Freeze-fracture study of human brain tumors.

This freeze-fracture study was performed in 3 astrocytomas, 6 glioblastomas, 2 ependymomas, 3 medulloblastomas, 1 cerebellar sarcoma, 3 germinomas, and 1 medulloepithelioma. The number of nuclear pores/mum2 nuclear membrane was not correlated with biological malignancy. Fracture faces A and B were discernible in nuclear, Golgi and rough endoplasmic reticulum (ER), mitochondrial surface, and plasma membranes. Fenestrae were evident in Golgi and ER membranes. The transitional zone of cristae from the inner surface membrane appeared as a circular hole and broken-off neck on faces A and B of the inner surface membrane, respectively. The decrease in number of membrane particles in the plasma membrane seemed to correlate with the frequency of metastases, and, in addition, the membrane particles appeared to cluster in glioblastoma, medulloblastoma, and medulloepithelioma. The gap junctions were abundant in astrocytomas, moderate in number in ependymomas and germinomas, and rare in glioblastomas, cerebellar sarcoma, and medulloepithelioma. Tight junctions were often found in germinomas and medulloepithelioma, and rarely in ependymomas.     

Fenestrae in golgi and endoplasmic reticulum cisternae of human brain tumours

Fenestrae were found in freeze-fractured cisternae of the Golgi apparatus and endoplasmic reticulum of glioblastoma, oligodendroglioma, ependymoma, medulloblastoma, medulloepithelioma, meningioma, cerebellar sarcoma, hemangioblastoma, and chromophobe adenoma. They were about 200--400 A in diameter and often diffusely distributed or concentrated in groups in Golgi cisternae, while they were around 300--600 A in size and scattered in distribution in cisternae of endoplasmic reticulum. They appeared as conical protrusions or circular broken-off necks of face A and as circular holes on face B in tangential fractures, and as several constrictions of cisternae in cross fractures.     

Small vesicle bouton synapses on the distal half of the lateral dendrite of the goldfish Mauthner cell: freeze-fracture and thin section study

To understand principles of synaptic integration, it is necessary to define the types of synapses on a particular neuron and their distribution. Thin sectioning and double replica freeze-fracture techniques were employed to characterize the small vesicle bouton (SVB) synapses on the distal half of the Mauthner (M) cell lateral dendrite, which probably mediate a remote dendritic inhibition. Three morphologically distinct SVB synapses, types A, B, and C, were found. These three SVB synapses form roughly 90% of the synapses on the distal half of the lateral dendrite, with types A and B being most common. The SVB A synapse is characterized by mostly oval and round synaptic vesicles, a discrete presynaptic active zone with a highly variable shape, and a postsynaptic active zone with no apparent particle aggregate in either the E or P face. At the SVB B synapse, most of the synaptic vesicles are flat. A very high particle density is present throughout the presynaptic P face, and vesicle attachment sites are dispersed over much of the presynaptic membrane. Postsynaptic P face particle aggregates are subjacent to the presynaptic vesicle attachment sites, and are often large and anastomosing. The SVB C synapse is characterized by synaptic vesicle profiles that vary from flattened to round. The SVB C cytoplasm was unclouded by the flocculent material that characterized SVBs A and B. The presynaptic active zones at the SVB C synapse are discrete, and macular or oblong. No particle aggregates are apparent in the postsynaptic active zone. Small, macular particle aggregates were found in nonactive zone regions of the postsynaptic E face of all three types of SVBs. Small subsurface cisterns were also observed underlying the M cell membrane at all three types of SVB synapses. Neither the postsynaptic E face aggregates nor the subsurface cisterns were ever observed directly subjacent to presynaptic active zones, but were often seen adjacent to active zones. Short, straight rows of particles and short cylinders were often seen in both pre- and postsynaptic surrounding zone regions of SVB A and C synapses. These structures are thought to represent tight junctions.     

