Ultrastructure of a fibroxanthosarcoma (malignant fibroxanthoma)

ULTRASTRUCTURE OF A FIBROXANTHOSARCOMA (MALIGNANT FIBROXANTHOMA) LEONARD P. MERKOW,RID, PHD," JOHN C. FRICH,JR., hlD, P H D , ~CONSTANTINE G. KYREACES,RID, A N D RIATIAS PARDO

MALCOLM SLIFKIN,

Primary a n d metastatic lesions from a fibroxanthosarcoma (FXS) originating i n the neck of a 29-year-old Caucasian woman a r e reported. Light a n d electron microscopic observations a r e correlated to define the various types of cells constituting this neoplasm. Ultrastructural examination verified that several types of cells can act as facultative fibroblasts. Since a malignant potentiality cannot be predicted solely from the histopathology of fibroxanthomas, a n attempt was made to categorize certain ultrastructural nuclear a n d cytoplasmic organelles associated with FXS cells. T h i s was done i n order to further define the malign a n t neoplastic cells of the FXS, so that distinguishing the malignant from henign forms of this neoplasm may become more feasible.

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tissue have been difficult to diagnose accurately. Distinguishing malignant from benign fibrous tumors has also been a problem.51p52 Stout antl co-workers have pointed out that a number of different cell types such as Schwann cells, reticulum cells, histiocytes, and mesothelial cells may act as facultative fibroblasts.",39,51-5~ This is also true of the cells' corresponding benign and malignant neoplasnis.52 Furthermore, the lattei '5 nomenclature attests to the resulting confusion i n classification. For example, the terms fibrous histiocytoma, xanthofibroma, fibrous xanthonia. xanthogranuloma, dermatofibroma, From the Sections of Surgical Pathology antl Elcctron Microscopy, Division of Experimental Pathology, William H. Singer Memorial Research Institute, and the Divisions of Radiation Therapy and Surgery, Allegheny General Hospital, Pittsburgh, Pa. * Adjunct Assistant Professor at Pennsylvania State University. Address for reprints: L. Merkow, MD, PhD, Head, Division of Experimental Pathology and Electron Microscopy, William H. Singer Memorial Research Institute, Allegheny General Hospital, 320 E. North Ave., Pittsburgh, Pa. 15212. The authors express their appreciation to Dr. Heniy L. Kazal and Dr. Joseph M. Stowell for contributing the original biopsy slides and referral of patient for radiotherapy, respectively. They are also gratefully indebted to Dr. Hernando Salazar and Dr. William E. Pod for constructive criticism of the manuscript, Miss Bernice Campbell, Miss Beth Heitzenroeder, and Mrs. ETelyn Gilbert for special histologic stains, Mrs. Jean Cox and Mrs. Pamela Schiller for their technical assistance in electron microtomy, Miss Florence AfcClure and Miss Linda Fischer for secretarial assistance, and members of the Department of Medical Illustration for the photographic reproductions. Received for publication January 15, 1971.

antl their malignant toiiiiterpi t b d l have been used to describe an essentiall) siniilar type of fibious neoplasm posessing a wide variation in cell types. T h e purpose of this report is to describe the ultrastructure of metastases from a malignant fibroxanthoma or fibroxan th osarmina (FXS). T h e information presented iiiay provide a more practical approach toward distinguishing malignant from benign neoplasms of this type.

CASEREPORT In June 1966, a 29-year-old Caucasian woman had a X.5 x 2.0 x 1.2 cni painless mass ienioved from the midline of her neck posteriorly. Grossly, the lesion was rubbery and ye]low-white. T h e histopathology h o w e d a predominance of histiocytes, fihiour tissue, and many multinucleatetl giant cells. I n A%iigiist 1966, a wbciitaneoiis neck nodule, 1.4 cin in diameter, recurred. T h e histopathology of this recurrent lesion was similar t o the original except that fewer giant cells were evident. Diagnosis at this time was locall) recurrent atypical fibrous xanthoma (probably malignant). I n June 1969, a biopsied metastatic nodule of the right chest wall showed a similar pattern to that of previous specimens, arid it was diagnosed as FXS. In February and March 1970, biopsies of the right and left lung, scalp, and left tonsil showed metastatic FXS. These lesions displayed a similar pattern to that noted previously except that multinucleated giant cells were qiiite rare. T h e patient h a d received 4,300 rads (total dose) to the chest wall to a depth of 6 cm,

