The involvement of calpain-dependent proteolysis of the tumor suppressor NF2 (merlin) in schwannomas and meningiomas.

Neurofibromatosis type 2 (NF2) protein, also known as merlin or schwannomin, is a tumor suppressor, and NF2 is mutated in most schwannomas and meningiomas. Although these tumors are dependent on NF2, some lack detectable NF2 mutations, which indicates that alternative mechanisms exist for inactivating merlin. Here, we demonstrate cleavage of merlin by the ubiquitous protease calpain and considerable activation of the calpain system resulting in the loss of merlin expression in these tumors. Increased proteolysis of merlin by calpain in some schwannomas and meningiomas exemplifies tumorigenesis linked to the calpain-mediated proteolytic pathway.

The role of hemodynamic factors in arterial wall thickening in the rat.

In a study on the pathogenesis of arterial wall thickening, hemodynamic factors in the common carotid artery of the rat were experimentally altered with an autograft. The relationship between flow pattern and wall-thickening was examined in a half-ring bypass model with an induced stenosis, using both flow-visualization in a corresponding in vitro model circuit and observation of the wall by microscopy. Wall-thickening was found in the neighborhood of bifurcations, junctions and curved segments, which corresponded to regions of low-shear in the flow-field. Marked histologic changes in the wall were observed in the post-stenotic segments where the flow field was very disturbed. Histologic changes in the arterial wall correlated well with flow patterns.

The diagnostic value of 123I-IMP SPECT in non-Hodgkin's lymphoma of the central nervous system.

UNLABELLED: N-isopropyl-p-[123I]iodoamphetamine (IMP) SPECT is of low diagnostic value in patients with brain tumors, because brain tumors are visualized as uptake defects. Some reports have described non-Hodgkin's lymphoma of the central nervous system (CNS) as showing high uptake on delayed 123I-IMP SPECT images, suggesting its usefulness in diagnosing CNS lymphoma. In this study, we investigated the clinical value of 123I-IMP SPECT as a diagnostic tool for CNS lymphoma. METHODS: Ninety-six patients with brain tumors, including 12 patients with CNS lymphoma, underwent 123I-IMP SPECT. Eleven patients had primary CNS lymphoma, and 1 had a parenchymal brain metastasis from a breast lymphoma. The total number of lesions was 18, 14 of which were in the cerebral parenchyma, 3 in the brain stem, and 1 in the ventricle. Early SPECT images were initiated 15-30 min after intravenous injection of 111 MBq 123I-IMP, and delayed images were collected 4 h later. SPECT images were visually analyzed with a color-grading scale. Tumor-to-normal activity ratio (T/N) and tumor-to-cerebellum activity ratio (T/C) were calculated for both early and delayed images for semiquantitative analysis. RESULTS: By visual estimation, more than a 3-cm3 volume of CNS lymphoma was detected as an obvious focus of increased accumulation on delayed images. All other brain tumors tested appeared as decreased accumulation on delayed images. T/Ns and T/Cs on delayed images of CNS lymphomas, including tumors less than 3 cm3 in volume, were 1.48+/-0.42 and 1.08+/-0.16, respectively. These ratios in patients with glioma (0.30+/-0.05 and 0.31+/-0.07 respectively) or meningioma (0.34+/-0.10 and 0.41+/-0.17, respectively) showed a significant difference from those in patients with CNS lymphoma (P < 0.0005). CONCLUSION: 123I-IMP SPECT is a helpful tool for diagnosing CNS lymphoma.

Effect of nicardipine on abnormal excitability of CA3 pyramidal cells in hippocampal slices of spontaneously epileptic rats.

