[CANCER RESEARCH 39, 811-816, March 1979]0008-5472/79/0039-0000$02.00

Enhancement of Gastric Carcinogenesis in Dogs Given N-Methyl-N'-nitroN-nitrosoguanidine following Vagotomy'

Masahide Fujita,2 Motohisa Takami, Masao Usugane, Shigeru Nampei, and Tetsuo Taguchi

Department of Oncologic Surgery, Research Institute for Microbial Diseases, Osaka University, Yamada-kami, Suita, Osaka 565, Japan

ABSTRACT

In an attempt to improve the method of induction ofgastric cancer in dogs, selective vagotomy without drainageprocedure was performed before administration of N-methyl-N ‘-nitmo-N-nitrosoguanidine(MNNG). Twenty dogswere divided into two groups of 10 dogs (five beagles andfive mongrels). The control group received MNNG alone.Dogs of the experimental group were vagotomized 2months before administration of MNNG. MNNG solution (50p.g/ml) was administered in the drinking water for i2months. In the vagotomized group, gastric adenocarcinomas were induced in all of eight effective cases, whereas inthe control group they were seen in six of eight cases.Gastric cancer was found to be multiple in many dogs, butthe majority of cancer lesions remained in the early stage.The total numbers of cancer lesions induced in the vagotomy and control group were 42 and 16, respectively. Vagotomy resulted in the production of more advanced cancers.One lesion in the control group and four lesions in thevagotomy group were advanced cancer invading the serosa. Advanced gastric cancers with metastases to the liver,lymph nodes, and lungs developed exclusively in two vagotomized dogs. Gastric acid secretion of the vagotomizeddogs measured after termination of MNNG was reduced, onthe average, to one-fourth of that of the control. Thepossible moleof vagotomy in the enhancement of gastriccarcinogenesis in dogs by MNNG is discussed in terms ofthe inactivation of MNNG in gastric juice and ulcerogenicaction of the carcinogen to the gastric mucosa.

INTRODUCTION

The experimental induction of gastric cancer in dogspermits one to investigate the developmental process ofgastric cancer by serial radiographic and endoscopic examination without sacrificing the animal.

A high incidence of adenocarcinomas in the stomach hasbeen demonstrated in rats (15) and dogs (2, 4, 17) followingP.O. administration of MNNG.3 However, dogs given MNNG

in their drinking water developed leiomyosarcomas in thesmall intestine (9) while the gastric cancers remained in theearly stage without metastasis at the time when the animalswere sacrificed because of the intestinal sarcomas (4, 14,17). In view of this difficulty, attempts have been made toimprove the methods for inducing more advanced carcino

I Supported in part by a grant-in-aid for Cancer Research from the

Ministry of Health and welfare, Japan.2 To whom requests for reprints should be addressed.

3 The abbreviations used are: MNNG, N-methyl-N'-nitro-N-nitrosoguani

dine; MAO, maximal acid output.Received June 27, 1978; accepted November 22, 1978.

mas with metastases (3, 10), such as the use of vagotomy toenhance gastriccarcinogenesisinmats(8,11).

The present study is designed to evaluate whether selective vagotomy performed before p .o. administration ofMNNG will enhance the production of gastric cancers indogs.

MATERIALS AND METHODS

Animals and Operation. Twenty dogs weighing 6 to 11kg were used in this experiment. Ten male beagle dogs and10 mongrel dogs (7 males and 3 females), 10 months old,were divided into 2 groups, each consisting of 5 beaglesand 5 mongrels. The animals were housed in individualcages and given about 300 g of dog chow (No. 1; CLEA,Japan) every morning except Sunday. In one group, selective vagotomy was performed under i.v. anesthesia withsodium pentobambiturate. After an upper median incision,the vagus nerves in the lesser omentum from the esophagogastric junction to the antrum were cut. The nerves aroundthe esophagus and to the extragastnic organs were preserved (13). No gastric drainage operation was carried out.No operative procedure was done in the other group, whichservedascontrols.