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completed in August 1969. Following this, she received 1,615 rads to the entire chest, completed in September 1969. Actinomycin D, 4.5 nig. and vincristine, 3 mg, were administered on 3 separate days i n September 1969. Her WBC decreased to 3. 100/mm3, the platelet cowit to 100,000/mm3, and she subsequently developed alopecia. Slic received 5,000 rack to the niecliastinwn between March 2 and March 27, 1970, anrl 3,465 rads to the left tonsil between March 16 and April 3, 1970. She died on .4pril 11, 19TO. An autopsy was not permitted. hfATERrhLS AND METHODS

Portions of metastatic tumor from the Iiiiiga, scalp. am1 left tonsil were fixed in neutral-buffered formaldehyde solution. After fixation, tissues were dehydrated, embedded in paraffin blocks, ant1 scictioned a t 5-6 p. Sections were stained with hematoxylin and eosin, PAS. Masson’s trichrome, reticulum, a n tl plios p h o t 11 ngs t i c acid lie m a t ox y 1i n (PTAH) st:iiiis.24 For electron microsropy, samples of the nirtastatir neoplasm from the scalp and left tonsil were plared in cold (4C), 3% glutaraldehydc and 0.1 b I phosphate buffer (PH 7.4) containing 1 calcium chloride. .4fter 4 Iiotirs, the tiinior tissue was washed in 0.1 M phosphate buffer (pH 7.4), cut into I-mm cubes. and post-fixed in 1% osmium tetroxide in 0.1 M phosphate huffer (pH 7.4) with S I I crose. T h e cubes of tissue were kept in fixative and refrigerated for 1 to 2 hours. Dehydration was performed through a graded sequenre of etlianols (1.5 minutes each in 50%, 70%. 95%. IOW!.;,. 100rJ;,. and 1 0 0 ~ )T. h e samples were then placed in propylene oxide for two 15minute rhanges. Infiltration of tissue was performed with one change of 500/, propylene oxitle: 50% epoxy resin for one h o w followed by pure epoxy resin overnight. Finally, the tissue was embedded in epoxy resin (Epon 812). Sections 1-p thick were prepared and stained with methylene blue for light microscoliy. Sections displaying silver or gold interfercnce rolors were obtained with an ultramicrotome (Porter-Blum M T 2 or Reichert OM-U2) equipped with glass or diamond kiiircs. IJltrathin sect.ions were stained with ;I 5y0 solution of uranyl acetate followed by lead citrate. T h i n sections were examined with 311 electron mirroscope (Philips lO0R or ; i n E~ 3 0 0 ) . Scoplxtic rells were cultured on WI-38 ant1

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HeLa indicator cells in an attempt to isolate virus. Cultures were incubated for 3 weeks in order to observe cytopathogenic effects (CPE), indicative of virus isolation.

