Studies on Bovine Lymphosarcoma - Cancer Research...BreedNo.Holstein Guernsey Jersey Ayrshire Angus...

Studies on Bovine Lymphosarcoma

This paper reports observations on the clinicalmanifestations and pathological alterations in bovine lymphosarcoma. During the past 2 yearsdetailed studies have been made on 48 cases. Wehave included partial data on eleven additionalcases currently under investigation. Studies onthe distribution of cases in six herds with a highincidence of lymphosarcoma are also reported.

a Supported by research grants from the American CancerSociety (E-210, E-211, E-111C, and E-211A); U.S.P.H.S.Grant CY 8845 (CS); and a grant from the World HealthOrganization.

t U.S.P.H.S.PostdoctoralResearchFellow(GrantCF10298).

@ Pennsylvania Plan Scholar.Â§AmericanCancerSocietyPostdoctoralResearchFellow

Grant E-210. Present address: School of Veterinary Medicine,University of California, Davis, California.

Received for publication August 17, 1961.

With the possible exception of ocular squamouscell carcinoma, lymphoid tumors are regardedas the most common neoplasms of domestic cattle(5, 11, 15, 38, 63). In this study, all cases fitinto the category of lymphosarcoma' and/or lymphatic leukemia. There were no cases of Hodgkin'sdisease, giant follicular lymphoblastoma, or reticulum-cell sarcoma (unless the latter be considered the same as lymphoblastic lymphosarcoma).

According to Karlson (26), lymphosarcoma inthe bovine species was probably first described

1 Many other names have been applied to lymphoid tumors

of cattleâ€”e.g., leukosis, lymphoblastoma, lymphocytoma,leukemia, lymphadenosis, pseudoleukemia, and malignantlymphoma. At a recent WHO conference on â€œComparativeStudies in Leukemias,â€•Feb. 27â€”Mar.4, 1961, Philadelphia,the term â€œleukaemiaâ€•was used to embrace all proliferativediseases of the hematopoietic system.

202

I . Clinical Aspects, Pathological Alterations, and Herd Studies*

R. R. MARSHAK,L. L. CORIELL,W. C. LAWRENCE,t J. E. CROSHAW,JR.,

H. F. SCHRYVER4 K. P. ALTERA,@AND W. W. NICHOLS

(School of Veterinary Medicine, Univerrity of Pennsylvania, Philadelphia, Pa.; andSouth Jersey Me4WOJ Re.,earch Foundaiion, Camden, N.J.)

SUMMARY

An attempt has been made to characterize lymphosarcoma of cattle with regard toclinical manifestations, pathological alterations, and familial distribution of cases inhigh-incidence herds.

According to data from a total of 59 cases it is evident that, in many respects, thecattle disease resembles lymphosarcoma of man. There is localized neoplastic proliferation of lymphoid tissue which results in diffuse infiltration of organs or formationof discrete solid tumor masses. These are locally destructive and invasive. The processprobably spreads by metastasis and, in time, becomes widely disseminated. Someanimals develop leukemic peripheral blood with massive bone marrow involvement;in many others, the blood and marrow appear normal throughout the course of the

disease.The common clinical picture is one of emaciation, pallor, a history of progressive

weight loss, and decreased milk production. Enlargement of palpable lymph nodes isan outstanding feature.

Analysis of pedigree data from cattle with lymphosarcoma in six multiple-caseherds indicates that the probability of chance occurrence of the disease in these relatedanimals is extremely remote. This familial aggregation of cases is highly suggestive ofgenetic susceptibility to lymphosarcoma. The data are equally compatible with aconcept of vertical transmission of an infectious agent.

on July 2, 2021. © 1962 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from

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BreedNo.Holstein

GuernseyJerseyAyrshireAngus

Total:43

7531

59

l@IARSHAKet al.â€”Studies on Bovine Lymphosarcoma 9203

by Leisering in Germany in 1871 ; since then,numerous case reports, mostly European (2, 6â€”8,10, 12â€”14, 19, 20, 24, 25, 28â€”30, 35â€”37, 39, 41,

44-46, 50â€”52,61â€”65,68, 69), have appeared in theliterature, but detailed studies have been few.

MATERIALS AND METHODS

Cattle with lymphosarcoma (diagnosis confirmed by tissue biopsy and hematological technics) were procured through the farm animalclinics, School of Veterinary Medicine, Universityof Pennsylvania, and through the cooperationof practicing veterinarians in a multistate area.In addition, six large herds with a high incidenceof lymphosarcoma were available for long-termexperimental study.

Animals with lymphosarcoma were hospitalizedand maintained for study at the School of Veterinary Medicine. Where possible, accurate and detailed histories were taken with particular reference to herd and family incidence. The clinicaland laboratory work-up included complete physical examination, measurement of lymph nodes,recording of an electrocardiogram, complete bloodcount (2â€”3X weekly), bone marrow biopsy, platelet count, reticulocyte count, sedimentation rate,urinalysis, fecal examination and determinationof blood urea nitrogen (BUN) (23), blood sugar(18), protein-bound iodine (1) and serum ketones,2

uric acid (40), alkaline phosphatase (4), totalprotein (66), inorganic phosphorus (17), calcium(16), magnesium (42), sodium,3 potassium,3 chlo

ride (@53),bilirubin (34), serum glutamic-oxalacetictransaminase (SGO-T) (47), serum glutamic-pyruvic transaminase (SGP-T) (47), and serum lacticdehydrogenase (LDH) (43). Most determinationswere done at least twice, except when animalsfailed to survive for more than a few days afterhospitalization. All blood and marrow films werefreshly prepared at time of sampling and stainedby Wright's method. The blood of all cattle wastyped to determine blood groups.4

At time of death or sacrifice, each animalwith lymphosarcoma was subjected to completegross and histopathological study. Histopathological studies were done on tissues fixed either in10 per cent buffered formalin or in acidified Zen

ker's solution and stained with hematoxylin andeosin, Giemsa stain, or Gridley's reticulum stain.Zenker's fixation was preferred for cytological

2 E. IL Stadtman, Acetoacetate and acetone-microdiffusion

method, unpublished data.

3 Chemical analysis for sodium and potassium by flame

photometry.

4 Cattle Blood Typing Laboratory, The Ohio State Univer

sity, Columbus, Ohio.

detail and for staining by the Giemsa technic.Formalin fixation was preferred for hematoxylinand eosin and for Gridley's reticulum stain. Lymphnode and tumor imprints made at necropsy wereair-dried and stained with May-Grunwald Giemsastain.

In the high-incidence herds, the familial distribution of lymphosarcoma was determined bypedigree analysis. These data were then analyzedto determine the probability of chance occurrenceof lymphosarcoma in related animals.

RESULTS

Clinical studies.â€”Ante mortem observation penods on cattle with lymphosarcoma ranged from1 to 180 days, with a mean study time of 24 days.Until time of transfer to the University, all animalshad been maintained under usual herd conditions.In each case diagnosis was confirmed by tissuebiopsy.

TABLE 1

BREED DIsTRIBUTIoN59 CASEs OF BOVINE

LYMPHOSARCOMA

Table 1 shows the breed distribution of animaLsin this series. It appears probable that the highincidence observed in Holsteins is simply a reflection of the greater numbers of this breed in thestudy area. The low incidence of lymphosarcomain the beef breeds of cattle (one case in thisseries) is at least partially accounted for by the

usual circumstances of early slaughter.Ages of affected cattle ranged from 1 to 17

years, with a mean age of 5.5 years (Chart 1).@The ratio of females to males was 55 to 4 and probably reflects the normally high ratio of cows tobulls in the dairy cattle population. Of 54 maturecows, 30 were nonpregnant, and 24 were pregnant(five in the first trimester of pregnancy,nine inthe second trimester of pregnancy, and ten in thelast trimester of pregnancy).

In the absence of enlarged palpable lymphnodes or positive blood or bone marrow findings,diagnosis was difficult. Lymphosarcoma may mim

5 Since this was written, we have acquired a 2-month-old

female calf with lymphosarcoma.



