THE ROLE OF EXFOLIATIVE CYTOLOGY IN THE ...August I96I McGREW: Role ofExfoliative Cytology in Diagnosis andTreatment ofMalignantDisease 4S9 over go,ooo patients coming to the Mayo

POSTGRAD. MED. J. (I96I), 37, 456

THE ROLE OF EXFOLIATIVE CYTOLOGYIN THE DIAGNOSIS AND TREATMENT OF

-MALIGNANT DISEASEELIZABETH A. McGRw, M.D.

Associate Professor of Pathology, University of Illinois College of Medicine, Chicago I2, Illinois, U.S.A.

THE study of cells derived from the variouslining and covering surfaces of the body datesback to the days of the first microscope, for surely,cells scraped from the lining of the mouth andfrom other areas were among the first wonders tobe observed with the new contrivance. Detailedobservation of exfoliated cells was first undertakenby Pouchet, however, who in i847, reported thechanges occurring in unstained vaginal smearsduring the female sex cycle. This kind of materialwas but little used thereafter, until I917, whenStockard and Papanicolaou published the first of aseries of articles on cyclic changes in vaginalsmears. These studies contributed materially toour present understanding of the mammalian sexcycle.The indentification of malignant cells in various

body fluids began with the report of Beale in i86o,that he had found particles from a malignantpharyngeal tumour in sputum, and in i 64, Sandersrecorded the discovery of malignant tumour parti-cles in the sediment of urine. 'Increasinginterest in the diagnostic possibilities of thismethod followed, particularly with reference tofluids from the pleural and peritoneal cavities.'A Handbook of Clinical Microscopy ' byBizzozero, appearing in its second GermanEdition in I887, mentions the cellular elements tobe found in sputum, urine, serous fluids and feceswith occasional reference to the possible identifica-tion of tumour cells. The development of tissuesectioning technique and improved stainingmethods in the early I900S obscured the potentialvalue of these observations, however, and the fieldof tissue pathology evolved.

Papanicolaou's announcement in 1928 thatabnormal cells could be identified in vaginal smearsfrom.women with cancer of the genital tract fell ondeaf ears generally until the classic monographdescribing his methods, criteria and results waspublished in I943. In the ensuing i8 years greatefforts have been extended throughout the world

to put this technique to the widest possible use inthe detection of malignancy of the female genitaltract and also to adapt it to the solution of diag-nostic problems'in other areas. The publicationof Papanicolaou's 'Atlas of Exfoliative Cytology'in 1954 marked the firm establishment of themethod as an accepted diagnostic adjunct and animportant branch of anatomic pathology.

Success in utilizing the criteria of malignancy sodeveloped depends upon obtaining suitable cells inthe fresh state, dispersing them properly on theslides and fixing immediately in the solution ofether and alcohol. If the cells are permitted todegenerate or to dry on the slide, the importantintranuclear features (Fig. i) differentiatingbenign from malignant cells cannot be visualizedand cell types are difficult to distinguish. Therather delicate, but not difficult, staining procedureresults in very pleasant clear pastel cytoplasmiccoloration, which quite reliably differentiatessquamous cells from histiocytic and other types.In vaginal smears of good quality the colour of thecytoplasm of the squamous cells assists in theevaluation of hormonal states as well, since estro-genic effect generally causes a high proportion ofthem to take the eosin stain. The nuclear stain ishlmatoxylin and in well fixed smears the chro-matin material has a very distinct network orgranular pattern.

Histopathologists are familar with many of thenuclear criteria for malignancy which can beapplied' to exfoliated cells, such as large nuclearsize, hyperchromatism, irregular shape, duplica-tion of nuclei and an increased ratio of nuclear tocytoplasmic area. Derangements of chromatinstructure, however, are rarely of value in formalin-fixed and imbedded material. In cytologicsmears, the normal. interphase nucleus is a smoothoval with a sharp regular wall of chromaticmaterial and an internal structure resembling atransparent cocoon. As the stain is relativelytransparent one can focus through the fine

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Aliugtst I96I McGREW: Role of Efoliative Cvtology in Diagnosis and Treatment of Malignant Disease 457

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Reprinted with permlission of the Yoiurnal of The Anierican Medical a'oitn's Association.A B

1IG. I.-Malignant cells from two areas of the same smear of pleural fluid sediment. (a) Drying occurred beforcimmersion in the fixing solution. (b) Well fixed area, Papanicolaou stain. x 2IO0.

fibrillar network of chromatin sharply stainedagainst a ground-glass background. In activelyproliferating cells the chromatin is more or lesscoarsely granular, but still evenly distributed.The nuclei of malignant cells, on the other hand,in addition to containing excessive amounts ofchromatic material and assuming large irregularforms, display marked derangement of thechromatin. The nucleus may be composed ofdense, opaque chromatin with no visible structure.Large nuclei generally allow resolution of thechromatin particles, which are coarse and variablein size, shape and distribution. In other instances,most of the chromatin is concentrated in a fewlarge, irregular masses leaving most of the nucleusempty. There is also clearing of the nuclearbackground substance, which may be focal as ifpart of the nucleus had been erased (Fig. 2).

