Megakaryocytes in pleural and prevalence, cytohistological … · Spleen:haematopoiesis witha NodulesofAMMonmesenteryand predominanceofmegakaryocytes serosaofsmallintestine, andin

J Clin Pathol 1980;33:1153-1159

Megakaryocytes in pleural and peritoneal fluids:prevalence, significance, morphology, andcytohistological correlationNB KUMAR AND B NAYLOR

From the Department ofPathology, The University of Michigan, Ann Arbor, Michigan, 48109, USA

SUMMARY Over a period of 22 years, 4844 pleural and peritoneal fluids from 3279 patients were

examined cytologically. Megakaryocytes were found in the fluids from five patients. The clinicaldiagnoses in the five patients were agnogenic myeloid metaplasia, chronic myeloid leukaemia, andlymphocytic lymphoma. All of these patients had persistent serous effusions. Megakaryocytes inserous fluids occurred in three forms: (1) a large type with abundant cytoplasm and multilobednuclei, (2) a smaller type with a high nucleocytoplasmic ratio and unlobed nuclei, and (3) anucleatecytoplasmic masses. Foci of agnogenic myeloid metaplasia found on the serous surfaces at necropsyof two patients contained megakaryocytes similar to those in the corresponding effusions. Theclinical course of our patients confirmed that the presence of megakaryocytes in serous fluids sig-nifies an advanced haematopoietic malignancy.

Megakaryocytes, normal inhabitants of bone mar-row, are occasionally seen in peripheral blood and ona rare occasion they may be found in sputum' due tothe passage of circulatingmegakaryocytes through thepulmonary alveolar walls. They have also been de-scribed in cerebrospinal fluid due to unintentionalpuncture of a vertebra during lumbar puncture.2

Megakaryocytes have also been described inpleural and peritoneal fluids'-4 in association withmyeloproliferative disorders, presumably due to thedevelopment of extramedullary haematopoiesis onthe pleura or peritoneum. Such foci of haema-topoiesis may develop in various organs wheneverthe haematopoietic activity of the bone marrow isinadequate to meet normal requirements. Thiscompensatory metaplastic phenomenon occurs, forexample, when the bone marrow is diffusely replacedby neoplasm such as lymphoma. On the other hand,extramedullary haematopoiesis may be a truly neo-plastic change as in agnogenic myeloid metaplasia(AMM). In addition to the extramedullary haema-topoiesis, AMM is also characterised by myelo-fibrosis and the proliferation of atypicalmegakaryocytes.The finding of megakaryocytes in pleural and peri-

Received for publication 8 April 1980

toneal fluids has, in our experience, been a rare event.This paper concerns the prevalence, significance, andmorphological features of megakaryocytes in pleuraland peritoneal fluids examined in our laboratory andin two cases correlates the cytological findings withthe findings at necropsy.

Material and methods

The records of the Cytopathology Laboratory in theDepartment of Pathology of The University ofMichigan from 1957 to 1979 were searched forspecimens of pleural, peritoneal, and pericardialfluids that contained megakaryocytes.The sediment obtained by centrifugation of the

fluids was smeared, fixed in 95% ethanol, and stainedby the Papanicolaou method. In four of the fivecases the sediment was clotted, and this, with anyspontaneously occurring clot, was fixed in formalin,embedded in paraffin, sectioned, and stained withhaematoxylin and eosin. Wright-stained specimens ofperitoneal fluid prepared in the Cytospin were avail-able from patients 1 and 2.The original histological sections of biopsy

material were reviewed and new sections were pre-pared when necessary. Tissues from the two patientswho came to necropsy were embedded in paraffin,

1153

copyright. on January 6, 2021 by guest. P

rotected byhttp://jcp.bm

j.com/

J Clin P

athol: first published as 10.1136/jcp.33.12.1153 on 1 Decem

ber 1980. Dow

nloaded from

http://jcp.bmj.com/

1154 Kumar and Naylor

Table I Megakaryocytes in pleural and peritoneal fluids: clinical and pathological features

Patient Age Sex Diagnosis Type of Biopsyfindings Follow- Necropsyfindings(yr) effusion up

1 70 F AMM Pleural and Bone marrow: myelofibrosis with Died Pleural effusion. Nodules ofAMM onperitoneal proliferation of megakaryocytes visceral and parietal pleura. Ascites.