Primitive neuroectodermal tumors including the medulloblastoma: glial differentiation signaled by immunoreactivity for GFAP is restricted to the pure desmoplastic medulloblastoma ("arachnoidal sarcoma of the cerebellum")

Immunoreactivity of tumor cells for glial fibrillary acidic protein (GFAP) is usually regarded as sign of astrocytic histogenesis and/or differentiation. The present study aimed at a systematic evaluation of the significance of GFAP-containing cells in primitive neuroectodermal tumors (PNETs) with special reference to the controversial entity of desmoplastic medulloblastoma (so-called "circumscribed arachnoidal sarcoma of the cerebellum"). Fifty-three PNETs, including 17 pure desmoplastic medulloblastomas were investigated, using GFAP antisera and the peroxidase-antiperoxidase (PAP) technique. Seventy percent of the pure desmoplastic medulloblastomas showed GFAP immunoreactive cells, in 47% indistinguishable from adjacent nonreacting tumor cells. Most immunoreacting cells were found in the reticulin free islands, showing in 6 cases a gradual transition of immunoreacting cells from tumor cells to larger cells shaped like astrocytes. The classical medulloblastomas showed only larger immunoreacting cells which were interpreted as reactive astrocytes. Therefore, the so-called circumscribed arachnoidal cerebellar sarcoma or pure desmoplastic medulloblastoma merits a separate place in the group of PNETs as a tumor with frequent signs of astroglial differentiation; this interpretation appears to be clinically correlated by a difference in age incidence and prognosis of that special tumor-type in comparison with classical medulloblastoma.     

Myelin figures associated with the cytoplasmic membranes in human medulloblastoma

Summary Membranous whorls or myelin figures were found in human medulloblastoma treated with glutaraldehyde-OsO₄ double fixation. They were closely associated with the nuclear membrane, the endoplasmic reticulum, the mitochondria, and the plasmalemma.In the nuclei, they appeared as outpocketings at focal areas of the nuclear envelope, or exhibited lamellar structures of varying complexity, sometimes extending to the centre of nuclei. Membranous whorls derived from the rough endoplasmic reticulum became partly electron dense, forming myelin figures. The apposing membranes of the smooth endoplasmic reticulum were closely approximated or fused with each other to connect with the peripheral lamellae of myelin figures. In the mitochondria, the myelin figures were most conspicuous and continuous with the surface or the cristal membranes, and protruded from the surface membranes, or were sometimes lacalized beneath the surface membrane, or closely associated with the cristae in the centre of mitochondria. At the surface of cells, the myelin figures were closely allied with numerous vesicles and connected with the plasma membrane, often displaying a protrusion into the extracellular space. The peripheral lamellae of myelin figures in the medulloblastoma might be continuous with the membranes of the cytoplasmic constituents. It might be suggested that the myelin figures in the medulloblastoma reflected an abundance of available phospholipid or lipoprotein possibly disorganized by glutaraldehyde.     

Morphological investigations on cerebellar “neuroblastoma” group

Summary Classic medulloblastoma is a relatively common and distinct clinicopathologic entity consisting of primitive multipotential cells with differentiating capacity to neuroblastic and/or glial cell lines. Desmoplastic medulloblastoma has some features in common with cerebellar neuroblastoma, in which ultrastructural evidence of significant neuroblastic differentiation is extremely rare. We studied three cerebellar tumors with evidence of neuronal differentiation as compared to four classical desmoplastic medulloblastomas. Two of three tumors contained the regions of different degrees resembling desmoplastic medulloblastoma and one consisted of neuroblastic cells exclusively. This spectrum of differentiation suggests a relationship between cerebellar neuroblastoma and medulloblastoma, especially of the desmoplastic type. The nature of cerebellar neuroblastoma and its nosology are briefly discussed.     