Exfoliated cells are easy to obtain from mostareas of the body but another important ad-vantage rests on the fact that body fluids generallycontain cells representing large areas, while abiopsy consists only of the tissue chosen. This isparticularly advantageous in urinary bladderlesions where the biopsy may represent only thesuperficial fronds of a tumour, and in bronchiallesions so often inaccessible to biopsy.The chief disadvantage of exfoliative cell study

in the diagnosis of malignancy is that the cells areusually isolated from one another and their growthpattern cannot be studied as it can in tissuesectionis. Consequenltly, it is best to conIfirnm thefinding of malignant cells in cytologic smnears bytissue study whenever possible.

Considerable experience must be attained in

order to differentiate between malignant cells anidthose rendered atypical in form, size and nuclearcharacteristics by chronic inflammation, radiationor other agents. Indeed, the range of atypia incells of benign lesions is great, and it is impossibleto draw a distinct line at a point where all on oneside are benign and all on the other side malignant.Thus, if one is very conservative, there may be nofalse positive cytologic interpretations, but thenumber of cancers missed will increase. Since,in most applications, the value of the method liesin its sensitivity in detecting otherwise silent andhidden malignancy, sacrificing this for greaterreliability would deprive many patients the op-portunity for successful treatment. Pathologistsand clinicians alike must be prepared, therefore, toaccept the consequences of an occasional falsepositive report.

The Female Genital TractAs carcinoma of the uterine cervix is tbc

second most common fatal malignancy in women,and since cervical and vaginal smears are easy toobtain, it is natural that this method has found itsgreatest application in the detection of femalegenital tract carcinoma. By the early 1950Sreports of the cytologic detection of early andclinically unsuspected carcinoma of the uterus hadbecome somewhat monotonous. Review of almostIoo of these articles (McGrew, Bibliography,1952), eliminating duplications, yielded a total of551,1 17 womeni in whom 7,042 cancers of thegcenital tract were found, atn iincidenice of 1.3%. (Jfthese tumours, 3,46I or 490/,, were not suspected oncliniical evidence, Seventy-nine per cent. of the

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FIG. 2.-(a-e) Malignant cells. There is marked variation in size. The nuclei may be vesicular (a), and distorted (b).The chromatin may be diffuse and coarsely granular (c), opaque (d), or separated by 'erased ' areas (e). (f) Nuclearenlargement and irregular chromatin distribution in chronic inflammation. (g) Abnormal cell from atypicalsquamous epithelial hyperplasia. Papanicolaou stain. All cells x 1400. A fewv polymorphonuclears are includedfor size reference.

unsuspected carcinomas were pre-invasive lesionsof the cervix and, therefore, curable by relativelysimple procedures. If these one-half millionwomen are divided according to their clinicalcircumstances at the time of examination, it isfound that 3.300 of women presenting themselvesfor examination in a gynacologist's office or clinichave carcinoma of the genital tract. Only 17%of these lesions are unsuspected and only 12%/, arein the pre-invasive stage. If smears are takeniroutinely in the offices of genieral practitioners anidinlterInists, I.5%1 of adult women will be found toliave carcinoma of the genital tract, If the general

female population of a community is surveyed,o.8o% have carcinoma, and if only women withoutsymptoms are screened, the incidence is 0.300.In these last three groups, 50 to 6o% of thetumours discovered are unsuspected on the basisof symptoms or physical examination, and 50 to8o%/ are in the pre-invasive stage. Even inyounger age groups, screening for cervical car-ciiloma shoukl not be overlooked. Slate, Martinanid Merrit, in 1957, reported anl incidence of twomalignancies per thousand in 5,935 pregnanitwomen. Seventy-eight per cent. of these werepre-invasive. Soule and Dahlin in I959, screened



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August I96I McGREW: Role of Exfoliative Cytology in Diagnosis and Treatment of Malignant Disease 4S9

over go,ooo patients coming to the Mayo Clinicwithout specific gynecologic symptoms and dis-covered 678 carcinomas, an incidence of 0.75"0,.A second examination was performed a year ormore later on I4,400 of these women with thedetection of only 27 carcinomas (o.I0/ incidence),thus demonstrating the value of periodic routinepelvic examinations and cytologic smears.

