Aspectos ultraestructurales de la plasticidad sináptica en el edema cerebral peritumoral en humanos / Ultrastructural features of the synaptic plasticity in peritumoral cerebral oedema in humans

Caracterizar ultraestructuralmente los fenómenos de plasticidad sináptica en el tejido cerebral peritumoralhumano. Pacientes y métodos. Se procesaron 16 biopsias de tejido peritumoral mediante las técnicas convencionales para microscopía electrónica de transmisión. Se revisaron y analizaron los datos clínicos y el seguimiento posquirúrgico. Resultados.En el tejido cerebral peritumoral predominó el edema cerebral del tipo celular u oncótico. Se observaron múltiples sinapsis perforadas con curvaturas convexas y, en menor frecuencia, con curvaturas irregulares. Generalmente, los terminales presinápticospresentaron aspecto edematoso, con mitocondrias edematosas con cristolisis y matriz mitocondrial electrón densa.Los terminales presinápticos mostraron numerosas vesículas sinápticas de diferentes tamaños y ocasionalmente dispuestas en pequeños acúmulos. Se evidenció la presencia de vesículas de endocitosis que sugieren una alta recaptación de neurotransmisores.Algunos terminales presinápticos mostraron cambios degenerativos. Los terminales postsinápticos se observaron edematosos y las espinas dendríticas mostraron diferentes formas y aparatos espinosos degenerados en grado variable. Las prolongacionesastrocitarias perisinápticas se observaron edematosas, con cantidad variable de gránulos de glucógeno. Se propone una posible secuencia de cambios estructurales en el proceso de formación de sinapsis perforadas en el tejido cerebral peritumoral.Conclusiones. Los hallazgos ultraestructurales observados en las sinapsis y su microambiente pueden considerarse relevantes en relación con las manifestaciones clínicas y las secuelas de los pacientes estudiados, y se vinculan con alteracionesfuncionales del tejido lesionado, cambios degenerativos y muerte neuronal

To provide an ultrastructural characterisation of the synaptic plasticity phenomena in peritumoral brain tissuein humans. Patients and methods. Sixteen peritumoral tissue biopsy samples were processed using conventional transmission electron microscope techniques. Clinical data and post-surgical follow-up were reviewed and analysed. Results. In the peritumoralbrain tissue there was a predominance of cellular or oncotic cerebral oedema. A large number of synapses were seen to be perforated with both convex and, although less frequently, irregular curvatures. Generally, the presynaptic terminals had an oedematous appearance, with oedematous mitochondria with cristolysis and electron-dense deposits in the mitochondrialmatrix. The presynaptic terminals showed numerous synaptic vesicles of different sizes and which were sometimes arranged in small accumulations. Observations also confirmed the presence of endocytosis vesicles, which suggest high neurotransmitterreuptake. Some presynaptic terminals displayed degenerative changes. The postsynaptic terminals were seen to be oedematous and the dendritic spines displayed different shapes and spiny processes that were degenerated to varying degrees. Perisynapticastrocytic prolongations were seen to be oedematous, with a variable number of glycogen granules. We propose a possible sequence of structural changes in the process of forming perforated synapses in peritumoral brain tissue. Conclusions. The ultrastructural findings observed in the synapses and their microenvironment can be considered to be significant with respectto the clinical manifestations and the sequelae in the patients who were studied. Moreover, they are linked to functional alterations in injured tissue, degenerative changes and neuronal death

Neuronal features of oligodendrogliomas--an ultrastructural and immunohistochemical study.