RESULTS G,-OFSpathology: Specimens of a metastatic lesion of the scalp, 2.0 cm in diameter, consisted oE several irregular fragments of pale gray, firm tumor invading the skin and subcutaneous tissue. T h e biopsy of the left tonsil consisted of several irregular fragments of 11ale gray, soft tissue, 2.0 cm in aggregate. I.ight micioscopy: Paraffin-embedded sections stained with heniatoxylin and cosin from the initial biopsy of the neck lesion showed numeyous neoplastic rells resembling histiocytes or spindle shaped fibroblasts, and two types of giant cells arranged in a n ill-defined sheet within the deep dermis (Fig. la). Mitoses were infrequent. Many neoplastic stromal cells resembled p l m i p fibroblast3 (Figs. la, b). Fibroblasts were often arranged in n storiform pattern as seen under low magnification. Other neoplastic cells more cIosely resenibletl foamy histiocytes (Figs. l a , b). Cleft-like spaces (probably artefactual) often surrounded the niultinucleated “Touton”-type giant rells (Figs. l a , b). These giant cells clisplayed an eosinophilic, finely ngranular,homogeneous. or sometimes vacuolated cytoplasm surrounding nuclei with prominent nucleoli (Figs. l a , 11). Within the scalp and tonsillar metastases (Fig. 1b), many histioryte-like giant cells displayed slightly to moderately hypcrchroinatic vesicular nuclei that were irregular or round, occasionally bilobed, ;ind contained prominent nucleoli (Fig. 1b). These Iiinucleated giant cells were smaller than the “Touton”-type rells and resembled SternlmgReed cells (Fig. 1b). hldtinucleated giant cells (“Touton” type) were less frequeiltly eiicountered anrl, when found, contained less 1111clei in the metastatic lesions (Fig. Ib) than i n the initial surgical specimens (Fig. 1a). Tlie fibruns stromal element again revealed a storiform pattern, and mitoses were infrequent. Some histiocytes rontained PAS-psi tive cytoplasmic material. T h e P T A H stain was negative. Large areas of the neoplasm were devoid of an affinity for the reticulum stain. Epoxy resin embedded 1-p thick sections of a metastasis showed a mixture of light and dark neoplastic cells (Figs. ?a, b). T h e cytoplasm of the light stainiiig cells was granular

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FIG. l a (top). Original biopsy of posterior neck mass in June 1966. Eosinophilic neoplastic histiocytes with poorly defined cell borders and vesicular nuclei are intcrrnixed with plump fibroblasts. Some cells display a relatively translucent cytoplasm (arrows). Three multinucleated "Touton"-typc giant cells consist of a dark homogencous granular cytoplasni and nuclei containing prominent nucleoli. Cleft-like spaces are noted about these giant cells. At low magnification, a storiform pattern was evident (Formalin-fixed, paraffinembedded, H and E, x260); b (bottom). Metastasis to scalp in March 1970. Poorly demarcated clusters of malignant neoplastic histiocytes display either slight to moderately hyperchromatic or vesicular nuclei surrounded by homogeneous cytoplasm. Other cells resemble fibroblasts. Some giant cells are binucleated, contain prominent nucleoli, a folded nuclear outline, and resemble Stcrnberg-Reed cells (arrows). Two "Touton"-type giant cells (arrows) with a homogeneous cytoplasm, rare in metastases, display less nuclei than in the primary site (Fig. la), formalin-fixed, paraffinembedded, H and E, x300.

and surrounded a large, irregular, vesicular, light-staining nucleus with one or more nucleoli (Figs. Za, b). T h e dark-staining neoplastic cells displayed a homogeneous, nongranular, dense cytoplasm with an occasional vacuole and either a densely staining nucleus or a nucleus displaying a prominent nuclear membrane and one or more nucleoli (Fig. 2a). A third, more infrequent cellular component was a xanthoma-type neoplastic cell containing many small, light cytoplasmic vacuoles