It is clear, therefore, that the mortality fromcarcinoma of the cervix, formerly around 70%,could be reduced to zero and the disease eradicatedas a threat to life or health if cytologic examinationwere used in all women to detect the disease at acurable stage. Lund (I96I) has written ' AnEpitaph for Cervical Carcinoma' in which hereports the mortality from carcinoma of thegenital tract reduced 2o% over a nine-year periodduring which a community cytology project wasconducted in Rochester, New York.One of the bonus benefits of these studies is the

understanding we have gained of pre-malignantand very early lesions of the cervix. The conceptof pre-invasive carcinoma of the cervix was pro-posed many years ago but was regarded withsuspicion by most pathologists who had nooccasion to see it. With the advent of routinePapanicolaou smears, however, abnormal cells shedby these lesions resulted in the taking of biopsieswhich presented the pathologist with an array ofunexpected and unfamiliar lesions. As a result,the concept of pre-invasive carcinoma has becomefirmly established and numerous studies havedemonstrated that this lesion usually either ac-companies or progresses to the invasive form. Wehave also come to recognize a similar, but generallyreversible lesion variously called atypical hyper-plasia, dysplasia, or anaplasia, which displaysabniormal individual cellular characteristics butdifferentiates well, has few mitotic figures, andforms an epithelium which is not as thick or ascrowded as pre-invasive carcinoma (Fig. 3).These interesting lesions may accompany pre-invasive and invasive carcinoma or may constitutethe only abnormality of the cervical epithelium.The cells exfoliating from these lesions are some-what better differentiated than those of pre-invasive and invasive carcinoma but cannot alwaysbe distinguished from them.A word of caution is therefore in order with

regard to the verification of positive cytologicfindings. Abnormal cells may be found in car-cinoma of the cervix in all stages of advancement,in pre-invasive carcinoma and in atypical epithelialhyperplasia. A single punch biopsy is adequatefor evaluation of a patient with positive or sus-picious cervical smears, only if it shows invasivecarcinoma. Four quadrant biopsies are onlyslightly more reassuring since many pre-invasive

and early invasive carcinomas arise in patchesscattered throughout the endocervical lining andglands. In order adequately to evaluate thepatient with positive cervical smears, a conizationof the entire external os and endocervical canalwithout the use of cautery is necessary. Themultifocal distribution of pre-invasive carcinomaand of hidden areas of invasion have been welldemonstrated by Przybora and Plutowa in 1959and Fidler and Boyd in I960.

It should also be made clear that vaginal andcervical smears cannot entirely rule out thepresence of carcinoma since necrosis of the surfacemay result in smears which contain no recognizabletumour cells. Therefore, whenever the history orappearance of the cervix is suspicious, biopsyshould be performed.As Koss has indicated (I96I), adenocarcinoma

of the endometrium is somewhat more difficult todetect on cervical and vaginal smears. The cellsdo not exfoliate as abundantly as those fromsquamous epithelial tumours, and are often de-generated by the time they appear in the vaginalpool or cervical mucus. Endocervical or endo-metrial aspiration smears disclose the presence ofthis lesion in 8o to go% of cases and may be usedif the patient's age or symptoms so indicate. Thepresence in cervical or vaginal smears of old bloodor necrotic cellular debris not otherwise explainedmay also suggest the need for endometrial smears.In large surveys, adenocarcinomas of the uterinebody may constitute 3 or 4% of the unsuspectedtumours found.

Carcinoma of the Fallopian tube and ovary, orperitoneal carcinomatosis may be represented invaginal smears. We have seen several instances ofthe latter in the course of rendering hormonalevaluations on vaginal smears of patients receivingtherapy for carcinoma of the breast.

The BronchusThe Papanicolaou technique was rapidly applied

to the diagnosis of pulmonary carcinoma as well.Nor were these efforts without precedent. SinceBeale found tumour fragments in sputum in i86o,many investigators have reported carcinoma cells insputum, and in I897, Hampeln urged wider use ofsputum examinations for this purpose. Increasinginterest was represented by Dudgeon and Wrigley(935) who reported a wet film method of demon-strating particles of malignant growth in sputum.In 1943, Gowar examined sputum in 93 patientsand correctly diagnosed 36 carcinomas, several ofwhich were operable. Wandall in I944 publishedthe largest series up to that time. Papanicolaoufirst reported the successful diagnosis of lungcarcinoma in 22 of 54 patients by sputum examina-tion in I946. With Watson, Cromwell and



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(b) In situ carcinoma. H+E. x 400.FIG. 3.