Carcinogen. At 12 months of age, all the dogs were givenan aqueous solution of MNNG (50 @g/ml;Aldrich ChemicalCo., Milwaukee, Wis.) ad libitum for 12 months and thenreturned to tap water. Stock solution of MNNG dissolved indeionized water at a concentration of 1 mg/mI was dilutedwith tap water just before use. The amount of MNNGsolution consumed was determined twice a week.

Morphological Observations. Routine endoscopic examination was performed under i.v. anesthesia every 3months. The time the first tumor was observed in each dogwas recorded.

All animals were autopsied, and entire organs weregrossly examined when the animals died or were killedwhen moribund. The vagus nerves in the experimentalgroup were examined to ascertain that vagotomy was doneproperly. The stomach was opened along the greater curvature and was step sectioned into 5-mm widths for therecognition of geographic relationship of each lesion (Fig.1). Other organs which showed grossly abnormal findings

were also sectioned. Sections were routinely stained withhematoxylin and eosin. The Alcian blue-periodic acid-Schiffmethod was also used to stain selected sections.

Assay of Gastric Acid Secretion. The estimation of gastmic acid secretion was performed 3 to 6 months after thetermination of MNNG administration. After the dogs had

been fasted for 36 hm,stomach tubes were inserted intotheir stomachs under iv. anesthesia with sodium pentobambiturate.Gastricjuicein15-mm aliquotswas collectedfor

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Effects of selective vagotomy on production of gastric cancer in dogs givenMNNGAll

dogs of the control and vagotomy group were given MNNG solution (50 @g/ml)p.o. for 12 months, with one exceptional case(Dog47)given the solution for 9 months because of heavy anorexia. In dogs of the vagotomy group, selective vagotomy was performed2months

before beginning of MNNGadministration.StomachcancerInva

MNNG Latent siveTotalBodywt intake peniodb Survival Preinvasive (Grade (Grades D Metastasis and

Dog (kg)― (g) (mos.)― (days) (GradeD) E) + E) vesselinvasionSarcomaControlgroup

57― 10 5.3 36& 0 1 1LYb60―10 6.1 38 1186― 1 0 1 ——62―8 4.0 — 1170 0 0 0 —SB(1)@'65@

11 6.5 — 802 2 1 3 —SB(3)457 5.2 30 1420 3 2 5 —SB(2)469 6.0 48 1457 1 1 2 — SB(1)

47h 6 1.6 — 2230― 0 0 0 ——499 4.2 28 1008 1 3 4 ——8.8

±O.6@ 4.9 ±0.6 1205 ±192 2.0 ±0.7Vagotomy

group58'@10 5.1 — 1094― 2 2 4 ——59k'9 6.3 24 967 7 3 10 LY,LN,H,L1YSB(1)63―8 6.8 36 1186 2 0 2 —SB(3)64―8 5.3 29 1850 3 5 8 LY(2) ST(1 ),@6(1)39

10 3.7 42 1922 1 0 1 — ST(2)41@l 8 3.6 54 l648e 4 3 7 LV SB(1)42h@ 5.4 22 1163 1 7 8 LY(2), LN, HST(1)48

7 3.0 — 1228― 2 0 2 ——8.6±0.4 4.9 ±0.5 1382 ±130 5.3 ±12

M. Fujita et a!.

30 mm before and 90 mm after s.c. injection of tetragastninat 2 p.g/kg body weight. Each aliquot was titrated to pH 7.0with 0.1 N NaOH. The peak acid concentration and MAO 1hr after injection were then calculated.

RESULTS

Gastric Acid Secretion. The peak acid concentration was76 ±17 mEq/Iiter (mean ±S.E.) in the vagotomized dogswhile it was 128 ±6 mEq/Iiter in the control dogs (p <0.025). The volume of gastric juice in the vagotomized dogswas reduced to about one-half of that in the control dogs(19 ±3 ml versus 42 ±6 ml, p < 0.01). MAO in thevagotomized dogs was 1.1 ±0.2 mEq/hm versus 4.3 ±0.7mEq/hr in the control dogs (p < 0.01), which represents areduction of gastric acid secretion by approximately 75% inthe vagotomized dogs, as compared to that in the controldogs.