AIMS: To assess neuronal differentiation in oligodendrogliomas (ODGs). METHODS AND RESULTS: An electron microscopic and immunohistochemical study of 41 consecutive cases was performed. In all cases, tumour cells with neuritic structures were identified ultrastructurally, including synapses and neurosecretory granules. For the immunohistochemical identification of synaptophysin, monoclonal antibody clones 27G12, Snp88 and SY38 and a polyclonal antibody were compared in optimized protocols on slides from a spectrum of tissues and 16 ODGs. 27G12 gave the best signal-to-noise ratio, while SY38 gave the poorest. When 27G12 was applied on all 41 ODGs, widespread immunoreactivity was obtained in 100%. Among three antibodies to chromogranin compared similarly, clone LK2H10 and a polyclonal antibody gave identical patterns of immunoreactivity, whereas clone DAK-A3 gave weaker reactions. When LK2H10 was applied on all tumours, staining was found in 12 (29%). All tumours but one stained strongly for glial fibrillary acidic protein and all for synapsin I. Fluorescence in situ hybridization analysis showed a concomitant 1p/19q deletion in 12/16 ODGs. CONCLUSIONS: Our study provides evidence for widespread neuronal differentiation in ODGs, suggesting that these tumours may be derived from progenitor cells with limited commitment. Antibody selection and protocol optimization are mandatory for reliable immunohistochemistry results.

[Histopathology of central nervous system cavernomas]. / Anatomie pathologique des cavernomes du système nerveux central.

Central nervous system cavernomas are vascular malformations, which occur in two circumstances: sporadic forms and familial autosomal dominant forms. The lesion consists of enlarged, closely packed vessels without interposition of brain parenchyma, surrounded by hemosiderin and gliosis, calcified in few cases. In 80% of sporadic forms the lesion is unique, multiple lesions are rare (median: 4). In familial forms the lesions are always multiple. Cavernomas are often associated with other vascular malformations, especially with venous developmental anomalies. The size of cavernomas is variable from 1 mm to several centimeters. About 70% of cases are supratentorial and 30% in the posterior fossa, particularly in the brain stem. Macroscopic and histopathological findings are typical and the diagnostic is generally easy.

Ependymomas with neuronal differentiation: a morphologic and immunohistochemical spectrum.

The category of mixed glioneuronal tumors of the CNS is rapidly losing its definition as encompassing tumors composed of histologically distinct neuron variants and glia. We encountered five ependymomas with neuronal differentiation seen in two by histology, in two by immunohistochemistry alone, and in one by electron microscopy. Antibodies against GFAP, S-100 protein, neurofilament protein, chromogranin, synaptophysin, Neu-N, and EMA were applied. Ultrastructural studies were also performed. In addition, 33 randomly selected ependymomas of various histologic types were screened for these same antigens. Cases 1 and 2 were anaplastic and showed clearly defined neuropil islands or pale islands as in nodular desmoplastic medulloblastoma, respectively. The tumors affected a 16-year-old male and a 5-year-old female and involved the right frontoparietal lobe and fourth ventricle, respectively. The islands were positive for synaptophysin and Neu-N (cases 1 and 2), and chromogranin (case 1). Cases 3-5, as well as 7 of the 33 screened ependymomas, showed a suggestion of neuronal differentiation by immunohistochemistry alone, including immunoreactivity for Neu-N (n = 8), synaptophysin (n = 4), neurofilament protein (n = 4), and chromogranin (n = 2). Five tumors each were WHO grade II and III. Electron microscopy performed on the two cases with neuronal islands demonstrated microtubule bundles and dense core granules (case 1) and poorly differentiated cells with high nuclear/cytoplasmic ratios, with intermediate filament accumulation and rare cilia (case 2). Cases identified by immunohistochemistry or electron microscopy demonstrated dense core granules (n = 5) and aligned microtubules (n = 3). Neuronal differentiation occurs in ependymomas but is less frequently definitive (histologic, ultrastructural) than merely a limited immunohistochemical finding. The clinical significance of these observations is unknown but deserves further exploration.

Atomic force microscopy of height fluctuations of fibroblast cells.

We investigated the nanometer scale height fluctuations of 3T3 fibroblast cells with the atomic force microscope under physiological conditions. A correlation between these fluctuations and lateral cellular motility can be observed. Fluctuations measured on leading edges appear to be predominantly related to actin polymerization-depolymerization processes. We found fast (5 Hz) pulsatory behavior with 1-2 nm amplitude on a cell with low motility showing emphasized structure of stress fibers. Myosin driven contractions of stress fibers are thought to induce this pulsation.

Telomere length and telomerase catalytic subunit expression in non-astrocytic gliomas.