小脑富脂质髓母细胞瘤临床病理学特征:病例报告及文献综述

目的 探讨小脑富脂质髓母细胞瘤临床病理学特征.方法 对一例小脑富脂质髓母细胞瘤患临临床表现、影像学特点,以及组织病理学和免疫组织化学特征进行回顾性分析,并复习相关文献.结果 男性患者,26岁.临床表现为反复头痛,伴头晕、恶心、呕吐.MRI显示病灶填允第四脑室,T1WI呈均匀低信号、T2WI呈不均匀高信号,增强T1WI扫捕病灶晕明显均匀强化,边界清晰.手术后光学显微镜下观察肿瘤细胞弥漫增生,大小一致,胞质淡染,呈弱酸性或嗜双色,核圆形或卵圆形,染色质细腻,核仁可见,核分裂象易见;间质中含有丰富的薄壁血管;富于脂质的细胞呈灶性分布.免疫组织化学染色肿瘤细胞CD56和突触素弥漫阳性,局灶表达神经微丝,弱表达少突胶质细胞系转录凶子-2;不表达巢蛋白、神经元核抗原、S-100蛋白、胶质纤维酸性蛋白和上皮膜抗原;TP53染色阳性(约10%).Ki-67抗原标记指数约为40%.结论 发生于小脑的富脂质髓母细胞瘤为临床罕见的中枢神经系统肿瘤,影像学呈现发生于小脑蚓部向第四脑室生长的占位性病变;病理学表现肿瘤细胞由高度密集的一致圆形细胞组成,具有灶性脂肪样细胞分化.应注意与小脑脂肪神经细胞瘤和室管膜瘤等相鉴别.     

Fine structure of myomedulloblastoma

Summary Electronmicroscopic studies in a case of myomedulloblastoma support the concept that the medulloblastoma and the myoblastoma component of the tumor derive from different cell lines. The myoblastoma component shows bundles of myogenic elements, whereby quite undifferentiated cells lacking myofilaments are tightly attached to differentiated elements having well organized myofilaments. These bundles are encompassed by a common basement membrane, and their cells are joined by junctions, helping the identification of the undifferentiated elements. No comparable structures were seen in the medulloblastoma component. The fine structure of the myoblastoma is reminiscent of aspects of normal fetal development of skeletal muscle.     

Subcellular localization of benzodiazepine/GABAA receptors in the cerebellum of rat, cat, and monkey using monoclonal antibodies

Two monoclonal antibodies, bd-17 and bd-24, specific for the beta- and alpha-subunit of the GABAA/benzodiazepine receptor/chloride channel complex, respectively, were used to determine the subcellular distribution of immunoreactivity in the cerebellum by electron microscopy. The 2 antibodies showed similar antigen distribution on the plasma membrane (except in the rat; bd-24 does not recognize the rat antigen), but intracellular immunoreactivity was more prevalent for the alpha-subunit. The plasma membrane of all neuronal types was immunopositive. The degree of immunoreactivity varied greatly between different types of cell, but it was stereotyped among individual cells of the same type. Granule cells showed the strongest immunoreactivity, not only on their dendrites which receive synapses from GABA-containing Golgi cell terminals, but also on their somata which do not receive synapses. Stellate and basket cells were somewhat weaker in immunoreactivity. Purkinje cells were only weakly positive on their somatic membrane but stronger on their dendritic shafts and spines. Golgi cells showed negligible if any immunoreactivity. Neurons of the deep cerebellar nuclei were strongly immunopositive along their plasma membrane. Immunoreactivity was strong in cisternae of the endoplasmic reticulum and in the Golgi saccules of stellate and basket cells, variable in Purkinje cells, while granule cells were rarely immunoreactive intracellularly. It is suggested that these differences reflect differences in the turnover of the receptor complex in the different cell types. The synaptic clefts established by boutons of the GABAergic stellate, basket, and Golgi cells were immunopositive, as were many synapses in the deep cerebellar nuclei. However, immunoreactivity was also present along the nonjunctional plasma membrane, and it was concluded that this reflected the distribution of the antigen. The synaptic clefts at the presumed glutamate-releasing parallel and mossy fiber terminals were almost always immunonegative. No immunoreactivity was detected on axons, nerve terminals, or glial cells. The results demonstrate that different neuronal types express the GABAA/benzodiazepine receptor/chloride channel complex to different degrees. The distribution of the receptor complex suggests that the cellular topography of GABAergic influence is not governed by the precise spatial arrangement of the receptors but by the precise placement of the GABA-releasing terminals, a characteristic of the cerebellar circuit.     

