[CANCER RESEARCH 39, 833-839, March 1979]0008-5472/79/0039@0000$02.00

Incidence and Pathological Features of Spontaneous Tumors inAthymic Nude Mice1

Francis E. Sharkey2 and Jorgen Fogh

Departmentof Pathology,Milton S. HersheyMedicalCenterof ThePennsylvaniaStateuniversity,Hershey,Pennsylvania17033(F. E. S.), and the HumanTumorCellLaboratory,Sloan-KetteringInstitutefor CancerResearch,Rye,NewYork10580(J. F.J

ABSTRACT

Systematic observation of 1141 nude mice (Swiss background strain) that received human tumor transplants mevealed 24 spontaneous tumors, 18 of lymphometiculamoriginand 6 pulmonary adenomas. Spontaneous tumors wereseen at an average age of 9.1 months, and 22 of the tumorswere seen only in that fraction of our group (324 mice)surviving for 5 months or more (22 of 324 x 100 = 6.8%).Transplantation of these tumors to other nude mice wassuccessful in three of five cases. Mice transplanted withadenocarcinoma of the colon and with tumors of the umogenital tract developed spontaneous tumors more oftenthan did mice receiving other types of human tumor transplants. Progressive growth of the human tumor transplantoccurred significantly less often in the mice that eventuallydeveloped spontaneous tumors than in the mice thatshowed no spontaneous tumor development. Nevertheless,the incidence of spontaneous tumors in these nude micewas similar to that reported for the thymus-beaning background strain.

INTRODUCTION

Early observers of athymic nude mice (4, 10) believed thatthese animals did not develop spontaneous tumors (6, 11).With the development of germ-free colonies, however,spontaneous tumomigenesis was indeed observed (9), usually at a rate and in a proportion similar to the backgroundstrain (15). Stutman (15) suggested that the chief limitationof those early observations was the brief life span of nudemice under conventional experimental conditions (8).

Although tumors originating in the lymphoreticular systern have been the most common type reported in nudemice, descriptions of their histopathological features havenot been available. We now report the results of ourobservations on over 1100 nude mice, 324 of which livedfor 5 months or more, detailing the appearance, chassification, and transplantability of spontaneously arising tumorsin these animals.

MATERIALS AND METHODS

NudeMice. Allmicedescribedinthisreportwereusedina project which sought to define the spectrum and growth

I Supported in part by USPHS Grant CA08748 from the National Cancer

Institute and USPHS Contract N01-CB43854 from the Division of CancerBiology and Diagnosis, National Cancer Institute. Presented in part (5) at theSixty-ninth Annual Meeting of the American Association for Cancer Research, Washington, D. C., April 1978.

2 To whom requests for reprints should be addressed.

Received August 29, 1978; accepted December 1, 1978.

characteristics of human tumors which can be grown innude mice (13). Male and female nude mice of Rex/TrembIer origin, backcrossed to outbmed Swiss 2 times, wereobtained from the breeding colony at Sloan-Kettering Institute (New York, N. Y.). Heterozygotic females were matedto homozygotic males in a scheme that favored outbreeding.3 At approximately 6 weeks of age, these mice wereinoculated s.c. with human tumors obtained directly fromsurgical specimens in the Memorial Hospital Department ofPathology. Tumors were obtained at the time of frozensection diagnosis, minced in a few drops of sterile, isotonicphosphate-buffered saline (pH 7.2), and injected via a 13-gauge trocam into the s.c. tissues of the flank. Recipientmice were housed in New York in a closed colony withautochaved cage materials and were covered with an autochavedfiberglass filter bonnet.

Tumor Transplantation Procedures. Mice were observedbiweekly and were allowed to survive until the tumor transplants developed progressive growth or until the micebecame terminally ill. Sick mice and mice bearing tumorsthat had reached a size of approximately 1 cu cm, weresystematically culled from the colony and sacrificed bycervical dislocation. Growing tumors, including some ofthe spontaneously developing tumors, were minced andreinoculated s.c. into fresh groups of 6-week-old nudemice, thus maintaining the tumors in continuous passage.Tumors were also frozen in liquid nitrogen and their viabilitywas tested by reinoculation into nude mice, some of theselatter being mice in which a previous tumor transplanttaken from surgical specimens had failed to grow.

