TUMOR JNDUCTION AND TUMOR GROWTH IN HYPOPHYSECTOMIZED MICE

Transcription

1 TUMOR JNDUCTION AND TUMOR GROWTH IN HYOHYSECTOMIZED MICE REMMERT KORTEWEG AND FRGDGRIC THOMAS (From he Laboraory of he Anoni van Leeuwenhoekhuis, Neherlands Insiue for ncer Research, Amserdam) Whereas numerous hough ofen conflicing papers have been wrien on he relaionship beween umor growh and he piuiary in general, only scany daa are available on he subjec of umor inducion and umor growh in hypophysecomized animals. Since, wih he excepion of wo papers, all he repors dealing wih his subjec have been published in his Journal, we will conen ourselves wih a brief summary of he conclusions which have been reached. Ball and his associaes have sudied he influence of hypophysecomy on he growh of 1 : 2 : 5 : 6dibenzanhracene sarcoma and grafed Walker carci noma in he ra. They were he firs (1, 2) o show ha he growh rae of hese umors is markedly slower in hypophysecomized animals, and hey furher esablished ha his reardaion is direcly dependen upon he absence of piuiary secreion (5, 7). McEuen and Thomson, in an excellen paper (3), likewise come o he conclusion ha hypophysecomy reards bu does no preven he growh of grafed Walker carcinoma, a conclusion incidenally confirmed by Franseen and McTiernan (6). McEuen and Thomson, however, are of opinion ha he reardaion is due no direcly o lack of piuiary hormones, bu o he ensuing cachexia. They observed he same reardaion of umor growh in grafed animals subjeced o sarvaion. Reiss, Druckrey and Hochwald (4), in a paper published simulaneously wih ha of he lasmenioned auhors, repor experimens carried ou on 250 hypophysecomized ras. The umor used was he Jensen sarcoma. The conclusions of his paper may be summarized as follows. (a) If hypophysecomy is performed in a umor carrier, he umor grows much more slowly from he ime of he operaion onwards. In some cases i sops growing alogeher or even regresses. (b) If he grafing is performed afer hypophysecomy, he umor aains a diameer of 1.0 o 1.5 cm. and hen in many cases regresses. (c) When growh hormone is given o he hypophysecomized umor bearer he umor coninues o grow, growh being proporional o he dose of hormone. (d) The regression of he graf is hasened if he hypophysecomized animal is given folliculin. (e) Exirpaion of he middle or poserior lobe alone furnished no conclusive resuls. (The auhors hink hemselves warraned in concluding from 1 Hypophysecomy designaes surgicd exirpaion of he hypophysis. Experimens performed on animals wih a view o suppressing piuiary funcion more or less compleely, eiher by irradiaion or adminisraion of folliculin, are open o criicism, and are no discussed in his paper. 36

2 TUMOR INDUCTION AND GROWTH IN HYOHYSECTOMIZED MICE 37 FIGS. 1 AND 2. SKULL OF A NORMAL MOUSE (LEFT) AND SKULL SHOWING REMOVAL OF BONE ANEL (RIGHT) The inse shows he hook used in cuing ou he bone. A separae phoograph of his wih scale is shown in Fig. 4. hese experimens ha he growh of umors depends on he exisence of an (( Inkremilieu, which may be one of he subsraa of he I Tumorkrankhei. ) As appears from he above review of he lieraure, he daa available on he subjec of he influence of hypophysecomy on umor growh are scany. All he experimens have been carried ou on he ra. Since here is no echnic which allows he operaor o bring he whole gland of his animal ino view before exirpaing i, here is always a chance of performing an incomplee operaion. Fragmens of he gland may be lef and be discovered only when hisologic examinaion of serial secions of he skull base is made. For his reason and ohers, ras are no ideal maerial for such experimens. One of us () has developed a echnic in he mouse which allows he removal of a large panel of bone a he base of he skull (Figs. 13). The enire gland is hus exirpaed under direc visual conrol. Since he publicaion of his echnic, we have furher simplified i in ha we use a single hook (Fig. 4) for he operaion insead of wo, as formerly. This mehod dispenses wih he necessiy of serial cuing of he skull base, making experimens on a large scale easily possible. The operaive moraliy is low and he period of survival is eigh o welve monhs, excepionally foureen. This echnic was used in he experimens described below, a preliminary repor of which was published in 1937 (9).

