Primitive Neuroectodermal Tumors: Ultrastructural and Immunohistochemical Studies

We report here ultrastructural and immunohistochemical studies of neuroblastic differentiation in the retrospective (n = 17) and prospective (n = 26) series of primitive neuroectodermal tumors (PNETs). By electron microscopy, neuritelike structures containing parallel-oriented microtubules, adhesive plaque junctions, and pleomorphic dense-core vesicles were found in the majority of tumor specimens while synaptic specializations were very rare. By immunohistochemistry, synapto-physin appeared to be the most reliable marker for neuroblastic differentiation present in the majority of tumors, while 200 kDa neurofilament protein was immunovisualized in a lower proportion of tumors. Glial fibrillary acidic protein (GFAP) was expressed in both reactive astrocytes and in a small proportion of otherwise typical neoplastic cells. We conclude that the majority of PNETs revealed diverse differentiation and that electron microscopy is still the most reliable tool for its detection followed by immunohistochemistry for synaptophysin.     

Primitive Neuroectodermal Tumors of Bone and Soft Tissue

Primitive neuroectodermal tumors (PNET) of the bone and soft tissue were reviewed by immunohistochemistry and partly by morphometry, focusing particularly on histologic changes in recurrent or metastatic foci, in order to elucidate their probable histogenetic relationship with Ewing's sarcoma (ES) and its extraskeletal counterpart (EES). Eleven cases of bone tumor (average patient age; 15.1 yr) and 12 cases of soft tissue tumor (average patient age; 22.1 yr) which disclosed unequivocal Homer Wright rosettes and/or at least foci of ganglion cell differentiation either in a given primary tumor or metastatic (or recurrent) foci were selected from small round cell tumors primarily categorized as ES or EES. Most of the cases for which follow up biopsy samples were available disclosed prominent Homer Wright rosettes in the metastases, whereas the primary tumors showed features of ES and lacked rosettes. In only one case, Homer-Wright rosettes were absent in the metastatic tumor. Most cases had been treated by combined intensive chemotherapy and radiotherapy, which might have influenced cell differentiation. Neural markers (neuron-specific enolase, neurofilament protein and others) were positive in most cases. Three cases with otherwise typical histologic features of ES or EES showed minute foci of ganglion cell differentiation, as confirmed by morphometry and neural markers. These results suggest that ES (or EES) and PNET are histo-genetically related, but represent different stages of cell differentiation.     

Central Nervous System Primitive Neuroectodermal Tumors: A Clinicopathologic and Genetic Study of 33 Cases

Central nervous system (CNS) primitive neuroectodermal tumors (PNETs) include supratentorial, brain stem, and spinal cord tumors with medulloblastoma‐like histopathology. The prognostic impact of various pathologic and genetic features has not been thoroughly investigated. After re‐diagnosis of three infantile cases as atypical teratoid/rhabdoid tumor (AT/RT), 33 remaining CNS PNETs were retrieved for clinicopathologic and fluorescence in situ hybridization studies. Anaplastic and/or large cell features were seen in 18 of 33 (55%) examples and survival was decreased in these patients (P = 0.036). MYCN or MYCC gene amplifications were noted in about half, with a trend towards decreased survival (P = 0.112). Polysomies of chromosomes 2 and 8 were each individually associated with decreased survival in children, with an even stronger association when combined (P = 0.013). Neither EWS gene rearrangements, nor AT/RT‐like 22q deletions were encountered. We conclude that in CNS PNET: (i) routine application of INI1 immunohistochemistry helps rule out AT/RT, particularly in infants; (ii) MYC gene amplifications (especially MYCN) are common; (iii) involvement of CNS parenchyma by Ewing sarcoma/peripheral PNET is rare enough that EWS gene testing is not necessary unless significant dural involvement is present; and (iv) both anaplastic/large cell features and polysomies of 2 and 8 are associated with more aggressive clinical behavior.     

Atypical Primitive Neuroectodermal Tumors Comparative Light and Electron Microscopic and Immunohistochemical Studies on Peripheral Neuroepitheliomas and Ewing's Sarcomas

