ cells (PDGF producers) and GFP+ cells (recruited progenitors). Time-lapse microscopy of slice cultures confirmed that both DSRED+ and GFP+ cells were highly migratory and proliferative. Furthermore, adding exogenous PDGF to slice cultures generated from nontumor-bearing brains (injected with control GFP retrovirus only) stimulated the migration and proliferation of GFP+ progenitors. These findings reveal the inherent growth factor responsiveness and tumorigenic potential of PDGFRalpha+ progenitors and highlight the importance of paracrine signaling in stimulating glioma growth and infiltration.

Selective destruction of glioblastoma cells by interference with the activity or expression of ATF5.

Glioblastoma multifome is the most common and most aggressive primary brain tumor with no current curative therapy. We found expression of the bZip transcription factor ATF5 in all 29 human glioblastomas and eight human and rat glioma cell lines assessed. ATF5 is not detectably expressed by mature brain neurons and astrocytes, but is expressed by reactive astrocytes. Interference with ATF5 function or expression in all glioma cell lines tested causes marked apoptotic cell death. In contrast, such manipulations do not affect survival of ATF5-expressing cultured astrocytes or of several other cell types that express this protein. In a proof-of-principle experiment, retroviral delivery of a function-blocking mutant form of ATF5 into a rat glioma model evokes death of the infected tumor cells, but not of infected brain cells outside the tumors. The widespread expression of ATF5 in glioblastomas and the selective effect of interference with ATF5 function/expression on their survival suggest that ATF5 may be an attractive target for therapeutic intervention in such tumors.

Expression of a truncated receptor protein tyrosine phosphatase kappa in the brain of an adult transgenic mouse.

Receptor protein tyrosine phosphatases (RPTPs) comprise a family of proteins that feature intracellular phosphatase domains and an ectodomain with putative ligand-binding motifs. Several RPTPs are expressed in the brain, including RPTP-kappa which participates in homophilic cell-cell interactions in vitro [Y.-P. Jiang, H. Wang, P. D'Eustachio, J.M. Musacchio, J. Schlessinger, J. Sap, Cloning and characterization of R-PTP-kappa, a new member of the receptor protein tyrosine phosphatase family with a proteolytically cleaved cellular adhesion molecule-like extracellular region, Mol. Cell. Biol. 13 (1993) 2942-2951; J. Sap, Y.-P. Jiang, D. Friedlander, M. Grumet, J. Schlessinger, Receptor tyrosine phosphatase R-PTP-kappa mediates homophilic binding, Mol. Cell. Biol. 14 (1994) 1-9]. The homology of RPTP-kappa's ectodomain to neural cell adhesion molecules indicates potential roles in developmental processes such as axonal growth and target recognition, as has been demonstrated for certain Drosophila RPTPs. The brain distribution of RPTP-kappa-expressing cells has not been determined, however. In a gene-trap mouse model with a beta-gal+neo (beta-geo) insertion in the endogenous RPTP-kappa gene, the consequent loss of RPTP-kappa's enzymatic activity does not produce any obvious phenotypic defects [W.C. Skarnes, J.E. Moss, S.M. Hurtley, R.S.P. Beddington, Capturing genes encoding membrane and secreted proteins important for mouse development, Proc. Natl. Acad. Sci. U.S.A. 92 (1995) 6592-6596]. Nevertheless, since the transgene's expression is driven by the endogenous RPTP-kappa promoter, distribution of the truncated RPTP-kappa/beta-geo fusion protein should reflect the regional and cellular expression of wild-type RPTP-kappa, and thus may identify sites where RPTP-kappa is important. Towards that goal, we have used this mouse model to map the distribution of the truncated RPTP-kappa/beta-geo fusion protein in the adult mouse brain using beta-galactosidase as a marker enzyme. Visualization of the beta-galactosidase activity revealed a non-random pattern of expression, and identified cells throughout the CNS that display RPTP-kappa promoter activity. Several neural systems highly expressed the transgene-most notably cortical, olfactory, hippocampal, hypothalamic, amygdaloid and visual structures. These well-characterized brain regions may provide a basis for future studies of RPTP-kappa function.

GGF/neuregulin induces a phenotypic reversion of oligodendrocytes.