Postmortem Examination of Experimental Mice. A cornplete external and internal gross examination was performed on 1141 (90%) of the 1264 mice observed , with theremainder being spontaneous deaths that were unsuitablefor postmortem examination due to severe decay. Theselatter have been eliminated from all subsequent calculations. Microscopic examination was performed on 74% ofall grossly examined mice, including all well-preservedspontaneously dead mice, all sick-looking mice, and mosthealthy-looking mice. Microscopic examination routinelyincluded samples of tumor, lungs, liver, spleen, pancreas,kidneys, and axillary lymph nodes. In cases of grosslyrecognized spontaneous hymphoreticulam tumors and alltheir passages, microscopic examination was also performed on multiple lymph nodes, bone marrow, brain,bowel, genital tract, and soft tissue tumor masses. Periphemalblood smears, air dried and stained with Wright's stain,were alsoprepared.

Tissues were fixed in 10% neutral buffered fommalin and

3 A. Csurny, personal communication.

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Nudemousemortality°Sacrificed for tumorSpontaneoustumorsNo.observed%

ofeachCumulativeMos.No.eachCumulativeNo. each monthlymonthlyincidencesurvivalmo.%mo.

mortalityinterval(%)134318

5323833736295331364289

92284877276875837960

72425661844167129748882758342826911350250935947202581021966

29111197655371122098945175131099330492143991

33152991961619917199189919992021001

100Totals1141918

8024mice

whowere unsuitable for gross examination due to

F. E. Sharkey and J. Fogh

processed for paraffin embedding and sectioning in theusual way. Sections of all blocks were stained with hematoxyhin and eosin, and lymphoreticular tumors were furtherstudied with methyl green-pyronine and chloroacetate-estemase stains. Other special stains (periodic acid-Schiff,mucin, reticuhin, Masson's tmichrome, Giemsa, and PenIs'

iron)were appliedasneeded.Survival of mice was calculated in months, with a month

being included only after it had completely passed. Birthdate was assumed to be 6 weeks prior to tumor inoculation.

Lymphometicular tumors were classified according to therecommendations of Dunn (1).

RESULTS

Nude Mouse Morbidity and Mortality. A total of 1141mice were autopsied during a continuous period of 26months (Table 1). Median survival was 3 months, and 72%survived less than 5 months, with most of these mice (91%)being terminated for removal of growing tumor transplants.The percentage of terminated mice bearing successfultumor transplants decreased progressively at later ages,although an increased incidence of tumor growth wasobserved at about 12 months of age because of the reuse ofmice, in which the initial tumor transplant had failed, to testthe viability of human tumors that had been frozen in liquidnitrogen.

Of those mice showing no evidence of transplant tumorgrowth, the median survival was 6 months. Late mortalitywas due mostly to mouse hepatitis virus and Sendai virusinfections (12).

Spontaneous Tumor Incidence. Twenty-four sponta

neous tumors were identified, mostly at initial gross examination (Table 2). Affected mice were killed at a mean age of9.1 months. The youngest was diagnosed at 3 months ofage, but 22 of the 24 (92%) were seen at ages of 5 monthsor more. Thus, spontaneous hymphoreticulamtumors developed in 2.1% (24 of 1141) of all evaluable mice and in 6.8%(22 of 324) of those surviving for 5 months or more. The sexincidence of the affected mice was nearly equal and similarto that of the entire colony.

Of those mice developing spontaneous tumors, 13 wereinoculated with human tumor transplants derived directlyfrom surgical specimens, and 6 were inoculated with subpassages of human tumor transplants (Table 2, originalhuman tumor type). In the other 5 cases, mice were inocuhated with cryopreserved human tumors (Table 2, survivaltumor type); 3 of these latter (Table 2, Cases 22 to 24) hadpreviously received a human tumor transplant direct fromsurgicalspecimensthathad failedtogrow.

