PATHOlOGY
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Signet-ring Cell Ependymoma: Case Report with Implications for Pathogenesis and Differential Diagnosis Istvan Vajtai 1, Zoltan Mucsi 2, Zsuzsanna Varga 3 and Istvan B6di 1 Departments of ' Pathology and ' Neurosurgery, Albert Szent-Gyorgyi University Medical School, Szeged, Hungary ' Institute of Clinical Pathology, Un iversity Hospita l Zurich, Zurich, Switzerland
Summary
Introduction
We describe light microscopic, immunohistochemical and ultrastructural features of a signet-ring cell ependymoma (WHO grade II) identified in a surgically resected left cerebellar cystic tumor from a 64-year-old man . Part of the tumor showed clear-cell differentiation. Immunohistochemical coexpression of glial fibrillary acidic protein and epithelial membrane antigen, characteristic of ependymoma, was detected in both components. Sinuous intermediate junctions, cytoplasmic lumina, and scant astroglial filaments were demonstrated by electron microscopy. Signet-ring cell change was shown to be induced by disproportionate cavitation of either microvillus-bearing cytoplasmic lumi na or microrosettes. The staining qualities of clear eells were mainly due to paucity and degeneration of subcellular organelles. Therefore, signet-ring cell ependymomas represent a unique anomaly of intra- and extracellul ar compartmentali zation to be distinguished from various unrelated forms of cytoplasmic volume increase, resulting in an optically simi lar "empty" appearance of tumor cell s. As a clinically relevant consequence, signet-ring cell ependymoma must be included in the differential diagnosis of primary or metastatic neoplasms of the central nervous system, having in common a phenotype characterized by overdeveloped optically lucent cell bodies.
Despite the uniform presentation of most ependymomas as pseudorosette-forming glial neoplasms evolving along the ventricular system and the spinal canal, some rare examples may occasionally show unusual features with regard to their topography or histOlogical composition [lJ. The latter involves a subset of tumors with optically empty-appearing cytoplasm including the c1earcell, the lipidized, and the recently described signet-ring cell variants [2, 4, 8, 14/. Such tumors must be differentiated from glial, neurocytic, meningothelial and angiogenic lesions as weB as from metastatic carcinoma [I, 3, 6, 10, 11 , l3J. In addition, despite the apparent similarities of their microscopic appearance, clear-cell and signet-ring ceB ependymomas themselves seem to represent distinct morphogenetic entities rather than mere variants of a common defect in cytoplasmic modeling. To our knowledge, only three cases of signet-ring cell ependymoma have been documented in the literature. Herein, we report a fourth example that also incorporates a clear-cell moiety. In the present case, electron microscopy and immunohi stochemical studies showed that signet-ring cells develop by excessive dilatation of cytoplasmic lumina and microrosettes, whereas the perinuclear cytoplasmic lucency, characteristic of c1earcell ependymoma, is rather due to paucity and distention of subcompartmentalized organelles.
Key words: Ependymoma - Signet-ring cell tumor Clear cell change - Subcellular compartments
Address for correspondence: Istvan Vajtai, M.D., Department of Pathology, Albert Szent-Gyorgyi University Medical School , Kossuth L. sgl. 40., H -6724 S7.eged, Pf.: 401, Hungary. Tel.: +36-62/455 878. Fax: +36-62/455 868. E-mai l:
[email protected]
Pathol. Res. Pract. 195: 853--858 (1999)
0344-0338199/1 95/12 -8 53 $ 12.00/0
854 . I. Vajtai et a!.
Case History The patient, a 64-year-old man, underwent two consecutive craniotomies at eight months interval for a left cerebellar cystic mass. The first exploration was non-diagnostic, yielding only acellular cyst content. The patient was readmitted to the hospital because of increasingly severe occipital headache, dizzy spells, and disabling gait ataxia. Neurological examination revealed left sided dysdiadochokinesis, deviation on tandem walking, and backward tilting in Romberg's posture. Computed tomography (CT) revealed a reexpanded left hemispheric cavity in the posterior fossa with a cystmural nodule configuration (Fig. I). Samples taken from the nodule by means of occipital craniotomy were submitted for histology.
Materials and Methods The surgical specimen was fixed in 6% buffered fonnalin, routinely processed to paraffin, and 4 J-Im serial sections were obtained from the tissue blocks. Histochemical stains includ-
ed hematoxylin and eosin (H&E), periodic-acid-Schiff (PAS)
with and without diastase pretreatment, and Gamori's silver impregnation for reticulin. For immunohistochemistry. the
following panel of primary antisera (all supplied by DAKO® Glostrup, Denmark) was applied: SIOO protein (polyclonal 1:20000), glial fibrillary acidic protein (GFAP, polyclonal I: 1(00), vimentin (monoclonal 1: 10(0), epithelial membrane
Fig. I. On postcontras! axial CT scan, a large left cerebellar cyst is evident with slightly hyperintense contours and a circumscribed enhancing muml nodule (arrowhead),
antigen (EMA, monoclonal 1:50), synaptophysin (polyclonal 1:100), neurofilament protein triplet (monoclonal 1:(00), and Ki-67 (polyclonal I: ISO). Immunolabeling was detected by a standard two-step procedure involving biotinylated swine-antirabbit or rabbit-antimouse immunoglobulins, the avidin-bi-
otin-complex method (EnVisionoo kit, DAKO®Glostrup, Den-
mark), and 3-aminoethylcarbazol as a chromogen. For ultrastructural study, selected areas were excised from the paraffin blocks and routinely reprocessed for ultrathin sectioning. Uranyl-acetate and lead-citrate contrasted grids were viewed using a Philips eM 10 transmission electron microscope.