Spleen: haematopoiesis with a Nodules ofAMM on mesentery andpredominance of megakaryocytes serosa of small intestine, and in

diaphragm, liver, kidney, periadrenaltissue, breast, and lymph nodes

2 66 M AMM following Pleural Bone marrow: progressive Died Pleural effusion and ascites. Nodules ofthrombocythaemia myelofibrosis with proliferation of AMM on diaphragmatic pleura and in

megakaryocytes pericholedochal, periaortic, andperiadrenal lymph nodes

3 52 F AMM following Peritoneal Bone marrow: trilineal hyperplasia Died No necropsypolycythaemia vera

4 70 M CML Pleural Pleura: leukaemic infiltrate Died No necropsyBone marrow: (aspirate) CML

5 60 F Lymphocytic Pleural Bone marrow: lymphocytic lymphoma Alivelymphoma Skin: lymphoma

Pleura (needle biopsy): lymphocyticpleuritis

sectioned, and stained with haematoxylin and eosin. pleural effusions for one year before death. Patient 2The overall cellularity of each smear was sub- with a diagnosis of AMM, had recurring massive

jectively graded from + to + + +. In addition, the pleural effusions for two years before he died andnumber of megakaryocytes on each smear was developed ascites one week before death. Patient 3counted. Fewer than five per slide was graded as +, developed AMM after a 13-year history of poly-six to 10 per slide as + +, and more than 10 per slide cythaemia vera. She had recurring ascites during theas + + +. three years before she died. The duration of pleuralThe clinical information in the patients' medical effusion in patient 4, who had CML, was not avail-

records was also reviewed. able from the medical records. He died two monthsafter admission to our hospital. Patient 5, who has

Results lymphocytic lymphoma and is still alive, has hadrecurrent bilateral pleural effusions for the last three

PREVALENCE OF MEGAKARYOCYTES years.

During 22 years, 4844 pleural and peritoneal fluidsfrom 3279 patients were examined and megakaryo- CYTOLOGICAL FINDINGScytes were found in such fluids from only five The cytological features of the pleural and peritonealpatients. fluids from the five patients are summarised in Table 2.

The cellular components of the sediments con-CLINICAL FINDINGS sisted of various proportions of mesothelial cells,A summary of the major clinical features for each histiocytes, erythrocytes, and megakaryocytes. Thepatient is presented in Table 1. Three of the five fluids from three of the five patients also containedpatients were women and two were men, all within an neoplastic cells in the form of immature granulocytesage range of 52-70 years. Three of the five patients (patients 1 and 4) or lymphoma cells (patient 5).had AMM, one had chronic myeloid leukaemia The number of megakaryocytes in the smears(CML), and one had lymphocytic lymphoma. Each ranged from + to + + +. They were not adherent topatient had a persistent serous effusion. Patient 1, each other. In the Papanicolaou-stained smears, thewith a diagnosis of AMM, had recurrent massive diameters of the megakaryocytes ranged from 20 to

Table 2 Megakaryocytes in pleural and peritoneal fluid: cytological findings

Patient Character ofeffusions Cellularity Background cells No. ofmegakaryocytes

I Cloudy yellow to Pleural: + to + + Immature granulocytesbloody Peritoneal: + + Histiocytes and mesothelial cells

2 Cloudy yellow to Pleural: + + Immature granulocytes, erythrocytes, histiocytes, mesothelial t- tobloody cells

Peritoneal (necropsy) Mesothelial cells3 Cloudy light red Peritoneal: -+- Erythrocytes and leukocytes -4 Cloudy orange to red Pleural: + to + + + Leukaemic cells and erythrocytes to5 Cloudy yellow Pleural: . to + + + Lymphoma cells, occasional histiocytes



j.com/

J Clin P


ber 1980. Dow

nloaded from

http://jcp.bmj.com/

Megakaryocytes in pleural and peritoneal fluids

Fig. 1 Patient 1. Cytospin centrifuge preparation ofperitonealfluid. A megakaryocyte with a multilobednucleus with lobes superimposed on each other. The cellhas a large amount of irregularly outlined cytoplasm.(Wright stain. Mark is 10tum.)