We have previously shown that glial growth factor (GGF), a member of the neuregulin (NRG) family of growth factors, is a mitogen and survival factor for oligodendrocyte progenitors in cell culture and blocks their differentiation at the pro-oligodendrocyte stage (P. D. Canoll et al., 1996, Neuron 17, 229-243). We now show that GGF is able to induce differentiated oligodendrocytes to undergo a phenotypic reversion characterized by loss of MBP expression, reexpression of the intermediate filament protein nestin, reorganization of the actin cytoskeleton, and a dramatic reduction in the number of processes per cell. TUNEL analysis demonstrates that GGF is not cytotoxic for mature oligodendrocytes, but rather enhances their survival. GGF also induces the rapid activation of the PI 3-kinase and MAP kinase signaling pathways. These results further support a role for the NRGs in promoting the proliferation and survival of and inhibiting the differentiation of cells in the oligodendrocyte lineage and demonstrate that oligodendrocytes that differentiate in culture retain a substantial degree of phenotypic plasticity.

GGF/neuregulin is a neuronal signal that promotes the proliferation and survival and inhibits the differentiation of oligodendrocyte progenitors.

We show that GGF/neuregulin is a mitogen for prooligodendrocytes (O4+/O1- cells), oligodendrocytes (O4+/O1+ cells), and type-2 astrocytes. Heregulin beta 1, another neuregulin isoform, is also mitogenic. The proliferative effect of glial growth factor (GGF) does not require, but is greatly potentiated by, serum factors. GGF also promotes the survival of pro-oligodendrocytes under serum-free conditions. High levels of GGF reversibly inhibit the differentiation and lineage commitment of oligodendrocyte progenitors and, in differentiated cultures, result in loss of O1 and myelin basic protein expression. All three erbB receptors are expressed by progenitors and are activated by GGF; the relative abundance of these receptors changes during differentiation. Finally, cortical neurons release a soluble mitogen for pro-oligodendrocytes that is specifically blocked by antibodies to GGF. These results implicate the neuregulins in the neuronal regulation of oligodendrocyte progenitor proliferation, survival, and differentiation.

Three forms of RPTP-beta are differentially expressed during gliogenesis in the developing rat brain and during glial cell differentiation in culture.

In situ hybridization and Northern analysis demonstrate that the three splicing variants of RPTP-beta have different spatial and temporal patterns of expression in the developing brain. The 9.5-kb and 6.4-kb transcripts, which encode transmembrane protein tyrosine phosphatases with different extracellular domains, are predominantly expressed in glial progenitors located in the subventricular zone (SVZ). The 8.4-kb transcript, which encodes a secreted chondroitin sulfate proteoglycan (phosphacan), is expressed at high levels by more mature glia that have migrated out of the SVZ. The three transcripts are also differentially expressed in glial cell cultures; O2A progenitors express high levels of the 9.5- and 8.4-kb transcript, whereas type 1 astrocyte progenitors predominantly express the 6.4-kb transcript. C6 gliomas also express high levels of the 6.4-kb transcript. Treating C6 cells with the differentiating agent dibutyryl cyclic-AMP (DBcAMP), induces a decrease in the 6.4-kb transcript and a corresponding increase in the 8.4-kb transcript. O2A cells grown in the presence of basic fibroblast growth factor (bFGF) and platelet-derived growth factor (PDGF) remain highly proliferative and undifferentiated, and continue to express high levels of RPTP-beta. However, when O2A cells are grown in conditions that induce oligodendrocyte differentiation, there is a marked decrease in the expression of the transmembrane forms of RPTP-beta, as determined by immunofluorescence. These results demonstrate that RPTP-beta expression is regulated during glial cell differentiation and suggest that the different forms of RPTP-beta perform distinct functions during brain development.

Expression of rat cathepsin S in phagocytic cells.