Pathological Findings Light microscopy showed an expansive glial neoplasm of variable and moderate cellularity evolving amidst cerebellar folia. Part of the lesion was composed of reticular sheets of irregularly round clear cells with central nuclei and nondescript processes (Fig. 2e). Occasionally, a rudimentary process formation was discernible near delicate blood vessels; however, perivascular pseudorosettes were not seen. A second component was conspicuous for its loose aggregates of large cells embedded in a meshwork of glial fibrils. The majority of these cells displayed irregularly ballooned contours, an empty-appearing or coarsely vacuolated cytoplasm, and peripheral displacement of the nucleus (Figs. 2a,b). Both crescent-shaped and bizarre giant nuclei were noted. Entangled with these signet-ring cells were labyrinthine-hyalinized vessels with focal rudimentary pseudorosette formation by slender processes. In some areas, scattered clear cells mingled with the signet-ring cell population. Mitotic figures were not encountered in either subset. The same was true for vascular proliferation and necrosis. On special stains, many of the clear cells had faint diastase-sensitive PAS positive granules. No glycogen could be demonstrated in the vacuoles of signet-ring cells. Only vascular basal lamina were highlighted by the reticulin impregnation, but areas populated by tumor cells of either type remained largely unstained (not shown). Immunohistochemically, most tumor cells showed a moderately intense nondescript staining for S 100 protein; in some of the vacuoles, diffuse serum-binding type positivity was noted (not shown). Single dotlike EMA signals appeared in the cytoplasm of many c1earcells (Fig. 21). In contrast, the same antibody tended to visualize the peripheral rim of signet-ring cells, sometimes also revealing subcompartments in very large vacuoles (Fig. 2c). In both components, the GFAP and vimentin reactions labeled mainly cell processes, a fainter positivity being randomly discernible along the inner surface of cell membranes (Fig. 2d). The Ki-67 labeling index averaged less than I % throughout the
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Fig. 2. Histology and immunophcnotype of the tumor. A: Signet-ring cell area of irregularly round cells randomly clustered amidst glial fibrils. Note coarse hyalinized vasculature in upper left; H&E: x lOO. B: Plump, empty-appearing cell bodies harboring peripheral crescentic nuclei; H&E: x400. C: EMA immunoreactivity outlines cytoplasmic margins of vacuoles and occasional dots in signet-ring cells (arrowheads); x400. D: Staining for GFAP is confined to scant coarse processes emanating from nonreactive cytoplasm of signet-ring cells; x400. E: The clear-cell population is characterized by cohesive sheets of tumor cells with translucent cytoplasm evoking a honeycomb texture; H&E: x4(X). F:A minority of clear cells exhibit dotlike EMA signals; x400.
856 . I. Vajtai et al.
tumor. Antibodies to synaptophysin and neurofilament proteins were nonreactive.
Ultrastructurally, despite poor preservation of the specimens, the ependymal nature of tumor cells was shown by the concomitant presence of "zipperlike" intermediate junctions, cytoplasmic lumina and microrosettes with microvilli and cilia, as well as by scant
glial filaments (Fig. 3a). Signet-ring cells either appeared to harbor distended cytoplasmic lumina or, less frequently, were attached pairwise by zonulae adherentes around oversized "microroseltes" (Figs. 3b, d). In either event, the vacuoles, lined by few degenerated microvilli, appeared to be devoid of any specific subcellular constituents (Fig. 3c). Clear cells, on the other hand,
Fig. 3. Ultrastructure of signet-ring cell ependymoma. A: Sinuo~s ("zipperlike") intermediate junctions and intracytoplasmic lumina crowded with microvilli were an ubiquitous finding in both clear cells and signet-ring cells; x2S.000. B: Paucity of organelies and distended rER (arrowheads) underlie the opticaliy empty appearance of clear celis; x2600. C: Detail of ballooned intracytoplasmic lumen with scant degenerate microvilli (arrowhead) in signet-ring cell; x25000. D: Pairwise coupling of tumor cells around dilatated intercellular lumen produces ependymal microrosette with signet-ring cell morphology; x870.