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August i96I McGREW: Role of Exfoliative Cytology in Diagnosis and Treatment of Malignant Disease 461

Craver he reported the results of sputum examina-tion in I,200 cases in I949. Meanwhile, Herbutin I946, and McDonald and Woolner in I947, alsoreported successful identification of malignantcells in bronchial secretions and in sputum,respectively, in large numbers of patients. Morerecently, Hampson (I956), Steele (I957) andUmiker (I960), have reported similar results.

It may be concluded from these reports thatlung cancer can be detected by examination ofsputum and bronchial secretions in approximately8o% of persons with the disease. Bronchialbiopsy, on the other hand, has been reported togive positive results in as few as 27%. A recentreview (Valaitis, Leader, Chomet and McGrew,I96I) of cases of pulmonary carcinoma at aVeterans' Hospital in Chicago indicates, in ad-tion, that io to I500o of the X-ray exminations onthese patients may fail to disclose the true nature ofthe disease. Large numbers of patients nowundergo thoracic exploration on the basis of X-rayand cytologic evidence without confirmatorypositive biopsy in the hope of improving theoperability and survival rate in this disease.

Care must be exercised in obtaining specimensof sputum and bronchial secretions for cytologicexamination. It has been found that tumour cellsmay be produced in sputum rather irregularly, andthat diagnostic accuracy is increased many foldwith the examination of multiple fresh morningsputum specimens. The patient must be observedand instructed in the production of a deep coughspecimen and the sputum must either be processedat once or mixed with 5o% alcohol in saline.Four to eight smears are prepared from eachsputum specimen. The value of direct smearsobtained at bronchoscopy or saline washings of thebronchi varies with the interest and care taken bythe bronchoscopist and the laboratory staff. Manyreports indicate difficulty in obtaining suitablematerial from bronchoscopy and better resultsfrom multiple sputum specimens. It has beennoted by several observers that sputum specimensobtained two to 48 hours after bronchoscopy aremore likely to contain tumour cells than either thebronchoscopic secretions or sputum taken prior tobronchoscopy. Bronchoscopy is not always doneuntil after suspicious or positive cytologic findingshave been reported, however. Techniques havebeen developed recently to obtain a better con-centration of malignant cells, both to lighten theburden of screening and to increase the detectionrate. Beckerman, Sproul and Barach (1958),reported a method of stimulating sputum pro-duction by means of an inhaled aerosol solution,and Klein, Pollack and Volk (1959), used variablepressure with an anesthesia mask. Mucolyticagents have been used by a number of investi-

gators to permit concentration of the epithelialcells by sedimentation or centrifugation andrecently Chang, Anken and Russell (I96I) de-veloped a method of enzymatic digestion andmillipore membrane filtration to concentratecells from sputum.

Although mass surveys of the population similarto those for the detection of female genital tractcarcinoma cannot be seriously predicted at thistime, the development of sputum-inducingmethods and practical concentration techniquesmay well result in the discovery of increasingnumbers of early, asymptomatic carcinomas of thelung in selected population groups.

The Urinary TractThere had also been attempts to recognize

tumours of the urinary tract from urine sedimentprior to the development of the Papanicolaoumethod. Sporadic reports following that ofSanders (I864) are adequately summarized byDeden in I954. The reports of Papanicolaou andco-workers in 1945 and I946 (Papanicolaou andMarshall), of Fremont-Smith, Graham and Meigs(1948) and Chute and Williams (1948) are repre-sentative of the early work. Deden's monographin 1954 reported definite tumour cells identified in78% of patients with carcinoma of the bladder andin three of six patients with carcinoma of theprostate. The largest series reported is that ofFoot, Papanicolaou, Holmquist and Seybolt inI958, reviewing 2,829 cases. They found malig-nant cells in urine sediment in 62% of transitionalcell carcinomas of the bladder, ureter and pelvis,in 8% of renal carcinomas and in i5% of prostaticcarcinomas. Many authors have stated that thestudy of urine sediment in suspected renalneoplasms is useless, but in I954, we (McGrew)reported successful identification of malignantcells in 50% of renal adenocarcinomas with nofalse positives. In a series now totalling 536patients, I5/ of the urinary tract malignancieswere missed and 6% of the positive cytologicinterpretations were erroneous. Seventy-two percent. of the bladder carcinomas yielded positivecells in the urine sediment and another i5% werereported as suspicious. It is of some interest thatof the 57 cases with positive cytologic reports,eight were not initially recognized as carcinoma oncystoscopic examination. Of 76 prostatic car-cinomas examined by means of direct smears ofprostatic secretions or post-massage urine sedi-ment, approximately half were detected cyto-logically. Of 40 renal neoplasms, 20 of 28 renalcell carcinomas had suspicious or positive urinesediment cytology. Five of six transitional cellcarcinomas of the pelvis, one of four Wilm'stumours and one sarcoma were correctly interpreted