Cysteine lysosomal proteases are essential for turnover of intracellular and extracellular proteins. These enzymes are strongly implicated in normal and pathological processes involving tissue remodeling. Among the cysteine proteases, cathepsin S seems to be best suited for such a process since it retains most of its enzymatic activity at neutral pH. In situ hybridization analyses of the adult rat brain, spleen, and lung reveal that cathepsin S mRNA is preferentially expressed in cells of mononuclear-phagocytic origin. After entorhinal cortex lesion of adult rat brain (a paradigm for neuronal degeneration and reactive synaptogenesis), cathepsin S mRNA is dramatically increased in activated microglia in the deafferented dentate gyrus and in macrophages at the wound site, suggesting a role in lesion-induced tissue remodeling. This possibility is further supported by the finding that cathepsin S degrades a number of extracellular matrix molecules at neutral pH and by the finding that inflammatory mediators stimulate its secretion from the microglia and macrophages. These data suggest that cathepsin S is an important player in degenerative disorders associated with the cells of the mononuclear phagocytic system.

Protein tyrosine kinase PYK2 involved in Ca(2+)-induced regulation of ion channel and MAP kinase functions.

The protein tyrosine kinase PYK2, which is highly expressed in the central nervous system, is rapidly phosphorylated on tyrosine residues in response to various stimuli that elevate the intracellular calcium concentration, as well as by protein kinase C activation. Activation of PYK2 leads to modulation of ion channel function and activation of the MAP kinase signalling pathway. PYK2 activation may provide a mechanism for a variety of short- and long-term calcium-dependent signalling events in the nervous system.

Expression of receptor protein tyrosine phosphatase-sigma (RPTP-sigma) in the nervous system of the developing and adult rat.

The expression of receptor protein tyrosine phosphatase-sigma (RPTP-sigma) mRNA during rat development was examined by Northern blot and in situ hybridization analyses. Northern blot analysis revealed that the two transcripts (5.7 kb and 6.9 kb) had different spatial and temporal patterns of expression. The 6.9-kb transcript was more abundant during embryonic development, whereas the 5.7-kb transcript was more abundant during postnatal development and in the adult. In situ hybridization revealed that RPTP-sigma mRNA was widely expressed throughout the central and peripheral nervous system during embryonic development. Very high levels were seen in the ventricular zone, subventricular zone, cortex, dorsal root ganglia, cranial nerve ganglia, olfactory epithelium, and retina. During postnatal development the level of expression decreased in most brain regions. However, high levels continued to be seen in the hippocampus. Emulsion autoradiography revealed that the majority of RPTP-sigma mRNA is expressed in neurons. Northern analysis showed that cultured glial cells expressed the 6.9-kb transcript, but not the 5.7-kb. RPTP-sigma mRNA expression profiles were clearly distinct from those of leukocyte antigen-related protein (LAR), a closely related RPTP. The spatiotemporal pattern of RPTP-sigma mRNA expression indicates that RPTP-sigma may play a role in the development of the nervous system.

Cloning of neurotrimin defines a new subfamily of differentially expressed neural cell adhesion molecules.

Previous studies in the laboratory indicated that glycosylphosphatidylinositol (GPI)-anchored proteins may generate diversity of the cell surface of different neuronal populations (Rosen et al., 1992). In this study, we have extended these findings and surveyed the expression of GPI-anchored proteins in the developing rat CNS. In addition to several well characterized GPI-anchored cell adhesion molecules (CAMs), we detected an unidentified broad band of 65 kDa that is the earliest and most abundantly expressed GPI-anchored species in the rat CNS. Purification of this protein band revealed that it is comprised of several related proteins that define a novel subfamily of immunoglobulin-like (Ig) CAMs. One of these proteins is the opiate binding-cell adhesion molecule (OBCAM). We have isolated a cDNA encoding a second member of this family, that we have termed neurotrimin, and present evidence for the existence of additional family members. Like OBCAM, with which it shares extensive sequence identity, neurotrimin contains three immunoglobulin-like domains. Both proteins are encoded by distinct genes that may be clustered on the proximal end of mouse chromosome 9. Characterization of the expression of neurotrimin and OBCAM in the developing CNS by in situ hybridization reveals that these proteins are differentially expressed during development. Neurotrimin is expressed at high levels in several developing projection systems: in neurons of the thalamus, subplate, and lower cortical laminae in the forebrain and in the pontine nucleus, cerebellar granule cells, and Purkinje cells in the hindbrain. Neurotrimin is also expressed at high levels in the olfactory bulb, neural retina, dorsal root ganglia, spinal cord, and in a graded distribution in the basal ganglia and hippocampus. OBCAM has a much more restricted distribution, being expressed at high levels principally in the cortical plate and hippocampus. These results suggest that these proteins, together with other members of this family, provide diversity to the surfaces of different neuronal populations that could be important in the specification of neuronal connectivity.