SignsNo.cases

showingsignsPer

cent ofcases show

ingsignsAdenopathy

(palpable nodes, includlag rectal palpation)

Asthenia, depressionMarked weight lossTach@rcardia (>100/min)Respiratory distressAnorexia44

272419181492

56504038

29Posteriorparesis

Cardiac abnormality on auscultation(arrhythmias, murmurs, etc.)

Fever(>102.5Â°F.)14111029

2321Exophthalmos817Urinary

abnormalityDiarrhea6 51310Constipation48Edema36

9204 Cancer Research Vol. 9292,February 1962

ic primary heart, renal, respiratory, nervous, orgastrointestinal disease. However, the commonpicture is one of emaciation, with pallor of the mucons membranes, a history of progressive weightloss, and decreased milk production. Enlargementof palpable lymph nodes is an outstanding feature.

Table 2 summarizes the most important physical findings at the time diagnosis was established.

Lymph node enlargement was the most commonchief complaint and was present in over 90 percent of cases. Nodes were usually several timesnormal size, firm, well circumscribed, and apparently painless (Fig. 1). Involvement was oftenwidespread and symmetrical. At times, palpablenodes attained enormous proportions. Chart 2indicates the frequency of involvement of palpablenodes in the living animal. Other signs attributableto enlarged nodes were related to pressure ofnodal masses on structures in the thoracic, abdominal, or pelvic cavities or to lymphosarcomatous infiltration of the heart, kidneys, abomasum,and other organs. The presence of enlarged palpable lymph nodes did not accurately reflect thedegree of involvement of inaccessible nodes orother internal organs. In two cases it appearsprobable that the primary sites of involvementwere extra-nodal. In one case only the heart wasinvolved ; in the second case involvement wasrestricted to the genito-urinary system. Regionallymph nodes were not affected.

Cardiovascular system.6â€”Heart lesions were com

6 Special examinations of the cardiovascular system were

conducted by Drs. D. F. Patterson and J. W. Buchanan, Comparative Cardiovascular Studies Unit (U.S.P.H.S. GrantH4885).

mon and contributed to the frequently observedsigns of asthenia, tachycardia, and dyspnea. Cliical examination of the heart consisted of auscultation of the valve areas, determination of the size ofthe area of cardiac dullness by percussion of the thorax, observation of the degree of jugular distensionand pulsation, and recording of a standard limblead, augmented unipolar lead, and unipolar precordial lead electrocardiogram. Phonocardiogramswere recorded in some cases.

In many animals the results of clinical examination of the heart were negative. In others the onlyremarkable finding was sinus tachycardia, withrates of 90â€”120per minute.

Seven animals had increased jugular venouspressures, suggesting early congestive heart failure; all but one showed abnormal jugular pulsations. Venous pressures in excess of 20 mm. ofmercury were recorded for three of these animals.Two were in failure; cardiac output was wellbelow normal limits in both.

Auscultation of the heart revealed presystolicand systolic murmurs in two cows with elevatedvenous pressures. One of these had a diastolic

TABLE 2

CLINICAL MANIFESTATIONS IN 48 CASES

OF BOVINE LYMPHOSARCOMA

â€”i9ge in @cars@

CHART 1.â€”Ages of cattle in 59 cases of lymphosarcoma

a Cases in which careful physical examination was per.formed on living animal.

gallop rhythm as well. Presystolic murmurs anddiastolic gallop rhythms were noted in two othercows, and an isolated systolic murmur in another.An audible fourth heart sound was present in twoanimals which had sinus tachycardia (the fourthheart sound is occasionally heard in normal cattle,but only at slow heart rates). A systolic galloprhythm was present in one cow.



MARSHAK et al.â€”Studies on Bovine Lymphosarcoma 9205

Percussion of the chest revealed an increasedarea of cardiac dullness in three cows, one of whichhad an elevated venous pressure.

There were few electrocardiographic abnormalities. Delayed atrioventricular conduction was present in five animals with sinus tachycardia. Oneof these and another cow had QRS axes whichfell somewhat outside the range usually observedin normal cattle (at the present state of knowledgeno interpretation can be given to this finding).Isolated@ findings in three other animals were:prolonged QT interval, electric alternans, andslight ST segment deviation.

Although clinicopathologic correlations havenot been completed, clinical studies indicate thatsigns of cardiovascular disease are often presentin cows with lymphosarcoma. Pathologic studiesto date have revealed cardiac involvement in89 per cent of the animals necropsied. Signs ofcongestive heart failure in six cows suggest thatcardiac failure might often be the cause of deathin cattle with lymphosarcoma when the diseaseis allowed to run its full course.

Respiratory system.â€”The lungs were neither frequently nor severely involved, although adenopathy of bronchial, mediastinal, and cervical nodeswas common and undoubtedly contributed to thehyperpnea and/or dyspnea noted as a prominentsign in eighteen animals. In one case, dyspneaand loud stenotic sounds on inspiration and expiration were due to marked tracheal constrictionby a ring of neoplastic lymphoid tissue.

Digestive system.â€”Involvement of the gastrointestinal tract, particularly the abomasum, andassociated lymph nodes was common, though related signs were not prominent. Marked anorexiawas present in 29 per cent of cases ; diarrhea andconstipation were noted occasionally. In one anima! with signs of melena, extensive infiltration ofthe rectum and colon was observed at necropsy.

Urogenital system.â€”Urinary abnormalities wereevident in six cases. In one of these there weremarked weight loss, anorexia, dysuria, and ohguria. A large pelvic mass involving the urinarybladder, uterus, cervix, and vagina was palpable.Significant laboratory findings were anemia, moderate elevation of BUN, and low urine specificgravity. At necropsy the large tumor involved theentire urogenital tract, which was considered asthe primary site of involvement. Regional lymphnodes and all other structures were unaffected.A second animal showed incontinence and posterior paresis associated with spinal nerve cornpression by tumorous tissue. In a third cow,BUN was elevated to 40 mg. At necropsy therewas extensive bilateral renal invasion and ureteral

obstruction. In the three remaining animals thepresence of large, obstructing pelvic masses wasassociated with signs of tenesmus, frequent passage of small quantities of urine, and, in one case,hematuria.

Nervous sytem.â€”In fourteen animals bilateralposterior paresis was present. This was most oftenthe result of cord compression, with or withoutmeningeal invasion; in several animals the paresisresulted from compression of spinal nerves bytumor.

Other signs.â€”Ten of the animals studied hadfever in the absence of clinically detectable infection. Moderate to severe exophthalmus, present

in eight cases, resulted from pressure by a rapidlygrowing retrobulbar tumor mass (Fig. 2) . Occasionally, small lymphosarcomatous nodules were

Cral7ial Cgrzi.

Caadal CQrzi.

Aortzc (Zcanh)

5&pzllvaImvRarq

Iliac

Prtl-IQ.moral

Pril-xapalar

5 10 15 20â€”BnÃ¼nals@

30 35 40

CHART 2.â€”Frequency of enlargement of palpable lymphnodes in 59 cases of lymphosarcoma at time clinical diagnosiswas established.

observed in the skin. These were usually small,firm, well delineated, single masses. In three animals extensive subcutaneous growths were presentover the rib cage. Hemorrhagic tendencies or boneinvolvement has not been noted.

Laboratory findings.â€”Laboratory findings, exclusive of hematology, are summarized in Table3. Many animals were sampled during the terminalstages of the disease.

Elevated BUN values were observed in fifteencases. Urinary tract involvement was present inthree animals ; eleven others were in the terminalstages of lymphosarcoma.