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POSTGRADUATE MEDICAL JOURNAL

as malignant. A metastatic ttumouir was missed.'l'here is some dloubt as to the value of prostaticsecretions obtained by massage in view of thepossibility of disseminating malignant cells into theblood stream by trauma. Prostatic massage as adiagnostic procedure has been largely abandonedin this hospital and is looked on rather unfavour-ably by many surgeons. None the less, threepositive prostatic secretions in our series were laterconfirmed in relatively young men with clinicallybenign prostatic disease. Herbut (I949) examinedprostatic secretions in 480 patients, 66 of whomlhad carcinoma of the prostate. Fifty-four ofthese, or 82%, had malignant cells in prostaticsecretion. Similarly, Frank and Scott (I958),reported 840,0 positive cytology in 100 cases ofprostatic carcinoma.

Cells exfoliated from renal cell carcinomas(hypernephromas or adenocarcinomas of thekidney) generally showed advanced degenerativechanges in our experience and were few in number.Unfortunately, most of those with positivecytologic findings were advanced cases withextensive involvement of the pelvis and vessels ofthe kidney. However, four of them had notdemonstrably invaded veins. Small and presu-mably curable renal adenocarcinomas have beendetected cytologically by Fremont-Smith et al.(I948), McDonald and Woolner (94S), Bungeand Kraushaar (1950), and Foot and Papanicolaou(I949)-

Thus, it may be anticipated that urinarycytology will disclose the presence of bladdermalignancies in a high percentage of cases and willbe almost as reliable in transitional cell carcinomasof the ureter or renal pelvis. If prostatic massageis utilized to obtain cells, clinically occult car-cinoma of the prostate may be detected also, andthe inclusion of urinary cytology in the study ofrenal diagnostic problems may have at leastconfirmative value.Although some authors recommend mixing

urine specimens with an equal volume of 70°Oalcohol, the cells adhere to the slide better andstain more reliably if the specimen can be obtainedfresh, and the sediment smeared and fixed at once.Better representation oftumours of transitional andrenal cell type is obtained by examining multiplespecimens, best obtained after the patient has beenup and moving about or after palpation of thearea. Greatly improved cytologic results may beobtained following antibiotic therapy of a grosslyinfected bladder. Millipore filtration of previously-fixed urine has been recommended by Solomon,Amelar, Hyman, Chaiban and Europa (I958) andHarpst, Ware and Eisenberg (I96I), but it isdoubtful that results in transitional cell car-

cinomas would be greatly improved by this metho(dsince they usually exfoliate abundantly.The cytologic examination of urinary sedlimen t

or prostatic secretion is usually requested for thosepatients with hrmaturia or other clinical signsleading to a suspicion of urinary tract malignancy.However, Crabbe, Cresdee, Scott and Williams(I956), and Melamed, Koss, Ricci and Whitmore(IjS5), have utilized urinary cytology to detectearly tumours in individuals exposed to bladdercarcinogens in industry.

The Gastrointestinal TractThere were attempts to identify malignanit cells

in secretions or washings from the stomach as longago as i882 (Rosenbach). Many false negativeresults were discouraging, however. Papanicolaoumentioned the detection of gastric malignancy inI946 and two years later Graham, Ulfelder andGreen (1948) reported malignant cells in salinewashings of the stomach in five resectable cases ofgastric carcinoma. An accuracy of around 6000,using vigorous lavage with Ringer's solution wasreported by Seybolt, Papanicolaou and Cooper(I95i) and with the use of a balloon covered withan abrasive netting which was inflated in thestomach, they were able to recognize 880/ of Sicarcinomas (Cooper and Papanicolaou, I953).A rotating brush which could be introduced intothe stomach and retracted into its sheath, wasdevised by Ayre and Oren (I93), and Rubin andBenditt's method for lavage with buffered chymo-trypsin (I955) enjoyed considerable popularity.Mlore recently results in the range of go to 9500accuracy have been reported. These are sum-marized and the technique and criteria well illu-strated by Schade (i959) and by Raskin, Palmerand Kirsner (I96I).Improvement in the detection of gastric

carcinoma by the cytologic method is due to in-creasing experience and to measures preventingdeterioration of the cells rather than to the use ofany specific method for obtaining specimens, sincethese excellent results are obtained from salinewashings as well as by the use of enzyme digestiontechniques. Pernicious anoemia may cause nuclearenlargement giving rise to errors in the interpreta-tion of gastric material and the cytopathologist isentitled to an intelligent clinical history in this, asin other types of specimens.