The effects of nicardipine, a Ca2+ channel antagonist, on the abnormal excitability of hippocampal CA3 neurons in spontaneously epileptic rats (SER), a double mutant (zi/zi, tm/tm), were examined to elucidate whether or not the abnormality was due to that of Ca2+ channels. An intracellular recording study was performed using brain slice preparations of SER 12-15 weeks of age, when SER showed both tonic convulsions and absence-like seizures. Bath application of nicardipine (10 nM) completely inhibited the depolarizing shifts lasting for 60-120 ms and accompanying repetitive firings on mossy fiber stimulation in SER. However, this drug did not affect the single action potential induced by the mossy fiber stimulation in CA3 neurons of SER and normal Wistar rats. In the CA3 pyramidal neurons of SER, the Ca2+ spikes induced by the depolarizing pulse applied in the cell in the presence of tetrodotoxin and tetraethylammonium had a different configuration from that in normal Wistar rats. Nicardipine also inhibited the Ca2+ spikes in SER CA3 neurons at a concentration (1 nM) that had no effect on those in normal Wistar rats, while the Ca2+ spikes in Wistar rat CA3 neurons were inhibited by 10 nM nicardipine. These findings suggest that the abnormal excitability of CA3 pyramidal neurons in SER might be attributed to abnormalities of the Ca2+ channels, and that the Ca2+ channel antagonist may be effective as an antiepileptic drug.

Consensus opinion on diagnosis of cerebral circulatory arrest using Doppler-sonography: Task Force Group on cerebral death of the Neurosonology Research Group of the World Federation of Neurology.

BACKGROUND AND PURPOSE: Oscillating flow or systolic spikes are typical Doppler-sonographic flow signals found in the presence of cerebral circulatory arrest, which if irreversible, results in brain death. The Neurosonology Research Group (NSRG) of the World Federation of Neurology (WFN) created a Task Force Group in order to evaluate the role of Doppler-sonography as a confirmatory test for determining brain death. METHODS: The available evidence from the literature has been reviewed and discussed by a group of experts, the members of the Task Force Group on cerebral death of the NSRG. RESULTS AND CONCLUSIONS: Extra- and intracranial Doppler-sonography is a useful confirmatory test to establish irreversibility of cerebral circulatory arrest as optional part of a brain death protocol. Doppler-sonography is of special value when the therapeutic use of sedative drugs renders electroencephalography unreliable. Doppler-sonographic criteria are defined and guidelines for the use of Doppler-sonography in this setting are presented.

Tentorial cavernous angioma with calcification in a neonate.

A cavernous angioma of the tentorium cerebelli, first disclosed by perinatal serial ultrasonographic studies, was extirpated totally without remarkable neurological deficit in a neonate. The tumor was accompanied by a calcified expansive hematoma in the posterior fossa. To our knowledge, this is the first case not only of cavernous angioma treated surgically and verified histologically in a neonate, but also of a calcified hematoma revealed on a conventional skull x-ray film at birth. This case suggests the possibility of hemorrhage from intracranial cavernous angioma early in life, even prenatally, and emphasizes the necessity for early diagnosis and early treatment of intracranial cavernous angiomas.

Intraoperative monitoring with pulse Doppler ultrasonography in transsphenoidal surgery: technique application.

We describe an intraoperative use of pulse Doppler ultrasonography in transsphenoidal surgery to prevent mechanical vascular injury, particularly to the intracavernous portion of the internal carotid artery. This system is integrated by connection to a video processor. The use of Doppler sonography provided real-time measurement of arterial or venous flow velocity and source of flow by both real-time sonograms and sound frequencies. With the use of a microprobe, 1 mm in diameter, vessels located within 7 mm from the tip of the probe could be easily, rapidly, and noninvasively detected, without disturbing operative field. Furthermore, both the size and the distance of a vessel could be measured by turning the dial of Doppler signal gain from initially waxing to waning Doppler sounds, because the acoustic sounds were adjusted to the axial flow of each vessel in 0.1-mm increments. Our intraoperative monitoring enhanced operative safety with technical simplicity and reliability.

Malignant melanoma metastatic to sphenoid bone with an unknown primary origin: report of a case with an immunohistochemical analysis of the infiltrating lymphocytes.