3 35 REMMERT KORTEWEG AND FR~D~RIC THOMAS The following experimens were carried ou on F, hybrids. The mohers of he animals belonged o he albino srain 020 Leeuwenhoekhuis, which has been produced by breeding broher o siser for weny generaions. The fahers belonged o he dilue brown MurrayLile srain. Animals of boh sexes were used. As no difference was observed in he resuls, no furher menion of sex will be made in his repor. Some of he animals had been hypophysecomized a he age of four o seven weeks, one o six monhs before he experimens were begun. Afer he operaion he body weigh decreased slighly, hereafer remaining consan FIG. 3. SERIES OF REMOVED BONE ANELS Scale in half millimeers for he res of he animal s life. These wo phenomena gave objecive evidence of he horoughness of he hypophysecomy, previously demonsraed by adequae exposure of he gland a he ime of operaion and finally conrolled in some insances by serial secions of he skull base. The mice were given a die of milk, barley, hemp and canary seed, as well as he sandard die of laying hens (conaining codliver oil, yeas and corn sprous). EXERIMENTS WITH TRANSLANTED TUMORS The umors used were T95379, T9554, and T95391, which are sponaneous mammary carcinomas occurring in females of he dilue brown MurrayLile srain. They had been ransplaned respecively 1, 6, and

4 TUMOR INDUCTION AND GROWTH IN HYOHYSECTOMIZED MICE imes in animals of he same srain, as well as in differen F, hybrids of which he faher was invariably a dilue brown. The grafing was done by injecing an emulsion of he umor in saline under he skin of he back on he righ side. As was o be expeced from heoreical deducions, he grafs succeeded in every case. Experimen A: Tumor T95379: (1) Nine animals, abou hree monhs of age, which had been hypophysecomized five weeks before, were grafed. (2) Eigh normal conrol animals of he same age were also grafed. These animals, of course, weighed more a he ime of grafing han he hypophysecomized animals. FIG. 4. HOOK USED FOR CUTTING OUT BONE ANEL Scale, above, in half millimeers (3) Eigh normal conrol animals of he same weigh as he hypophysecomized mice, and herefore much younger, were also grafed. Two weeks afer he grafing, when he firs animal died, all animals were sacrificed. The umors were disseced and weighed. Experimen B: Tumor T9554: Three series of animals, as in experimen A, were injeced wih umor emulsion. As umor T9554 grows more slowly han T95379, he animals were allowed o live wenywo days afer grafing. Exerimen C: Tumor T95379: (1) Eigh mice, abou seven monhs of age, which had been hypophysecomized six monhs before, were grafed. (2) Six normal conrol mice, hree o four monhs of age, were also grafed. These animals were sacrificed seveneen days afer grafing. Experimen D: Tumor T95391: (1) Five mice, abou eigh monhs of age, which had been hypophysecomized seven monhs before, were grafed. (2) Two normal conrol mice, abou four monhs of age, were grafed. These animals were sacrificed seveneen days afer he grafing. The resuls of hese four series of experimens are presened in Table I. The weigh in grams represens he average weigh for each group of animals. Discussion: In all 31 hypophysecomized mice he umors grew. In 13 of hese animals he hypophysis had been exirpaed a leas six monhs before he experimen, which eliminaes he possibiliy of a posoperaive acion of he hormones. There were no differences in umor growh and srucure eiher grossly or microscopically beween he hypophysecomized animals and he conrols. Equal numbers of mioic figures were observed in boh series of umors.

5 40 REMMERT KORTEWEG AND FRBDBRIC THOMAS TABLE I : Experimens wih Transplanable Tumors Experimen A H ypophysecomized Conrols same age Conrols same weigh Toal number 9 Body weigh before graf (gm.) Final body weigh (m Final umor weigh (m.) I ncreasc of body weigh during experimen 0% 0% 40 % ercenage of final umor weigh o original body ueigh 7% 6% 13% ercenage of final u mar weigh o final body weigh 7% 6% 9% Experimen B H ypophysecomized Conrols same age Conrols same weigh o % 13% 76% 7% 5% 17% 6% 4% 1070 Experimen C Hypophysecomized Conrols yc 3 9% 3 % Experimen D Hypoph ysecomized Conrols 5 2 _ 14? ~. 4% 4% 1.4% The percenage figures in he las wo columns of Table I show ha he umors were relaively larger in he conrols of he same weigh han in he hypophysecomized mice or in he conrols of he same age for experimens A, B and C. This does no hold rue for experimen D, bu since only 2 conrols were used in his experimen his figure may no be represenaive for he group. Excep in experimen B, he body weigh of he operaed animals did no increase during he experimen. The increase of weigh noed in group B is so sligh ha, apar from possible errors in weighing, i can be aribued o he growh of he umor. In adul animals he weigh of he umor was approximaely proporional o he body weigh. In oher words, he acual weigh of he umor had he same relaionship o he body weigh in he hypophysecomized animals as in he conrol animals of he same age. As was o be expeced, he rae of umor growh was much greaer in he series of younger conrols. Conclusions: ( 1) These experimens, wih hree ypes of ransplanable umor, show ha he umors grow perfecly well in animals hypophysecomized one o seven monhs previously. (2) Comparison of he figures showing he final weigh of he umors in he hypophysecomized and in he conrol mice shows reardaion of growh in he operaed series. When hese figures are considered in relaion o he respecive body weighs of he grafed mice, however, here is no evidence of a difference in rae of growh beween he hypophysecomized and he conrol mice of he same age.