4F-:

*::...''...:..~: .............. :.:.

Fig. 3 Patient 2. Smear ofpleural fluid. Amegakaryocyte with an eccentric bilobed nucleus. Thechromatin is coarsely clumped, leaving a clear areabetween the clumped chromatin and the nuclearmembrane. The nuclear membrane is indented andappears to be delicate. (Papanicolaou stain. Mark is15 um.)

90

I.A.

.:; *.!!

Fig. 2 Patient 5. Smear ofpleural fluid. Amegakaryocyte with a multilobed nucleus. Thesuperimposed lobes of the nucleus accentuate thehyperchromasia. (Papanicolaou stain. Mark is 15 ,um.)

Fig. 4 Patient 1. Smear ofperitoneal fluid. Twoanucleate megakaryocytes. Both contain residualnuclear material. (Papanicolaou stain. Mark is 19 ,um.)

80 microns. Their shapes were round to ovoid.Occasional megakaryocytes were fusiform or irregu-lar in shape, which may have been an artifactcreated by the smearing of the sediment. Most of themegakaryocytes had abundant cytoplasm. With thePapanicolaou stain the cytoplasm was eitheracidophilic, cyanophilic, or amphophilic. In somemegakaryocytes the cytoplasm was finely granular, afeature that was better seen in the Wright-stainedpreparations. Essentially, three basic forms ofnmegakaryocytes were present: (1) megakaryocytes

with multilobed nuclei and a large amount ofcytoplasm, (2) anucleate cytoplasmic masses, and (3)megakaryocytes with unlobed nuclei and a smallamount of cytoplasm.The multilobed megakaryocytes contained nuclei

with two or more lobes that were frequently super-imposed, thereby accentuating their hyperchromasia(Figs 1 and 2). Frequently, the chromatin wasirregularly clumped, generally toward the centre ofthe nucleus, leaving a clear area between the clumpedchromatin and the nuclear membrane (Fig. 3). A

llSS

0 *



j.com/

J Clin P


ber 1980. Dow

nloaded from

http://jcp.bmj.com/

Kumar and Naylor

Nucleoli, when present, were not unduly prominent.Anucleate masses of megakaryocytic cytoplasm

were found in the fluids from three of the patients.About the same size and shape as maturemegakaryocytes, they appeared as distinct cyano-philic anucleate masses (Figs 4 and 5). Sometimesbare nuclei, apparently from megakaryocytes, laynext to these anucleate masses. What we interpret asa form of megakaryocyte intermediate between thesetwo forms consisted of a cytoplasmic mass with itsnucleus 'compressed' at the periphery, imparting adistinctive signet-ring appearance to the cell (Figs5 and 6).Megakaryocytes with unlobed nuclei were com-

monly seen. These cells had well-defined cyto-plasmic borders, high nucleocytoplasmic ratios, anda more uniform chromatin pattern than the mega-karyocytes with lobed nuclei (Fig. 7). Thesemegakaryocytes with unlobed nuclei may representless mature forms.