Telomerase activation has been implicated as a major factor in the development of cancer. In our previous study we reported on the telomerase activity of a variety of gliomas. To further investigate the role of telomere and telomerase regulation in the pathogenesis of non-astrocytic gliomas, we examined the telomere length and the mRNA expression of telomerase reverse transcriptase gene (hTERT) and telomerase-associated protein (hTEP) in a series of 27 oligodendroglial and 18 ependymal tumors in this study. No statistical difference was found between the mean telomere length in telomerase-positive and telomerase-negative tumors (11.5 kb vs 13.1 kb; p = 0.424), although a slightly shorter length was observed in telomerase-positive oligodendroglial tumors. mRNA expression of hTERT was highly correlated with the telomerase activity status. hTERT was expressed in 8/8 (100%) and 2/2 (100%) telomerase-positive oligodendroglial and ependymal tumors, respectively, whereas 3/6 (50%) telomerase-negative oligodendroglial tumors and no telomerase-negative ependymal tumors showed expression. In contrast, hTEP1 mRNA was widely expressed in both telomerase-positive and telomerase-negative oligodendroglial and ependymal tumors. Our data support the notion that hTERT plays a critical role in determining the enzymatic activity of human telomerase. It has recently been proposed that both p16(INK4)/Rb pathway inactivation and telomerase activity were required for immortalization of epithelial cells. Although lack of p(16INK4a) expression was detected in a substantial proportion of tumors, no correlation between the p16(INK4a) or pRb protein expression and telomerase activity was observed in our series of non-astrocytic tumors.

Class III beta-tubulin immunoreactive intranuclear inclusions in human ependymomas and gangliogliomas.

We have observed intranuclear inclusion bodies immunoreactive for the cytoskeletal protein class III beta tubulin (C3betaT) in neurons and ependymal cells of post-mortem human brain. The relationship of these inclusions, detected by light microscopy, to the intranuclear rodlets described by the classical microscopists is unknown. The present study was conducted to determine whether these proteinaceous inclusions (C3betaT-NIIs) exist in the neoplastic counterparts of these cell types. Immunohistochemical staining for C3betaT revealed intensely stained, predominantly rod-shaped intranuclear inclusions in a variable proportion of tumor cells in five of ten ependymomas. In addition, C3betaT-NIIs were encountered in less than 1% of neuronal cells in two of five gangliogliomas. This study represents the first report of tubulin-containing intranuclear inclusions in brain tumors. The functional significance of these inclusions in normal human brain and in cerebral neuroepithelial neoplasms remains to be determined.

Electron microscopy in diagnosis of tumors of the nervous system.

An abbreviated catalogue of major categories of brain tumors and ultrastructural findings regarding these neoplasms are presented in short review.

Computerized morphometric analysis of human leukemic and lymphomatous cells in various histological environments of central nervous system.

The leukemic and lymphomatous cells appear within the central nervous system (CNS) in 5 different environments: in CNS vessels, perivascular spaces, meninges, nervous tissue and in CNS hemorrhages. A computerized analysis of geometric and densitometric parameters of neoplastic cells in these compartments were done for better recognition of penetration and spreading of leukemia and lymphoma within the CNS. A post-mortem neuropathological investigations were carried out on 16 patients deceased due to acute myeloblastic leukemias (M1, M2), blastic phase of chronic myelogenous leukemia, lymphoblastic lymphoma and acute lymphoblastic leukemia. Following nuclear parameters of neoplastic cells were analyzed: area, "form factor", mean, minimal and maximal density. An evident differentiation of nuclear parameters within the CNS environments was found. The nuclei within the perivascular spaces and especially in CNS hemorrhages were significantly shrunken and dense (p < 0.01), but not evidently deformed. The intracerebral infiltrates appeared to be most differentiated group (p < 0.01). Morphometric values of leukemic and lymphomatous cells show regressive changes of neoplastic cells within the CNS perivascular spaces, nervous tissue and in CNS hemorrhages. These changes depend on unfavorable factors in the mentioned CNS environments, and also on time of cell persistence in these regions. Meninges were found to be the only CNS structure facilitating the survival and proliferation of leukemic and lymphomatous cells.