Recently, primitive neuroectodermal tumors (PNETs) have been shown to cover a wide spectrum of small round cell sarcomas, probably including some Ewing's sarcomas (ESs) and extraskeletal Ewing's sarcomas (EESs), in addition to classical peripheral neuroepitheliomas (PNs). In studies of small cell sarcomas, we found a group of undifferentiated tumors resembling PNETs with some features of neuroectodermal differentiation, but possessing areas of relatively large, pleomorphic cells. To clarify the nature of these tumors and their relationship to PNETs, we examined the variety of histological, immunohistochemical and ultra-structural features of 11 small cell sarcomas. Five of these tumors were composed of uniform, small round cells and were classified as PNs because of the presence of definite Homer-Wright rosettes and fibrillary processes. The presence of well developed neurite like processes containing neurosecretory granules and immunore-activities for various neural markers suggested that these PNs showed more advanced neuronal differentiation. Two tumors, with the classical features of ES, showed no ultrastructural evidence of neuronal differentiation, although only gamma gamma neuron specific enolase (NSE) positivity was detected. Four undifferentiatied tumors with atypical features, included in this study as an atypical PNET group, showed certain neuroectodermal characteristics, such as ganglion cell differentiation, perivascular pseudorosettes, and gamma gamma NSE reactivity. It is concluded from this study that PNETs may include small round cell tumors showing different degrees of neuroectodermal differentiation and some histological variations. Acta Pathol Jpn 41: 444–454, 1991.     

Primary Cerebral Neuroectodermal Tumors: Neuroblastoma,Differentiated Neuroblastoma,and Composite Neuroectodermal Tumor

Seven cases of primary cerebral neuroectodermal tumors with predominant neuroblastic features were studied ultrastructurally and five were evaluated immunohistochemically. The fine structural features were indicative of neuroblastic differentiation by the presence of elongated cytoplasmic processes, electron-dense neurosecretory granules, and neurotubules. Five of the seven cases had the morphologic findings of classic cerebral neuroblastoma, and the sixth case, originally diagnosed as an oligodendroglioma, had the features of a differentiated neuroblastoma. Desmoplastic and/or stromal foci were intermingled with neuronal-ganglionic cells and neuroblasts in the seventh case. In addition to strong immunoreactivity for S-100 protein and glial fibrillary acidic protein in the desmoplastic areas, the spindle cells had fibroblastic and Schwannian features by electron microscopy in the latter case. The neuroblastic cells and fibrillary network were immunoreactive for neuron-specific eno-lase and neurofilament in the five study cases. It is concluded that cerebral neuroectodermal tumors may express an range of phenotypic features from the exclusive neuroblastic stage to a neuronal and stromogenic phase analogous to the classic neuroblastoma of the sympathetic nervous system.     

Primary Cerebral Neuroectodermal Tumors: Neuroblastoma, Differentiated Neuroblastoma, and Composite Neuroectodermal Tumor

Seven cases of primary cerebral neuroectodermal tumors with predominant neuroblastic features were studied ultrastructurally and five were evaluated immunohistochemically. The fine structural features were indicative of neuroblastic differentiation by the presence of elongated cytoplasmic processes, electron-dense neurosecretory granules, and neurotubules. Five of the seven cases had the morphologic findings of classic cerebral neuroblastoma, and the sixth case, originally diagnosed as an oligodendroglioma, had the features of a differentiated neuroblastoma. Desmoplastic and/or stromal foci were intermingled with neuronal-ganglionic cells and neuroblasts in the seventh case. In addition to strong immunoreactivity for S-100 protein and glial fibrillary acidic protein in the desmoplastic areas, the spindle cells had fibroblastic and Schwannian features by electron microscopy in the latter case. The neuroblastic cells and fibrillary network were immunoreactive for neuron-specific eno-lase and neurofilament in the five study cases. It is concluded that cerebral neuroectodermal tumors may express an range of phenotypic features from the exclusive neuroblastic stage to a neuronal and stromogenic phase analogous to the classic neuroblastoma of the sympathetic nervous system.     

Loss of INI1 Protein Expression Defines a Subgroup of Aggressive Central Nervous System Primitive Neuroectodermal Tumors

Pediatric embryonal brain tumors can be difficult to classify. Atypical teratoid rhabdoid tumors (ATRT) contain rhabdoid cells, while primitive neuroectodermal tumors (PNETs) are composed of “small round blue cells.” Loss of INI1 is a common event in ATRT; therefore, we investigated if the loss of INI1 protein expression was also observed in central nervous system (CNS) PNET and pineoblastoma. A histological review of 42 CNS PNETs and six pineoblastomas was performed. INI1 expression was assessed by immunohistochemistry. Sequencing was performed on the mutational hotspots of INI1. INI1‐immunonegative tumors were further investigated using fluorescence in situ hybridization. Epithelial membrane antigen (EMA) protein expression was assessed in six CNS PNETs to further define the phenotype. Five CNS PNETs without rhabdoid cell morphology were immuno‐negative for both INI1 and EMA. Of these primary CNS PNET patients, three died <11 months postdiagnosis, which was dissimilar to the INI1‐immunopositive primary CNS PNETs where 18/24 (75%) patients were alive 1 year postdiagnosis. We have identified a small subgroup of CNS PNETs which lack INI1 protein expression, but have no evidence of rhabdoid cell morphology. INI1 protein loss may occur through mechanisms other than gene deletion. INI1 immunohistochemistry should be performed for all CNS PNET cases.     