6 TUMOR INDUCTION AND GROWTH IN HYOHYSECTOMIZED MICE 41 (3) The fac ha he grafed umors grew perfecly well in hypophysec omized mice leads o he conclusion ha he influence of he piuiary is cerainly no qualiaive. If any influence exiss, i can be only quaniaive in naure. EXERIMENTS WITH INDUCED TUMORS In anoher series of experimens 34 benzpyrene in 0.5 benzol was used as he carcinogenic agen. The animals were F, hybrids, described above. Experimen E: A he beginning of his experimen he animals were hree monhs old. Eigh of hem had been hypophysecomized four weeks previously. Eigh ohers were used as conrols; heir weigh was herefore much greaer han ha of he hypophysecomized animals. The skin of he neck was ouched hree imes a week by a glass rod dipped in he benzpyrene soluion. The hair was no removed before applicaion of he benzpyrene. Afer a few weeks, however, he hair disappeared from he area of applicaion in boh series of animals. In boh he conrols and hypophysecomized animals here was laer a regrowh of hair, more rapid in he former han in he laer. eriods of loss of hair alernaed hereafer wih periods of regrowh. A papilloma was observed 259 days afer he beginning of he experimen in one of he hypophysecomized mice. A carcinoma, wihou a preliminary papilloma, was observed in anoher mouse of his series a 300 days. The hypophysecomized animals died beween nine and en monhs afer he beginning of he experimen, ha is en o eleven monhs afer hypophysecomy. Each of he conrol animals developed a papilloma wihin 54 o 126 days from he onse of he experimen, and a carcinoma wihin 141 o 255 days. Experimen F: The following mice were used in his experimen. (1) Fifeen mice of abou seven monhs of age ha had been hypophysecomized a he age of six weeks. (2) Seven mice of abou five monhs of age, which had also been hypophysecomized a six weeks. The animals of boh hese groups were small bu acive. They showed he ypical habius of adul mice hypophysecomized when young. (3) Twelve conrol animals which a he beginning of he experimen had he same weigh as he animals of Groups 1 and 2. These animals were hen abou six weeks old. (4) Twelve conrol mice, beween hree and six monhs of age. These were, of course, much heavier han he mice of Groups 1 and 2. Since he animals of Groups 3 and 4 gave he same experimenal resuls, no differeniaion beween hem will be made. In none of he animals in experimen F was he hair removed. Two drops of he carcinogenic agen (3:4benzpyrene in 0.5 per cen benzol) were dropped from a dropper upon he skin of he neck, wice weekly. Since his soluion evaporaes quickly, a large porion of he benzpyrene remained in he hair and did no come in conac wih he skin. Afer baldness had occurred only one drop of he carcinogenic agen was given. The resuls of experimen F are given in Chars I and 11. The hypophysecomized mice are represened in Char I and he conrol mice in Char 11. The animals are arranged from above downward according o he order of appearance of he papillomas. In one animal (number 19) a carcinoma developed wihou a preliminary papillomaous sage. This animal s posiion in he able is deermined by he dae of appearance of he carcinoma. In he conrol series all of he animals developed papillomas beween 44 and 100 days, and carcinomas of he skin beween 93 and 14 days. Eigh of he hypophysecomized animals died beween 90 and 1 days wihou showing any umor. The firs four of hese belonged o Group 1. Their deah occurred before any of he oher hypophysecomized animals showed a papilloma. These animals had survived hypophysecomy eigh and a half monhs. Foureen of he hypophysecomized mice developed a umor. Thireen of hem de