BIOPSY FINDINGSBone marrow biopsies were performed on patients1, 2, 3, and 5. In patients 1 and 2, the specimensshowed myelofibrosis with proliferation of mega-karyocytes. In patient 3, the marrow showed tri-

Fig. 5 Patient 1. Smear ofplural fluid. lineal hyperplasia, and in patient 5 it containedMegakaryocytes. The largest is bibbed with delicate lymphocytic lymphoma. Patient 4 underwent bonenuclear membranes and central chromatic clumpingBelow it are two anucleate megakaryocytic masses, one marrow aspIratlonwh2ch revealed CML.containing residual nuclear fragments. The two above it Patients 1 and 2 underwent splenectomy, whichare bilobed. The uppermost megakaryocyte has a signet- showed extramedullary haematopoiesis includingring appearance. (Papanicolaou stain. Mark is 19 )um.) marked proliferation of megakaryocytes. The pleural

biopsy specimen from patient 4 contained a leukaemicdistinctive feature of megakaryocytic nuclei was their infiltrate. Additional biopsies in patient 5 showeddelicate and indented nuclear membranes (Fig. 3). lymphocytic lymphoma in skin, a lymphoreticular

4I~~~~~~I.S_ _

Fig. 6 Patient 5. Cell block ofpleural fluid. Amegakaryocyte with an eccentric multilobed nucleus,imparting a signet-ring appearance to the cell.(Haematoxylin and eosin. Mark is 15 gum.)

Fig. 7 Patient 2. Smear ofpleural fluid. Amegakaryocyte with an unlobed nucleus.(Papanicolaou stain. Mark is 15 1m.)

1156

.6-Afimk.

.qw. #W.040-1.

0.

....0 V.

::AF.



j.com/

J Clin P


ber 1980. Dow

nloaded from

http://jcp.bmj.com/

Megakaryocytes in pleural and peritoneal fluids 1157

Fig. 8 Patient 1. Small intestine. Confluent nodules ofAMM in the mesentery. (Mark is 12 mm.)

Fig. 9 Patient 1. Lung. Slightly raised nodules ofextramedullary haematopoiesis on the visceral pleura.(Mark is 7-7 mm.)

infiltrate in the liver, and lymphocytic pleuritis.

NECROPSY FINDINGSPatient 1 had massive ascites (5300 ml of sero-sanguineous fluid) and a right pleural effusion. Theomentum and mesentery contained masses of grey-white, firm tissue (Fig. 8). Similar masses were foundin peripancreatic and periadrenal soft tissues. Theliver weighed 3140 g and contained numerous grey-white nodules. Smaller nodules were found on thevisceral pleura (Fig. 9) and in the kidneys, and a mass12 cm in diameter invaded the left hemidiaphragm.The bone marrow was dry, grey-white, and sclerotic.

Microscopical examination revealed that thenodules were composed of fibrous connective tissuemingled with haematopoietic cells, includingatypical and bizarre megakaryocytes (Fig. 10). Suchfoci of haematopoiesis were found in lymph nodes,visceral and diaphragmatic pleura, peritoneum,adrenal glands, peripancreatic and periadrenal adi-pose tissue, kidney, liver, gallbladder, pulmonaryparenchyma, heart, breast, and stomach. The bonemarrow was fibrotic and contained scattered en-larged and atypical megakaryocytes in addition toother haematopoietic cells. Each of the threemorphological types of megakaryocyte seen in thecytological preparations was readily identified in thehistological sections of the nodules, bone marrow,and lymph nodes.The left and right pleural cavities of patient 2

contained 2700 ml and 800 ml of serosanguineousfluid, respectively, and the peritoneal cavity con-tained 2000 ml of serous fluid. Other pertinent grossfindings were a granular appearance of the pleural

I Fig. 10 Surface of lung., Nodule ofAMM consisting oft fibrous tissue containing

haematopoietic cells, with apredominance of

4E megakaryocytes.~. ,,~ Haematoxylin and eosin.~~~~~ Mark is 300 pim.)

Iyll. .:1

7V3'.g. .iFz.z By Xj..__s~~~~~~~~~~A fl ..1,



j.com/

J Clin P


ber 1980. Dow

nloaded from

http://jcp.bmj.com/

1158

Fig. 11 Patient 2. Diaphragm. Pleural surface showingirregularly shaped plaques ofAMM. (Mark is 3-6 mm.)

Fig. 12 Patient 2. Diaphragmatic pleura.Extramedullary haematopoiesis includingmegakaryocytes. (Haematoxylin and eosin. Mark is 48

'Um.)

surface of the diaphragm (Fig. 11), enlarged lymphnodes in the porta hepatis, patchy pneumonicconsolidation of the lungs, and sclerotic bonemarrow.