Mediastinal Tumors: Ultrastructural and Immunohistochemical Evaluation of Intermediate Filaments as Diagnostic Aids

The histogenesis of six mediastinal tumors was investigated ultrastructurally and immunohistochemically using monospecific antibodies against intermediate filament proteins. Four of the tumors, showing different appearances by light microscopy, displayed desmosomes and cytoplasmic tonofilaments, by electron microscopy, compatible with an epithelial thymoma. These cases also showed keratin positivity by immunofluorescence microscopy. One spindle cell tumor showed zonula adherens-type junctions, prominent collections of intermediate filaments, and abundant cytoplasmic neurosecretory granules consistent with a neuroendocrine tumor. In this tumor, neurofilaments could be demonstrated by immunofluorescence microscopy, a feature also consistent with a neuroendocrine tumor. One malignant tumor, lacking tonofilaments and desmosomes but showing a few primitive junctions, did not contain keratin but showed vimentin positivity. This suggests a mesenchymal origin and a diagnosis of primitive sarcoma. These cases illustrate the diagnostic usefulness of electron microscopy and immunohistochemical evaluation of intermediate filaments.     

Sinonasal Teratocarcinosarcoma: Ultrastructural and Immunohistochemical Evidence of Neuroectodermal Origin

The authors report a case of sinonasal teratocarcinosarcoma in a 37-year-old man, which was located in the anterior skull base and extended to the right nasal cavity and paranasal sinuses. The tumor was surgically resected twice, but it could not be removed completely. Microscopically, it was mainly composed of primitive cell nests within a moderately cellular stroma. The components of squamous cell epithelia with focal teratoid appearance and adenocarcinomatous differentiation were observed. There were many rhabdomyoblasts scattered in the nests and stroma. Ultrastructurally, the primitive cells had many neural processes with parallel microtubules, resembling olfactory neuroblastoma. Rhabdomyoblasts showed various degrees of skeletal muscle differentiation. Some of the stromal spindle cells had actin filaments with dense patches and dense core granules. Immunohistochemically, the primitive cells were positive for epithelial markers, neuron-specific enolase, synaptophysin, and myogenic regulatory proteins. The rhabdomyoblasts showed immunoreactivity for myoid markers, cytokeratin, epithelial membrane antigen, and synaptophysin. Most of the stromal spindle cells were positive for smooth muscle actin, neuron-specific enolase and synaptophysin. The immunohistochemical and ultrastructural findings suggest that primitive cells had the most primitive phenotype of placodes, and support the possibility that sinonasal teratocarcinosarcoma is essentially a neuroectodermal tumor with divergent differentiation.     

Post‐chemotherapy Maturation in Supratentorial Primitive Neuroectodermal Tumors

Maturation in central nervous system embryonal tumors is an uncommon phenomenon that is mainly reported in the context of specific histological subgroups of medulloblastoma. In this report we describe two cases of histological maturation in patients with supratentorial primitive neuroectodermal tumor with strikingly different outcomes. We discuss the potential impact of such findings on treatment and outcome.     

Intracerebral Ganglioneuroblastoma with Intracytoplasmic Microtubular Aggregates: Case Report and Ultrastructural Study

Cerebral ganglioneuroblastoma is a rare entity that is not well characterized at the ultrastructural level. We report a case of such a neoplasm with some unique fine structural features. The tumor contains both small cells with little evidence of differentiation and large, bizarre multinucleate cells, many of which contain intracytoplasmic aggregates of 22- to 24-nm micro-tubules that displace the nucleus and cytoplasmic organelles to the periphery of the cell. Inclusions of this type have not previously been described in this entity.     

Intracerebral Ganglioneuroblastoma with Intracytoplasmic Microtubular Aggregates: Case Report and Ultrastructural Study

Cerebral ganglioneuroblastoma is a rare entity that is not well characterized at the ultrastructural level. We report a case of such a neoplasm with some unique fine structural features. The tumor contains both small cells with little evidence of differentiation and large, bizarre multinucleate cells, many of which contain intracytoplasmic aggregates of 22- to 24-nm micro-tubules that displace the nucleus and cytoplasmic organelles to the periphery of the cell. Inclusions of this type have not previously been described in this entity.     