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(Fig. 2a). Occasional neoplastic cells contained a solitary, large and/or several small cytoplasmic, deeply osmiophilic lipid bodies (Fig. 2b). Multinucleated giant cells were rare (Fig. 2b). Numerous irregular, elongated, vascular channels were noted between groups of tumor cells (Fig. 2a). Electron microscopy: A survey of neoplastic cells showed both light- and dark-staining cells (Figs. 3a-e) comparable to those observed in thick sections (Figs. 2a, b). Light tumor cells contained large, lobular nuclei with a nucleolus and one or more nuclear projections (Fig. 3a). T h e cytoplasm contained scattered solitary cisternae of rough endoplasmic reticulum (RER), several small mitochondria, and a relatively translucent, lightly granular background (Fig. 3a). Dark-staining tumor cells also had large, irregular, lobular nuclei with slightly darker staining nucleoplasm frequently containing a spheroidal nuclear body (Fig. 3a). T h e cytoplasm of dark cells contained more abundant RER and ribosomes, but only a few small mitochondria (Fig. 3a). These neoplastic cells were of a histiocytic type and frequently exhibited numerous variable sized lysosomes, lipid bodies, and a prominent Golgi complex (Figs. 3a, b, d). Cells closely resembling a typical fibroblast exhibited a n irregular, lobular nucleus, abundant parallel arrayed and interconnecting RER, many clusters of loose ribosomes, a Golgi complex, dense bodies, and bundles of delicate cytoplasmic filaments (Fig. 3c). T h e slightly dilated cisternae of RER in these fibroblasts contained osmiophilic material (Fig. 3c). Lysosomes were often entrapped between nuclear projections (Fig. 3c). Infrequently, xanthoma-type neoplastic cells were packed with clear as well as osmiophilic cytoplasmic vacuoles, in association with lysosomes, myelin figures, and cytoplasmic filaments (Fig. 3d). Some xanthoma cells contained interconnecting clusters of smooth endoplasmic reticulum (SER). Another type of dark-staining neoplastic cell was a giant cell characterized by cytoplasm nearly filled with small mitochondria and a few lysosomes (Fig. 3e). In most neoplastic cells, bundles of delicate filaments were noted most frequently in close proximity to the nuclear envelope (Figs. 3c, 4a, b, 5a, b, d, 6b, c). Frequently, groups of filaments resembled the twists and coils of a skein of yarn (Fig. 5b). Banding with a periodicity of approximately 50A was observed in focal regions of the filaments (Fig. 4b). Lat-

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era1 or cross bridging between adjacent parallel filaments was observed (Fig. 4b). Different neoplastic cells exhibited a variety of cytoplasmic organelles (Figs. 5a, 6d). Many neoplastic histiocytes displayed nonparallel-arrayed RER (Figs. Sa, b), lysosomes (Figs. Sb, d), prominent Golgi (Fig. 6b), a centriole, and multivesicular bodies. Cytoplasmic annulate lamellae were noted in some neoplastic histiocytes (Figs. 5a, b). These organelles displayed periodic constrictions in register, interlamellar granular material and, in fortuitous sections, interlamellar tubules (Fig. 5a). Annulate lamellae were associated with ribosomes at their lateral margins and showed continuity with both RER and SER (Figs. 5a, b). An enface plane of section revmled that the annulate lamellae consists of “pores” composed of numerous microcylinders (Fig. 5b). Both cytoplasmic annulate lamellae and filaments were often in close proximity to one another since both were usually found adjacent to the nuclear envelope (Fig. 5a). Frequently, clusters of closely aligned endoplasmic reticulum with smooth and/or rough components were arranged in a dense or whorled pattern (Fig. 5c). T h e whorled arrangement often circumscribed dilated vesicles. Giant mitochondria with elongated cristae traversing the entire diameter of their lurnina (Fig. 5d) were noted. Some giant mitochondria contained rod-like inclusions (Fig. 5d) or were converted by their elongated cristae to whorls of membranous elements (Fig. 5e). An occasional neoplastic cell contained a very large cytoplasmic osmiophilic lipid body surroundecl by numerous smaller lipid bodies. Infrequently, a neoplastic cell contained considerable cytoplasmic glycogen (Fig. 5f). In nearly completed dividing cells, the socalled Flemming body was observed, represented by a dense osnliophilic band with lateral processes across the mictotubules of the constricted mitotic spindle (Fig. 6a). T h e plasmalemma of adjacent neoplastic cells showed numerous interdigitation and enfoldings (Figs. 6b-d). Some finger-like projections formed microvilli (Figs. 6b, c), or projected as elongated structures from one cell into the cytoplasm of another (Fig. 6c). Where 3 tumor cells converged, the arrangement of microvilli resembled a bile canaliculus (Fig. 6d). This space sometimes contained filaments. Small desmosome-like structures arid less differentiated attachment sites adjoining neoplastic cells were also seen (Fig. 6d).