Decreased serum protein levels were observedin emaciated and severely anemic animals. Ele

vated serum uric acid values, noted in ten nonterminal cases, were interpreted as evidence ofincreased breakdown of nucleoproteins and/or failure of excretion. The marked elevation of SGO-T,observed in twenty out of 21 cases of lymphosarcoma tested, is probably related to widespreadinvolvement of tissues rich in this enzyme or tocirculatory derangements resulting from the pres



ConstituentNo.

animals withabnormal values/

Total no.mals testedAbnormal

valuesRemarks

TABLE 3

LABORATORY STuDIEs IN BOVINE LYMPHOSARCOMA5

15/40

5/40

9/39

18/39

0/29

20/21

4/11

29/40

5/32

1/821/82

3/821/82

4/87

6/35

6/35

9/87

0/81

0/22

2/222/21

0/11

Blood urea nitrogen(high)

Blood sugar (low)

Serum total protein(low)

Serum uric acid (high)

Serum bilirubin

SGO-T (high)

SGP-T (high)

Serum LDH (high)

Serum Nat (low)

Serum K'@(high)(low)

Serum Cl(high)(low)

Serum Ca@(low)

Serum Mg@(low)

(high)

Inorganic phosphorus(high)

Serum alkaline p'tase

Range: 82â€”175mg.%Mean: 65 mg.%

Range: 25â€”34mg.%Mean: 28 mg.%

Range: 4.8-5.7 gm.%Mean: 5.3 gm.%

Range: 2.4â€”6.6mg.%Mean: 4.2 mg.%

Range: 110-770 S.F unitat

Mean: 310 S-F units

Range: 41-47 S-F unitsMean: 44 S-F units

>s.5x103 units

Range: 104â€”188mEq/l.Mean: 128 mEq/l.

8.8 mEq/l.1.5 mEq/l.

Range: 118.5â€”123 mEq/l.81.5 mEqjl.

Range: 2.5â€”3.5mEq/l.

Range: 0.61â€”1.2 mEq/l.Mean: 0.94 mEq/l.Range: 3â€”5mEq/l.Mean: 3.9mEq/1.

Range: 9.5â€”18.7mg.%Mean: 12.0 mg.%

11 cases were terminal at time of sampling; in 3 casestumor involved urinary tract.

4 cases were terminal at time of sampling.

5 animals were severely anemic; 6 cases were terminalat time of sampling.

5 animals were leukemic; 8 were terminal at time ofsampling.

Normal values for adult cattleâ€”48 .8 Â±5.7 S-F Unith4

Normal values for adult cattleâ€”19 .7 Â±12 .6 S-Funits4

Normal values for cattle usually

CowNo.No.

nays5TUDIEDNo. COUNTSL@[email protected] (psn cu.sa.)Hb

(gm/100ml)HctHb

(gm/100ml)HctNb

(gm/100ml)Hct2

3101112131516181921222826A29803237443

1734484

102050473218272168079491

4288251527201678214333245.8

@5.39.18.78.89.49.29.98.812.98.39.85.98.910.98.29.75.38.417

1928252730272631352329192831253017254.6

8.75.56.88.24.23.47.59.66.96.68.28.010.48.29.44.211.417

27222623181325272120152529282614345.0

9.27.67.89.06.87.87.69.27.26.74.58.210.38.39.14.89.418

282726.52822.628.624.929.622.822.117.2263024271628400,000

60,800

220,000370,500178,75061,710660,190557,220

MARSHAK et al.â€”Studies on Bovine Lymphosarco'ma 9207

ence of tumor masses. SGP-T levels were highin only four out of eleven animals tested.

Elevated serum LDH levels (3.5 X 1O@unitsto 4.0 X 10@units) were recorded for 29 out of40 cases in this series. Similar findings for cattle

with lymphosarcoma have been reported elsewhere (59). In normal cattle, serum LDH levels7are usually below 3.5 X [email protected] Though somerise in LDH activity appears to occur in cattleover 5 years of age, 32 out of 33 normal serumsamples were found to contain less than 3.5 X 1O@units.

not unusual. Hematocrit readings paralleled hemoglobin levels, reflecting the degree of anemia.Morphologically, the red cells were normocyticand norrnochromic.

In anemic animals the bone marrow was eitherleukemic or showed marked reduction in erythroidelements, indicating that decreased red cell production rather than increased destruction of redcells or blood loss was the major factor. Thoughanemia tended to be more severe in animals witha leukemic blood picture, it was a common findingin cattle without leukemia.

TABLE 4

TYPICAL HEMOGLOBIN, HEMATOCRIT, AND PLATELET VAiuEs IN BOVINE LYMPHOSARCOMA

Most electrolyte abnormalities appear to beassociated with the terminal stages of lymphosarcoma.

Hematology.â€”Tables 4 and 5 summarize cornplete blood counts on nineteen animals with lymphosarcoma. These were selected to represent thevaried hematological configurations encounteredin our series of 48 cases.

It is evident that anemia is a common finding.

Of 34 animals with normal initial hemoglobinvalues, 24 were in the lower portion of the physiologic range (8.0â€”14gm. per cent) (54). Hernoglobin values in most serially followed animalsdeclined steadily. Values below 5 gm/100 ml were

7 There was little or no difference in LDH values in serum

and plasma from the same animal.5 J. C. Cardeilhac and P. T. Cardeilhac, Preliminary Studies

on Lactic Dehydrogenase Activity in Blood and Tissues fromNormal and Leukemic Cattle (in press).

Platelet numbers were low in only four out ofsixteen cases in which counts were made. All

were terminal cases at time of sampling.In four animals, the average total white cell

count ranged from 81,500 to almost 300,000 percu. mm., with greater than 90 per cent lymphocytes (Figs. 9 and 10). In ten other aimals inwhich counts ranged from 20,000 to 48,800 perCu. mm., lymphocytes were the predominant celltype. These cells were usually very large withabundant cytoplasm; the nuclei were often bizarrein shape and contained a lacy or spongy chromatin,with several bodies resembling nucleoli. Mitoticfigures were common (Fig. 10), giving the impression of immature, rapidly proliferating cells.

In five animals in which initial counts werenormal, daily sampling showed that they ultimately entered a leukemic phase. In two animaLs,average total white counts were low, and in several



others, followed serially, leukopenia appeared at usualâ€• or â€œabnormalâ€•lymphocytes appeared inone time or another. the peripheral blood (Fig. 8) ; however, these were

The occurrence and extent of palpable lymph difficult to classify and seldom exceeded S ornode involvement did not appear to be correlated 4 per cent of total lymphocytes.with the presence or absence of leukemic peripheral Studies on several hundred healthy cows mdiblood. cate that many normal animals may have a small

In many animals followed serially, white and percentage of unusual-appearing lymphocytes indifferential counts remained essentially normal their peripheral blood (Figs. 5, 6).until the end (Fig. 12). Intermittently, some â€œun- Table 6 illustrates eight representative differen

TABLE 5

TYPICAL LEUKOCYTE AND DiFFERENTIAL COUNTS IN BOVINELYMPHOSARCOMA*Cow

NO.DAYSSTUD

â€˜EDNo.COUNTSINITIAL

%

@ â€”

WBC N E B M L ULFINAL

%

@â€” â€”â€”â€” â€”â€”

WBC N E B M L ULAv.

â€” â€”

WBC %L %ULAv.

anSOLUTE

LTMPES2

S101112131516181921222826At29803287448

173448

4102050473218272163079491

428825

1527201678214S3324293.0

18.58.420.728.914.699.46.78.118.92.47.3

282 .010.77.6

256.015.619.541.30

733533

934

31142341731

21826

1238510

0810102612251211

1310

0000000101000

S3403045805031100

2450629142688244387556977061

744746many

040

6820341285S

many71

100114S11.9

7.424.243.242.355.65.314.835.05.65.4

240.08.79.8

240.036.518.291.473

856899585675046

33069

2554480

400100002001

1

50

400100000000

311

21031202601

525

52769034438932844647976830

7325482

27

625448005901

many

1100116414.7

7.028.736.028.284.27.213.184.74.97.2

261 .012.37.5

249.021.518.943.819

51722544545939444654977142

7836431

15

658287223812

2many

S2

10021542.9

8.618.232.421.476.64.45.826.72.04.0

253.09.13.8

249.016.19.620.6

CEu@TYPEANnEAL

NO.1015161819215515Blasts(stemcells)

RubriblastsErythroblastsNormoblastsPromyelocytesNeutrophilicmyelocytesNeutrophilicmetam.yelocytesNeutropbilicbands(stabs)NeutrophilsEosinophilicgroupBasophilicgroupLymphocytesUnusuallymphocytesPlasmacellsMonocytesMitoticfiguresOther1.5

04.08.02.08.58.522.02.012.0029.0

1.51.02.01.07.02.0

06.02.501.01.02.54.00021.060.0

00002.3

2.010.04.01.38.66.611.86.31.0085.67.00.60.82.03.01.6

08.35.30.38.04.318.013.620021.8

1.000.70.761.5

09.56.501.03.512.04.52.0041.020.5

00003

02319

1362051017

010201.5

1.017.518.00.56.04.512.56.05.50.527.5

0.5000.500.5

011.013.50.55.05.015.58.01.5034.5

2.00.501.50.5


* WBCâ€”white blood count; Nâ€”neutrophil; Eâ€”eosinophil; Bâ€”basophil; Mâ€”monocyte; Lâ€”lymphocyte; ULâ€”unusua

lymphocyte.t Mastitis.