Freeze-fracture of developing neuronal plasma membrane in postnatal cerebellum

The structure of the plasma membrane of two neuronal cell types was quantitatively assessed in freeze-fracture replicas of rat cerebellar cortex from 2 to 73 postnatal days. The measurement of the number and size of intramembrane particles (IMP) in different regions of the Purkinje cell and of the granule cell showed that between 2 and 21 postnatal days there was a differential increase in the number of IMP; several regions of the cell characterized by a different pattern of synaptic interconnections showed differences in particle size and in particle number: neuronal processes had lower numbers of IMP than perikarya, whereas dendritic spines contained fewer IMP than dendritic trunks. These differences in IMP content appeared at the time at which specific synaptic contacts are established in these various regions of the cells.     

Tumour type and size are high risk factors for the syndrome of "cerebellar" mutism and subsequent dysarthria

OBJECTIVE: "Cerebellar mutis" and subsequent dysarthria (MSD) is a documented complication of posterior fossa surgery in children. In this prospective study the following risk factors for MSD were assessed: type, size and site of the tumour; hydrocephalus at presentation and after surgery, cerebellar incision site, postoperative infection, and cerebellar swelling. METHODS: In a consecutive series of 42 children with a cerebellar tumour, speech and neuroradiological studies (CT and MRI) were systematically analysed preoperatively and postoperatively. Speech was assessed using the Mayo Clinic lists and the severity of dysarthria using the Michigan rating scale. RESULTS: Twelve children (29%) developed MSD postoperatively. The type of tumour, midline localisation, and vermal incision were significant single independent risk factors. In addition, an interdependency of possible risk factors (tumour>5 cm, medulloblastoma) was found. CONCLUSION: MSD often occurs after paediatric cerebellar tumour removal and is most likely after removal of a medulloblastoma with a maximum lesion diameter>5 cm.     

Freeze-fracture studies of gap junctions of normal and neoplastic astrocytes

Summary Gap junctions of normal and neoplastic astrocytes were studied by the use of freeze-fracture or-etching technique. The gap junctions between normal astrocytes generally revealed a structure composed of small particles on fracture face A and associated pits on fracture face B. The particles and pits had presumably a complementary correlation: however, the pits were usually more regularly oriented—often in hexagonal array—than the particles. In the astrocytoma, the gap junctions were abundant and the pits were occasionally in tetragonal array. In the glioblastoma multiforme, the number of gap junctions was markedly decreased and the space between the particles sometimes increased, indicating a loose arrangement of the particles. The gap junctions in normal and neoplastic astrocytes demonstrated discontinuous macular plaques and did not exhibit continuous zones around the astrocytes. There was no evidence of tight junction in the fracture replicas of astrocytes. The present study might suggest that the malignancy of neoplastic astrocytes is represented by the decrease of the gap junction or by the loose orientation of the particles in the gap junction.This study was partly supported by a grant-in-aid for scientific research from the Ministry of Education, Japan.     

Diffusion-weighted MR imaging and pathologic findings in adult cerebellar medulloblastoma

OBJECTIVES, MATERIALS AND METHODS: The authors present the diffusion-weighted MR imaging and pathologic findings in two adult patients with cerebellar medulloblastoma. RESULTS: Both presented with a vermian mass of the posterior fossa with low signal on SE T1 weighted images, and moderate enhancement of the mass after gadolinium injection. The tumors were of high intensity on diffusion-weighted images with low ADC value. The ADC values (x10(-3) mm2/s) were respectively 0.60 +/- 0.06 and 0.59+/-0.11 (tumor), and 0.65 +/- 0.04 and 0.67 +/- 0.07 (cerebellar white matter). Tumors were highly cellular and composed of densely packed small round cells with hyperchromatic nuclei and scanty cytoplasm. CONCLUSION: diffusion-weighted MR imaging may be useful for the diagnosis of cerebellar medulloblastoma, due to their high cellularity and high nuclear-to-cytoplasmic ratio.     