The distribution of human tumor types with which themice developing spontaneous tumors were inoculated (Table 2) was significantly different from that observed in the324 mice that survived for 5 months on more (Table 3)@analysis, 0.05 > p > 0.02). (Since 92% of the spontaneoustumors developed in mice surviving for 5 months or more,these mice were considered for analytical punposes to bethe group ‘‘atrisk.―x2 analysisdetected no significantincrease in the incidence of spontaneous tumors in micereceiving transplants of 2 different tumors, and they weretherefore counted twice in Table 3, as if each different typehad contributed independently to the development of spontaneous tumors.) Among the individual tumor types, x2analysis revealed significant ovemmepresentationof colonic

Table1Nudemousemortality and spontaneoustumor development

a Does not include 123

postmortem decay.

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Table2Characteristicsof nude mice developing spontaneoustumorsMice

were inoculated at 6 weeksof agewith humantumors previouslygrown innudemiceor taken directly from surgical specimens (‘‘originalhuman tumor type'‘)and/orinoculated

at a later age with nude mouse-grown human tumors that had beenpreservedinliquid nitrogen (“survivaltumor type―).Mice were observed biweekly andsacrificedwhen

progressivetumor growth developedor when mice appearedill.Humantumor type

Age SpontaneoustumorCase (mos.) Sex Original (1) or survival (2)type1

3 M 1. Duct carcinoma of breast, primary Pulmonaryadenoma23 F 2. Adenocarcinoma of colon, meta-Lymphosancomastatic3

5 M 1. Carcinoma of bladder, metastaticLeukemia45 M 1 . Adenocarcinoma of ovary, meta-Leukemiastatic5

5 F 1. Ewing's sarcoma of bone, meta-Lymphosarcomastatic6

5 F 1. Osteogenicsarcoma, recurrent RCS,typeA@'76 F 1. Thymoma, primary RCS, typeA87 M 1 . Renal cell carcinoma, primary Pulmonaryadenoma97 M 1. Adenocarcinomaof lung, primary RCS,typeA107 F 1. Adenocancinoma of colon, meta- Pulmonaryadenomastatic1

1 8 F 1. Renal cell carcinoma, metastatic RCS, type A12 9 F 1. Malignant melanoma, primary RCS, typeB―13

9 M 1. Adenocarcinoma of breast, meta- RCS, typeAstatic14

9 M 1. Adenocarcinomaof ovary, primary RCS,typeA1511 F 1. Adenocancinoma of stomach, pni- Plasma celltumormary16

11 M 1. Adenocarcinomaof colon, primary Pulmonaryadenoma1711 M 1. Adenocarcinoma of lung, primary Pulmonaryadenoma1812 M 1 . Adenocancinoma of endocervix, Pulmonaryadenomaprimary19

13 F 1. Carcinomaof bladder, metastatic RCS,typeB2013 M 2. Adenocarcinoma of colon, meta- RCS,typeBstatic21

13 M 1. Malignant teratoma, metastaticLeukemia2213 F 1. Adenocancinoma of breast, pni-Lymphosancomamary2.

Malignant melanoma,metastatic2315 F 1. Embryonalcarcinoma, metastatic RCS,typeB2.

Adenocarcinoma of colon, metastatic24

20 M 1. Spindle cell sarcoma of forearm,Leukemiaprimary2.

Oat cell carcinoma of lung, metastatic

Spontaneous Tumors in Athymic Nude Mice

a Reticulum cell neoplasm, type A.

b Reticulum cell neoplasm, type B.

adenocarcinomas and of tumors from the urinary andgenital tracts (0.05 > p > 0.02 for each type). By x2 analysis,no significant underrepresentation of any tumor type wasseen, nor was the inoculation of cryopreserved humantumors significantly associated with increased developmentof spontaneous tumors.

In only 2 instances was growth of the human tumortransplant observed in a mouse that developed a spontaneous tumor (Table 2, Cases 2 and 20). Both of those micereceived transplants of the same cryopreserved humantumor but developed histologically different lymphoreticulam human tumors at different ages. In considering thegroup of 324 mice that survived for 5 months or more (Table1), spontaneous tumor development was seen in only oneof 133 mice that showed growth of the human tumortransplant, whereas the remaining 21 spontaneous tumorsthat developed in this age group occurred among the 191mice that showed no growth of the human tumor transplant

at all. By x2 analysis, this increased incidence of spontaneous tumors among mice that yielded no growth of thehuman tumor transplant was highly significant, 0.01 > p >0.001.