TABLE 6

REPRESENTATIVE BONE MAmtow DIFFERENTIAL Cou@ (NEAREST 0.1%) IN BOVINE LYMPHOSARCOMA

S Reticuloendothelial cells, fat cells, unclassified, etc.



OrganPercentLymph

nodes95Heart89Kidney

andureter59Abomasum57Uterus

andcervix51Spinalcord49intestine46Liver37Spleen

Urinary bladderRumen, reticulum, omasum27

2119Gallbladder11Lung8Ovary

Adrenal medulla5 S

Animalno.Bone

marrow. Peripheralblood101

ErythroidelementsNormal15LeukemicThrombocytopenia,

anemia,leukemia161

Lymphocytes@ ErythropoiesisAnemia181

ErythropoiesisAnemia19LeukemicLeukemia21MyelopoiesisLeukopenia22NormalNormal121

Lymphocytes,notleukemicLeukemia


tial counts of costal bone marrow from cattle withlymphosarcoma. As depicted, animals with lymphosarcoma may have a normal marrow (animal#10, #12, #16, #18, #21, #22)@ or leukemic marrow

(animal #15 and #19) (Fig. 11). In addition, wehave encountered cases in which areas of metastasis were superimposed upon otherwise normalappearing marrows. Generally speaking, the peripheral blood picture (anemia, leukemia, etc.) accurately reflected the condition of the bone marrow (Table 7).

Necropsy findings.â€”Detailed examinations ofall organs and systems were made on 63 casesof bovine lymphosarcoma.'Â°

As seen at necropsy, lymphosarcoma was usually a widespread disease. The lymphatic organs,especially the lymph nodes, were involved mostprominently and most frequently, but organswhich usually contain small lymphatic foci, suchas the stomach, intestine, kidney, and liver, oftenparticipated in the disease. In addition, certainorgans normally devoid of organized lymphatictissue, such as the heart, were frequently infil

TABLE 7

CoMPARIsoN OF BONE MARRow AND PERIPHERALBLOOD IN SIX CAsES OFBOVINE LYMPHOSARCOMA*

crete, nodular tumors were usually yellow or greyish white, uniformly patternless masses whichwere soft and mushy to moderately firm in consistency. Areas of necrosis and/or areas of hemorrhage were seen only in the larger tumors. Thediffuse, infiltrative tumors showed variable characteristics depending on the organ involved andthe extent of infiltration. Infiltrations of paren

TABLE 8

ORGAN INVOLVEMENT IN 63 CAsEsOF BOVINE LYMPHOSARCOMA9

chyrnatous organs such as liver, kidney, or heartproduced a markedly enlarged, friable organ whichwas very soft and pale. Despite great increasein bulk, the general configuration of affected organs was usually retained.

Lymph nodes.â€”The number of lymph nodesaffected in any particular case varied from fewto many to all nodes examined. In all cases therewas marked asymmetry of nodal involvement.

The lymph nodes of the pelvis, mammary gland,and mesentery were most frequently and mostseverely affected. The lymph nodes of the limbsand thoracic cavity were commonly, but less severely, involved. The nodes of the head and neckwere seldom more than slightly affected.

The degree of change within a node variedfrom a barely perceptible gross alteration to extreme distortion. By examination of many lymphnodes a progression of gross changes was observed.Initially, the medulla appeared to be slightlyswollen and there were one or more unusuallyprominent follicles in the cortex. Sometimes therewas a single, discrete, bulging, greyish-white nod

ule, with the remainder of the node retainingits usual appearance. The normal nodal components were crowded out by the progressively enlarging tumor nodule. Continuing growth produced a greatly enlarged lymph node which was

S I means increase;@ means decrease.

trated by the tumor. Table 8 illustrates the frequency of organ involvement in this series of 63cases.

Lymphoid tumor masses were either discreteand nodular or diffuse and infiltrative. The dis

aThe marrow of Cow #16 shows a lymphocyte count bordering on the leukemic side.

10@ seriesincludesseveralanimalsnecropsiedat theSchool of Veterinary Medicine prior to the start of the currentproject.




often 5 or more times its usual size. All normalarchitecture was obliterated with replacement bysoft, yellow or greyish white, moist, bulging tissue.Streaks of hemorrhage and foci of necrosis appeared in the largest nodes. For the most part, thenodes remained well encapsulated, but eventuallythe neoplastic cells broke through the capsuleand invaded the surrounding tissue. Neighboringnodes often coalesced to form bulky lobulatedstructures. In one instance both the para-aorticand iliac lymph nodes were fused to form an enormous, lobulated tumor which weighed 47 kg. (Fig.4). This large mass infiltrated into most of the pd

vie organs and through the ventral abdominalmusculature into the subcutaneous tissues of theflank.

The progression of gross changes can be followed histologically. The sinusoids, especially themedullary sinusoids, of otherwise normal lymphnodes become packed and distended with large,anaplastic lymphoid cells (Fig. 3). Many mitoticfigures were seen among these cells, and many ofthem had large eosinophilic nucleoli. All had deeply basophihic cytoplasm. Rapid growth of thesecells in one focus produced a grossly visible noduleor a diffuse infiltration with replacement of medullary structures. The cortical tissue was compressedinto a thin rim under the capsule of the lymphnode (Fig. 3). Continued proliferation eventuallypushed aside all the normal elements until thestructure of the node was obliterated. Neoplasticcells finally invaded through the capsule into theadjacent tissues.

Heart.â€”Although the heart is usually considered devoid of organized lymphatic tissue, it wasa site of tumor development in 89 per cent ofcases in this series. In one case the heart was theonly organ which appeared to be involved.â€•

On gross inspection, the lesions appearedas poorly defined, yellowish, swollen streaks or blotches.The right atrium and associated epicardial fatwere most frequently involved. Infiltration of theright atrium sometimes increased the thicknessof the atrial wall to over 5 times its normal dimension. However, the general configuration of theatrium was always well preserved.

Discrete, nodular, sometimes polypoid masseson the endocardial surface were observed (Fig.18) much less often than infiltrative tumors. The

right atrium was also the common site for polypoid tumors. In one case, a saddle embolus of theaortic bifurcation resulted when a left atrial lymphoid polyp broke free and entered the circulation.

11 We have recently necropsied another cow in which the

only abnormal finding was lymphosarcomatous infiltration ofthe heart.

The pericardium was often involved by nodularor sheetlike masses of tumor tissue. Frequently,tumors of the pericardium spread to or fromadjacent structures such as the cranial and caudalmediastinum and the pleural surfaces. Pericardiallymphoid tumors were seldom associated withmyocardial tumors.

The endocardium was commonly infiltrated.Loose, bulging masses of tumor cells, not coveredby endothelium, were often seen on the endocardialsurface.

When the heart was examined histologically,a widespread infiltration by neoplastic cells wasobserved. The invading cells infiltrated widelythrough the interstitial spacesâ€”particularly thoseassociated with Purkinje fibers. Individual muscleand conduction fibers became widely separatedbut seemed to retain their integrity until theinfiltration was severe.