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FIG.2a (top). Light- and dark-staining neoplastic cells are intermixed with capillaries. Light-staining tumor cells contain irregular nuclei with light nucleoplasm and 1 or more nucleoli (arrow). The cytoplasm of these cells is granular. Dark cells show a dense, coarse, granular cytoplasm (arrow) and a nucleus with prominent nuclear membrane and nucleolus (arrows). A few xanthoma-type cells (circles) display numerous small cytoplasmic vacuoles (glutaraldehyde-fixed, eponembedded 1-p thick section stained with methylene blue, ~ 4 0 0 ) ; b (bottom). Cells similar to Fig. 2a. A neoplastic cell displays one large osmiophilic lipid body. Several “Touton”-type multinucleated giant cells are evident (arrows). One is closely associated with the lipid body. Epon-embedded, 1-CL thick sectioii stained with methylene blue ( ~ 3 5 0 ) .

Virus-like particles were frequently noted within neoplastic cells (Figs. 7a, b). These particles displayed an envelope and were either free within the cytoplasm (Fig. 7a) or within a cisterna of RER (Fig. 7b). Most neoplastic cells displayed a prominent nuclear envelope (Figs. Sc, 5e, 6a, d, 8a, b) due to the density of the inner nuclear membrane (Fig. Sa). In addition, spheroidal nuclear bodies (Figs. Sa, 8a) and lipid bodies (Fig. 8a) were observed within the nucleus. T h e nucleolus also infrequently contained an osmiophilic lipid body. Rarely, a nucleus contained several foci of membrane-bound delicate filaments within its nucleoplasm (Fig. 8b). These

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FIG.3d (left). A xantlioma-type cell between other neoplastic cells. This cell contains numerous clear or lightly os~niophiliclipid bodies Lb), myelin figures (Mf), lysosomes (Ly), antl filaments (Fi) within its cytoplasm (~7,500); e (rig t). A neoplastic giant cell with increased numbers of mitochondria (M) nearly filling the cytoplasm. Numerous Ijsosomes (Ly) and a nucleus (N) containing a coiled niicleolus (Nc) are e\ ident (~10,500).

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nuclciir fi1;inieiitous bodies as well as soni:: enc.ounteretl within the cytoplasm resembled a “rodlet” depending upon the plane of section. .4niorphous material resembling hy a 1‘ine replaced sinall foci of cells. Dense intercellular fibrillar material representing fibrin was observed (Figs. 3a, 5c). .lfter 3 weeks ol observation, no CPE W;IS ollserved in vitro in any indicator-tumor cell cilltures.

DISCUSSION This is the first tlescriptioii o f the fine structure of a metastatic fil~ro?tanthos~trcot~i~i (FXS). T h e metastases contained n variety oL fibrous, histiocytic, antl xanthomatow types of cells. 111 contrast, Fisher antl VuLei.ski12 found only spindle and romidecl fornis of iieoplastic cells with “histiocytic features” in it clerniatofibrosarcoma. Althoiigh cross-bantletl interstitial

Fic. Sa (fop).Surwv view of light- and dark-staining neoplastic rells. T h e light cells (lispla) proniir~rnt,lobulatcd. irregular nuclei (N) with nuclear projections (arrows). Their cytoplasm contains a relatire pancity of organelles. T h e dark tumor cells show inore rougli endoplasmic reticulum (Rer), lysosomes (Ly), arid irregular nuclei (N) containing spheroidal nuclear bodies (circlcs). Nucleoli (Nc) are dense in both types of cells. Focal deposits of fibrin (Fb) are present 1)ctwct.n cells. This, arid all sulwquent figures, represents glutaraldeliytle ~,oatosmiuin-hsetlepox) resi1i-rmhcddet11ded material stained with uranyl acetate and lead citrate (~8,000); b (bottom, [ e f t ) . .4 neoplastic his1 xocyte displaying a nucleus (N), variable-sized lysosoines (Ly), lipid bodies (Lh). :I prominent Chlgi complex (Go), small mitochondria (M), aud scattered cisternae of rough critloplasmic r c t i d u m (arrows, x15,oOO; c (botlom, ~ i g h t .A neoplastic cell resembling a fibroblast. pal allel-artayed, intcrcorinecting cisteriiae of rougli endoplastnic reticulum Rer), ribosomes ( R n p ) . lysosomes (L)), a n d cytoplasmic filaments (Fi) arc evident. T h e nuclrus (Sutlisplayr ) 1ol)uliitioil. niiclcar p q r c t i o n s , and a dense nuclrar envelope (~17.000).