In no case did a spontaneous tumor develop at the site(flank) of human tumor inoculation, although the dorsalmidline growth of an early case of reticulum cell neoplasm,type A, was initially confused for growth of the humantumor transplant, and the hepatic involvement was grosslythought to be metastasis of the transplant. Microscopicstudy, however, revealed the correct diagnosis, and in nocase did the spontaneous tumor beam any histologicalresemblance to the human tumor transplant.

Other lesions seen in these animals were hepatitis in 19cases, and one case each of Sendai virus pneumonia andPneumocystis pneumonia. These pathogens were cornmonly seen in older mice in our colony (12).

Gross and MicroscopicFeatures of SpontaneousTu

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Numberof each human tumor type transplantedinto allnudemice,into nude mice surviving 5 months or more, and intonudemice

that developed spontaneous tumorsSpontaneoustumor

type(no.)No.

ofmiceNo. of Pul

Total no. surviv- sponta- Lym- monof tumor ing for 5 neous phone-anytrans-

mos. or tu- ticular adeTumor type plantsa mon&' mons@' tumorsnomaLung

carcinoma 218 67 3 21Breastcarcinoma 161 70 3 21Coloncarcinoma 115 24 5C 32Urinarytract

96 20 4C3carcinomaMalignant

79 11 22melanomaSofttissue

75 19 11tumorsBonetumors

67 26 22Genitaltract 54 28 5C 41carcinomaLymphoneticular

37 19 11Gastrointestinal34 12 11tract

carcinoma(noncolon)Other

22447Totals

1160a 34341 27― 21 6

TotalTissues

involvedRetropenito

Blood andSubpasno. ofLymph nodesneum andbonesage(caseTumor

typecasesand spleenmediastinumLivermarrowOther (seetext)no.Y'Reticulumcell6—++—Local6,7neoplasm,(nodular)invasiontype

AReticulum cell4++ in1+—Lung,NA―neoplasm,case

only(nodular)kidneytypeBStemcell4++++Kidney,3leukemia(diffuse)lung,

leptomeningesLymphosar

3+++BoneLeptoNAcoma(nodular)marrow

onlymeninges,ovaryPlasmacell1Mesentenic———NAtumornodes

only

F. E. Sharkey and J. Fogh

mors. Eighteen of the spontaneous tumors that developedin this experimental group were of the lymphoma-leukemiavariety. All were initially identified at the time of grossexamination. Their classification, gross features, and histological features are outlined in Table 4.

Table 3

The most common tumor, as seen in 6 cases, histologically corresponded to Dunn's reticulum cell neoplasm, typeA (1). Tumor was noted on external examination as a flats.c. mass on the dorsal thorax, and internal examinationrevealed confluent mediastinal and retropenitoneal involvement. The liver was grossly enlarged by pale, fleshy tumornodules. No ovarian, splenic, or lymph nodal involvementwas seen, although occasional involvement of other organsresulted from direct extension of retropenitoneal tumor.Histologically, the tumor was composed of large, undifferentiated, moderately pleomomphic cells having cleaved nuclei and prominent nucleoli. Methyl green-pymonine stainshowed cytoplasmic pymoninophihia. Prominent tumor cellnecrosis was present, giving a ‘‘starry-sky―appearance onlow-powerview (Fig. 1).

Four mice developed reticulum cell neoplasm, type B (1).All 3 showed peripheral and internal lymphadenopathy, aswell as hepatomegahy, and multiple 1- to 2-mm pale nodulesin an enlarged spleen. Only one had confluent retroperitoneal tumor. Microscopic examination revealed a pleomorphic cellular infiltrate that consisted of a mixture ofatypical reticulum-like cells, lymphocytes, occasionalplasma cells, and rare granulocytes. The reticulum-likecells had large, irregular nuclei with large nucleoli, andoccasionally resembled Reed-Stemnberg cells (Fig. 2). Twoof the cases also had multinucleated giant cells in thetumor. Methyl green-pyronine showed cytoplasmic pyroninophihia in the atypical reticulum cells. Hepatic infiltrateswere periportal, and the splenic infiltrates characteristicallysurrounded the arterioles, often with a mass of matureplasma cells in the center. The pulmonary infiltrates weretypically penivascular, and quite massive in 2 of the cases(Fig. 3). No tumor cells were seen in peripheral blood orbone marrow.