A rare case of hemorrhagic malignant melanoma from an unknown primary origin to the right sphenoid bone in a 37-year-old woman is presented. The tumor occupied an extensive intracranial extradural space with a mild orbital infiltration, but little involvement of the leptomeninges and parenchymal tissues. Immunohistochemical analysis of the tumor-infiltrating lymphocytes showed that the majority expressed pan-T or Leu-1 surface antigens and that the lymphocytes bearing the Leu-3 antigen, or helper phenotype, were predominant compared to the Leu-2 population representing both suppressor and cytotoxic T cells. Furthermore, most of the T lymphocytes stained with HLA-DR antigens. Because few B cells were seen, this observation indicates the presence of T cells in an activated state. The literature pertinent to associated organ involvement of metastatic melanomas and the correlations between the tumor and immunological cellular responses are discussed.

Analysis of intra- and intercellular calcium signaling in a mouse malignant glioma cell line.

Intra- and intercellular calcium signaling in glioma cells was examined by mechanical stimulation of a monolayer cell line of methylcholanthrene-induced mouse ependymoblastoma, 203-glioma, with a fine round-tip glass needle. A fura-2 fluorescence image of the glioma revealed a four- to eightfold increase in the cytosolic calcium ion concentration in directly stimulated signal cells. The increased calcium spread to surrounding cells at a speed of 20 microns/sec for a distance of up to 200 microns. Calcium was transmitted between adjacent cells and even in cells up to 200 microns distant from the initially stimulated cell. Microinjection of Lucifer yellow dye showed no gap junctional communication between cells. Depletion of extracellular calcium ion inhibited both cytosolic calcium elevation and propagation to neighboring cells by mechanical stimulus. An intracellular calcium blocker, TMB-8, eliminated the cytosolic calcium mobilization in a mechanically stimulated cell, but had no effect on calcium diffusion to surrounding cells. Nifedipine and verapamil, antagonists of voltage-dependent calcium channels, did not act on the mechanically induced calcium response. This suggests that some stimulating factor may trigger transmission of calcium, which may be ejected directly from single stimulated cells and mediated via a membrane receptor but not through a gap junction. The calcium signaling in a mechanically stimulated cell may be related to both an influx and a redistribution of intracellular calcium from internal stores, while calcium propagation to neighboring cells may involve calcium influx alone.

Intraoperative use of Doppler ultrasound and endoscopic monitoring in the stereotactic biopsy of malignant brain tumors. Technical note.

An intraoperative monitoring tool is described that prevents mechanical injury to intracerebral vessels during stereotactic surgery. The method, which combines pulse Doppler ultrasonography and fiberendoscopy, allowed stereotactic biopsy to be performed without serious intracerebral bleeding in 25 patients with hypervascular malignant brain tumors, 13 with glioblastoma multiforme, five with anaplastic astrocytoma, five with metastatic tumor, and two with malignant lymphoma. The ultrasound apparatus has a built-in fast-Fourier transformation system analyzer and an improved filtering system that provide real-time measurement of blood flow velocity. The source of flow (arterial or venous) could be identified by both real-time sonography and acoustic signal frequencies. It was possible to measure the size and distance of a vessel by adjusting the Doppler signal gain dial from initially waxing to waning sounds, because the acoustic signal was adjusted to the axial flow of each vessel in 0.1-mm steps. Each of three Doppler probes (1 mm, 2 mm, and 3 mm in diameter) fit through the outer cannula of the biopsy needle. Vessels located within 7 mm from the tip of these probes could be detected easily and rapidly, so the biopsy needle could be advanced safely to the desired target in 7-mm steps. If sonograms revealed blood flow, indicating the presence of larger vessels in the intended stereotactic trajectory, the angle of the needle was changed slightly to avoid vascular injury. Because the fiberendoscope was connected to a video processor, the vessel could be visualized at a higher magnification on the video display, unless there was active bleeding. This technically simple and reliable system enhances operative safety while maintaining accuracy.