Digestive tract.â€”Involvement of the digestivetract was common. In order of frequency, lymphoid proliferation was observed in the wall ofthe abomasum, colon, small intestine, omasum,rumen, and reticulum.

The abomasum was affected in more than halfthe cases in this series. Part or all of the organwas thickened by tumor cell proliferation, butthe pyloric region was most frequently involved.Infiltration of the pyloric region always haltedabruptly at the pyloric sphincter, never extendinginto the duodenum. Infiltration of all layers of theabomasal wall (Fig. 14) was uniform, so that thegeneral configuration of the organ was retaineddespite massive enlargement and almost total replacement by neoplastic lymphoid cells. Becauseenlargement of the organ was always annular,complete obstruction was never encountered. Diffuse loss of mucosa was not observed althoughdeep, bleeding ulcers were often present. Themucosa was the only element of the abomasumwhich appeared intact when examined histologically (Fig. 15). Occasionally, the omentum wasaffected by localized tumor invasion from theabomasum (Fig. 14).

Involvement of the forestomachs and intestinesdiffered in character from involvement of theabomasurn. Discrete, plaque-like nodes or nodulesof tumor were observed on the serosal surfacesand on the omentum and mesentery. These nodelike structures commonly invaded the deeper layers of the adjacent digestive tract, impinging upon,but never obstructing, the intestinal lumen. Plaquesor nodes in the mesentery often fused, traÃ¼sforming the mesentery into a thick, dense, aÃ¼drigidsheet of tumor tissue.

Kidney.â€”Lymphosarcoma of the kidney was




either nodular and solid or diffuse and infiltrative. The nodular form, which occurred mostoften, usually involved the renal pelvis or thecapsule. Tumor foci were also commonly observedwithin the renal parenchyma. Tumors of the capsule or pelvis infrequently invaded the kidneyparenchyma, whereas tumors in the renal pelvis,which probably originated in the renal lymphnodes, often surrounded the ureter (Fig. 16), invading its outermost coats, and obstructing thelumen sufficiently to produce some degree of hydronephrosis. Tumor nodules involving the renalcapsule were usually small; in two cases, however,many such nodules coalesced to form a thick,dense sheet of neoplastic tissue which completelyencased the kidney.

In the case of diffuse infiltration, particularlyinvolving the cortex, the kidney appeared as avery large, uniformly pale and friable organ (Fig.17) . The cortical architecture was usually obscured(Fig. 18) ; on histological examination the infiltrating tumor cells caused wide separation of tubulesand glomeruli. Nevertheless, the appearance ofthese structures indicated that they had remainedviable.

Uterus and cervix.â€”Lymphosarcorna of the uterus and cervix was frequently encountered andappeared to originate in the endornetrium. Infiltration of the endornetrium, myometrium, and serosaresulted in annular enlargement of the uterus(Fig. 19) ; segments of one horn or the entireorgan were involved. In one case, in which thecervix was the only structure in the body affected,a diffuse, infiltrative neoplasm produced an annular enlargement of over 30 cm. The regionallymph nodes were unaffected.

Eleven cows in this series were pregnant, andthe uterus contained a normal, viable fetus. Theuterus of five pregnant cows showed evidence oflymphosarcoma; the remaining six were not involved. There was no gross or histologic evidenceof lymphosarcoma in the fetal tissues. Microscopicexamination of the uterus and placenta indicatedthat the placenta was an effective barrier againstinvasion by the tumor. Infiltration by neoplasticcells seemed to halt abruptly at the sites of placental attachment (Fig. 20).

Liver.â€”Twenty-three of 63 cases showed somedegree of liver involvement. This varied frommicroscopic foci of tumor cells to extensive, diffuseinfiltration of the entire organ. No solid tumorsoccurred.

The entire liver or portions of it were swollenand pale. Lobular markings in moderately affectedareas were unusually distinct; markings were obliterated in areas of extensive infiltration.

Histologically, the infiltration appeared to originate either in the connective tissue of the portaltriads or within the sinusoids at the peripheryof the lobule (Fig. 21).

Spinal cord.â€”Extradural masses of lymphosarcoma were observed at the origin of the ventrallumbar nerve roots (Figs. 22, 23) in 31 of 63eases in this series. Multiple tumors were presentin most cases. Tumors were nodular in character

and varied in size from a few millimeters to largemasses which filled the vertebral canal and cornpletely encircled the spinal cord (Fig. 24). Theneoplastic cells infiltrated widely through the epidural fat of the lumbar area but invaded only themost superficial layers of the dura.

Peripheral nerves which traverse the pelvicwall and supply the pelvic limbs were sometimescompressed or infiltrated by large tumors involvingthe pelvic lymph nodes.

Spleen.â€”Lymphosarcoma of the spleen was encountered in seventeen of 63 cases. In these thespleen was extensively and diffusely infiltrated.There was marked splenornegaly, with loss ofarchitecture and replacement of splenic tissue bytumor cells.

General histology and cytology.â€”A special cellular pattern or arrangement was not noted. Sheetsof patternless lymphoid tissue appeared to growwidely through tissue spaces, pushing aside andeventually displacing the normal structures. Invasion of serosal surfaces, organ capsules, andblood vessels was commonly observed, and in somecases was massive. In nearly every animal tumor,cell emboli were found in arteries, veins, bloodsinusoids, or lymph vessels (Fig. 25).

The stroma of the neoplastic tissue was scanty.Tumor masses often overgrew the scanty stromaso that hemorrhage and necrosis were frequentlyencountered. Reticulum stains showed that neoplastic cells did not produce reticulum. Pre-existing reticulum was compressed by the advancingtumor and appeared as a dense zone surroundingtumor masses.

Basically, two types of cells were present. Onecell type corresponded to the lymphocyte; theother cell type corresponded to the lymphoblast.In each case the cellular composition was mixed(Fig. 26) ; however, one cell type predominated.In all cases the cellular pattern and compositionwas consistent for all organs involved in the neo

plastic process. Serial biopsies were not performedto determine if changes in cell type developed overa period of time.

In the case of lymphocytic lymphosarcorna,varying numbers of lymphoblasts and an occasional reticulum cell were interspersed singly or



FamilyNo. studiedNo.affectedM18SA80N61D101K120Others5

Totals:Si0805

92192 Cancer Research Vol. 9292,February 196@

in clumps among the preponderant lymphocytelike cells. In sections the lymphocytes appearedas small, round cells (8â€”10 ji in diameter) withdiscrete cell boundaries. The cytoplasm was homogeneous and nongranular and formed a thin rimaround the nucleus. The nucleus was round, centrally located, and deeply staining. Large blocksof chromatin were found within the nucleus. Be

CHART3.â€”Simplified composite pedigree of lymphosarcomacases in herd #1. Solid circles are affected females; open circlesare normal females; open squares are normal males; X, Y, andz representsires whichare commonancestors.Light solidlines show descent from sire Y; broken lines show descentfrom sire X; dotted lines show descent from sire Z. Note that,with exception of one female, all cases descend from all threesire ancestors.

cause the chromatin tended to clump at the nuclear membrane, the appearance of the membranewas very distinct. Occasionally, a weakly eosinophilic nucleolus could be found. One to four mitotic figures per high-power field were usually observed.

In tissue sections from cases of lymphoblasticlymphosarcoma the cells were larger than lymphocytes, ranging in size from 12 to 15 @.iin diameter.The cells were usually round, but irregular shapeswere relatively common. The cytoplasm was moreabundant than in the lymphocytes; many cellshad irregular tags or streamers of cytoplasm.The nucleus was usually spherical, but oval, beanshaped, or coarsely lobed nuclei were sometimesseen. Chromatin clumping@ was much less pronounced than in lymphocytes, but a few smallclumps were found around the nuclear membrane.The nucleus had a pale-staining, vesicular appearance. Nucleoli were always present as large singleor small multiple structures.