The 4 cases of stem cell leukemia (1) demonstratedmarked axillary, inguinal, and cervical lymphadenopathy,as well as confluent involvement of the retropemitoneum

Table4

a Mice inoculated with 2 different tumor types (n 19) arecounted once under each tumor type.

b Tumors arising in recipients of 2 different tumor types (Table2, Cases 22 to 24) are counted once under each tumor type.

C Significant ovennepresentation, 0.02 > p > 0.01.

Characteristics of spontaneous lymphoreticular tumors in nude mice

Tumorswere classified according to the methodof Dunn (1).

a Refers to case numbers in Table 2.

b NA, not attempted.

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SpontaneousTumorsinAthymicNude Mice

and posterior mediastinum . Occasionally, anterior mediastinal tumor was noted, but thymic involvement was notpossible due to the congenital absence of that organ (10).The spleen showed replacement of the usual penivasculamlymphoid tissue by tumor. The bone marrow was almostcompletely replaced by tumor cells, and extensive infiltrateswere seen in liver, kidney, and leptomeninges (Figs. 4 and5). Histological sections and peripheral blood smearsshowed primitive, undifferentiated mononuclear cells.Methyl green-pyronine was markedly positive for cytoplasmic pyroninophilia.

The 3 cases of hymphosarcoma (1) showed peripheral andinternal lymphadenopathy, nodular tumor infiltrates in theliver, and massive enlargement and replacement of thespleen. Microscopically, tumor was also seen in the leptomeninges and bone marrow but not in the peripheral blood.The tumor consisted of monotonous sheets of large, poorlydifferentiated lymphocytes, some having cleaved nuclei.Methyl green-pyronine showed focal cytoplasmic pyroninophilia.

A single case of plasma cell neoplasm (1), primary insmall bowel, also showed involvement of local peritoneumand of rnesentenic lymph nodes (Fig. 6). Small bowel obstruction by the tumor produced severe nutritional debihitation in the affected animal. Small bowel mucosa adjacentto the tumor displayed atrophy and marked plasma cellinfiltrate, and increased numbers of plasma cells were alsonoted in the spleen and peripheral lymph nodes.

Six pulmonary adenomas were identified, one by grossexamination of the lung and the other 5 only after microscopic examination. All were solitary and ranged in sizefrom 0.5 to 2 mm. All consisted of an expansile growth ofwell-differentiated epithelium that resembled bronchial hining cells. None was large enough to cause any apparentharm to the affected animal.

Passage of Spontaneous Lymphoreticular Tumors.Transmission of spontaneous tumors in nude mice wassuccessful in 2 of 3 attempts of type A reticulum cellneoplasm and in the only case attempted of stem cellleukemia, but was unsuccessful in one case of type Breticulum cell neoplasm (Table 4). Passaged tumor grewrapidly in the new host, reaching a diameter of 1 cu cm in 3to 4 weeks. In each case, tumor in the recipient nude micefaithfully recapitulated the topological and histological features found in the mouse of origin, with the addition oflocal tumor growth at the site of s.c. tumor inoculation.

The 2 successfully passaged cases of reticulum cellneoplasm, type A (Table 2, Cases 6 and 7) were maintainedin passage for 6 and 3 generations, respectively, and case 3(stem cell leukemia) was maintained in passage for 5generations. In only one case (Case 7) did the tumor die outin passage; the other 2 tumors were terminated voluntarily.