Primary intracerebral and aneurysmal subarachnoid hemorrhage in Izumo City, Japan. Part I: incidence and seasonal and diurnal variations.

OBJECT: The purpose of this community-based study was first to estimate the incidence rates of primary intracerebral hemorrhage (ICH) and aneurysmal subarachnoid hemorrhage (SAH) in Izumo City, Japan, and second to investigate whether there were seasonal and diurnal periodicities in their onset. METHODS: During 1991 through 1996, 267 patients with primary ICH and 123 with aneurysmal SAH were treated in Izumo City. The crude and the age- and sex-adjusted annual incidence rates per 100,000 population for all ages were 52 and 48 for ICH and 24 and 23 for SAH, respectively. These incidence rates were higher than those previously published for any other geographical region. The incidence rates of both ICH and SAH increased almost linearly with age. For ICH, a significant seasonal pattern was observed in men and in patients younger than 65 years, with a peak in winter and a trough in summer. However, no significant seasonal fluctuation was found in women or in individuals aged 65 years or older. There was no significant seasonal periodicity for SAH, even when patients were analyzed according to sex and age. Diurnal variations in the onset of both ICH and SAH were significant (except in men with SAH), with a nadir between midnight and 6:00 a.m. CONCLUSIONS: The actual incidence rates of both primary ICH and aneurysmal SAH seem to be much higher than have been reported so far. In addition, the data indicate the existence of seasonal periodicity for men and younger patients with ICH, and that the risk of both ICH and SAH is lower during nighttime.

Role of histocompatibility antigen gene and protooncogene expressions in intracerebral tumorigenicity of mouse neuroblastoma.

The role of N-myc, c-src, and major histocompatibility complex (MHC, H-2 in the mouse) class I antigen gene expressions in dimethyl sulfoxide (DMSO)-induced differentiation and intracerebral tumorigenicity was examined using a mouse MNB85 neuroblastoma cell line. A fluorescence-activated cell sorter disclosed cell-surface MHC enhancement by DMSO, causing an increase in cytotoxic T-lymphocyte sensitivity. Southern blot analysis verified a single copy of the proto-oncogenes and MHC deoxyribonucleic acids in both untreated and DMSO-treated MNB85 cells. Northern blot analysis indicated that DMSO treatment induced a decrease in N-myc and an increase in c-src and MHC messenger ribonucleic acids. Nuclear run-off transcription assay revealed down-regulation of N-myc at a posttranscriptional level, contrasted with primary up-regulation of c-src at a transcriptional level. Immunoprecipitation after treatment with enzyme endo-beta-N-acetyl-glycoseamidase H proved that the terminal glycosylation of MHC heavy-chain gene products normally occurs in the Golgi apparatus of MNB85 cells. Intracerebral tumorigenicity assay showed that cells highly MHC-expressed by DMSO were less tumorigenic than untreated cells in association with DMSO-augmented cytotoxic T-lymphocyte susceptibility. These results suggest that proto-oncogenes may be linked to cellular differentiation, while cell-surface MHC gene expression influences intracerebral immunosurveillance.

A morphological and ultrastructural investigation of normal mouse brain tissue after intracerebral injection of tumor necrosis factor.