The expression of a novel receptor-type tyrosine phosphatase suggests a role in morphogenesis and plasticity of the nervous system.

Analysis of the localization of receptor-type protein tyrosine phosphatase-beta (RPTP-beta) by in situ hybridization and immunocytochemistry indicates that it is predominantly expressed in the developing central nervous system (CNS). RPTP-beta is highly expressed in radial glia and other forms of glial cells that play an important role during development. The immunoreactivity localizes to the radial processes of these cells, which act as guides during neuronal migration and axonal elongation. The pattern of RPTP-beta expression changes with the progression of glial cell differentiation. In the adult, high levels of RPTP-beta are seen in regions of the brain where there is continued neurogenesis and neurite outgrowth. The spatial and temporal patterns of RPTP-beta expression suggest that this receptor phosphatase plays a role in morphogenesis and plasticity of the nervous system.

The cloning of a receptor-type protein tyrosine phosphatase expressed in the central nervous system.

We have isolated cDNA clones and deduced the complete amino acid sequence of a large receptor-type protein tyrosine phosphatase containing 2307 amino acids. The human gene encoding this phosphatase, denoted RPTP beta (or PTP zeta), has been localized to chromosome 7q31-33. RPTP beta is composed of a large extracellular domain, a single transmembrane domain, and a cytoplasmic portion with two tandem catalytic domains. We have also cloned a variant of RPTP beta lacking 859 amino acids from the extracellular domain but with intact transmembrane and cytoplasmic domains. Interestingly, the amino-terminal region of the extracellular domain of RPTP beta contains a stretch of 266 amino acids with striking homology to the enzyme carbonic anhydrase. Immunoprecipitation experiments from a human neuroblastoma cell line indicate that the apparent molecular mass of the core and glycosylated forms of RPTP beta are approximately 250 and 300 kDa, respectively. Northern blot analysis shows that RPTP beta is strictly expressed in the central nervous system. In situ hybridization was used to further localize the expression to different regions of the adult brain including the Purkinje cell layer of the cerebellum, the dentate gyrus, and the subependymal layer of the anterior horn of the lateral ventricle. Hence, RPTP beta represents the first mammalian tyrosine phosphatase whose expression is restricted to the nervous system. The high level of expression of RPTP beta transcripts in the ventricular and subventricular zones of the embryonic mouse brain suggests the importance of this tyrosine phosphatase in the development of the central nervous system.

Identification of a carbonic anhydrase-like domain in the extracellular region of RPTP gamma defines a new subfamily of receptor tyrosine phosphatases.

The tyrosine phosphatase RPTP gamma is a candidate tumor suppressor gene since it is located on human chromosome 3p14.2-p21 in a region frequently deleted in certain types of renal and lung carcinomas. In order to evaluate its oncogenic potential and to explore its normal in vivo functions, we have isolated cDNAs and deduced the complete sequences of both human and murine RPTP gamma. The murine RPTP gamma gene has been localized to chromosome 14 to a region syntenic to the location of the human gene. Northern (RNA) blot analysis reveals the presence of two major transcripts of 5.5 and 8.5 kb in a variety of murine tissues. In situ hybridization analysis reveals that RPTP gamma mRNA is expressed in specific regions of the brain and that the localization of RPTP gamma changes during brain development. RPTP gamma is composed of a putative extracellular domain, a single transmembrane domain, and a cytoplasmic portion with two tandem catalytic tyrosine phosphatase domains. The extracellular domain contains a stretch of 266 amino acids with striking homology to the zinc-containing enzyme carbonic anhydrase (CAH), indicating that RPTP gamma and RPTP beta (HPTP zeta) represent a subfamily of receptor tyrosine phosphatases. We have constructed a model for the CAH-like domain of RPTP gamma based upon the crystal structure of CAH. It appears that 11 of the 19 residues that form the active site of CAH are conserved in RPTP gamma. Yet only one of the three His residues that ligate the zinc atom and are required for catalytic activity is conserved. On the basis of this model we propose that the CAH-like domain of RPTP gamma may have a function other than catalysis of hydration of metabolic CO2.