Microscopically, foci of haematopoiesis withprominent atypical megakaryocytes were seen in thepulmonary parenchyma, lymph nodes, kidney, liver,periadrenal and periaortic tissues, and on the pleuralsurface of the diaphragm (Fig. 12). The bone marrowwas fibrotic and contained numerous atypicalmegakaryocytes.

Discussion

Megakaryocytes are rarely found in routinelyprepared smears of peripheral blood in normalstates, although they can be found in very smallnumbers in smears of white cell concentrates of peri-

Kumar and Nay/or

pheral blood from normal persons.5 They are rarelyseen in pleural and peritoneal fluids. This is borneout not only by the small number of reportedexamples in the literature but also by our ownexperience, in which we found only five examples ofmegakaryocytes in almost 5000 pleural and peri-toneal fluids.Whenever megakaryocytes have been seen in

pleural or peritoneal fluids it has been in associationwith AMM or leukaemia or lymphomatous involve-ment of bone marrow. Their presence in pleural fluidhas been reported by Spriggs and Boddington2 inassociation with CML, and in peritoneal fluids byCalle3 and by von Haam4 in association with AMM.Similarly, our patients all had some form of myelo-proliferative disorder or neoplastic involvement ofthe bone marrow. Three had AMM, one had CML,and one had lymphocytic lymphoma.

Since grossly visible nodules of AMM mayresemble metastatic neoplasm,6 it is important not tomisinterpret atypical megakaryocytes as malignantmesenchymal or epithelial cells in histological sec-tions of such nodules or in any associated serouseffusion. They have been mistaken for malignantcells in peripheral blood from patients known to havecancer, and numerous reports have appeared regard-ing the difference between megakaryocytes andmalignant cells in peripheral blood.7 8 Furthermore,since AMM could manifest itself by tumour forma-tion, as exhibited in our case 1 and as previouslydescribed,6 9-11 it is conceivable that one may bepresented with a cellular sample obtained byaspirating a nodule of AMM or a lymph nodeinvolved with AMM. In such a situation, it would beextremely important to be familiar with the mor-phological features of megakaryocytes and to be ableto distinguish them from a variety of neoplastic andnon-neoplastic cells.

Generally, it should not be difficult to identifymature megakaryocytes in a serous fluid. Many ofthem are outstandingly large, much larger thantypical carcinoma cells or cells of lymphomas, andthey do not adhere to each other as do carcinomacells. Those megakaryocytes which have abundantcytoplasm do not exhibit the marked vacuolisationthat is frequently seen in adenocarcinoma cells.Mature megakaryocytes contain multilobed nucleiwith nuclear membranes that are not undulyprominent. In contrast, the nuclei of carcinoma cellsmay be multiple but they are not usually lobed. Inaddition, the nuclei of carcinoma cells are usuallyangulated and have sharp, heavy membranes. Thenucleoli of carcinoma cells are frequently veryprominent, whereas the nucleoli of megakaryocytesare usually small and may be difficult to perceive.Those megakaryocytes with unlobed or only bilobed

0I. 44!: I. .-m' ::... IlkK::x.,'k..-W

-Y...

t:.01 *R.



j.com/

J Clin P


ber 1980. Dow

nloaded from

http://jcp.bmj.com/

Megakaryocytes in pleural and peritoneal fluids 1159

nuclei and megakaryocytes with degenerated pykno-tic nuclei are, perhaps, the ones that are more likelyto be confused with malignant cells. Reed-Sternbergcells could be confused with megakaryocytes. How-ever, they are generally smaller than megakaryocytesand have a high nucleocytoplasmic ratio, well-defined, heavy nuclear borders, and prominentnucleoli. Benign multinucleated mesothelial cellsbear some resemblance to megakaryocytes although,unlike megakaryocytes, they contain distinctlymultiple rather than lobed nuclei, which aresmoothly round to ovoid with sharply definedborders. In addition, mesothelial cells frequentlyadhere to each other.The explanation for the presence of megakaryo-