Morphological and ultrastructural changes in normal mouse brain tissue were investigated after intracerebral stereotactic injections of tumor necrosis factor (specific activity: 2.0 x 10(6) U/mg protein) into the right frontal lobe. The mice received either a single infusion or multiple tumor necrosis factor infusions in three different dose groups (10, 100, or 500 U). Compared with sham-treated control mice that received adjusted intracerebral injections of purified albumin, the tumor necrosis factor-treated mice in all dose groups did not show any specific in vivo behavioral abnormalities during the 2 months of study following the infusions. Histological studies revealed hemorrhage attributable to the mechanics of the intracerebral infusions, a thickening of the arachnoid membranes, a reactive gliosis, and neutrophilic and/or mononuclear cell infiltration along the infusion pathway. A local neutrophilic response was prominent 1 day after tumor necrosis factor injection. An immunohistochemical analysis indicated that the mononuclear cell infiltration consisted of lymphocytes and macrophages. Except for the transient neutrophilic infiltration, these histological alterations did not differ from those seen in the sham-treated control groups, and most nonspecific reactive changes disappeared within 8 weeks after the injections. Furthermore, an ultrastructural study showed no apparent pathological changes in the cytoplasmic organelles of neuronal, glial, and endothelial cells in the tumor necrosis factor-injected mouse specimens. These results suggest that the tumor necrosis factor injections caused no specific toxicity and did not alter the parenchymal and stromal cells comprising normal mouse brain tissue.

Effects of cerebrovascular autoregulation and CO2 reactivity in experimental localized brainstem infarction.

Using the previously reported method of experimental localized brainstem infarct in dogs, we designed this study to elucidate sequential changes of regional cerebral blood flow (rCBF) in three separate regions of the central nervous system: the cerebral cortex, thalamus, and midbrain. The data obtained were referred to in subsequent investigations of cerebrovascular autoregulation and vasomotor reactivity to CO2. Localized brainstem infarct was produced by permanently occluding the perforators of the posterior cerebral arteries between the bilateral origins of the posterior communicating arteries. The hydrogen clearance method was applied to measure rCBF. Cerebrovascular autoregulation and CO2 reactivity were assessed in three regions 1, 3, and 5 h after vascular occlusion, respectively. Vascular occlusion resulted in a decrease of rCBF that was 65% in the midbrain and close to 30%-40% in the thalamus. However, no significant change was seen in the cerebral cortex even 5 h after vascular occlusion. Induced hypertension impaired autoregulation in the thalamus, while it was preserved in the cerebral cortex. Induced hypotension did not alter autoregulation in any of the three regions. A marked loss of CO2 reactivity was observed in the ischemic brainstem, although it was well preserved in the cerebral cortex. The results suggest that noradrenergic fibers originating from the cervical sympathetic ganglia play a main role in the cerebrovascular autoregulation in the cerebral cortex, while noradrenergic fibers possibly originating from the autonomic centers in the brainstem are responsible in the thalamus; that the noradrenergic neuron probably is not involved in the maintenance of cerebral blood flow during hypotension; and that the effect of CO2 is mediated by its direct effect on the arteriolar wall in the central nervous system.

Experimental study on quantitative flow measurement by a Doppler flowmeter with a sound-spectrograph.

A bidirectional ultrasonic Doppler flowmeter equipped with a sound-spectrographic analyzer has enabled us to determine blood-flow direction and to obtain useful information from the flow pattern in assessing the cerebral blood flow transcutaneously or intraoperatively. Using a regulated flow system produced in a dog, we performed an experimental study in order to establish the optimal mode for its use and to study the reliability in the quantitative cerebral blood-flow measurements. Although we could not measure the absolute blood-flow volume directly with this apparatus, we were able to obtain useful, quantitative information on blood flow and flow pattern by placing the ultrasonic probe in an appropriate position with respect to a blood vessel.

Quantitative analysis of microstructural components of human cerebral arteries.

Microstructural components were analyzed quantitatively by means of a microphotometric technique applied to three kinds of major cerebral arteries and two kinds of extracranial arteries of comparable size obtained from human autopsy subjects. No statistically significant difference was found in the fractions of three structural components, elastin (Fe), collagen (Fc), and smooth muscle (Fm), in the total of fractions of these three components and in the ratio between extracranial arteries. The Fe-values were significantly less in cerebral arteries than in extracranial arteries. The Fc/Fe ratios in cerebral arteries were about triple those in extracranial arteries. Although there was no statistically significant difference in the Fe- and the Fm-values between the two age groups, under 49 and over 50 years old, the Fc/Fe ratio were higher in the older age group. This change of microstructural composition with age was more prominent in cerebral arteries than in extracranial arteries.