DISCUSSION

The spontaneous tumors identified in this group of nudemice all developed in a colony that was used experimentallyfor the development of nude mouse-grown lines of mahignant human tumors. As such, these observations cannotprovide a definitive answer as to the true incidence ofspontaneous tumor development in these immune-deficient

animals. This conclusion is reinforced by the followingobservations: (a) the spontaneous tumors tended to appearin nude mice in which the tumor transplant had failed togrow; (b) development of spontaneous tumors appeared tooccur in association with exposure of the mice to certainhuman tumor types (Table 3). However, the apparent association of spontaneous tumors with certain human tumorsshould be interpreted with caution, since such groups as“genitaltract tumors―actually include a wide variety ofhistologically and clinically separate tumors. Breakdown ofthese convenience groups into more homogeneous subgroups, however, resulted in aggregate numbers that weretoo small for analysis.

Sampling technique may well have underestimated thetrue incidence of pulmonary adenomas. Since most werenot seen by gross examination, several may have beenmissed in the 26% of the mice not examined microscopically. We also made no effort to section the lungs seriallyfor microscopic study, thus potentially missing some microscopic examples. However, we do not feel that a significantnumber of lymphoreticulam tumors were missed, since almost all were first noted upon external examination of theaffected animals.

Despite all these caveats, our observed incidence ofspontaneous tumor development and the types of tumorsseen were quite similar to that reported for the thyrnusbearing background strain (Swiss) thatwe used. Pulmonaryadenomas and lymphoreticulan tumors are the most cornmon benign and malignant tumors, respectively, reportedin this strain (14), and the report by Toth et a!. (16) of a 6.5%incidence of malignant tumors occurring by 17 months inSwiss mice is not unlike our observed incidence of 6.8% (22of 324) in those of our group that survived for 5 months ormore. Stutman (15) has also reported that the incidence ofspontaneous and induced tumors in nude mice of bothBALB/c and CBA/H backgrounds was the same as that ofthe thyrnus-bearing background strain. Thus, althoughsome of our data suggest a relationship between exposureto malignant human tumors and the development of spontaneous tumors, the effect would seem to have been small.

The development of spontaneous tumors in nude micethat had received transplants of certain tumor types did notappear to be explicable on the basis of variations in expenimental technique. In particular, use of cryopreserved tumortransplants, as opposed to transplants of fresh tumors, didnot significantly affect the incidence of spontaneous tumoms.

The spontaneously developing tumors were clearly ofmunine origin, bearing no histological resemblance to theoriginal human tumor transplants. Successful transplantation of some of the lyrnphoreticular tumors to other nudemice, with maintenance of their essential pathological featunes, would seem to confirm their neoplastic origin (2, 3,7). The tumors also showed pathogenic effects that weretotally unlike that of human tumor transplants, which rarelymetastasize (13).

Our observations confirm that the vast majority of thesetumors develop at ages well in excess of the mean age (1 to2 months) of conventionally maintained nudes (8). Similarly,Outzen et a!. (9) reported development of 22 lymphoneticulamcancers in a group of 261 male and female germ-free

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F. E. Sharkey and J. Fogh

nudes at a mean age of 13.1 months. This age factorprobably explains the discrepancy of our observations withthose of Rygaamdand Poylsen (11), who reported completeabsence of spontaneous tumors in 2 large groups of nudemice having average life spans of only 4 and 3 months,respectively.

In summary, a systematic study of a large number ofathymic nude mice has revealed a number of spontaneoustumors. These occurred mostly in mice that had survived 5months or more, and the tumors were readily classifiedaccording to standard histological descriptions. Tumorscould be passed into other nude mice and showed pathological effects characteristic of malignant tumors. Althoughexposure of the mice to human tumor transplants may havehad some effect on spontaneous tumor development, theincidence, type, and age of onset of spontaneously arisingtumors in our nude mice seemed similar to that reported byother authors for the thymus-beaning background strain.Routine histological survey of all nude mouse-grown tumorlines would seem to be indicated in order to avoid contamination by spontaneous muminetumors.

ACKNOWLEDGMENTS

The authors wish to express their appreciation forthe technical assistanceof Marcia Bains-Grebner, Thomas Orfeo, Kay Y. Tai, and John A. Tiso.

REFERENCES

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2. Dunn,T. B. Morphologyandpathologyof reticularneoplasmsin themouse-relationship to man. Ann. N. Y. Acad. Sd. , 76: 619—628,1958.