Ultrastructural Pathology of Glial Brain Tumors Revisited: A Review

The ultrastructural pathology of primary brain tumors of glial origin is examined. These are divided into two major groups. The first category comprises astrocytoma with the variants: fibrillary, protoplasmic, gemistocytic, and anaplastic. These are biologically aggressive tumors of a relatively high proliferative potential and include a substantial proportion of cases that transform into the most malignant secondary glioblastoma. The second category, comprised of rather benign tumors of a limited proliferative capacity and a reasonable good prognosis, includes such clinico-pathological entities as pilocytic astrocytoma, pleomorphic xanthoastrocytoma, and subependymal giant cell astrocytoma of tuberous sclerosis. There is no ultrastructural feature, however, which makes it possible to discriminate between major subclasses of astrocytes; but secondary glioblastoma cells, while still retaining the stigmata of neoplastic astrocytes, are characterized by nuclei that seem to be more indented, cisterns of the endoplastic reticulum may be distended, and intranuclear pseudoinclusions are frequently observed. Primary glioblastoma, which probably originates de novo, is characterized by poorly differentiated cells with a paucity of subcellular organelles and no obvious features of astrocytic origin. Granular cell tumor also belongs to neoplasms of astrocytic lineage and the hallmark of this entity is a cell characterized by the presence of numerous membrane-bound, electron-dense autophagic vacuoles. Its malignant analogue is the granular cell glioblastoma. Two subtypes of granular cell glioblastoma have been distinguished. The first is characterized by the presence of numerous granular, electron-dense bodies which correspond to autophagic vacuoles. The second type is characterized by numerous electron-dense, amorphous masses within cellular processes. These electron-dense inclusions are virtually indistinguishable from minute Rosenthal fibers. The pilocytic astrocytoma is virtually indistinguishable at the ultrastructural level from fibrillary astrocytomas but cells tend to be more elongated. Besides Rosenthal fibers, two types of distinctive structures are relatively common in pilocytic astrocytomas: eosinophilic hyaline droplets and round granular bodies, which are composed of large aggregates of electron-dense secondary lysosomes or small electron-dense bodies, respectively. Pleomorphic xanthoastrocytoma is characterized by astrocytes surrounded by basal membranes. It belongs to a peculiar category of astrocytic “desmoplastic” brain tumors occurring in younger patients, the common denominator for which is the presence of basal lamina. The last category in this group is subependymal giant cell astrocytoma, a tumor of bivalent (glial and neuronal) differentiation, the cells of which are characterized by the presence of peculiar crystalloids. The hallmark of oligodendroglioma is the presence of concentric arrays of membranes (so-called membrane laminations, whorls, or scrolls). A fragment of the cytoplasm sequestrated within a particular whorl may contain mitochondria, lysosomes, or abundant glycogen granules. Ependymomas are characterized by a florid picture dominated by the presence of microlumina, cilia with basal bodies (blepharoplasts), microvilli, and long, interdigitating intercellular junctions of the zonulae adherentiae type. Ganglioglioma, the last category covered by this review, is a mixed glio-neuronal tumor. While glial cells are indistinguishable from their counterparts encountered elsewhere (mostly pilocytic astrocytes), the ganglion cells are characterized by abundant intracytoplasmic dense-core vesicles, absence of intermediate filaments, and numerous microtubules. Occasionally a close apposition of ganglion cells and Rosenthal fibers is seen. Dense-core vesicles are pleomorphic and ranged in a diameter from small synaptic vesicles to large lysosome-like neurosecretory granules.     

Ultrastructural and Immunohistochemical Features of Common Lung Tumors: An Overview

Lung cancer is rapidly becoming the most commonly diagnosed malignant neoplasm in the world. Pathologists play a key role in the care of patients with lung cancer by accurately classifying and staging these neoplasms. Immunohistochemistry and electron microscopy have become important tools in the diagnosis of lung tumors, especially those which are histologically undifferentiated. This review discusses the ultrastructural and immunohistochemical features of common lung tumors, with an emphasis on their diagnostic usefulness.     

Paragangliomas of the Head and Neck: Ultrastructural and Immunohistochemical Analysis

Eighteen head and neck paragangliomas were studied by light microscopy and light microscopic immunohistochemistry by the peroxidase technique for the presence of NSE (neuron-specific enolase), serotonin, and a battery of neuropeptides. Seven of these tumors were also studied by electron microscopy.