Anderson, MIcDonald and Olsen (1949) reportedthe use of cytologic examination of washings ordirect smears from the cisophagus but had manyfalse negative results. However, Johnson, Koss,Papanicolaou and Seybolt (I955) reported 70%N,correct positive diagnoses in malignant tumours ofthe esophagus from washings and Rubin (I960),stated that 94 to 9800 accuracy is possible.

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Lemon and Byrnes (1949) reported some successwith the detection of carcinomas of the biliarytract anid pancreas using duodenal drainagespecimens. Others, including Raskin, WVenger,Sklar, Pleticka and Yarema (I958), have obtainedduodenal fluid with malignant cells in a very highpercentage of cases.

Malignant tumours of the colon and rcctum havealso been subjected to cytologic detection utilizingdirect smears obtained through the proctoscope orlavage specimens. WVisseman, Lemon an{dLawNrenice (1949), Bader and Papanicolaou (1952)and Chapman (I959), reported good results.Oakland (I96I) was able to identify malignant cellsin saline lavage specimens from 8300 of coloniccarcinomas. Ulcerative colitis and polyps mayproduce atypical cells but with experience thesecan be distinguished from true malignancy.

Since gastrointestinal diagnostic procedures arerarely performed on patients w\ithout clinicalevidence of disease, and since many of the neo-plasms in question are notoriously silent in theirearly phases, the number of patients benefited bythis means of diagnosis may be small. However,Shahon and Wangensteen (I960) reported a cancerdetection study in which gastric analysis and stoolexamination were part of the general examinationof asymptomatic patients. Nine thousand andthirty-five patients were examined in this way andof 296 with cancer, the gastrointestinal tract wasthe site of the lesion in 70. One-third of thesepatients were found on review to be truly asympto-matic and the five-year survival rate for these was500 . Compared with the dismal survival figuresusually reported for gastrointestinal malignancies,it would seem that procedures to detect asympto-matic cancer of the stomach and intestine would atleast double the present cure rate.

Carcinoma in Other SitesThe diagnostic aspects of exfoliative cytology

have been investigated in relation to many otherareas of the body. Brill and Koprowska (I955)and Papanicolaou, Bader, Holmquist and V'alk(1956, 1958) published articles reflecting interest inthe cytology of secretions from the breast. Spriggs(I954) and McCormack, Hazard and Belovich(I954) and Naylor (I96I) have reported cytologicfindings of value in cerebrospinal fluid. Lesionsof the pharynx, nose and ear have been investigatedby Friedman (I95I. Umiker and Lampe (I960)and Salndler (I96I) have recently studied cytologyof the oral mucosa anid skini lesions have been in-vestigated by Woodburne, Philpott anid Philpott(1960). lTltmanin, Koprowska anid Engle (1958)anid otlhers have rccomimncided the study of imprintsmears of tissues, includiing lymplh nodes, for the

purpose of making more definitive or morerapid diagnoses.

Exfoliative Cytology as an Aid to Therapy'T'here are several area-, in whiclh the study of

exfoliated cells may be helpful in directing therapyof malignant disease. Vaginal smcars obtainedfrom patients with carcinoma of the breast mayyield information of value in planning the course ofendocrine therapy. A woman whose vaginalsmear displays marked estrogenic effect is morelikely to benefit from oophorectomy and andro-genic therapy than one with little estrogenic effect.Studies of changes in vaginal smears of patientstreated by radiation for carcinoma of the cervixwere made by Graham (1947). In I95I, she re-ported that patients could be divided into thoseshowing good and poor responses to radiation.The patients displaying a good response wereeight or ten times more likely to survive five yearsthan those showing a poor response. This in-formation has been utilized in determiningwhether surgery should be performed in additionto radiotherapy. Present concepts of the criteriaand significance of cellular radiation response aresummarized in a 1959 symposium of the Inter-national Academy of Gynecological Cytology(Wied-Editor).