cytes in serous fluids was obvious in two of our fivepatients: nodules of AMM on the surface of thepleura or peritoneum and in various other abdominalorgans. Although we are not aware of any reportswhich demonstrated such a histocytological cor-relation, the existence of this situation was hypo-thesised by Calle.3 The presence of megakaryocytesin the peritoneal fluid of patient 3, who had AMM,was most likely due to nodules ofAMM on the peri-toneum. In patient 4, diagnosed as having CML,AMM was also considered in the differentialdiagnosis, and, therefore, the source of the mega-karyocytes in his pleural fluid may have been pleuralfoci of AMM.

In patient 5, who has lymphocytic lymphoma,there are two likely explanations for the presence ofmegakaryocytes in her pleural fluid. The more likelyis the presence of compensatory extramedullaryhaematopoiesis on the pleura, although it is possiblethat an increased number of circulating mega-karyocytes in peripheral blood, which is known tooccur in the presence of various malignant neo-plasms, may account for the presence of the mega-karyocytes in the pleural fluid.

Finally, the question whether any blood in theserous fluids contributed to the presence of themegakaryocytes should be considered. Against this,the number of megakaryocytes in different specimensof serous fluid from the same patient did not increasewith increasing bloodiness of the specimen.

The finding of megakaryocytes in a serous fluidshould be reported because it may be a manifestationof some type of haematopoietic malignancy. Theirpresence seems to denote an advanced serious dis-order and may be an ominous sign as four of our fivepatients died within four to 90 days after the findingof megakaryocytes in the serous effusions.

We gratefully acknowledge the assistance of PatriciaM Novak, who took the photomicrographs.

References

Koss LG. Diagnostic Cytology and Its HistopathologicBases. 3rd ed. Philadelphia: JB Lippincott Company,1979.

2Spriggs AI, Boddington MM. The Cytology of Effusionsand of Cerebrospinal Fluid. 2nd ed. London: WilliamHeinemann Medical Books, 1968.

3 Calle S. Megakaryocytes in an abdominal fluid. Acta Cytol1968 ;12:78-80.

4 von Haam E. Cytology of transudates and exudates.Monographs in Clinical Cytology. Basel, New York: SKarger, 1977.

Alexander RF, Spriggs AI. The differential diagnosis oftumour cells in circulating blood. J Clin Pathol 1960;13:414-24.

Estenez JM, Enrique EG, Moran TJ Acute megakaryo-cytic myelofibrosis. Case report of an unusual myelo-proliferative syndrome. Am J Clin Pathol 1974;62:52-9.

7 Kierszenbaum AL, Tres LL. Differential diagnosis ofmegakaryocytes from cancer cells in peripheral blood.Acta Cytol 1964;8:91-4.

8 McGrew EA, Romsdahl MM, Valaitas J. Differentiationof hematopoietic elements from tumor cells in blood.Acta Cytol 1962 ;6:551-4.

9 Glew RH, Haese WH, McIntyre PR. Myeloid metaplasiawith myelofibrosis: The clinical spectrum of extra-medullary hematopoiesis and tumor formation. JohnsHopkins Med J 1973;132:253-70.

10 Leiberman PH, Rosnoli RV, Ley AB. Extramedullarymyeloid tumors in primary myelofibrosis. Cancer 1965;18 :727-36.

French HE, Yates DR. Myelofibrosis and myeloid tumorsas a surgical disease. South MedJ 1970;63:387-91.

Requests for reprints to: Dr NB Kumar, Box 45, Univer-sity of Michigan Medical Center, Ann Arbor, Michigan,48109, USA.



j.com/

J Clin P


ber 1980. Dow

nloaded from

http://jcp.bmj.com/

Documents

Megakaryocytes in pleural and prevalence, cytohistological … · Spleen:haematopoiesis witha NodulesofAMMonmesenteryand predominanceofmegakaryocytes serosaofsmallintestine, andin