All 18 cases demonstrated immunostaining for NSE; 10 of the 11 carotid body tumors had immunostaining for multiple hormones. Considering all 18 cases, the most frequently demonstrated hormonal substances were in order: serotonin, leuenkephalin, gastrin, substance P, vasoactive intestinal polypeptide (VIP), somatostatin, bombesin, calcitonin, and alpha MSH. In several tumors, adjacent-step sections stained for different hormonal substances strongly suggested reactivity for more than one hormone in given tumor cells.

By electron microscopy, all 7 cases studied displayed considerable heterogeneity of the neurosecretory granules with respect to size, shape, and electron density. This demonstrated that branchiomeric paragangliomas are capable of producing a spectrum of neuropeptides in addition to their known amine content. The presence of immunoreactive serotonin in most of these neoplasms was confirmed. In addition to these findings, neurofibrils within the substance of carotid body paragangliomas demonstrated immunoreactivity for somatostatin and a gastrinlike neuropeptide. The significance of the neuropeptides in these neoplasms and their possible presence and role in normal and hyperplastic paraganglia remain to be defined.     

Paragangliomas of the Head and Neck: Ultrastructural and Immunohistochemical Analysis

Eighteen head and neck paragangliomas were studied by light microscopy and light microscopic immunohistochemistry by the peroxidase technique for the presence of NSE (neuron-specific enolase), serotonin, and a battery of neuropeptides. Seven of these tumors were also studied by electron microscopy.

All 18 cases demonstrated immunostaining for NSE; 10 of the 11 carotid body tumors had immunostaining for multiple hormones. Considering all 18 cases, the most frequently demonstrated hormonal substances were in order: serotonin, leuenkephalin, gastrin, substance P, vasoactive intestinal polypeptide (VIP), somatostatin, bombesin, calcitonin, and alpha MSH. In several tumors, adjacent-step sections stained for different hormonal substances strongly suggested reactivity for more than one hormone in given tumor cells.

By electron microscopy, all 7 cases studied displayed considerable heterogeneity of the neurosecretory granules with respect to size, shape, and electron density. This demonstrated that branchiomeric paragangliomas are capable of producing a spectrum of neuropeptides in addition to their known amine content. The presence of immunoreactive serotonin in most of these neoplasms was confirmed. In addition to these findings, neurofibrils within the substance of carotid body paragangliomas demonstrated immunoreactivity for somatostatin and a gastrinlike neuropeptide. The significance of the neuropeptides in these neoplasms and their possible presence and role in normal and hyperplastic paraganglia remain to be defined.     

Ependymoma: Ultrastructural Studies of Two Cases

Several unusual ultrastructural findings in two ependymomas are reported. In case 1, a grade I ependymoma of the fourth ventricle, there were rosettes, perivascular pseudorosettes, and tumor cells having unusual intracytoplasmic vacuoles by light microscopy. Ultrastructurally, these vacuoles were frequently microrosettes as well as scattered, degenerated cytoplasmic processes of tumor cells. The lumina of some of the microrosettes were bordered by abnormally long and malformed zonulae adherentiae. In case 2, a recurrent grade III ependymoma of the third ventricle, there were rosettes and perivascular pseudorosettes as well as more cellular and anaplastic areas by light microscopy. Ultrastructurally, the cytoplasmic processes of tumor cells in perivascular pseudorosettes contained frequent dense-core vesicles and occasional parallel arrays of microtubules. These structures do not occur in normal mammalian ependymal cells but do occur in the ependymal tanycyte, a related cell that is plentiful in the walls of the third ventricle. Thus some of the tumor cells of this third ventricle ependymoma appear to have differentiated as tany-cytes.     

Ependymoma: Ultrastructural Studies of Two Cases

Several unusual ultrastructural findings in two ependymomas are reported. In case 1, a grade I ependymoma of the fourth ventricle, there were rosettes, perivascular pseudorosettes, and tumor cells having unusual intracytoplasmic vacuoles by light microscopy. Ultrastructurally, these vacuoles were frequently microrosettes as well as scattered, degenerated cytoplasmic processes of tumor cells. The lumina of some of the microrosettes were bordered by abnormally long and malformed zonulae adherentiae. In case 2, a recurrent grade III ependymoma of the third ventricle, there were rosettes and perivascular pseudorosettes as well as more cellular and anaplastic areas by light microscopy. Ultrastructurally, the cytoplasmic processes of tumor cells in perivascular pseudorosettes contained frequent dense-core vesicles and occasional parallel arrays of microtubules. These structures do not occur in normal mammalian ependymal cells but do occur in the ependymal tanycyte, a related cell that is plentiful in the walls of the third ventricle. Thus some of the tumor cells of this third ventricle ependymoma appear to have differentiated as tany-cytes.