In three cases of lymphoblastic lymphosarcomathere were many large cells about 20â€”30jz indiameter. These were usually round, with abun

dant, weakly eosinophilic cytoplasm. The nucleuswas large and coarsely lobulated; multiple nucleiwere commonly observed (Fig. 27). Multinucleated cells contained two or more nuclei, whichwere rather like lymphocytes in chromatin pattern.

The frequency of mitotic figures was the samefor lymphoblastic and lymphocytic lymphosarcoma.

Herd studies.â€”Pedigree analyses of cattle insix large herds with multiple cases of lymphosarcoma showed that affected individuals within eachherd were genetically related. In any particularherd, animals from affected and unaffected familieswere maintained under as nearly identical environmental circumstances as can be found under fieldconditions.

Herd #1 : Chart 3 depicts a simplified compositepedigree of cattle with lymphosarcoma in thispurebred Jersey herd. It is evident that all affectedanimals were related through at least two commonancestors in five generations and that cases aregrouped into three cow families. In one instance,lymphosarcoma appeared in two daughters of onecow and in a daughter of one of these affecteddaughters. In another instance a cow had twoaffected granddaughters, one through a daughterand one through a son. The latter case involvedanother cow family on the dam's side, but thiswas the only case in that family.

For statistical purposes an attempt was madeto separate members of this herd into familiesand to tabulate the numbers of cattle involved

TABLE 9

INCIDENCE OF LYMPHOSARCOMABYCow FAMILIESIN HERD#1

x

* includes all families smaller than six

in number, as well as purchased cows.

against the cases of lymphosarcoma in each family.This information is presented in Table 9. Statistical analysis by the binomial method resulted ina P value of 0.0283 for the â€œMâ€•family, indicatingthe unlikelihood of chance occurrence of lymphosarcoma in this family.

Herd #2 : During the last 5 years a clinicaldiagnosis of lymphosarcoma was confirmed in



Sire groupNo.studiedNo.affectedSireY

Sire L (son of sire Y)SireHSireWLOthers*

Totals:12

1228

5464

210

0987

Sire groupNo.studiedNo.affectedSireR

SireCOthers5

Totals:35

556

765

00

5

MARSHAK et al.â€”Studies on Bo@vine Lymphosarcoma 9213

seven animals in this purebred Jersey herd. Therelationship of affected animals is striking (Chart4). Four were sired by bull â€œY,â€•two were sired

by a son of bull â€œY,â€•and one was a daughter ofone of the affected daughters of bull â€œY.â€•Statistical analysis of the data (Table 10) gives aP value of 0.008657 for the daughters of bullâ€œY',alone; the relationship of bull â€œYâ€•to theother three animals reduces the probability ofchance occurrence of lymphosarcoma in these cattle still further.

Herd #3 : In herd #3, approximately equal nurnbers of registered Guernsey and registered Holsteincattle have been maintained together for nearly20 years. At present, however, there are 150Holsteins and only 35 Guernseys.

Within a 3-year period, five Holstein cows suecumbed to lymphosarcoma, but no cases appearedamong the Guernseys. Although affected animalswere related to one another, none of the cowfamilies or sire groups had more than one case.It is noteworthy that herd #3 and herd #4 (below)

are geographically close and have been artificiallyinseminated from the same bull stud for manyyears.

Herd #4, composed of approximately 300 gradeHolstein milking cows, has been maintained as aclosed herd for at least ten generations. Since itis a state institution herd, its records are remarkably complete and accurate. Out of six confirmed

CHART 4.â€”Simplified composite pedigree of lymphosarcomacasesin herd@ Solid drcles are affectedfemales;open circlesare normal females; open squares are normal males. Brokenarrows show descent from bull Y (see text); solid arrows represent descent from bull D. Bull D is another common ancestorfor S of the 7 cases in this herd.

cases of lymphosarcorna in the last 2@ years,five occurred within a 4-month period in late1960. Three cases, involving a cow and her two

daughters, were in a small cow family consistingof only five members. In addition, one of the daughters had the same sire as one of the cases in herd#3. Statistically, the probability of lymphosarcoma

affecting three out of five cows in this single familyin a herd numbering 300 head is approximately

0.000441.Herd #5 : Herd #5 is a purebred Holstein herd

in which there have been five confirmed casesof lymphosarcoma in less than 2 years. As depictedin Table 11, all affected animals were sired by thesame bull. The probability that this was a chance

occurrence is 0.0192.

TABLE 10

INCIDENCE OF LYMPHOSARCOMA BY SIRE

GROUPS IN HERD @2

* includes all sire groups smaller than five in

number, those whose daughters were entirelyunder 2 years of age, and purchased animals.

TABLE 11

INCIDENCE OF LYMPHOSARCOMA BY

SIRE GRouPs, IN HERD #5

0S Includes all sire groups smaller than

five in number, those whose daughterswere entirely under 2 years of age, andpurchased animals.

Herd @6:This purebred Ayrshire herd has beenin existence for over 40 years. Although recordsare remarkably complete, cases prior to 1951 havebeen omitted, since there is some doubt regardingdiagnosis. As depicted on Chart 5, all seven casesof lymphosarcoma trace at least once to a singlesire and all but one to a single foundation female

as well. Five of the seven cases also trace to another foundation sire. In addition, it is noteworthythat one cow had an affected daughter (#12)and granddaughter (#80) and that these two animals are also daughter and granddaughter of sireâ€œS.',The most recent case involved a @3-monthold heifer (#208). Examination of this animal's




pedigree indicates that two of her direct ancestorsdied of lymphosarcoma.

The blood lines of nearly all animals in herd#6 are similar to the blood lines of individualsdepicted on Chart 4. Further study of this recentlylocated herd, including statistical analysis, is under way.

We have discovered at least two additionalherds in which there appears to be a familialaggregation of lymphosarcoma cases. These willprovide material for future reports.

Dc

CHART5.â€”Simplified composite pedigree of lymphosarcomacases in herd j'6. Solid circlesare affectedfemales;open circlesare normal females; open squares are normal males. Onlypertinent relationships are shown 5, RF, DF, NQ, PER,and PAR are coded names of foundation animals or importantpedigree figures. Numbers above solid circles are the herdnumbers of affected animals. Lines joining top or side of acircle or square represent descent from generation above. Linesleaving bottom of circle or square represent descent to generation below.

DISCUSSION

As in man, lymphosarcoma of cattle is characterized by localized neoplastic proliferation oflymphoid tissue which results in diffuse infiltrationof organs or formation of discrete solid tumormasses. These consist of a monotonous growth ofcells resembling either lymphocytes or lymphoblasts or, more often, both cell types in combination.

Lymphosarcoma appears to arise from lymphatic tissue or from tissues with lymphoid potential anywhere in the body. It may begin inone focus or in many foci simultaneously and thenspread rapidly by metastasis. At necropsy, dissemination is usually widespread, making it impossible to ascertain the primary site or sites oforigin.

A leukemic phase may be seen at some stage

in the disease. In such cases, all lymphoid organsand the bone marrow are massively involved. Thismust be differentiated from cases without massiveinvolvement, in which a few lymphosarcoma-likecells from affected lymph nodes and/or focal bonemarrow metastases appear in the blood. Sincemost leukemic animals are seen late in the disease,it is admittedly difficult to distinguish betweenthe leukemic phase of lymphosarcoma and primary

. lymphatic leukemia.

Lymphocytes in normal cattle, as comparedwith those in man and the dog, are more numerousin the peripheral blood and tend to show a greaterproportion of immature forms (Figs. 3, 4). Thus,the investigator who is unfamiliar with the bovinehemogram is apt to gain an erroneous impressionof abnormality when examining the blood of anormal cow. This fact assumes particular importance in the diagnosis of lymphosarcoma in theabsence of bone marrow or peripheral lymph nodeinvolvement. In such cases it is difficult to becertain that â€œunusualâ€• or â€œatypicalâ€• cells ohserved in peripheral blood are related to tumor,merely represent normal variation, or reflect thepresence of other disease.