Electron microscopy in tumor diagnosis: indications for its use in the immunohistochemical era.

The electron microscope reveals much more information on tumor cell structure than can be obtained with the light microscope, and some of the data are useful in diagnostic studies. In this review, major contributions of electron microscopy in the main tumor categories are discussed. To select between immunocytochemistry and electron microscopy, the probable contributions of each in the context of the particular case must be assessed. Usually, electron microscopy will only be requested after a battery of immunostaining procedures has been performed and found to be insufficient, but there are occasions when ulstrastructural study is logically the first choice after routine light microscopy. It is worth taking tissue for possible electron microscopy from any tumor that is known or anticipated to be a diagnostic problem.

Ganglioglioma: an ultrastructural and immunohistochemical study.

BACKGROUND: Ganglioglioma is a rare, mixed neuronal-glial neoplasm of the central nervous system that occurs in young patients and has a benign clinical course. METHODS: To define the immunophenotypic and morphologic features of ganglioglioma precisely, 27 specimens were studied by routine histochemistry, 21 specimens by immunochemistry, and 14 specimens were examined at the ultrastructural level. RESULTS: The age of the 27 patients, 14 males and 13 females, ranged from 3 to 52 years (mean, 22 years). The most commonly affected site was the temporal lobe (13 patients). Three patients experienced a local recurrence. Microscopically, the tumors were comprised of well differentiated, somewhat abnormal neurons as well as glial cells, the latter including astrocytes of fibrillary (59%) and pilocytic (41%) type. Scant mitotic activity was observed in 2 tumors (7%). Glial cells of all tumors were immunoreactive for glial fibrillary acidic protein, S-100 protein, and vimentin. Ki-67 labeling indices (LI) ranged from 0.6 to 10.5% (mean, 2.7%) and p53 LI from 1.1 to 42.4% (mean, 15.6%). Ki-67 and p53 LI in recurrent tumors were significantly higher than those of nonrecurrent ones (P = 0.036 and 0.026, respectively). No examples of anaplastic transformation were encountered. Immunohistochemically, many neuronal cells were positive for synaptophysin (100%), Class 3 beta-tubulin (100%), neurofilament protein (90%), and chromogranin A (86%), in addition to S-100 protein (71%) and, occasionally, vimentin (24%). Ultrastructural characteristics of neuronal cells included the presence of numerous, 100-230-nanometer dense core granules within both perikarya and cell processes, well developed rough endoplasmic reticulum, microtubules within cell processes, and synapses associated with clear vesicles. Astrocytic cells usually contained abundant intermediate filaments; their cell membranes, when abutting the stroma, were covered by basal lamina. CONCLUSIONS: Gangliogliomas are comprised of well differentiated neuronal cells and glial cells that are very often of pilocytic type. No cells with features intermediate between neurons and glia were observed. Neuronal cells are characterized by prominent neurosecretory features distinct from those of normal neurons in the central nervous system. Higher Ki-67 and p53 LI may indicate more aggressive behavior.

Ultrastructural characterization of oligodendroglial-like cells in central nervous system tumors.

Cells with uniform, small-round nucleus and clear cytoplasm (oligodendroglial-like cell, OLC) are commonly observed in central nervous system (CNS) neoplasm of glial and neuronal lineage, such as oligodendroglioma, clear-cell ependymoma, and central neurocytoma. Immunohistochemistry does not always contribute to the characterization of OLC because of (1) loss of antigen expression; (2) lack of specific markers for oligodendrogliomas; and (3) occasional coexpression of neuronal and glial antigens. An ultrastructural analysis associated with an immunohistochemical study of 20 cases of CNS tumors largely constituted by OLCs has been performed. Neurocytomas (12 cases), medullocytomas (2 cases), cerebral neuroblastoma (1 case), and ganglioglioma (1 case) showed OLCs with ultrastructural features of neuronal differentiation (neuritic processes, dense-core granules, synaptic structures). Oligodendroglioma (3 cases) OLCs were characterized by mitochondrial-rich cytoplasm, and ependymoma (1 case) OLCs showed microrosettes and scattered cilia. The electron microscopic analysis can provide a more precise diagnosis of these OLC-containing tumors despite their uniform morphological appearance.