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logical cell types. The ability of an array of different cell types to act as facultative fibre blasts in tumors with the “fibrosarcoma” pattern was described by Stout and co-workers.88. 39~61-63 Ozzello et al.39 showed that malignant histiocytomas and fibrous xanthomas possess the ability to phagocytize lithium carmine. Thus, as Kauffman and Stoutla indicated, all tumors of histiocytes can exercise their fibre blastic potentiality to a greater or lesser degree. By electron microscopy, certain types of organelles characterized the type of neoplastic cells within the FXS. For example, the predominating histiocyte contained numerous lysosomes and irregular, short solitary cisternae of RER. Conversely, the commonly observed fibroblast contained considerably more and parallel-arrayed RER with less lysosomes. Both types of cells usually contained cytoplasmic filaments and a lobular, irregular, enlarged nucleus. Thus, the electron microscope has confirmed and further elucidated various types of neoplastic cells in the FXS as histiocytes which possess certain attributes of the fibroblast. Cytoplasmic fibrils are almost universally observed in mesodermal cells and their n e e FIG.4a (top). A bundle of filaments (Fi) is adjacent plastic counterparts. The cytoplasmic fibrillar and in contact with the nuclear envelope (Ne). A sug gestion of periodicity is noted in focal regions (arrows), material in our case displayed areas of pe~42,000;b (botfom).At higher magnification, the periodicity (rectangle) and cross bridging (arrows) of fila- riodicity which suggest a form of primitive collagen. Cytoplasmic virus-like particles and ments (Fi) is more clearly defined (x118,OOO). their relationship, if any, to human neoplastic cells have as yet not been resolved. Indeed, fibers of long spacing collagen and unmyeli- verification that virus-like particles in our nated axons were neither observed in the FXS, tumor cells or those described by others from dermatofibroma,l* or derma tofibrosarcoma,l* human m a t e r i a l ~ ~ 9 . * 6 . 9 8 - s ~are ~~~~ actually 4~~~O we observed specialized attachment sites (des- true replicative virus particles has not been mosome-like structures) between neoplastic achieved. N o virus was isolated from in vitro cells of the FXS. growth of the FXS. By light microscopy, fibrous xanthomas, The lack of definite features by light midermatofibromas, and their malignant coun- croscopy prevented O’Brien and Stouts8 from terparts are diagnosed on the basis of morpho- distinguishing between the malignant and beb

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FIG 5a (top, left). .4 cytoplasmic annulate lamellae Al) displays the periodic constrictions in register, interlamellar granular material and interlame lar tubules (arrows). Rough endoplasmic reticulum is contiguous with this organelle a t its lateral margins (arrows). Numerous bundles of filaments (Fi) are adjacent to the annulate lamellae and nucleus (N),X25,OOO; b (top. right). An annulate lamellae (Al) in an enface plane of section shows “pores” consisting of microcylinders (arrows) and contiguity with rough endoplasmic reticulum. Many loose ribosomes are scattered between the intertwining bundles of filaments p i ) , ~ 3 4 , 0 0 0 ;c (center, leftj. a cluster of closely aligned smooth endoplasmic reticulum (Ser) is partially circumscribed by rough endoplasmic reticulum (arrows). T h e intercellular dense material represents fibrin (Fb), ~15.000; d (center, right). T w o giant mitochondria (M)display elongated aistae traversing the diameter of their lumina. T h e larger mitochondrion also contains a rod-shaped inclusion (arrow), ~23,000;e (botfom, Icff). A giant mitochondrion (M)shows remaining elongated cristae peripherally (arrow) and a central whorl of double membranes. Projections (arrows) from the nucleus (N), rough endoplasmic reticulum (Rer). and a li id body (Lb) are present (xl1,OOO); f (bottom, rtght). A neoplastic cell displays abundant cytopfknic glycogen (Gly), small mitochondria (M), a lipid body (Lb), and nucleus (N), ~7,000.