MNNG Intake and TumorIncidence.Dogsthat survived12 months or more were judged as effective cases becausethe first case of gastric cancer was noted in a dog that diedaccidentally from anesthesia for endoscopic examination

12 months after the beginning of MNNG administration.Comparison of body weight, MNNG intake, latent period,survival, stomach cancer incidence, and metastasis in thecontrol and vagotomy groups is presented in Table 1. Theaverage body weight gain during the experimental period inthe 2 groups of dogs was similar. Cumulative MNNG intakewas not different. Of the 8 control dogs at risk, 6 developedgastric cancer, whereas all the dogs in the vagotomy grouphad cancer. Endoscopic observation also indicated thatstomach cancer appeared earlier (latent period) in thevagotomy group than in the controls.

Gastric Cancer. Multiple epithelial lesionswhich showedvarious degrees of atypia were found in almost all of thestomachs examined. In our previous report (4), these lesions were classified histologically into 5 grades rangingfrom A (minimal atypia) to E (severe atypia) according tothe degree of departure from the normal structure. Thesame criteria were used in the present study, especiallyGrade E and D lesions, which were considered to becarcinomatous. Grade E lesion refers to typical adenocarcinoma, which consists of neoplastic glands showingmarked cellular and structural atypism with clear evidence

Table 1

a At the beginning of MNNG administration.b Month in which tumor appearance was detected by endoscopy.‘. Beagle.

d Not detected.

a) Died accidentally from anesthesia for endoscopy.

I The abbreviations used are: LY, lymphatic vessel invasion; SB, small bowel;

lung metastasis.9 Numbers in parentheses, number of lesions.

h Female.

@ Mean±SE.J Significantly different from the control group at p < 0.025.

LN, lymph node metastasis; H, hepatic metastasis; LU,

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Macroscopicappearancesof dog gastriccancersClassificationsystemsfor earlygastric cancerestablishedbytheJapan

Gastroenterological Endoscopic Society (12) and for advancedcancer by Bomrmannwereused.Early

cancer AdvancedBonn

I Ia @b ll@a Illmann(poly-(ele- (de- (exca- (typesIIpoid)

vated) (flat) pressed) vated) andIll)Control(l6)b

0 7 5 3 10Vagotomy(42)

3 6 12 16 1 4

Distribution of dog gastric cancers according todiameterNo.

of lesionswith diametersof@23-5

6-10 11-2021-60@61―mmmmmm mmmmmmControl

(16)b34 6 300Vagotomy(42)1 41 1 8 5 22

Vagotomy and Dog Gastric Carcinogenesis by MNNG

of invasion regardless of the depth of invasion. The lesionwhich is composed of the same nature of neoplastic glandsas Grade E lesion, but which is confined to the mucosawithout definitive evidence of invasion, has been listed asGrade D.

Multiple gastric cancers (Grade D + E), ranging from 2 to10 lesions, were found in all of the dogs of the vagotomygroup except one. On the other hand, gastric cancer wasseen in 6 of 8 dogs of the control group. Four of the 6 dogshad multiple (2 to 5) cancers. The vagotomy group had 20Grade E invasive cancers, whereas the control group had 8.The total number of grade D cancers which were pmeinvasive reached 22 in the vagotomy group, but themewere only8 found in the control group. The average number of cancerlesions in the vagotomy group and the control group was5.3 ±1.2 and 2.0 ±0.7, respectively.