The symptomatology of lymphosarcoma is essentially the same in both humans and cows. Themost common presenting complaint is the presenceof a massâ€”usually an enlarged lymph node. Allnodes and organs may be affected. Though thepicture is similar for humans and cattle, the veryhigh incidence of cardiac involvement and thelow incidence of cutaneous involvement in thebovine group, as compared with the human (33),are noteworthy.

In both human and bovine cases of lymphosarcoma and leukemia, the best diagnostic toolsare lymph node biopsy and bone marrow aspiration. Except for anemia of variable severity, theperipheral blood is usually normal in all respectsin lymphosarcoma until late in the disease. Occasionally, what appears to be a tumor cell may befound in the peripheral smear.

In the human disease, serum alkaline phospliatase and serum uric acid are frequently elevated,and the sedimentation rate is rapid. In cattle,serum uric acid, SGO-T, and serum LDH levelsmay be elevated, but other laboratory findingsare not remarkable or appear to be related to theanimal's terminal status.

Death from lyinphosarcoma in cattle and humans (32) usually results from tumor cachexia,infection, anemia, and mechanical encroachmenton vital organs. In humans, the first two are themost common causes, accounting for death in38.6 per cent and 35.8 per cent of cases, respective



@%IARSHAKet al.â€”Sludies on Bovine Lymphosarcoma 9215

ly. In cattle, cardiac failure is a major cause ofdeath.

Estimates of the number of cases of cancer inmen put leukemia and lymphoma in positions 10and 9, respectively, in order of incidence (9).In women, the incidence figures for leukemia andlymphoma are 13 and 10, respectively. It is estimated that there are 14,000 cases of lymphomaand 11,000 cases of leukemia in humans in theUnited States each year.

In children, lymphosarcoma is the most cornmonly encountered lymphoma, with Hodgkin'sdisease, reticulum-cell sarcoma, and giant follicularlymphoblastoma being relatively rare (71). Inadults, lymphosarcoma and Hodgkin's disease arethe two most common lymphomas and appearto affect about equal numbers of patients (48,71). The onset of lymphomas in humans has beenreported from infancy to 107 years.

In the bovine species there are no reliablestatistics on over-all incidence, but lymphosarcoma is thought to be the most common malignancy affecting cattle. Available statistics indicatethat the national rate in 1958 was approximately17.6 animals per 100,000 brought to slaughterat federally inspected abattoirs (59). It is questionable whether lymphomas other than lymphosarcoma exist in cattle (57).

Lymphosarcoma is principally a disease of mature cattle (Chart 1), but it has been reported incalves5 (8, 12, 41, 50, 61, 62) and even in fetuses(22, 27). Sex incidence is impossible to determine,

because few bulls are kept in dairy herds.In a masterly review, Karison (26) reported

that European workers had recognized the occurrence of lymphosarcoma in related animalsas early as 1897. In 1915 it appeared to attain

almost epizootic proportions in East Prussia, mostly affecting animals of the Schwartzbunten Niederungsvieh breed (28). Later, the appearance ofbovine lymphosarcoma in Silesia was attributedto introduction of these East Prussian cattle (56).Schaper (55), citing evidence to indicate the importance of hereditary factors, suggested in 1938that affected cattle of either sex, as well as theirparents and grandparents, should not be usedfor breeding purposes. The results of a â€œleukosisâ€•survey of Mecklenburg cattle in 1941, indicatinga higher incidence in certain large herds, wasattributed to widespread use of particular bullswithin these herds (31). In an extraordinary account published in 1944, Weischer (67) describedan outbreak of â€œcattle leukosisâ€• in which 149out of 150 daughters of an affected bull succumbedto the disease.

In the United States, Starr and Young (60)

reported a case in which a normal Holstein cowhad two daughters, a granddaughter, and a grandson with lymphosarcoma. They also described aGuernsey family in which lymphosarcoma appeared in four successive female generations.

Karlson (26) reported on a milking Shorthornherd, numbering 25 animals, in which there werefive cases within a 6-month period. He states onlythat two affected cows were half-sisters, but pedigrees in such herds usually indicate that all animals trace more or less directly to one or morefoundation ancestors. In a large herd of Anguscattle which had continually numbered over 100head during a 25-year period, Karlson reported onthe occurrence of lymphosarcoma in three highlyinbred individuals. Two of the affected animalswere full sisters; the third was a daughter of oneof the sisters. Finally, Karlson described a Jerseyherd in which six cases of lymphosa.rcoma occurredin one family during a 10-year period All affectedindividuals had more than one common ancestor;in most cases, common ancestors appeared morethan once in each pedigree. One unrelated cow inthis herd also succumbed to lymphosarcoma.

Numerous other investigators have suggestedthe importance of hereditary factors in bovinelymphosarcoma. An extraordinarily high mcidence of lymphosarcoma in cattle from Southeastem Sweden (5KB breed) and an almost negligibleincidence in ?@orthern Swedish cattle (5KB breed)provided an interesting experiment when attemptswere made to upgrade milk production of theNorthern cattle through importations of SRBbreeding stock in the late 1940's (70). During a5-year study of Northern herds in which lymphosarcoma was diagnosed, the majority of cases appeared among the imported Southern Swedishcattle or their offspring.

A recent view among European workers, the

Danes and Germans in particular, tends to discount the importance of hereditary factors andfavors a concept of lymphosarcorna as a slowlyadvancing infectious disease (3, 49). They preferthe term â€œleukosisenzootica boys,â€• which mdicates this concept. Nevertheless, it is generallyacknowledged that the spread of the disease parallels the exchange of breeding animals.

The familial aggregation of cases in the sixherds in our study is highly suggestive of geneticsusceptibility to lymphosarcoma. The data areequally compatible with a concept of verticaltransmission of an infectious agent.

To suggest that the familial aggregation ofcases in herd #4 could be a chance occurrence(P = 0.0000441) stretches credulity. The highest

probability for chance occurrence of lymphosar



Cancer Research Vol. 9292,February 196929216

coma in any of our herds was only 0.0283 (M Family, herd #1).

If genetic susceptibility were indeed a factor in

lymphoscarcoma of cattle, the mode of inheritancewould almost certainly be a recessive one. Probably multiple genetic factors would be involved.

One might postulate a dominant mode of inheritance of susceptibility in herd #2. However,the presence of unaffected daughters of matingswhich produced individuals in which lymphosarcoma developed weakens such a hypothesis.

Analysis of cattle pedigrees for mode of inheritance is difficult, since many â€œsusceptibleâ€•cowsmay leave the herd prior to the usual age of onsetof lymphosarcoma. There does seem to be a tendency for descendants of affected individuals to develop signs of lymphosarcoma at an earlier agethan their ancestors.

In conclusion, it seems evident that lymphosarcoma of cattle could provide a research tool of imrnense importance to oncologists. The establishedviral etiology of the avian leukosis complex andlymphocytic mouse leukemia (21) lends impetus tosimilar studies of lymphoid tumors in other spe

cies. Bovine lymphosarcoma is particularly interesting from the comparative standpoint, since itclosely resembles the disease in man. Parallelstudies on etiology, incidence (particularly as thisrelates to areas of high ambient radioactivity),geographical variation, epidemiology, and otherbasic and clinical aspects of lymphosarcoma inhumans and cattle should be profitable.

ACKNOWLEDGMENTSThe authors wishto expresstheirappreciation to Drs. W. D.

Malherbe, W. Medway, R. M. Dutcher, J. C. and P. T. Cardeilhac, J. Ipsen, J. L. Hyde, D. F. Patterson, J. W. Buchanan,J. T. McGrath, W. C. D. Hare, J. E. Prier, and A. M. Wallbank for their contributions to various aspects of the project;and to Mrs. F. Muldovan, Mrs. A. Williams, and Mr. IL Murray for their technical assistance.

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1. BARKER, S. B.; HUMPHREY,M. J.; and SOLEY,M. H. The@nicalDetermination of Protein-Bound Iodine. J. Clin.

Investigation, 30:55â€”62, 1951.