Cell membrane structure of human giant-celled glioblastoma

Summary A giant-cell glioblastoma was examined by electron microscopy and by the freeze-fracture technique. The cell membranes bordering the extensive extracellular space often showed complicated undulations and peripheral vacuoles as well as occasional microvilli or filopodia. The undulations were mainly composed of plasmalemmal vesicles as well as of large (400–800 nm in diameter) and small (30–50 nm in diameter) localized protrusions and invaginations of the cell membrane. Deep invaginations of the cell membrane apparently resulted in two separate cytoplasmic portions. Locking of protruded cytoplasmic tongues and adherens junctions were sometimes seen in closely approximated cell membranes. The average number of membrane particles per m² was 630±130 on the P face and 180±30 on the E face. The membrane particles were occasionally aggregated to form clusters about 30 to 150 m in diameter. Gap junctions were occasionally found, but there were no tight junctions. Large particles about 30 nm in diameter were found in places.     

A freeze-fracture study of the soma membranes of monoaminergic neurons in the hypothalamic nucleus recessi posterioris of the rainbow trout (Salmo gairdneri Rich.)

Monoaminergic neurons in the hypothalamic nucleus recessi posterioris of the rainbow trout were studied by freeze-fracturing. The size and distribution of intramembrane particles in the nuclear and plasma membranes and the density of nuclear pores in the nuclear envelope were determined. The density of nuclear pores in these cells was significantly greater than in the non-granulated neurons of this nucleus. The density of IMPs in the P face of the inner nuclear membrane was significantly greater than in the P face of the outer nuclear membrane; however in the E faces the density is greater in the outer nuclear membrane but the differences are not statistically significant. In both membranes, the proportion of large particles (greater than 9.6 nm) was greater in the E than in the P faces. The coefficient of partition between the plasma membrane faces was 1.97 +/- 0.24 while in non-granulated neurons was 4.45 +/- 0.51. There are significant differences among these values. Interestingly, the coefficient of partition of the plasma membrane in monoaminergic neurons was lesser than in some other types of neurons. Postsynaptic areas or other membrane specializations were not found on the cell body plasma membrane, which corroborates the results of conventional transmission electron microscopy.     

Cerebellar liponeurocytoma: a newly recognized clinico-pathological entity

The term "cerebellar liponeurocytoma", recently adopted by the World Health Organization Working Group (WHO), replaced many other different terms used up to now to give name to this rare tumor. To our knowledge, less than 20 cases have been related up to now under different names like as "lipomatous medulloblastoma, lipidized medulloblastoma, neurolipocytoma, medullocytoma and lipomatous glioneurocytoma". The new nomenclature eliminates the word "medulloblastoma", reinforces its benign character, and includes it in the category of glioneuronal tumors. We describe an addictional case of this distinct clinico-pathological entity removed from the right cerebellar hemisphere of a 53-year-old woman. With the present case report, we hope to contribute to the knowledge on the diagnostic and prognostic implications derived from the finding of mature adipose-like tissue within a medulloblastomatous tumour.     