Gross Features. In both groups, the majority of thegastric cancers were found in areas where atrophy of thegastric mucosa was frequently seen (the anterior wall of theupper body of the stomach and the gastric angle along thelesser curvature). These gastric cancers in dogs showed avariety of sizes, gross appearances, and histological types.As shown in Table 2, the gastric cancers ranged in sizefrom microscopic foci under 2 mm to large lesions oven 60mm in diameter. In the control group, all of the 16 lesionsfound were within 20 mm in diameter, and only 3 lesionswere between 11 and 20 mm. On the contrary, 9 of 42lesions in the vagotomy group were over 11 mm, and 4 ofthem were more than 21 mm in diameter.

Macroscopic features of gastric cancers obtained in thisexperiment showed that only 4 of the cancers found wereapplicable to Borrmann classification for human advancedcancer, but the rest of the cancers had macroscopic featunes fairly similar to those of human early gastric cancers.Therefore, these cancers were classified by the systemestablished for the classification of early gastric cancer atthe Japan Gastroenterological Endoscopic Society (11).Table 3 shows the frequency of appearance of the differentmacroscopic classes of gastric cancer in the 2 groups.Although the superficial type II was commonly prevalent inboth groups, the relative incidence of the various subtypesshowed some differences. More type ll@elevated cancerswere observed in the control group, whereas incidence oftype ll@ depressed cancers was higher in the vagotomygroup. The Bommmanntype of advanced cancers was foundexclusively in 4 lesions of the vagotomy group.

Histological Types. The distribution of the cancers byhistological types in both groups, together with the degreeof penetration, is given in Table 4. The cancers were

Table3

a Combination types of lI@ + ll@ and ll@ + Ill are included in type

b Number in parentheses, total number of lesions.

classified by the predominant histological type. Every typeOf gastric cancer except mucoid carcinoma was found in

both groups. The majority of lesions in both groups werehighly or moderately differentiated adenocarcinomas with apapillary or tubular pattern. Signet ring cell cancers inwhich cells completely lost their polarity and possessed anabundant amount of mucus were found in one of thecontrols and 2 of the vagotomy group. In both groups,poorly or moderately differentiated adenocarcinomasshowed a more conspicuous tendency to penetrate thestomach wall than did well-differentiated adenocarcinomaor signet-ring cell cancer. One lesion of advanced gastriccancer with moderate differentiation in which the invasionof cancer cell nests reached the semosawas found in thecontrol group, whereas 4 lesions of this type were seen inthe vagotomy group, 2 of them moderately differentiatedadenocarcinomas, one well differentiated, and one poorlydifferentiated. Carcinomas with vessel invasions (Fig. 2)were revealed in 7 lesions, 1 in the control group and 6 inthe vagotomy group.

The cancer lesions were without exception surroundedby mucosa which showed marked atrophy or loss of chiefcell and parietal cell. Intestinal metaplasia was not observed. Vagotomy in dogs had not been shown to affectsignificantly the histological structure of normal gastricmucosa.

Metastases. Furthermore, distant metastasesof advanced gastric cancers were revealed in 2 of the vagotomized dogs. One of these cases was a male beagle dog(Dog 59) that was killed on Day 967 because of heavy weightloss and weakness. In the stomach, a Bomrmann type IIpoorly differentiated adenocarcinoma (2.5 x 2.5 cm) waspresent on the anterior wall of the upper body, in additionto multiple lesions of superficial type gastric cancers. Metastases of the advanced cancer to the liver and lungs werefound in addition to the pemigastric lymph nodes metastases. The details were described in our recent report (3). Itwas considered to be the first report of hematogenic metastases of chemically induced gastric cancer.

The other advanced cancer with metastases was seen ina female mongrel dog (Dog 42) that was killed whenmoribund on Day 1163. At autopsy, 3 Bomrmann type IIItumors with deep craters and a Borrmann type I tumor wereseen on the fundic gland area of the stomach (Fig. 1). Theme

Table 2

a Even the lesions with the largest diameters were measured inmillimeters.

b Number in parentheses, total number of lesions.

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M. Fujita et a!.

Table 4

Histological types and degree of penetration of dog gastric cancers

Classification system adopted by the Japanese Research Society for Gastric Cancer (7)has been used.