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nigri form of fibroxanl homas. Indeed, these authors found that the mitotic rate in 50/11PF was entirely unreliable.38 Kempson and KyriakosIs reported that 3 of 30 FXS metastasized and stated that, though the histology was quite suggestive of malignancy, the biological behavior of these neoplasms was of a low grade. These authors considered a storiform pattern with plump stromal cells, frequent mitoses, and multinucleated giant cells with foamy or glassy cytoplasm and bizarre, enlarged, abnormal nuclei as essential histopathologic features for a diagnosis of FXS.ls Al-

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though a storiform pattern was observed in our biopsies, the multinucleated “Touton”type giant cells did not display bizarre, enlarged nuclei. T h e “Touton”-type giant cells were similar to those Iound in a metastasizing fibroxanthoma by Rosas-Uribe et a1.44 A second type of giant cell in our case was smaller, bizarre, usually binucleated, displayed a wrinkled o r folded nuclear membrane, and resembled Sternberg-Reed cells. Neither type of giant cell showed mitotic activity. It becomes apparent then that one cannot distinguish by light microscopy between benign, atypical,*”

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FIG.Ga. Two daughter cells are connected by thc Flcmming body which is a dense region across the remaining microtubules of the mitotic spindle (arrows). 'This osmiophilic band shows 2 lateral processes. T h e nucleus (N) shows a dense inncr nuclear membrane (>: 17,500). b

FIG. 61) (top). Prominent interdigitating finger-like processes form the microvilli (Mv) between two adjacent neoplastic cells. Projections (arrows) of the nucleus (N), adjacent cytoplasmic filaments (Fi), and a Golgi complex (Go) are evident in upper cell (~14,000). FIG. 6c (center). An elongated, finger-like projection of one microvillus (Mv) is noted to extend deep into the cytoplasm of a n adjacent neoplastic cell (~20,000). FIG.6d (bottom). T h e plasmalemma displays a desmosome-like attachment site (arrow) adjoining two neoplastic cells. Smooth endoplasmic reticulum (Ser) is clustered, and inicrovilli (Mv) are sectioned so as to simulate a bile canaliculus (~15,000).

or malignant fibroxanthomas until metastases have occurred. Furthermore, the potentially aggressive final stages of FXS are not predictable on the basis of histopathology. I n contrast, electron microscopy showed enlarged nuclei with considerable lobulation. Complex spheroidal nuclear bodies,35947 nuclear proje~tions,l0.3~ and intranuclear lipid bodies36 and fibrillar elements,49 though not unique to malignant neoplastic cells, are a

manifestation of an altered nucleus frequently associated with the neoplastic process. It has been suggested that spheroidal nuclear bodies may be associated with a state of nuclear hyperactivi ty.17 T h e appearance and alteration of certain cytoplasmic organelles may also suggest malignancy. Although annulate lamellae are observed in some normal c ~ I ~ s , ~ J ~and J ~ JnonG neoplastic altered cells,3 increased numbers of

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ULTRASTRUCTURE OF A FIBROXANTHOSARCOMA Merkow et al.