7 42 REMMERT KORTEWEG AND FRI?D$RIC THOMAS D%!l I 1 I I I I 1 I I , I CHART I. EXERIMENT F: HYOHYSECTOMIZED MICE. apilloma.. rcinoma. Died wihou Tumor veloped a papilloma beween 10 and 14 days; 10 of hem a carcinoma beween 121 and 190 days. In one of hese 10 umors hisologic differeniaion beween papilloma and carcinoma was difficul. The 9 ohers showed he ypical hisologic picure of skin carcinoma. Animals 24, 23 and 30, which developed papillomas beween 17 and 192 days afer he beginning of he reamen, died nine and a half monhs afer hypophysecomy wihou carcinoma. The following four animals did no develop a umor. No. 22, dying 177 days afer he beginning of he experimen and nine and a half monhs afer hypophysecomy; No. 23, dying 1 days afer he beginning of he experimen and eleven and a half monhs afer hypophysecomy; No., dying 143 days afer he beginning of he experimen and en and a half monhs afer hypophysecomy; No. 27, dying 153 days afer he beginning of he experimen and eigh and a half monhs afer hypophysecomy. Discussion: The skin umors induced by 3 :4benzpyrene developed in general much earlier in normal han in hypophysecomized mice. A number of he hypophysecomized mice did no develop a umor. Bu since many of hose ha did develop a umor showed i only a few days before deah, i is p ci b I +

8 TUMOR INDUCTION AND GROWTH IN HYOHYSECTOMIZED MICE \ \ likely ha, had hese nonumor animals lived longer, a umor would have appeared. The average survival period of he hypophysecomized mice reaed wih benzpyrene was en o welve monhs. This ime was evidenly oo shor o allow all animals o develop a umor. The difference in resul beween experimens E and F, i.e. he low percenage of umors in he hypophysecomized mice in E and he high percenage in F, nowihsanding he fac ha in E he benzpyrene had been applied hree imes and in F wice a week, can be explained by he difference in he mehod of applicaion of he carcinogenic agen. In Group F. he carcinogenic agen was dropped on he neck. In Group E i was applied wih a glass rod, and herefore in his group here was a possibiliy ha he soluion did no always come in conac wih he skin when he hair was abundan. This is subsaniaed by he observaion ha he umors of he conrol series of ex

9 44 REMMERT KORTEWEG AND FR~D~RIC THOMAS perimen F developed more rapidly han hose of experimen E. In F he papillomas developed in an average of 75 days and he carcinomas in 127 days, while in E he papillomas developed in 104 days and he carcinomas in 207 days. Conclusion: (1) These experimens prove ha skin umors can be induced in hypophysecomized mice. (2 Wih regard o he response o carcinogenic agens, he difference beween hypophysecomized mice and conrols is no of a qualiaive naure. I is enirely quaniaive. SUMMARY In his experimenal sudy he influence of he hypophysis on umor inducion and umor growh was sudied in mice. When he geneic consiuion of he mice was compaible, ransplanaion was successful in every case. The umor grafs grew more slowly in hypophysecomized animals han in conrols of he same age, bu he relaion of he final umor weigh o he body weigh of operaed and conrol animals of equal age was he same. apillomas and carcinomas induced by 3 : 4benzpyrene appeared markedly laer in he hypophysecomized mice han in he conrols. Wih regard o he growh of umor ransplans and he response o carcinogenic agens, no qualiaive difference was found beween hypophysecomized mice and conrols. The observed differences were enirely of a I) quaniaive naure. LITERATURE 1. BALL, H. A., SAMUELS, L. T., AND SIMSON, W.: Am. J. ncer 16: , SAIKUELS, L. T., BALL, H. A., AND SIMSON, W.: Am. J. ncer 1: 3032, MCEUEN, C. S., AND THOMSON, D. L.: Bri. J. Exper. ah. 14: 34391, REISS, M., DRUCKREY, H., AND HOCHWALD, A,: Klin. Wchnschr. 12: , BALL, H. A., AND SAMUELS, L. T.: Am. J. ncer 26: , FRANSEEN, C. C., AND MCTIERNAN, C.: Am. J. ncer 26: , BALL, H. A., AND SAMUELS, L. T.: Am. J. ncer 32: 5056, THOMAS, FR~D.: Endocrinology 23: 99102, KORTEWEG, R., AND THOMAS, FR~D.: Rappor de la 5"" Conference de la Leeuwenhoekvereeniging enue & aris en Sep. 1937, pp