Hi hI diff n n- Moderately Poorly differ@g,@ ,@ee differentiated entiated ad- Signet ringa e a eno adenocarci- enocanci- cell cancercarcinoma noma noma

rn― sm s m sm s m sm s m sm s

Control (16)'@ 5 1 0 4 2 1 1 1 0 1 0 0Vagotomy (42) 18 5 1 7 3 2 1 2 1 2 0 0

‘IThe abbreviations used are: m, mucosa; sm, submucosa; s. serosa.

h Number in parentheses, total number of lesions.

also were a polypoid lesion and 3 tiny lesions with centraldepression resembling superficial Il(. depressed-type earlygastric cancer in humans. Grossly multiple metastatic fociwere seen in the liver and perigastric lymph nodes. Themost advanced carcinoma, which was composed of moderately differentiated tubular glands and situated on theposterior wall near the cardia, revealed invasion to thepancreas tissue and metastases to pemigastric lymph nodes.Microscopic findings confirmed that the liver metastasesconsisted of tubular structures which were similar to thoseoftheadvancedcancer(Figs.3 and 4).

Sarcoma. Sarcomas of the small intestine were inducedin 4 dogs in each group. In the vagotomy group, sarcomasof the stomach were also seen. Histologically, these lesionswere leiomyosamcomas composed of interlacing bundles ofspindle cells. The intestinal sarcomas frequently causeddeath from obstruction and bloody diarrhea.

DISCUSSION

Selective vagotomy performed in dogs before the p.o.administration of MNNG solution resulted in the productionof more advanced gastric cancers with distant metastases,as well as an increase in the number of gastric cancerscompared to that of the controls. The results in the controlshowed sufficient similarity to our previous study (4) inwhich the early stage of the gastric cancer was found in 2of 6 dogs given MNNG (60 @.tg/ml).Shimosato et a!. (14)also reported that 4 dogs given MNNG developed adenocamcinomas with no metastasis. In these experiments andanother report (9), sarcomas, in addition to gastric adenocarcinomas, were frequently found in the gastrointestinaltract. These sarcomas induced simultaneously by MNNGseemed to interrupt further development of the gastriccancer.

In the present study, selective vagotomy without drainageprocedure was performed mainly with 2 aims: (a) to reducethe gastric acid secretion with resulting retardation ofMNNG inactivation in the stomach to increase the gastriccancer production; and (b) to reduce the transportation ofactive MNNG into the duodenum to result possibly in anincrease of tumor incidence in the stomach and a decreaseof the intestinal tumor production. Eventually, gastric acidsecretion expressed by MAO after tetmagastnininjection wasreduced by approximately 75% in the vagotomized dogs. Itis known that the inactivation of MNNG occurs rapidly

under the strong acidity of the normal stomach (16). Thesignificant reduction of the gastric acid secretion inducedby selective vagotomy is conceivably the main cause of theenhancement of the gastric cancer production by MNNG.While this type of vagotomy was suggested to be associatedwith delay of gastric emptying, the procedure did notreduce the induction of the intestinal sarcoma in dogs.

There have been some reports which attempted to evaluate the relationship between vagotomy and experimentalgastric cancer in rats. Kowalewsky (8) indicated that vagotomy contributed to the development of gastric adenocarcinoma in rats fed 2,7-diacetylaminofluorene. Morgenstern(11) reported that gastrojejunostomy combined with vagotomy enhanced the methylcholanthrene-induced gastriccancer in rats. These investigators have speculated that theeffect of vagotomy on carcinogenesis may be correlatedchiefly with artificially induced hypochiorhydria.