Malignant Cells in the BloodOf considerable interest is the recent work on

the isolation and demonstration of malignanttumour cells in the circulating blood. Althoughthere were several reports between I869 and 1950of malignant cells found in the blood of patientswith cancer, Engell (I955) was the first to succeedin isolating and identifying such cells. He lysedthe erythrocytes with saponin and studied hTma-toxylin and eosin sections of cell blocks of theremaining sediment. Two years later Moore,Sandberg and Shubarg (I957) also identifiedtumour cells in haematologic preparations of bloodafter red blood cells had been removed by fibrino-gen sedimentation. Roberts, Watne, McGrath,McGrew and Cole (1958), used fibrinogen sedi-mentation and separation of the remaining cells byflotation on bovine albumin of high specificgravity. MIalmgren, Pruitt, Del Vecchio and Potter(I958) lysed erythocytes and polymorphonuclearleukocytes with streptolysin-O, concentrating theremaining cells on millipore filters. Seal (I959)devised a silicone flotation method and collectedthe cells thus obtained on millipore filters.Numerous variations on these isolation methodlslhave been directed toward more coinplete recoveryamid better preservationi of cells. Recently, Kuper,Bignall and Luckcock (I96I) reported an ingeniousmetlhod of eliminating polymorphonuclear leuko-



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Reprinted with permission of The Yoturnal of Urology.FIG. 4.-Cells from adenocarcinoma of the kidney: (a) Direct smear of cut surface of the

tumour. (b) Blood from antecubital vein. Papanicolaou stain. x I400.

cytes writh a magnet after they have ingested ironparticles.

In general it has been found that malignantcells can be demonstrated in the blood in about30/ of patients with cancer (Fig. 4). Earlysmall lesions rarely, if ever, yield positive bloodsamples and the majority of patients with positivesamples have advanced, incurable disease. It isevident, therefore, that this technique has littlevalue as a detection method.

If serial blood samples are drawn during opera-tive procedures a definite correlation betweenmanipulation and release of tumour cells into thecirculation can be demonstrated. Pelvic examina-tion, curettage, prostatic massage and vigorousscrubbing of skin overlying a tumour have resultedin ' showers ' of cells. Fig. 5 shows the relativenumbers of cells per cubic centimetre of bloodin samples removed during resection of an ovariancarcinoma. From a pre-operative negative level,38 cells per cubic centimetre suddenly appearedduring pelvic examination after the patient hadbeen anxsthetized. Twenty to 120 cells percLubic centimetre were found in samples takendluring the surgical procedure. It is of initeresttlhat, wvitlhin a few minutes after removal of atumour, cells almost always disappear from subse-

quent samples, even if metastases are known to bepresent at the time. Patients with advanced dis-seminated disease sometimes have a fairly constantnumber of malignant cells in blood samples. Insuch cases, reduction in the number of cells can bedemonstrated during chemotherapy and also forseveral days following surgical procedures such asgastrostomy or exploration.

Speculation as to the prognostic significance ofthe presence or absence of tumour cells in the bloodis rather premature at this time. Engell (I959)reported that patients with and without circulatingmalignant cells had essentially the same five tonine year survival rate, but Long, Roberts,McGrath, McGrew and Cole (I960), noted 38%two to four year survival in patients whose bloodsamples were negative during their operations,while only 20(o0 of those with positive samplesduring surgery survived.

It is hoped that this kind of information may beuseful in the future in determining plans forclhemotlherapy or other measures. Modificationsof diagniostic and surgical procedures to reduce thenumLber of cells released into the circulation wouldseem to be clearly indicated at present.The search for malignant cells in saline washings



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August I96I McGREW: Role of Exfoliative Cytology in Diagnosis and Treatment of Malignant Disease 465

120

100 TUMOR REMOVED

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FIG. 5.-Cancer cells in the peripheral blood (antecubital vein) during resection of ovarian carcinoma.

of surgical wounds and of body cavities before andafter operations for cancer have been carried outby Smith, Thomas and Hilberg (1958) and Burke

and Sako (I960). These findings may also be ofprognostic value and lead to improvement intherapy.

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and Cardia of the Stomach, Proc. Mayo Clin., 24, 245.AYRE, J., and OREN, G. (1953): A New Rapid Method for Stomach Cancer Diagnosis: The Gastric Brush, Cancer,

6, I I77-BADER, G. M., and PAPANICOLAOU, G. N. (1952): Application of Cytology in Diagnosis of Cancer of the Rectum,

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2, 44.BECKERMAN, H. A., SPROUL, E. E., and BARACH, A. L. (I958): An Aerosol Method of Producing Bronchial Secretions

in Human'Subjects, Dis. Chest, 33, 347.BIZZOZERO, GIULIO (1887): Handbuch der Klinischen Mikroskopie, 2nd German Edition. Erlangen: Eduard Besold.BRILL, R., and KOPROWSPA, I. (1955): Diagnosis of Early Carcinoma of the Breast by Papanicolaou Technique,

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Tumours Amongst Dyestuff Workers, Brit. J. Industr. Med., 13, 270.DEDEN, CARL (1954): Cancer Cells in Urinary Sediment, Acta radiol. (Stockh.), Suppl. II5.DUDGEON, L. S., and WRIGLEY, C. H. (1935): Demonstration of Particles of Malignant, Growth in Sputum by Means