Ultrasonic bruits in the circle of Willis due to a large nonfunctioning pituitary adenoma.

In a man with a histologically verified non-functioning pituitary adenoma with suprasellar extension, Doppler signals resembled those associated with bruits (ultrasonic bruits). These signals were detected in the anterior circulation of Willis both preoperatively and intraoperatively. The large tumor was resected subtotally via a right orbitozygomatic approach. The use of microvascular sonography for intraoperative monitoring can provide information on the potential cerebrovascular complications of surgery. No previous studies on the presence of ultrasonic bruits associated with pituitary adenomas have been reported. The clinical implications for the surgical treatment of pituitary adenomas are discussed.

Synergistic antitumor effects of human interferon-beta and interferon-gamma on human gastric adenocarcinoma cells.

The combined effects of different classes of interferon (IFN) on human gastric adenocarcinoma cell line HPE-GAC-2 (GAC-2) were investigated in vitro. Synergistic effects of IFN-beta and -gamma, causing cell death, were observed in a dose-dependent manner. IFN-gamma modulated the sensitivity of GAC-2 cells to the cytocidal effect of IFN-gamma. The modulatory effect was independent of the cytostatic or cytocidal activity of IFN-beta since brief contact with IFN-beta did not influence the growth kinetics of the cells and was effective in modulate the cellular sensitivity to IFN-gamma. A brief pretreatment of the cells with IFN-alpha or-beta resulted in a variable level of modulation, whereas coculture of the cells with IFN-beta or -alpha and IFN-gamma had a similar cytotoxic effect indicating a different level of activity induced by IFN-alpha and -beta with the common type I receptor. These results suggest that IFN-beta-induced modulation was receptor-mediated. Phase contrast microscopy showed evidence of apoptotic cell death induced by one or more IFN agents. The morphological changes included chromatin condensation and nuclear fragmentation which were apparent at as early as 6 hr of culture with a high concentration of IFN(s). Our results indicate that IFN-beta and IFN-alpha enhance the in vitro apoptotic effects of IFN-gamma against GAC-2 tumor cells.

[Glial tumor cell proliferation and immune response in the brain].

The mechanism of glial tumor cell proliferation and the immune response to glioma cells in the brain were examined both in vitro and in vivo experimental systems, using mouse malignant glioma cell line, 203-glioma. A fura-2 fluorescence image showed marked rise in the intracellular calcium ion concentration in mechanically stimulated single cells. The increased calcium spread to adjacent cells, probably due to some stimulating factor released from cells. Dye microinjection revealed no gap junction between cells. Antagonists of voltage-dependent calcium channels did not act on the calcium response. These suggest that calcium signaling in the glioma cells may be mediated via a membrane receptor but not through a gap junction. Depletion of extracellular calcium ion and addition of intracellular calcium blocker demonstrated that calcium signaling in stimulated cells may be related to both an influx of extracellular calcium and a redistribution of intracellular calcium from internal stores, whereas calcium transmission to adjacent cells may involve calcium influx alone. The splenic cytotoxic T lymphocyte (CTL) activity in intracerebral tumor-bearing hosts increased with a peak 2 weeks after tumor cell inoculation, but rapidly decreased concurrently with increased intracranial pressure. The major histocompatibility complex, MHC, class I antigen expression on tumor cells grafted intracerebrally was found to enhance markedly, resulting in an increase in susceptibility to CTL. It was suggested that there may be a positive correlation between the cell surface MHC class I antigen expression and sensitivity to CTL in glioma cells.

Cavernous angioma in the fourth ventricle.

A cavernous angioma in the fourth ventricle of a 47-year-old man is reported. Because of a progressive clinical course and a ring-like enhancement on computed tomography, a brain tumor, rather than a vascular disease, was suspected. The lesion was removed totally and verified histologically as a cavernous angioma.