Signet-ring Ependymoma - 857
were found to be less voluminous, with a central nucleus located in a poorly osmiophilic cytoplasm with scant organelles and moderate amounts of glycogen (Fig. 3b). Small densely ciliated cytoplasmic lumina were also evident in these cells. luxtanuclear blepharoplasts could be identified only in rare clear cells.
Discussion The term "signet-ring cell" customarily denotes ballooning of the cell body by excessive accumulation of secretory products, metabolites, or hyperplastic organelles with concomitant displacement of the nucleus. With mucinous gastric adenocarcinoma being the prototype, the same phenomenon has been shown to playa role in a variety of visceral and brain neoplasms [3, lOJ. The latter groups include well-known lesions such as lipomatous medulloblastoma of adults, lipidized astrocytic tumors, the lipoblastic meningioma, signet-ring cell oligodendrogliomas, and rare examples of ependymoma with profuse storage of neutral fat in tumor cells [5-8, II, 13]. A peculiar mechanism of signet-ring cell development in ependymomas was described by Hirato et al. and Zuppan et al. [2, 14J. The tumor reported by the former group was identified in the left occipital lobe of a 2-year-old girl. Although it recurred following subtotal resection and radiotherapy, radical surgery eventually resulted in apparent cure. The lesions documented by Zuppan et aI., on the other hand, developed in the peri ventricular area of the left parietooccipital region of a 12-year-old girl, and in the 4,h ventricle of a 44-yearold woman, respectively. No further clinical data have been provided by these authors. In these cases, unlike the examples mentioned above, signet-ring like modeling of the cell body is not due to an increase in cytoplasmic volume. It rather proceeds by excessive sequestration of part of the extracellular space normally enclosed within cytoplasmic lumina and microrosettes of neoplastic ependymal cells. This assumption is supported by the observation in the aforementioned papers as well as by our own finding of cavities studded with degenerated microvilli and, occasionally, cilia, the normal components of ependymocytic lumina and microrosettes. The dissimilarities in the patterns of EMA immunostaining of ordinary and clear-cell ependymomas, on the one hand, and those seen in signet-ring cells, on the other hand, substantiate this hypothesis. Dotlike in the former two, while peri vacuolar in the latter, EMA immunoreactivity indeed identifies ependymal lumina and microrosettes of various sizes as a specialization of the apical membrane in normal and neoplastic ependymal cells [12/. Certainly, cavities of either origin may subsequently enlarge through a process of fusion, as also suggested by the observation of multilocular cells in the
present case [9J. In addition, Hirato et al. noted that distended lumina may also communicate with the endoplasmic reticulum (ER) system [2]. Also noteworthy in this respect is that distended cisterns of ER were found to underlie the signet-ring morphology of tumor cells in a case of anaplastic oligodendroglioma, as described by Mikami et al. [7]. The participation of the intracellular compartment in signet-ring cell development should therefore not be entirely ruled out in ependymomas. It seems, however, that cavitation of cytoplasmic lumina and microrosettes causes the basic alteration, possibly initiated by a defect in transmembrane fluid traffic or apical memhrane instability. Whatever their pathogenesis is, signet-ring cell ependymomas may be a source of differential diagnostic difficulties. Prognostic implications associated with this phenotype are also at issue. As regards the first point, the most notable "Iookalikes" include metastatic adenocarcinoma, hemangioblastoma, lipomatous medulloblastoma, the lipoblastic and clear cell variant of meningioma, and even pilocytic astrocytoma - if the vacuoles are misinterpreted as microcysts. Negative PAS staining readily excludes metastatic adenocarcinoma along with clear cell meningioma [11. Unlike lipomatous medulloblastoma, ependymomas as a group are negative for neuronal markers /1, 11/. The lack of an intricate reticulin pattern in signet-ring cell ependymoma may be used to differentiate it from hemangioblastoma [1]. Iffeasible, fat stains may further enhance diagnostic accuracy [6, 8]. Lastly, signet-ring cell ependymoma will be diagnosed because of the unique constellation of its ultrastructural hallmarks [9, 14J. Data relevant to the biological potential of signetring cell ependymoma have been provided in only one report: the tumor described by Hirato et al. [2J recurred nine months after subtotal resection, but radical second surgery appeared to be curative [2]. It may be prudent to consider prognostic connotations associated with the signet-ring phenotype in the broader context of an inconsistent correlation between histology and clinical behavior, for which ependymomas are notorious [I]. Unlike signet-ring cell oligodendroglioma, some examples of which have been reported to be equally aggressive, signet-ring cell ependymoma can be regarded as a histologically low-grade tumor. Because it is less known, it carries a great risk of being a diagnostic pitfall.
Acknowledgements, We wish to thank Ms. K. Pinter, Ms. M. Labdy, and Mrs. M.L. Elhardt for their help with histology, immunohistochemi stry, and electron microscopic procedures. The illu stration s have been technically processed by Mr. M. Dezso. We arc grateful to Professor A. T6szegi and Professor M. Bodosi for their support.
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Received: June 9,1999 Accepted in revised version: August 2, 1999