髓母细胞瘤CyclinD1、P16的表达及其与临床病理特点和预后的关系

目的检测髓母细胞瘤CyclinD1和P16的表达,分析它们与临床病理特点之间的关系,探讨其对髓母细胞瘤预后的影响。方法通过免疫组化方法并用图像分析系统检测CyclinD1和P16在髓母细胞瘤和正常小脑组织中表达的积分光密度(IOD),分析它们与临床病理特点及预后的关系。结果(1)正常小脑组织CyclinD1表达较低,髓母细胞瘤表达增高;P16在正常小脑组织中呈现高表达,而髓母细胞瘤出现降低;(2)髓母细胞瘤CyclinD1和P16表达情况与患者年龄、肿瘤部位、病理类型、是否伴坏死、分化程度及分化方向有关(P<0.01或P<0.05);(3)经单因素分析和多因素分析显示肿瘤分化程度、CyclinD1和P16表达与患者生存期存在密切关系(P=0.0000)。结论(1)髓母细胞瘤中CyclinD1和P16表达改变,反映了肿瘤细胞增殖活性和细胞周期活性发生了改变,并且不同临床病理特点的髓母细胞瘤,增殖活性和细胞周期活性改变的程度并不一致。(2)髓母细胞瘤临床病理特点的不同更深层次的原因之一是肿瘤细胞CyclinD1和P16发生了不同变化从而导致肿瘤细胞形成各自不同的细胞周期改变。(3)CyclinD1和P16表达改变,为髓母细胞瘤的预后判断和临床治疗提供了新的思路。     

Anatomo-functional study of the cerebellum in working memory in children treated for medulloblastoma

Introduction

Medulloblastoma is the most common malignant cerebral tumor during childhood, arising in the posterior fossa. Children treated for medulloblastoma often experience working memory (WM) deficits, affecting their quality of life and school performance. The aim of the present study undertaken to describe the cerebellar involvement in WM deficits observed in these children.

Duchenne型肌营养不良症的冷冻断裂电镜观察

应用冷冻断裂电镜技术对Duchenne型肌营养不良症(DMD)的骨胳肌标本4例(异常组),正常标本2例(正常组)肌细胞质膜的蛋白颗粒进行电镜观察。结果表明,正常组和异常组的EF(extracellular face)面膜蛋白颗粒比PF(protoplasmic face)面少;异常组与正常组比较,其EF面和PF面的膜蛋白颗粒均明显减少,并具有显著的统计学意义。     

RBP-J is not required for granule neuron progenitor development and medulloblastoma initiated by Hedgehog pathway activation in the external germinal layer

Background

The Notch signalling pathway plays crucial roles in neural development, functioning by preventing premature differentiation and promotion of glial cell fates. In the developing cerebellum Notch pathway components are expressed in granule neuron progenitors of the external germinal layer (EGL) but the precise function of Notch in these cells is unclear. The Hedgehog pathway is also crucial in cerebellar development, mainly via control of the cell cycle, and persistent activation of the pathways leads to the cerebellar tumour medulloblastoma. Interactions between Hedgehog and Notch have been reported in normal brain development as well as in Hedgehog pathway induced medulloblastoma but the molecular details of this interaction are not known and we investigate here the role of Notch signalling in the development of the EGL and the intersection between the two pathways in cerebellar granule neuron progenitors and in medulloblastoma.

Results

RBP-J is the major downstream effector of all four mammalian Notch receptors and the RBP-J conditional mouse facilitates inactivation of canonical Notch signals. Patched1 is a negative regulator of Hedgehog signalling and the Patched1 conditional mouse is widely used to activate Hedgehog signalling via Patched1 deletion in specific cell types. The conditional mouse lines were crossed with a Math1-Cre line to delete the two genes in granule neuron progenitors from embryonic day 10.5. While deletion of only Patched1 as well as Patched1 together with RBP-J leads to formation of medulloblastoma concomitant with disorganisation of cell layers, loss of RBP-J from granule neuron progenitors has no obvious effect on overall cerebellar morphology or differentiation and maturation of the different cerebellar cell types.

Conclusions

Our results suggest that even though Notch signalling has been shown to play important roles in cerebellar development, signalling via RBP-J is surprisingly not required in granule neuron progenitors. Furthermore, RBP-J inactivation in these cells does not influence the formation of medulloblastoma initiated by Hedgehog pathway activation. This may suggest a requirement of Notch in cerebellar development at a different developmental stage or in a different cell type than examined here - for example, in the neural stem cells of the ventricular zone. In addition, it remains a possibility that, in granule neuron progenitors, Notch may signal via an alternative pathway without the requirement for RBP-J.