In the present study, atrophic changes were observed inthe mucosa surrounding the gastric cancers in both thecontrol and the vagotomy group. It has been demonstratedthat MNNG acts not only as a strong carcinogen but also asan ulcerogenic agent to the gastric mucosa. Our recentstudy (18) revealed that the atrophic area where gastriccancers were frequently induced was closely related to theparticular region of the stomach where diffuse erosivechanges or shallow ulcers had been observed by endoscopyduring MNNG administration. The ulcerogenic action ofMNNG will influence the gastric carcinogenesis in 2 different fashions. It is possible that ulceration may provide morecells which will be susceptible to the carcinogen, resultinginan increaseof the number of transformedcellsintheregenerating epithelium. On the contrary, it may decreasethe cancer formation by sloughing off the mucosal cellspreviously transformed by MNNG. The strong acid in thegastric juice may also act as a promoting factor influencingthe ulcemogenic activity of MNNG. It was found in thepresent study that, in addition to the formation of moreadvanced gastric cancers, a significant increase in thenumber of minute cancer foci was seen in the vagotomizeddogs in which gastric acid secretion was considerablysuppressed. The present results suggest that the decreaseof gastric acidity in the vagotomized dogs has resulted incertain reduction of sloughing off of the mucosal cellstransformed during administration of MNNG. This factorseems to be another reason why vagotomy has enhancedgastric carcinogenesis in dogs given MNNG.

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Vagotomy and Dog Gastric Carcinogenesis by MNNG

Recently, long-term intake of carcinogen or procarcinogen present in food has been considered to be responsiblefor gastric cancer in man. It is also well-known that chronicatrophic gastnitis accompanying achlorhydnia is associatedwith gastric cancer (6), but the etiological correlation between these conditions remains unclear. On the other hand,it is recognized that gastric cancer is seldom found in theduodenal ulcer patients with hypemacidity. The results ofthe present study indicate that strong acidity in the stomachcould be one of the factors which reduces the incidence ofgastric cancer. Some clinical reports (1, 5) imply that acausal relationship exists between vagotomy for benigndiseases and gastric cancer. The present study suggeststhat patients undergoing vagotomy without gastric resection should be carefully followed up to evaluate their riskfor gastric cancer.

ACKNOWLEDGMENTS

The authors wish to thank Professor Emeritus Shigeru Shiba for hissupport and encouragement, Dr. Takeo Nagayo (Aichi Cancer Center, Aichi,Japan) and Dr. Haruo Taniguchi (the Center for Adult Diseases, Osaka,Japan) for the valuable suggestions, Professor Tadashige Murakami (TokyoMedical Dental University, Tokyo, Japan) for helpful discussion, and Dr.Norio Hirota and Dr. RIchard H. C. San (Naylor Dana Institute for DiseasePrevention, New York, N. Y.) for their help in preparing this manuscript.

REFERENCES

1. Capper, W. M. , and Johnson, H. D. Vagotomy and carcinoma of thestomach. Lancet, 2: 1063—1064,1964.

2. Fujita, M. , Taguchi, T., and Shiba, S. Production of gastric tumor indogs administered N-Methyl-N'-nitno-N-nitrosoguanidine (in Japanese).Proc. Jpn. Cancer Assoc., 29: 66, 1970.

3. Fujita, M., Taguchi, T., Takami, M., Usugane, M., and Takahashi, A.Lung metastasis of canine gastric adenocarcinoma induced by N-Methyl-N'-nitro-N-nitrosoguanidine. Gann, 66: 107-108, 1975.

4. Fujita, M., Taguchi, T., Takami, M., Usugane, M., Takahashi, A., andShiba, S. Carcinoma and related lesion in dog stomach induced by oraladministration of N-Methyl-N'-nitro-N-nitrosoguanidine. Gann, 65: 207-214, 1974.

5. Grahame, E. W. Vagotomy and carcinoma of the stomach. Lancet, 1:109, 1965.

6. Hitchcock, C. A., Mac Lean, L. D., and Sullivan, W. A. The secretory andclinical aspects of achlorhydria and gastric atrophy as precursors ofgastric cancer. J. NatI. Cancer list. , 18: 795—811, 1957.