Central nervous system neoplasms: indications for electron microscopy.

Diagnostic dilemmas of biopsy specimens in the central nervous system (CNS) tumors are often the result of multiple factors, including fixation artifact, biopsy size, lack of immunohistochemical techniques to distinguish cell types, and unawareness of rare entities. Correct diagnosis and confirmation of diagnosis of primary CNS neoplasms is imperative and may require electron microscopic examination. In some instances, use of electron microscopy may be the only approach for accurate recognition of an entity. Although diagnostic electron microscopy is expensive and cost cutting is encouraged in today's practice of medicine, cost must be weighed against the consequences of even 1 patient developing CNS treatment-related necrosis or a radiation-induced neoplasm secondary to misdiagnosis of a benign entity. This study reviews the ultrastructural differences of three groups of diagnostically difficult CNS lesions: clear cell neoplasms (ependymoma, oligodendroglioma, central neurocytoma), rare entities containing astrocytes invested by a basal lamina (pleomorphic xanthoastrocytoma, the desmoplastic neuroepithelial tumors of infancy), and benign entities characterized by transitional cell forms (subependymoma, subependymal giant cell astrocytoma).

Intracytoplasmic lumina in ependymomas: an ultrastructural study.

Intracytoplasmic lumina (ICL) are rarely described in tumors of the central nervous system. The morphogenesis of ICL remains incompletely characterized. Ultrastructural features of ICL in six ependymomas (one from lateral ventricle, three from fourth ventricle, and two from spinal cord) and three myxopapillary ependymomas of the filum terminale were analyzed. Two types of ICL were identified: ICL with both microvilli and cilia, and ICL with only microvilli. Some ICL also contained granulofibrillary or condensed material. Ciliated ICL were common in ependymomas of the ventricles, whereas nonciliated ICL were frequently seen in the myxopapillary variant. Various stages of formation and development were observed in ciliated ICL. They seemed to originate from distended periciliary cisterns, to enlarge by fusion with cytoplasmic vesicles or other ICL, and subsequently to open into the intercellular space. The last process may be the mechanism by which the intercellular microrosettes are formed. Ciliated ICL have not been described in other neoplasms. They may represent a characteristic ultrastructural feature of ependymomas. The morphogenesis of nonciliated ICL remains unknown. They may represent a degenerative form of ciliated ICL or pseudolumina resulting from invagination of the extracellular space within the cytoplasm.

Proliferating cell nuclear antigen and nucleolar organizer regions in CNS tumors: correlation with histological type and tumor grade. A comparative study of 82 cases on paraffin sections.

We investigated the expression of proliferating cell nuclear antigen (PCNA) and the number of nucleolar organizer regions (NORs) in 82 cases of CNS tumors. PCNA is a nuclear protein maximally elevated in the S phase of the cell cycle and recognized immunohistochemically in paraffin sections by the monoclonal antibody PC-10. On the other hand, NORs are loops of DNA that carry the rRNA genes and can be demonstrated in paraffin sections using an argyrophilic method (AgNORs). The present study shows a significant correlation of PCNA index and of AgNOR number with the histological grade (PCNA: I versus II, p < 0.01; II versus III, p < 0.01; and III versus IV, p < 0.05; AgNORs: I versus II, p < 0.001; II versus III, p < 0.05; and III versus IV, p < 0.001). Higher values of PCNA index (0.01 < p < 0.05) were found in recurrent tumors. Metastatic carcinomas were characterized by high PCNA indices and AgNOR numbers, similar to grade IV tumors, whereas in CNS lymphomas the malignancy grade was reflected in PCNA indices and AgNOR numbers. A wide range of PCNA and AgNOR values has been observed within each histological type and grade, probably reflecting variations in the biological behavior, but little overlap in PCNA values was present between grades II and III. The latter finding might be of importance in distinguishing between low- and high-grade CNS tumors. The linear regression coefficient between PCNA index and AgNOR number was excellent (0.91). We suggest that PCNA and AgNORs may be successfully applied in routine material to assess the growth potential of CNS tumors. Their prognostic value, however, must be validated with clinical studies.