3. Evans, C. H., and Dipaolo, J. A. Comparison of nude mice with the host

species for evaluation of the tumorigenicity of guinea pig and hamstercells transformed in vitro by chemical carcinogens. Cancer Res., 36:128—131,1976.

4. Flanagan, S. P. “Nude―, a new hairless gene with pleiotropic effects inthe mouse. Genet. Res. (Camb.), 8: 295-309, 1966.

5. Fogh, J., Filippa, D., and Sharkey, F. E. Spontaneous tumors and humantumor metastases in nude mice. Proc. Am. Assoc. Cancer Res., 19: 211,1978.

6. Gershwin, M. E., Merchant, B., Gelfand, M. C., Vickers, J., Steinberg, A.D., and Hansen, C. T. The natural history and immunopathology ofoutbred athymic (nude) mice. Clin. Immunol. Immunopathol., 4: 324—340, 1975.

7. Giovanella, B. C., Stehlin, J. S., and Williams, L. J., Jr. Heterotransplantation of human malignant tumors in “nude―thymusless mice. II.Malignant tumors induced by injection of cell cultures derived fromhumansolid tumors.J. NatI.CancerInst.,52:921-930,1974.

8. Gullino, P. M., Ediger, R. D., Giovanella, B., Merchant, B., Outzen, H.C., Jr., Reed, N. D., and Wortis, H. H. Guide for the care and use of thenude (thymus-deficient) mouse in biomedical research. ILAR News, 19:MI-20, 1976.

9. Outzen, H. C., Custer, A. P., Eaton, G. J., and Prehn, R. T. Spontaneousand induced tumor incidence in germ-free “nude―mice. J. Reticuloendothel. Soc., 17: 1-9, 1975.

10. Pantelounis, E. M. Absence of thymus in a mouse mutant. Nature•(Lond.),217: 370—371,1968.

11. Rygaard, J., and Povlsen, C. 0. The mouse mutant nude does notdevelop spontaneous tumors. Acta Pathol. Microbiol. Scand. (B), 82:99-106, 1974.

12. Sharkey, F. E. Histopathologic observations on a nude mouse colony.In: J. FoghandB.C.Giovanella(eds.),TheNudeMousein Experimentaland Clinical Research, pp. 75—93.New York: Academic Press, Inc., 1978.

13. Sharkey, F. E., Fogh, J. M., Hajdu, S. I., Fitzgerald, P. J., and Fogh, J.Experience in surgical pathology with human tumor growth in the nudemouse. In: J. Fogh and B. C. Giovanella (eds.), The Nude Mouse inExperimental and Clinical Research, pp. 187-214. New York: AcademicPress, Inc., 1978.

14. Sher, S. P. Tumors in control mice—literature tabulation. Toxicol. AppI.Pharmacol.,30: 337-359,1974.

15. Stutman, 0. Spontaneous, viral and chemically induced tumors in thenude mouse. In: J. Fogh and B. C. Giovanella (eds.), The Nude Mouse inExperimental and Clinical Research, pp. 411-435. New York: AcademicPress, Inc., 1978.

16. Toth, B., Nagel, D., Erickson, J., and Kuppen, R. Tumorigenicity oftetramethylhydrazine in Swiss mice. J. NatI. Cancer Inst., 57: 1179—1183,1976.

Fig. 1. Reticulum cell neoplasm, type A. ‘Starry-sky―appearance. H & E, x 100.Fig. 2. Reticulum cell neoplasm, type B. Pleomorphic infiltrate, Reed-Sternberg-like cell (arrow). H & E, x 480.Fig. 3. Reticulum cell neoplasm, type B. Penivascular infiltrate in lung. H & E, x 200.Fig. 4. Stem cell leukemia. Brain with meningeal infiltrate. H & E, x 300.Fig. 5. Stem cell leukemia. Peniportal infiltrate in liver. H & E, x 100.Fig. 6. Plasma cell neoplasm, replacing small bowel mucosa. H & E, x 200.

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on March 15, 2020. © 1979 American Association for Cancer Research.cancerres.aacrjournals.org Downloaded from

1979;39:833-839. Cancer Res   Francis E. Sharkey and Jørgen Fogh  Athymic Nude MiceIncidence and Pathological Features of Spontaneous Tumors in

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