this organelle are quite coininon in malignant neoplasms.7,",~,"6.".'" It is interesting that annulate lamellae have been encountered frequcntly in vivo in viral-induced tumor cells containing virus-like particles*7-?9J1 and in vitro in \.iral-infected"',~O.~O,~9 or presumably norcinfected continuous cell lines." T h e coexistence of these two structures appears to be more than coincidence. Cytoplasmic annulate lamcllae are frequently continuous with endoplasmic reticulumJ,11,~2,.I'aswas found in the FXS. T l i i i s , this organelle has been considered a s~xrializedform of endoplasmic reticulum.5. 11,43.44 T h e presence of interlamellar (intertubular) ~onnections,3(~ a dense matrix,'B ribosonies,21 and sinall granules21 in conjunction with its location, iisually in close proximity to the niiclear envelope, suggest a role in protein s y ~ t l i e s i s *and/or ~ ~ ~ ~ possible translation or transcription from the nucleus to the cytoT h e fine structure of this organelle in the FXS was quite similar with respect to its iticitlence, proximiry to the nucleus, and continuit). with the REK. Since annulate 1nniell;ie were always in close proximity to both tlie nuclear envclope and fibrillar eleiiients (also juxtapositionecl to the nucleus), a relationship may exist between these three org;tnelles. Distortion of mitochondria and nltcration of their cristae is another common manifesta tion associated with malignant transformation."; Giant mitochontlria in FXS cells (1ispl;iyetl menibranoils whorls of cristae superficially rcsembliiig the concentric lamellar foimations noted in adrenal cortical cells follorc-ing horinonal treatment in several spe-

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Xanthoma cells were infrequent and probably represent an altered histiocyte. T h e xanthoma-type cell in our investigation was similar to certain other types of benign xanthoma cells'J7 in that both contained lysosomes and clear lipid droplets. However, the xanthoma cells of the FXS consisted of membrane-bound, clear (highly unsaturated lipid) and lightly osmiopliilic lipid vacuoles. Abundant glycogen deposits woiild account for the PAS-positive histiocytes seen by light microscopy. Table 1 summarizes the main ultrastructural nuclear and cytoplasmic features which characterize the different neoplastic cell constituents of tlie FXS. These features were not described in benign neoplasms of this type." Moreover, S U C ~ Lultrastructural features as annulate lamellae, giant mitochondria, intranuclear vacuoles, and membrane-bound filaments were not observed in fibrous xanthomas or dermatofibromas (unpublished observations). It is not known at present whether the non-

( iesl',::"

Fic. i a ( f o p ) . Thrce cytiiplasinic virus-like particles (arrows) arc adjacent to a nucleus (N), x13,OOO. FIG.7b (bottom). Within a cistema of rough endoplasmic retirulum (Rer) thcre is an irregular viruslikc partitlc displaying P wrrowiiling capsid (arrow), Y42,OOo.

FIG. 8a (top). An intranuclear lipid body (Lb) and a spheroidal nuclear body (Snb) are adjacent to nuclear piojections. The inner nuclear membrane is quite dense and consists of two membranes and a dense corc ( ~ 3 3 , 0 0 0 ) . FIG. 8b (boffonz).Within the more translucent nurleoplasin (N), several membrane-bound bundles of delicate filaments (Fi) are evident ( ~ 1 4 , 5 0 0 ) .

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TABLE1. Ultrastructural Features Associated with Fibroxnnthosarcoma (FXS) Cells 1. 2. 3. 4.

5. 1. 2.

3. 4.

5.

IVucleus Hyperchromatic, enlarged, lobular Intranuclear vacuoles Intranuclear membrane-bound filaments Spheroidal nuclear bodies Nuclear projections Cytoplasm Frequent annulate lamellae Giant mitochondria with whorled cristae and inclusions Clusters of SER

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metastasizing atypical fibroxanthoma20 passesses at least some of these characteristics. Bednir? coined the word “storiform” which is synonymous with the terms spiral cartwhee1,jG twisted strip,50 whirligig,66 thumbscrew,ZO and pin wheel patterns. H e described this light microscopic pattern in neurofihromas and therefore regarded the dermatofibroma as neurogenous. However, ultrastructural studies on dermatofibromas or dermatofibrosarcomaslz and our FXS indicate that these tumors are not of neural origin. T h e FXS in children has been reported to be radioresistant.’* Similarly, radiotherapy as well as chemotherapy appeared to have little, if any, effect on this neoplasm in the adult. as was evident in our case. ,

Abundant delicate filaments especially near the nucleus Infrequent deqmosome-like structures

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