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FRANK, T., and SCOTT, W. W. (1958): Cytodiagnosis of Prostatic Carcinoma: A Follow-up Study, J. Urol., 79, 983.FREMONT-SMITH, M., GRAHAM, R. M., and MEIGS, J. V. (1948): Early Diagnosis of Cancer by Study of Exfoliative

Cells, _7. Atner; me'd. Ass., 138, 469.FRIEDMAN, I. (I95I): Exfoliative Cytology as Aid in Diagnosis of Tumours of Throat, Nose and Ear, 7. Laryng., 65, I.GOWAR, F. J. S. (1943): Cancer of the Lung, Value of Sputum Examination in Diagnosis, Brit. Y. Surg., 30, I93.GRAHAM, R. M. (I947): Effect of Radiation on Vaginal Cells in Cervical Cancer, Surg. Gynec. Obstet., 84, I53.- , ULFELDER, H., and GREEN, T. H. (1948): Cytologic Method as an Aid in Diagnosis of Gastric Carcinoma, Ibid.,

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HAMPELN, P. (I897): Uber den Auswurf Bein Lungencarcinom, Z. klin. Med., 32, 247.HAMPSON, F. (1956): Exfoliative Cytology in the Diagnosis of Lung Cancer: Examination of One Laboratory's

Results, Brit. med. Y., ii, 1461.HARPST, H. C., WARE, R. E., and EISENBERG, R. B. (I96I): Exfoliative Cytology of the Urinary Tract: Evaluation of

the Millipore Technique, Acta Cytologica, 5, 195.HERBUT, P. A. (1946): Bronchogenic Carcinoma: Diagnosis by Cytologic Study of Bronchial Secretions, 37. Amer.

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Evaluation of 364 Cases, Cancer, 8 (5), 951.KLEIN, M. R., POLLACK, A., and VOLK, H. (1959): A Machine to Collect Sputum for Screening for Lung Cancer.

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Blood, Lancet, i (7 I82), 852 .LEMON, H. M., and BYRNES, W. W. (I949): Cancer of the Biliary Tract and Pancreas, 7. Amer. med. Ass., 141, 254.LONG, L., ROBERTS, S., MCGRATH, R., MCGREw, E., and COLE, W. H. (I960): Cancer Cells in the Blood Stream,

Arch. Surg., 8o, 639.LUND, C. J. (I96I): An Epitaph for Cervical Carcinoma, Y. Amer. med. Ass., 175, 98.MALMGREN, A., PRUITT, J. D., DELVECCHIO, P. R., and POTTER, J. F. (1958): A Method for the Cytologic Detection of

Tumor Cells in WVhole Blood, 7. nat. Cancer Inst., 20, 1203.MCCORMACK, L; J., HAzARD, J. B., and BELOVICH, D. (1954): Neoplastic Cells in the Cerebrospinal Fluid. Trans.

Second Annual Meeting of I.S.C.C., p. ii6.MCDONALD, J. R., and WOOLNER, L. B. (1947): Bronchogenic Carcinoma: Diagnosis by Microscopic Examinations

of Sputum and Bronchial Secretions, Proc. Mayo Clin., 22, 369.-, ~(I949): Cytologic Diagnosis of Cancer from Body Secretions and Fluids, S. Dak. 37. Med., 2, 216.MCGREW, E. A.: Bibliography on request.- (1954): Cytology of Urine Sediment in Renal Cell Carcinoma. Trans. Second Annual Meeting of I.S.C.C., p. I i i.MELAMED, M. R., Koss, L. G., RICCI, A., and WHITMORE, W. F. (I960): Cytohistological Observations on Developing

Carcinoma of the Urinary Bladder in Man, Cancer, 13, 67.MOORE, G. E., SANDBERG, A., and SHUBARG, J. R. (I957): Clinical and Experimental Observations of the Occurrence

and Fate of Tumor Cells in the Blood Stream, Ann. Surg., 146, 58o.NAYLOR, BERNARD (I96I): Cytologic Study of Intracranial Fluids, Acta Cytologica, 5, 198.OAKLAND, D. J. (I96I): Diagnosis of Carcinoma of the Large Bowel by Exfoliative Cytology, Brit. Y. Surg., 48, 353.PAPANICOLAOU, G. N. (1928): New Cancer Diagnosis. Proc. Third Race Betterment Conf., p. 528.- and TRAUT, H. F. (1943): Diagnosis of Uterine Cancer by Vaginal Smear. Commonwealth Fund. Cambridge,

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