7. Jinnai, D. , Kajitani, T. , and Akasaki, K. (eds.). The general rules for thegastric cancer study in surgery and pathology, Ed. 9, pp. 27—61. Tokyo:Kanehara Press, 1974.

8. Kowalewsky, K. Relationship between vagotomy, peptic ulcer and gastnic adenocarcinoma in rats fed 2,7-diacetylaminofluorene. Can. J.Sung.. 16: 210-217, 1973.

9. Koyama, V., Omoni, K. . Hirota, T. , Sano, A. , and Ishihara, K. Leiomyosarcomas of the small intestine induced in dogs by N-Methyl-N'-nitroN-nitrosoguanidine. Gann 67: 241-251 , 1976.

10. Kurihara, M., Shirakabe, H., Murakami, T., Yasui, A., Izumi, T., andIgarashi, A. A new method for producing adenocarcinomas in thestomach of dogs with N-Ethyl-N'-nitro-N-nitrosoguanidine. Gann, 65:163-177, 1974.

11. Morgenstern, L. Vagotomy, gastrcenterostomy and experimental gastriccancer. Arch. Surg., 96: 920-923, 1968.

12. Murakami, T. Pathomorphological diagnosis. Gann Monogr., 11: 53-55,1971.

13. Nundy, S., and Baron, J. H. An Evaluation of the Hollander test bygraded vagotomy in the dog. Gut, 14: 665-668, 1973.

14. Shimosato, Y., Tanaka, N., Kogure, K., Fujimura, S., Kawachi, T. . andSugimura, T. Histopathology of tumors of canine alimentary tract produced by N-Methyl-N'-nitro-N-nitrosoguanidine. J. NatI. Cancer Inst.,47:1053-1070,1971.

15. Sugimura, T., and Fujimura, S. Tumor production in glandular stomachof rat by N-Methyl-N'-nitno-N-nitrosoguanidine. Nature, 216: 943-944,1967.

16. Sugimura, T., Fujimura, S., Kogure, K., Baba, T., Saito, T., Nagao, M.,Hosoi, H. , Shimosato, Y. , and Yokoshima, T. Production of adenocarcinomas in glandular stomach of experimental animals by N-Methyl-N'-nitro-N-nitrosoguanidine. Gann Monogr. , 8: 157-196, 1969.

17. Sugimura, T., Tanaka, N., Kawachi, T., Kogure, K., Fujimura, S., andShimosato, Y. Production of stomach cancer in dogs by N-Methyl-N'-nitro-N-nitrosoguanidine. Gann, 62: 67-68, 1971.

18. Takami, M., Fujita, M., Takahashi, A., Usugane, M. , and Taguchi, T.Long-term observation of canine stomach cancer induced by N-MethylN'-nitro-N-nitrosoguanidine (in Japanese). Stomach Intestine, 12: 1375-1386,1977.

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Fig. 1. Mucosal surface of the stomach of Dog 42, which had multiple tumors. Huge Borrmann type III tumors with deep ulcer can be seen in the posteriorwall and the anterior wall near cardia, as well as a Borrmann type I tumor and several small lesions with slight elevation on depression. The whole stomachwas step sectioned into sections about 5 mm wide along the lesser curvature. Metastatic foci were found in penigastric lymph nodes and the liver.

Fig. 2. Section of poorly differentiated adenocarcinoma with vessel invasion (Dog 57). Mucosal lymphatics contain clusters of anaplastic cells. H & E, x115.

Fig. 3. Section of metastatic carcinoma in the liver of the dog in Fig. 1.Arrows, metastatic foci. H & E, x 7,Fig. 4. Higher magnification of metastatic carcinoma shown in Fig. 3 (moderately differentiated adenocarcinoma). H & E, X 100.

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816

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1979;39:811-816. Cancer Res   Masahide Fujita, Motohisa Takami, Masao Usugane, et al.  

-nitrosoguanidine following VagotomyN-nitro-′N-Methyl-NEnhancement of Gastric Carcinogenesis in Dogs Given

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