2. BENDIXEN,H. J. Untersuchungen Ilber die Rinderleukosein DÃ¤nemark. II. Deutsch. TierÃ¤rztl. Wchnschr., 67:4-7,1960.

S. . Untersuchungen tiber die Rinderleukose in Danemark. III. I&id., pp. 57â€”65.

4. BruSHY, 0. A.; LOWRY,0. H.; and BRocK, M. J. A Methodfor the Rapid Determination of Alkaline Phosphatase withFive Cubic Millimeters of Serum. J. Biol. Chem., 164:521-29,1946.

5. BLOOD, D. C., and HENDERSON,J. A. Veterinary Medicine,pp. 946â€”49.Baltimore: Williams & Wilkins Co., 1960.

6. Boyn, W. L. Clinical Observations of Lymphoid Hyperplastic Diseases of Cattle. Cornell Vet., 24:40â€”56, 1954.

7. Boyi, W. L.; KARLSON,A. G.; JASPER,D. E.; SELLERS,A. F.; and COLLIER,J. R. Leucemic Lymphoblastoma in aCow with Involvement of the Udder. Am. J. Vet. Research,8:550â€”57,1947.

8. Buss, W. Leukamie bei einem Kalb. TierÃ¤rztl. Rdsch., 44:755â€”56,1958.

9. CA3IERON,C. S. Cancer Statistics: Incidence, Mortalityand Results of Treatment. Med. Clin. N. Am., pp. 581â€”590,1956.

10. CLAUSSEN, L. Em Fall hochgradigen Milzinduration beichronischer aleukÃ¤mischer Lymphadenose des Rindes.Deutsch. tieritrztl. Wchnschr., 45:575â€”74, 1957.

11. COTCHIN,E. Neoplasms of the Domestic Animalâ€”AReview, pp. 25-Si. Farnham Royal, Bucks, England: Com

monwealth Agricultural Bureaux, 1956.12. CREEd, G. T., and HINSEL,E. Malignant Lymphocyto

ma in a Young Calf. Vet. Med., 37: 118â€”19,1942.is. DOBBERSTEIN,J. Betrachtungen Uberdie Lymphadenose

des Rindes (Rinderleukose). Deutsch. tierarztL Wchnschr.,42:289â€”95,1954.

14. ERRINGTON,B. J., and BARBER, C. W. Pseudoleucaemia(Lymphocytoma). Cornell Vet., 25:67-69, 1955.

15. FELDMAN, W. H. Neoplasms of Domesticated Animals,Mayo Clinic Monograph, pp. 202-20. Philadelphia: W. B.Saunders Co., 1952.

16. FISKE, S. H., and LOGAN,M. A. Determination of Calciumby Alkalimetric Titration. J. Biol. Chem., 93:211â€”26, 1931.

17. FISKE, S. H., and SUBBAROW, Y. The Colorimetrie Determination of Phosphorus. J. Biol. Chem., 66:575-400, 1925.

18. Foux, 0., and Wu, H. A Simplified and Improved Methodfor Determination of Sugar. J. Biol. Chem., 41:367â€”74,1920.

19. Fnai@ix,E. R., and THOMPSON,W. W. Lymphoblastoma inBovines. N. Am. Vet., 21:217â€”21,1940.

20. GoTz, R. ; ROSENBERGER, G. ; and ZIEGENHAOEN, G. DieLeukose des Rindes. Ihre hamatologische mid klinischeDiagnose. Mb. Vet. Med., 9:517â€”26,1954.

21. GRoss, L. Oncogenic Viruses. New York: Fergamon Press,1961.

22. HATzIor.os, B. C. Lymphoblastic Lymphoma in a BovineFetus. J. Am. V. M. A., 136:569â€”75, 1960.

Fia. 1.â€”Markedlyenlargedmandibularand atlanta! lymphnodes in cow with lymphosarcoma.

Fm. 2.â€”Bilateral exophthalmos in a young bull withlymphosarcoma. This resulted from pressure of rapidly growingtumor within the orbits.

Fia. 5.â€”Section of lymph node showing replacement ofnormal architecture except for a thin rim of cortical tissue.H.&E., X15.

FIG. 4.â€”Macroscopic appearance of 47-kg. abdominal massinvolving paraaortic and lumbar lymph nodes. The openedaorta passes through the center of the photograph.



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FIG. 5.â€”Blood film prepared from a normal mature Jersey

cow. The â€œunusualâ€•cell with a tri-lobed nucleus is not aneutrophil. Similar cells have been observed in the blood ofboth normal cattle and cattle with lymphosarcoma. The othercell is a typical bovine lymphocyte. Wright stain, X450.

FIG. 6.â€”Large, â€œunusualâ€•mononuclear cell from peripheralblood of a normal mature Jersey cow. It is characterized byabundant cytoplasm and a nucleus containing a loose, spongy,chromatin network. Wright stain, X45O.

FIG. 7.â€”Lymphocytes from leukemic blood of cow withlymphosarcoma. Morphologically, the cells appear normal.Wright stain, X450.

FIG. 8.â€”Lymphocyte from blood of cow with lymphosar

coma. This animal's white count was within normal limits.Similar morphologic types are frequently observed in bloodfilms from normal cattle. Wright stain, X450.

FIG. 9.â€”Leukemic peripheral blood from bull with lymphosarcoma. White count >200,000/cu mm; 98 per cent lymphocytes. Wright stain, X 100.

FIG. 10.â€”Same as Fig. 7. X225. Note bizarre nuclearforms. A mitotic figure may be seen in center.

FIG. 11.â€”Aspirated costal marrow from leukemic bull(see peripheral blood, Figs. 9, 10). On differential count over90 per cent of cells were very large with an abundant, finelygranular, darkly staining cytoplasm. Bizarre nuclear formspredominate. Nuclei contain a pale staining, finely reticularchromatin. Wright stain, X450.

FIG. 12.â€”Blood film from cow with lymphosarcoma. Anerythroblast lies between two normal-appearing lymphocytes.Cell at lower left is a neutrophil. Total white and differentialcounts were within normal limits. Wright stain, X450.



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FIG. 15.â€”Polypoid mass of lymphosarcoma in the rightatrium. Note diffuse infiltration of atrial wall.

FIG. 14.â€”Wall of abomasum showing almost completereplacement of normal structures by tumor tissue. The mucosais intact.

F:o. 15.â€”Section of wall of abomasum showing replacementof normal structures. Note intact mucosa. H. & E., X50.

FIG. 16.â€”Ureter completely surrounded by lymphosarcomatous mass.



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Fia. 17.â€”Diffuse infiltration of the kidney in lymphosarcoma. Kidney is about twice usual size.

FIG. 18.â€”Close-up view of kidney lobule inlymphosarcoma.Normal corticalstriations are blurred by the diffuse infiltration.

FIG. 19.â€”Diffuse annular infiltration of the wall of theuterus.

FIG. 20.â€”Diffuse sheet of lymphocytes invading the wallof the uterus. Note that infiltration does not extend into thearea of placental attachment. H. & E., XiS.



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FIG. 21.â€”Perilobular infiltration of liver. H. & E., XiS.FIG. 22.â€”Tumor surrounding lumbar spinal nerve root.FIG. 25.â€”Lymphosarcomatous infiltration of epidural fat

at lumbar spinal nerve root. H. & E., XiS.Fxo. 24.â€”Envelopmentof spinal cord by lymphosarcoma

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FIG. 25.â€”Tumor embolus in artery. Giemsa, X280.Fia. 26.â€”Sheetof moderately well differentiated lympho

cytes and lymphoblasts in lymphosarcoma. Giemsa, X500.Fio. 27.â€”Multinucleated giant@cell@in@lymphosarcoma.

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MARSHAK e@al.â€”Studies on Bovine Lymphosarcoma 9217

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1962;22:202-217. Cancer Res R. R. Marshak, L. L. Coriell, W. C. Lawrence, et al. Pathological Alterations, and Herd StudiesStudies on Bovine Lymphosarcoma: I. Clinical Aspects,

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